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"Case-control study of shock index among women who did and did not receive blood transfusions due to postpartum hemorrhage.",
"\nTo compare shock index (SI) values between women who required blood transfusion due to postpartum hemorrhage (PPH) and women who did not. ",
"In a case-control study, clinical data were assessed from the medical records of women requiring blood transfusion for PPH at a center in Brazil between 2012 and 2015 (n=105). ",
"A control group was randomly selected from women who did not receive blood transfusion (n=129). ",
"Compared with women who did not receive a transfusion after delivery, women who did receive one had significantly higher SI values 10 minutes after delivery (0.81 ± 0.27 vs 0.72 ± 0.16; P=0.012), at 30 minutes (0.83 ± 0.26 vs 0.71 ± 0.15; P<0.001), and at 2 hours (0.84 ± 0.27 vs 0.70 ± 0.14; P=0.032). ",
"For vaginal deliveries, SI values were significantly different at 30 minutes (0.88 ± 0.26 vs 0.71 ± 0.14; P<0.001) and 2 hours (0.90 ± 0.23 vs 0.72 ± 0.14; P=0.001). ",
"No significant differences were found for cesarean delivery. ",
"The SI might be useful to identify early vital sign changes due to PPH. ",
"Increased SI values were associated with need for transfusion in vaginal deliveries."
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"Even at their most avant-garde, Charleston's best restaurants are inseparable from the town's historic charms.",
"\n\nCharleston, South Carolina is practically a living history lesson, evidenced by rows of pastel-colored mansions and Charleston City Market – one of the oldest of its kind in the country. ",
"The city's Lowcountry cuisine also draws significant influences from French, English and African origin.",
"\n\nBut of course for us the spotlight has turned to the food scene, two early standouts are Husk and McCrady's who helped put the city on the national culinary map. ",
"At both Husk and McCrady's, executive chef Sean Brock uses heirloom Southern crops he grows on his nearby farm, including the James Island Red Corn that goes into Husk’s beloved shrimp and grits. ",
"At McCrady’s, Brock nods to Southern cultural roots with dishes like beef tartare with hay-roasted beet, farro, and lichen, bringing back historic heirloom grains that hadn't been seen in decades. ",
"This kind of old-meets-new is simply the norm here—see for yourself on our tour of the town's culinary hot spots, below.",
"\n\nCharleston is literally surrounded by water—the town is perched scenically on a peninsula where two rivers meet the Atlantic. ",
"It’s this location that furthers the region’s nickname, The Lowcountry. ",
"For food hounds, all this water brings one thing to mind: seafood! ",
"The hottest new oyster-shucking mecca is The Ordinary, opened last year by Mike Lata of farm-to-table favorite FIG. ",
"At the restaurant’s digs inside a former bank, plonk down beneath the soaring ceiling at the long wooden bar and you can opt for an icy seafood tower piled with locally-sourced raw bar favorites, or a shrimp burger on squid-ink brioche. ",
"Recently, Lata launched a changing daily special dubbed \"the Ordinaries,\" a prix fixe special that includes salad and dessert. ",
"Tuesdays, it’s a lobster roll and Thursdays, Caribbean fish stew.",
"\n\nThere's hardly a notable restaurant in town that isn't housed in some kind of historic something-or-another, but lately a mini-trend seems to be eateries inside former gas stations. ",
"One exciting disused petrol outpost is Home Team BBQ’s original West Ashley location. ",
"The honky tonk-style joint has earned a rightful reputation for barbecue, including their famous smoked chicken wings and ribs. ",
"Equally known are its sauces—the restaurant has six of them, in flavors like Carolina Mustard and Alabama White.",
"\n\nNewcomer Rutledge Cab Company occupies another former gas station and has the roll-up doors to prove it. ",
"Their Southern-eclectic menu is loaded up on all-day breakfast (grits!) ",
"and a filling station charcuterie platter, featuring fried bologna with pepper jelly-mustard.",
"\n\nOne new place, the Grocery, started life—as you might guess—as a market. ",
"Executive chef-owner Kevin Johnson works with local farmers and fisherman for his brunch and dinner menus. ",
"For further provisions, you'll find local favorite Caviar & Bananas inside the Charleston City Market. ",
"The café and shop was founded by Dean & Deluca alums. ",
"Grab some edible souvenirs or pick up a sandwich or sweet treat to snack on.",
"\n\nIf you want to hang with the staff, the Butcher & Bee, is a sandwich and snack joint hidden in a former industrial warehouse, beneath a highway overpass. ",
"Gritty setting aside, the place has many things going for it, including its BYOB status. ",
"Typical to the place, you'll probably find your waiter from lunch sipping his own cans of beer after hours (the place stays open till 3am) with a lamb pita or banh mi sandwich. ",
"Be sure to check out their daily-changing chalkboard menu on Facebook or Twitter.",
"\n\nAnd if all this running around means you haven't yet tried Charleston's signature dish, shrimp and grits, pop over to Hominy Grill, in a beautiful, historic former home downtown, where it’s made with sautéed mushrooms and bacon. ",
"James Beard Award-winning chef-owner Robert Stehling also serves many other Lowcountry and Southern classics, like she crab soup, okra-and-shrimp beignets, and fried chicken. ",
"The food has all the appeal of Grandma's cooking—with a fresh spin. ",
"Much like the fresh yet historic feeling of Charleston itself."
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"Despite an increasing number of reports and the media attention to the issue, little supportive forensic evidence remains, potentially as a result of late presentation of victims, reluctance to seek police involvement and the short half‐lives of the drug agents implicated. ",
"2 - 5 For those presenting to ED in Australasia, there is generally no routine collection of clinical or laboratory data, thus the true extent of drink spiking is unknown. ",
"Currently, there appears to be no clear consensus regarding the management of these cases.",
"\n\nThe public perception of drink spiking is that sedative drugs are placed (usually by men) into the drinks of others (usually women). ",
"In recent years, there appears to have been an increase in the number of cases being reported within the media, although the anecdotal reports outnumber formal complaints to the police by an estimated factor of 10. ",
"1 A recent national report estimated that as many as 3000–4000 suspected incidents of drink spiking occurred in Australia between 1 July 2002 and 30 June 2003; 1 however, the authors acknowledge that these estimates should be taken as a rough guide only.",
"\n\n‘Drink spiking’ refers to drugs or ethanol being added to a drink (alcoholic or non‐alcoholic) without the consent of the person consuming it. ",
"1 It is purportedly done for purposes, such as sexual assault, rape, assault and robbery.",
"\n\nPatients were contacted by the Clinical Toxicology Service once laboratory results became available. ",
"The patients' condition and views about the results were recorded and they were invited to obtain police involvement if applicable.",
"\n\nWe defined a patient to be a ‘plausible drink spiking case’ if the following criteria were met: (i) patient believed that they had had their drink spiked; (ii) patient denied ingesting any agent detected by laboratory screening; (iii) patient signs and symptoms during the drink spiking incident were consistent with agents detected by laboratory screening.",
"\n\nThe number of standard drinks consumed was calculated from the reported numbers and types of drinks consumed (e.g. full‐strength beer, wine, spirits etc.), ",
"assuming that one standard drink contains 10 g of ethanol. ",
"6 If a person was unable to accurately describe the volume of an alcohol drink consumed, then it was assumed that the volume was one standard drink.",
"\n\nA preformatted data collection tool was used in all cases. ",
"Information collected included patient demographic data and details about the incident (place, company kept, when and what symptoms developed). ",
"A detailed history of drugs and alcohol ingested (time of first drink, number of drinks) before the incident was sought. ",
"A member of the SCGH ED‐based clinical toxicology service confirmed the history by direct patient interview either during their emergency presentation or within three working days by telephone. ",
"This information was correlated with findings following a full physical examination and laboratory results.",
"\n\nPatients aged 16 years and over who either self‐presented or were brought to the ED by police, paramedics, friends or relatives believing that they or the patient had had their drink spiked were recruited. ",
"For inclusion, all cases had to be within 12 h of the alleged drink spiking event or still exhibit signs or symptoms of intoxication at the time of presentation. ",
"Clinical staff also enrolled cases with altered conscious state or behaviour where they had reasonable suspicion that drink spiking might be implicated. ",
"Retrospective consent was obtained from these patients; however, if they chose not to continue to participate in the study, biological samples collected earlier were destroyed.",
"\n\nAs a key stakeholder, the WA Police Service conducted a concurrent drink spiking awareness and prevention campaign. ",
"Members of the public were encouraged to attend either the SCGH or JHC ED if they or their friends believed that they had been a victim of drink spiking. ",
"The police provided assurance to the public that detection of voluntary illicit drug taking as part of the drug screen would remain confidential to the study project and not involve prosecution.",
"\n\nED staff were briefed about the study. ",
"They were alerted to the time‐critical nature of specimen collection and procedures for maintaining a forensic ‘chain‐of‐evidence’. ",
"Staff were also reminded that particular patients might require additional assistance from sexual assault services and police.",
"\n\nThis was a prospective observational study of patients presenting with suspected ‘drink spiking’. ",
"The study was carried out in the ED of Sir Charles Gairdner Hospital (SCGH) and Joondalup Health Campus (JHC) in Perth, Western Australia (WA). ",
"Both departments serve a metropolitan area and have a combined annual census of approximately 75 000 adult patients. ",
"Other metropolitan hospitals were invited to redirect patients if they met the inclusion criteria. ",
"The Human Research Ethics Committees at SCGH and JHC approved the study protocol.",
"\n\nThe majority (87%) of patients were discharged from the ED. ",
"Other than the case admitted to ICU ( Box 1 ), all other patients were admitted to the ED observation ward overnight. ",
"Eleven per cent were referred to the Sexual Assault Referral Centre. ",
"The WA Police Service was involved in one in four of our patients; however, only 8% reported that they were still pursuing police proceedings at follow‐up. ",
"At the time of manuscript preparation, no police prosecutions had taken place. ",
"No significant sequelae were reported by any patient at follow‐up, and nearly all reported that they were completely symptom‐free by 24 h; however, 35% still believed that they had been the victim of a drink spiking incident irrespective of the results.",
"\n\nIllicit drugs were detected in 28% (27/97) of patients, with 85% of these having two or more drugs (including ethanol) detected. ",
"Amphetamines and cannabis (tetrahydrocannabinol) were the most common substances found. ",
"In 10 patients, amphetamines were detected in the urine, even though the patients denied ingesting amphetamines. ",
"None had symptoms consistent with acute amphetamine intoxication, and the median blood ethanol level at time of presentation was 98 mg/dL. With the exception of the plausible drink spiking case (case three in Table 1 ), all opiates and benzodiazepines detected in urine had been prescribed to the patients.",
"\n\nOnly 7% (7/97) of patients denied having consumed alcohol, with a further three patients not being able to recall if alcohol had been consumed; two of these returned a negative blood ethanol reading. ",
"Of the 87 who reported drinking alcohol, 76% had consumed more than four standard drinks, with the mean number of standard drinks consumed being 7.7 ± 3.9 SD (range 1–21). ",
"On presentation, 13 of the 87 patients recorded zero blood ethanol concentrations. ",
"Of the 74 with measurable blood ethanol levels on presentation, the estimated median blood ethanol concentration at the time of presentation (BAC) was 0.096% (96 mg/dL).",
"\n\nThe patient had reported to the police that her mobile phone and money had been stolen; however she did not wish to lay charges against the man involved, as she was fearful of the repercussions.",
"\n\nHer friend told her that the male client contacted her ∼17.00 hours to say that the patient had collapsed, and was vomiting and fitting on the floor. ",
"The friend collected the patient and took her to hospital.",
"\n\nThe patient was a 23‐year‐old sex worker who had left home at ∼14.00 hours to travel to a private residence of a man who was a known drug dealer to provide services. ",
"She had two vodka and orange drinks and finished her work at ∼16.15 hours. ",
"The man offered to pay her to stay for another 3 h, and she contacted a friend to say she would be staying. ",
"He served her another drink that tasted odd. ",
"Approximately 15 min later she felt weak, unable to stand and collapsed. ",
"She was dragged into another room where there were two other men, and had no further memory of events until she awoke in the ICU at SCGH.",
"\n\nA 23‐year‐old woman was transferred from another metropolitan hospital to the ICU at SCGH ventilated for coma and seizures of uncertain aetiology. ",
"She was enrolled in the study 17.5 h after the onset of symptoms once the history of drinking in a bar with unknown men became available. ",
"GHB was detected in her urine. ",
"When later contacted to feedback laboratory results and confirm the reported history, the patient changed her account of the event.",
"\n\nWe identified nine cases we believed were plausible drink spiking incidents. ",
"Of these, four patients denied ingesting the drugs that were detected in urine ( Table 1 ). ",
"Amphetamines were detected in three patients. ",
"One was a 20‐year‐old man who thought that his friends placed a tablet in his drink as a prank, and another was a 19‐year‐old woman who developed signs and symptoms after leaving a nightclub. ",
"She was subsequently lost to follow‐up. ",
"The third case was a 23‐year‐old sex worker who had voluntarily ingested ecstacy (methylenedioxymethylamphetamine) but had high levels of GHB detected 17.5 h after the onset of symptoms ( Box 1 ). ",
"The fourth case was a 30‐year‐old woman with a past history of migraines, anxiety and pseudoseizures, who developed symptoms (∼1000 h) 30 min after drinking from an unattended drink resting on her infant's pram in a shopping centre. ",
"Benzodiazepines and opiates (consistent with codeine ingestion) were detected.",
"\n\nOne hundred and one patients were enrolled over a 19 month period. ",
"Four patients were subsequently excluded from analysis, three because of late presentation to the ED (>12 h from onset of symptoms and not showing signs of intoxication) and one patient admitted to taking an intentional oral hyosine overdose. ",
"All had blood and urine samples collected. ",
"Of the remaining 97 cases, 88% were female (median age 23 years, interquartile range [IQR] 19–28) with 59% less than 25 years of age. ",
"Only 25% of patients presented between Monday and Thursday. ",
"Of those presenting, the majority (72%) reported onset of symptoms of drink spiking in pubs or nightclubs. ",
"Most patients self‐presented (53%) or arrived by ambulance (39%), and just 8% were brought to the ED by police. ",
"Presentation to the ED was between 22.00–04.00 hours for 69% of the study group. ",
"The median time from onset of symptoms to presentation was 2.5 h (IQR 0.3–20.2 h) and sample collection 4.5 h (IQR 2.5–7.8 h).",
"\n\nDiscussion\n\nMuch of the research on drink spiking to date has concentrated on laboratory analysis of urine and/or blood samples for the detection of drugs in patients who allege either drink spiking or sexual assault.7-10 Usually, no correlation is taken with the patients' symptoms or their drug taking history. ",
"To our knowledge, this is the first prospective Australian study where clinical, historical (including drug and alcohol ingestion) and laboratory data have been collated on patients who believe that they have had their drinks spiked.",
"\n\nOur findings do not support the public perception that sedative drugs are being used to spike people's drinks. ",
"We did not identify a single case where a sedative drug had been placed in a drink in a pub or nightclub setting. ",
"No cases of ketamine or flunitrazepam ingestion were detected. ",
"The only case where GHB was used as a drink spiking agent was in a situation not commonly associated with drink spiking (refer to Box 1). ",
"A detailed history enabled this case to be identified. ",
"In the other three plausible cases involving a drug agent, a detailed history did leave some doubt in the authors' minds as to whether these patients had their drinks spiked as alleged. ",
"For example, in one plausible case involving benzodiazepine and opiates that took place at a shopping centre, the patient's medical record revealed that she had access to these drugs. ",
"She had a known history of migraines, which might have been the cause of her symptoms.",
"\n\nWe identified five plausible drink spiking cases whose high blood ethanol levels (median 174 mg/dL) were inconsistent with their drink history. ",
"This supports previous findings that ethanol might be an agent being used in drink spiking.11-13 In a Welsh 1 year study of patients allegedly having their drink spiked presenting to an ED, 34/75 patients enrolled had blood ethanol concentrations measured, with 65% of those patients having a blood ethanol concentration >160 mg/dL.12 Relying on these patients' recall of alcohol consumption, with high blood ethanol levels, makes it difficult to determine whether their drinks were truly spiked. ",
"For example, one case in our study was a woman who awoke naked in a motel room. ",
"She had recollection of only consuming one standard drink at a bar during a job interview. ",
"Collateral history from the bar owner confirmed that this woman had consumed at least 10 ‘shooters’ within a short period of time, before leaving the bar. ",
"Although initially appearing to be a plausible drink spiking case, collateral history excluded her. ",
"From our study it appears that drink spiking does occur but is rare, and if it occurs ethanol is the likely agent used.",
"\n\nOf greater concern was the high level of self‐reported alcohol ingestion and blood ethanol concentrations, as previously reported in two UK studies.12, 13 Although many patients were surprised at their blood ethanol levels, most were unconcerned at their high levels of alcohol consumption.",
"\n\nThe present study also found a significant use of illicit drugs, mainly amphetamines and cannabis. ",
"In those where an illicit drug was detected, 85% had an additional agent (including ethanol) detected. ",
"This has been the finding in other studies.8-10, 13 Scott‐Ham and Burton found in their series of 1014 cases of drug‐facilitated sexual assault that cannabis was detected in 26% of cases, cocaine 11% and amphetamines 7%.10 The finding of 10 patients whose symptoms were inconsistent with amphetamine intoxication (despite amphetamine detection in their urine) demonstrates the difficulty in interpreting laboratory studies, and performing studies, such as ours, that are reliant on patients giving accurate histories, especially when it involves the use of illicit drugs.",
"\n\nThe present study demonstrates the difficulty in investigating alleged drink spiking incidents. ",
"Emergency departments do not routinely collect blood and urine specimens for drug levels as they usually have no impact on patient management. ",
"Furthermore, they do not routinely collect forensic specimens, as was required in the present study. ",
"As a result of the sophisticated laboratory testing required, there was a significant cost involved. ",
"As drink spiking is a crime, it would be better dealt with by the police service (as happens for drunk driving); however, only one in four of our patients had involvement of the WA Police Service.",
"\n\nWe would recommend that any patient presenting alleging that they have had their drinks spiked should have their symptoms treated on their merits. ",
"Any biological specimens that need to be collected for forensic purposes should be collected and processed by the police services.",
"\n\nThe present study has several limitations. ",
"A concurrent awareness campaign by the police and universities might have led to an increase in reporting, especially in younger people. ",
"Our inclusion criteria might have limited cases enrolled, or staff might have enrolled patients who were purely intoxicated and not a victim of drink spiking. ",
"We did not record patients who declined to participate in the study. ",
"We relied on patients for a history of alcohol and drug consumption and the time of onset of symptoms, which might have been unreliable with the level of alcohol and drugs being detected. ",
"The absolute blood ethanol concentration is difficult to interpret because of inter‐individual variability in tolerance to ethanol. ",
"The low prevalence of sexual assault means that the present study might be too small to be used to comment on the use of illicit substances in drug‐facilitated sexual assault. ",
"Further, we made assumptions that any drink consumed was a ‘standard’ drink."
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"Harden steps on David Lee's foot while eurostepping past him in transition, turned it pretty bad. ",
"He can barely walk and went straight to the locker room. ",
"Horrible news for Houston and their playoff chances if its anything even close to bad.",
"\n\nAnd it comes right before he made his first allstar game, as his city hosted the game no less.",
"\n\nLeBron slowly separating himself from Durant every day in the MVP race (the last 6 games in particular). ",
"Being able to watch him do his thing on a nightly basis is so, so cool. ",
"He's probably already a top 10 all-time great and is somehow still getting better. ",
"I wasn't really old enough to appreciate Jordan's greatness when he was in Chicago, so being able to watch LeBron is pretty awesome.",
"\n\nNot sure how I feel about these Brandon Jennings to mavs rumors. ",
"Volume shooters ftw?",
"\n\nI want to try and keep our backcourt locked up and just make a push for Howard in the off-season. ",
"Like you said that backcourt isn't anything special, but I think it works really well with Carlisle and his offense if we have a super star on the front court somewhere.",
"\n\n__________________\n\nQuote:\n\nOriginally Posted by Scott Wright\n\nI guarantee that if someone picks Cam Newton in the Top 5 they will regret it.",
"\n\nYeah, by shooting more. ",
"Jennings has talent, and I would like to believe what other Mavs' fans believe that Carlisle can wave his magic wand and turn him into efficient PG. ",
"But I don't see that happening.",
"\n\nI want to try and keep our backcourt locked up and just make a push for Howard in the off-season. ",
"Like you said that backcourt isn't anything special, but I think it works really well with Carlisle and his offense if we have a super star on the front court somewhere.",
"\n\nI'd be okay with having the same starting back court next season. ",
"Chemistry has been a problem for us this season, so having some sort constant going into next year will be a plus. ",
"Just have to hope a year under Carlisle and his system will lead to a improved year next year for both Collison and Mayo."
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"Action demanded after 1,100 dead dolphins wash up in France French marine researchers say a record 1,100 dead dolphins, their fins mutilated, have washed up on France's Atlantic coast since January\n\nPARIS -- The dolphins' bodies were horribly mutilated, the fins cut off.",
"\n\nBut what shocked French marine researchers wasn't just the brutality of the deaths of these highly intelligent mammals, but the numbers involved — a record 1,100 have landed on France's Atlantic coast beaches since January.",
"\n\nThe mass deaths, widely blamed on industrial fishing, have alarmed animal welfare groups and prompted France's ecology minister to launch a national plan to protect them.",
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"he asked.",
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"\n\nActivists say it's common for fishermen to cut body parts off the suffocated dolphins after they are pulled up on the nets, to save the nets.",
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"\n\nRugy has come up with some plans, including bolstering research into existing acoustic repellent devices in place in 26 two-vessel trawlers off the Bay of Biscay, an industrial fishing hub in the Atlantic Ocean. ",
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"\n\nIt claims many of the trawlers they watch in the region don't activate the repellent devices, fearing they will scare off valuable fish as well, and only turn them on if they are being checked fishing monitors.",
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"\n\n\"The government needs to take responsibility and act — especially Macron, who said he wanted to protect ecology,\" Lamya Essemlali, President of Sea Shepherd France, told The Associated Press.",
"\n\nShe cited scientists who predict that the current rates of fishing will likely drive the dolphin population to extinction.",
"\n\n\"The spotlight has been put on the trawlers that fish for sea bass ..., which is a scandal. ",
"But they were not the only ones responsible,\" she said.",
"\n\nShe suggested that aggressive hake fishing, which was given the green light three years ago after a long ban, was a major factor. ",
"The spike in dolphin deaths also began three years ago.",
"\n\nHer group says the ecological crisis stems from unprecedented demand for low-cost fish.",
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"Q:\n\nSet image width dynamically in MonoTouch\n\nI am trying to set the width of an image dynamically using MonoTouch. ",
"I have added the image to the view using Xcode Interface Builder.",
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"I have tried to set the images' width dynamically like this:\nimg.",
"Bounds.",
"Width = 150;\n\nand like this\nimg.",
"Bounds.",
"Size.",
"Width = 150;\n\nI have also tried to create a new RectangleF and setting the bounds equal to that Rectangle like this\nRectangleF fillrect = new RectangleF(50, 50,\n150, img.",
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"Bounds = fillrect;\n\nThe above method sizes the image but then moves the image to the incorrect place (X and Y values).",
"\nI have also tried various methods using SizeF but to no avail.",
"\nHow can I just set the width of the image and keep it in the same place I placed it on interface builder?",
"\nAnd also how to change the X and Y values so that I can move another image based on the growth of the aforementioned image.",
"\n\nA:\n\nSeems like changing of Bounds applies around the Center of the view.",
"\nTry to set new Frame instead:\n\nimg.",
"Frame = fillrect;\n\nLocation will be the same, but Size will be changed.",
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"Body composition and hormonal responses to a carbohydrate-restricted diet.",
"\nThe few studies that have examined body composition after a carbohydrate-restricted diet have reported enhanced fat loss and preservation of lean body mass in obese individuals. ",
"The role of hormones in mediating this response is unclear. ",
"We examined the effects of a 6-week carbohydrate-restricted diet on total and regional body composition and the relationships with fasting hormone concentrations. ",
"Twelve healthy normal-weight men switched from their habitual diet (48% carbohydrate) to a carbohydrate-restricted diet (8% carbohydrate) for 6 weeks and 8 men served as controls, consuming their normal diet. ",
"Subjects were encouraged to consume adequate dietary energy to maintain body mass during the intervention. ",
"Total and regional body composition and fasting blood samples were assessed at weeks 0, 3, and 6 of the experimental period. ",
"Fat mass was significantly (P <or=.05) decreased (-3.4 kg) and lean body mass significantly increased (+1.1 kg) at week 6. ",
"There was a significant decrease in serum insulin (-34%), and an increase in total thyroxine (T(4)) (+11%) and the free T(4) index (+13%). ",
"Approximately 70% of the variability in fat loss on the carbohydrate-restricted diet was accounted for by the decrease in serum insulin concentrations. ",
"There were no significant changes in glucagon, total or free testosterone, sex hormone binding globulin (SHBG), insulin-like growth factor-I (IGF-I), cortisol, or triiodothyronine (T(3)) uptake, nor were there significant changes in body composition or hormones in the control group. ",
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"\n\nDamian Green also set up a clash with police chiefs begging for an end to damaging funding cuts when he insisted: “There are no police cuts.”",
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"\n\nThe talks have hit a wall - 11 days after Ms May claimed a deal was done – which deprives the Tories of a guaranteed majority for any of the Bills to be unveiled today.",
"\n\nDUP sources have taken aim at the lack of “negotiating experience” on the Conservative side, a devastating criticism, just days after the separate Brexit talks also got underway.",
"\n\nAsked if an agreement was close, Mr Green replied: “There is still the possibility – there’s every possibility – of a DUP deal.”",
"\n\nThe First Secretary of State, the Prime Minister’s effective deputy, insisted the talks went “much wider” than just the Northern Ireland party’s demands for extra spending and tax breaks.",
"\n\nBoth sides saw the value of a “confidence and supply” arrangement, with the DUP backing the Government on key votes while preventing it being brought down by motions of no confidence.",
"\n\nBut, Mr Green added: “All talks of this kind take a long time - and they are still continuing.”",
"\n\nToday’s legislative programme is expected to be stripped of key Tory election manifesto pledges, because of the need for a minority government to avoid controversial votes.",
"\n\nMoves to open new grammar schools, end universal free school meals for infants, end the “triple lock” on pension payments and means-test winter fuel payments are all likely to be abandoned.",
"\n\nBut, speaking to BBC Radio 4’s Today programme, Mr Green insisted” “This is not a thin Queen’s Speech,” – pointing to measures on the economy, infrastructure and tackling injustices.",
"\n\nHe acknowledged it would be dominated by Brexit, with up to eight Bills expected to prepare for the complications of departure day, in March 2019.",
"\n\n“The biggest element of what’s in are Brexit bills, because we have got two years to do this deal,” Mr Green said.",
"\n\n“We need to be in a position, after we’ve done the negotiations, to make sure that our agriculture is secure, our immigration system is secure and so on.”",
"\n\nHe denied changes to social care had been “abandoned”, with a consultation over the summer, adding: “It will include a cap on costs.”",
"\n\nAnd Mr Green dismissed warnings, including from Metropolitan Police Assistant Commissioner Mark Rowley - Britain’s leading anti-terror police chief – that funds diverted to fight terrorism could leave the public at risk.",
"\n\n“There are no police cuts – we have protected police budgets in this parliament,” he insisted."
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"Sirio-class torpedo boat\n\nThe Sirio class (also known as the Saffo class) was a class of six sea-going steam-powered torpedo boats of the Italian Regia Marina (Royal Navy) built by the German shipyard Schichau-Werke from 1904–1906. ",
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"A third ship, Saffo was lost after running aground off Turkey on 2 April 1920. ",
"The surviving ships were disposed of in 1923.",
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"Mike Novogratz, CEO of crypto investment firm Galaxy Digital Capital Management, has again corrected his Bitcoin (BTC) price forecast. ",
"The billionaire investor said he doesn’t expect BTC to break the $9,000 barrier in 2018, CNBC confirmed Wednesday, October 3.",
"\n\nNovogratz, a former Wall Street manager, took part in Economist Finance Disrupted conference in New York on Oct. 2. \"",
"I don't think it breaks $9,000 this year,\" he has said from the stage.",
"\n\nNovogratz also linked the currently declining prices to the actions of companies in the industry who are selling crypto \"just to fund the burn rate of the industry.\" ",
"He stressed that these companies are not earning “anywhere near” the revenues from last year's \"crypto boom,\" which means that they are barely managing to cover salaries and operational costs.",
"\n\nBack in Nov. 2017, when cryptocurrency prices skyrocketed, Novogratz predicted that Bitcoin could climb as high as $45,000 within 12 months.",
"\n\nIn summer 2018, Novogratz slightly changed his mind, stating that mass adoption of cryptocurrencies and blockchain technology was still five to six years away. ",
"Despite that, he still expressed confidence that many institutions will come into the industry “in the next two to three years.”",
"\n\nIn mid-September he opined that crypto prices have hit the bottom. ",
"Later that month he reiterated this point of view, saying that crypto markets are experiencing “seller fatigue.” ",
"However, he noted that Bitcoin’s price was still quite stable, potentially attracting investor attention.",
"\n\nBitcoin has dropped almost 54 percent this year alone, while other currencies such as Ethereum and Ripple have lost even more. ",
"At press time BTC is trading at $6,524."
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"The minimally conscious state: definition and diagnostic criteria.",
"\nTo establish consensus recommendations among health care specialties for defining and establishing diagnostic criteria for the minimally conscious state (MCS). ",
"There is a subgroup of patients with severe alteration in consciousness who do not meet diagnostic criteria for coma or the vegetative state (VS). ",
"These patients demonstrate inconsistent but discernible evidence of consciousness. ",
"It is important to distinguish patients in MCS from those in coma and VS because preliminary findings suggest that there are meaningful differences in outcome. ",
"An evidence-based literature review of disorders of consciousness was completed to define MCS, develop diagnostic criteria for entry into MCS, and identify markers for emergence to higher levels of cognitive function. ",
"There were insufficient data to establish evidence-based guidelines for diagnosis, prognosis, and management of MCS. ",
"Therefore, a consensus-based case definition with behaviorally referenced diagnostic criteria was formulated to facilitate future empirical investigation. ",
"MCS is characterized by inconsistent but clearly discernible behavioral evidence of consciousness and can be distinguished from coma and VS by documenting the presence of specific behavioral features not found in either of these conditions. ",
"Patients may evolve to MCS from coma or VS after acute brain injury. ",
"MCS may also result from degenerative or congenital nervous system disorders. ",
"This condition is often transient but may also exist as a permanent outcome. ",
"Defining MCS should promote further research on its epidemiology, neuropathology, natural history, and management."
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"Email Marketing 101: 5 Ways to Avoid a Spam Filter\n\nNobody likes rejection. ",
"But unfortunately, your carefully crafted emails are rejected by spam filters and firewalls, sometimes without your knowledge. ",
"How can you ensure that your emails don’t “talk to the hand” and instead are greeted at your recipients’ inboxes with open arms? ",
"Here are five ways to avoid a spam filter:\n\nUse a solid subject line\n\nYour subject line is like a pick-up line; will that first line encourage your subscribers to listen to what you have to say? ",
"Try writing the subject line after you’ve written the email – and don’t speed through this process. ",
"Good subject lines are usually 5-8 words (35-40 characters) and should not contain any special characters or capital letters (those are big spam red flags!)",
"\n\nAvoid trigger words\n\nAvoid using spam trigger words in your subject line and email body. ",
"Take these words out of your vocabulary if you want to avoid a spam filter:\n\nFree\n\nClick here\n\nRead more\n\nGuarantee\n\nCall/act now\n\nSpam\n\nNo obligation\n\nManage your lists\n\nDon’t just hit ‘delete’ on those subscriber reports that your email marketing software sends you after sending an email. ",
"Pay attention to how many people opened, clicked, forwarded, and unsubscribed from your email. ",
"It’s important to remove emails from your list that bounced or could not be delivered. ",
"However, depending on the type of bounce, you may not always want to remove an email as it could possibly be delivered in the future.",
"\n\nKeep it short\n\nWe are bombarded with heaps of information daily. ",
"Not only do shorter emails increase readability, but they also reduce the chance of getting caught by the filter. ",
"Depending on the nature of the email, we recommend keeping the length around 200-300 characters to avoid a spam filter flag.",
"\n\nTest, test, and test again.",
"\n\nSend a draft preview of your email before sending out to your subscribers. ",
"Be sure to test the email on multiple email platforms (desktop, mobile, tablet) as well as various email providers (Gmail, Yahoo, Outlook).",
"\n\nThe No. ",
"1 Email Marketing No-No\n\nOne of the worst things you can do is send bulk emails from your personal email account, such as Gmail or Yahoo. ",
"If your email provider suspects that you are sending mass emails, your account could be frozen! ",
"Use email marketing software such as Constant Contact or MailChimp to send emails to your clients, customers, and prospects.",
"\n\nMake an Impact with Your Emails\n\nNeed help with email marketing? ",
"Give us a call – we’ll design a custom email template for your business and write the emails for you, so you don’t have to keep track of all these rules on how to avoid a spam filter. ",
"Or shoot us an email here (don’t worry; we will welcome your email with open arms!) ",
"Sarah Lane - Marketing Specialist"
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"High titers of IgG antibodies against Plasmodium falciparum serine repeat antigen 5 (SERA5) are associated with protection against severe malaria in Ugandan children.",
"\nPlasmodium falciparum serine repeat antigen (SERA5) is a promising asexual blood stage malaria candidate vaccine. ",
"However, there is a paucity of information about natural immune responses to SERA5 in children from malaria-endemic regions. ",
"We undertook a hospital-based case-control study of severe malaria in Apac District, Northern Uganda, in children 6-59 months of age. ",
"The commonest symptoms observed in children with severe malaria (SM) were respiratory distress (53.4%) and prostration (40.4%) followed by circulatory collapse (7.4%), severe anemia (Hb < 5 g/dL, 7.0%), and seizures (2.6%). ",
"None of the SM children had impaired consciousness, coma, or cerebral malaria. ",
"We measured serum IgG antibodies using a recombinant construct of SERA5 (SE36) in enzyme-linked immunosorbent assays. ",
"High titers of IgG anti-SE36 were associated with protection against severe malaria in children under 5 years old."
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"To link to the entire object, paste this link in email, IM or documentTo embed the entire object, paste this HTML in websiteTo link to this page, paste this link in email, IM or documentTo embed this page, paste this HTML in website\n\nPAGE THREE:\nUniversity of Southern California\nPAGE FOUR:\nrour Preps Will Delight T\"V TT ^ I IT /\\ \"^^T\" ^°)an ^w'mrners Trojan Audience Soon I m / \\ I I . \\ ",
"wEll I 1%. % #",
"^l /\"% 1^1 Test Trio of Northern Powers\nVol. ",
"XVI LOS ANGELES, CALIFORNIA, THURSDAY, FEBRUARY 11,1965 No. ",
"61\nROW RUNNERS—G reek Week preliminary relays at- final competition. ",
"One weary runner takes a short nap tracted both participants and onlookers. ",
"Coeds watch before his next race. ",
"Phi Psis captured top honors yes-as their fraternity neighbors vie for a place in today's terday. ",
"KAs, Kappa Sigs and Sig Eps gained the finals.",
"\nCREEK RELAY FINALS\nFour Houses to Battle\nPhi Kappa Psi, Kappa Sigma, Kappa Alpha and Sigma Phi Epsilon fraternities will compete in the finals of the Greek Relays today at 2 p.m. on the Row.",
"\nThe four entrants in the fraternity relay finals were determined yesterday in the preliminary heats. ",
"Selection was on a time basis, with Phi Kappa Psi posting the top mark of 1:03.9.",
"\nKappa Sigma was second with 1:04.2, Kappa Alpha was a tenth of a second behind with 1:04.3 and Sigma Phi Epsilon was fourth with 1:05.0.",
"\nOther Greek Activities\nThe Greek Relays are a part of Greek Week activities, which will also feature a Grecian Ball tomorrow night from 8 to 12 midnight in the ballroom of the International Hotel.",
"\nActivity on the Row will begin this afternoon at 2 p.m. with the sorority gunnysack race on the lawn of the Alpha Delta Pi house.",
"\nThe fraternity relay finals will follow on 28th Street between University Avenue and Figueroa Street.",
"\nThe sorority suitcase relay will be run last.",
"\nWomen in the suitcase relay will be required to wear men’s clothing over their own. ",
"They must take off the men’s clothes and pack their suitcases before passing them on to their teammates.",
"\nDelta Gamma is the defending champion of the suitcase relay. ",
"Sigma Chi was the winner of last year’s fraternity relay rnce. ",
"The gunnysack race is new this year.",
"\nDefending champion Sigma Chi ran the course today in 1:05.0, but was disqualified for passing the baton too far out on the first pass in front of the KA house.",
"\nNumerous protests were lodged by the brothers of Sigma Chi, who felt that an injustice had been done to their fraternity.",
"\n“Look, this race is a big thing with us,” a Sigma Chi pleaded.",
"\n“After all,” added another, “we also dropped the baton. ",
"If we hadn’t dropped the baton, we would have the best time here. ",
"Why, that would have knocked two whole seconds off our time.”",
"\n. ",
"Despite their pleading Sigma Chi remains disqualified.",
"\nImpromptu Race\nAbout 100 row residents witnessed the preliminary relay races which were interrupted by a momentary, impromptu motorcycle race.",
"\nThe prelims were run from University Avenue to the Kappa Alpha house; the second leg was made from the KA house to Woody’s; the third leg from Woody’s to the KA house and finally from the KA house back to University Avenue.",
"\nA street dance climaxed yesterday’s Greek Week activities. ",
"The Parleys played at the event held on the Row.",
"\nTrophies for the relays will be given at the Grecian Ball Friday night.",
"\nOi\ns\n*\nDIFFICULTIES OVERCOME\nOn Nerve Relief\n$20,000 Language Lab Aids Foreign Students.",
"\nBetinis Resigns Position As Convention Chairman\nIR Panel\nQuestions\nClergymen\nBy CHUCK CONYERS\nThree representatives of campus religion last night tried to convince members of Sigma Gamma Sigma, international relations sorority, that U. S. missionaries do not forcibly impose the Ju-deo-Christian ethic upon their charges.",
"\nPanel members for the discussion in the YWCA included Charles Doak, campus Presbyterian pastor; Milton R. Hughes, director of the Southern Baptist-sponsored Baptist Student Union; and Ben L. Cohen, director of the B’nai B'rith Hillel Foundation on campus.",
"\nMission Defended\nTne principal discussion began after Rev. Mr. Hughes stated his belief that Christian mission enterprises have won friends for the United States. ",
"He added that several young nations have gained responsibility and sovereignty through mission upbringing and education.",
"\nWhen the importance of contemporary missions in foreign societies was challenged, Rev. Mr. Hughes admitted the place of such outposts has changed because of post World War II anticolonialism feelings abroad.",
"\n“Now missionaries are there as advisers while nationals do the work. ",
"The job of a missionary is now to work himself out of a job,” Rev. Mr. Hughes said.",
"\nHe was then asked if missionaries force Western tradition upon their flock.",
"\nGive Peace “They try not to force Western tradition upon people but to give them the peace of God. ",
"Missionaries teach religion but also try to teach English and reading. ",
"The missionary does not think his way is the only way as far as the cultural customs go,” he replied.",
"\nOne sorority member argued that missionaries try to bring the Judeo - Christian ethic of Western civilization to the countries rather than accepting their ethics.",
"\n“I would say that most missionaries try to share their way of thinking. ",
"They give the people the chance to accept or reject their beliefs,” he said.",
"\nMARCEL BOLOMET\n... French lecturer\nNew Study System Can Save Time\nStudents who feel pressured by a constant lack of time to accomplish all their goals might find their tension unnecessary if they adopt the study system of Marcel Bolomet, former USC scholastic advisor and French lecturer.",
"\nGamma Phis Test\nBolomet is currently trying out his system with Gamma Phi Beta sorority.",
"\nThe purpose behind his method is to help students organize their time and discover their potentials. ",
"He wants them to clearly see their goals for the future, to decide what type of person they want to be and from this step work toward successfully accomplishing their well-marked ideals.",
"\nConstruct Chart\nThe mechanics of his system begin with constructing a chart containing 24 slots on one side and the days of the week on the adjoining side. ",
"At the beginning of the week the student plots the use of the entire 168 hours in the week. ",
"He indicates thei time devoted to individual studies, social activities, conferring with friends and all other actions for 24 hours. ",
"The insertions should be written in pencil so the time allotted for one event can be interchanged with another on, that same day.",
"\nAs a result the student is able to account for each hour of his time and see how much free time he has.",
"\nAccording to Bolomet, the student becomes the master of his time rather than its slave when he has some knowledge of how his time b spent.",
"\nSudden Move Stuns Members\nBy ELLIOT ZWIEBACH\nASSC President John Betinis resigned his chairmanship of the Constitutional Convention last night.",
"\nHis unexpected resignation followed a statement in which he said “it would be hypocritical of me to chair this convention and im-'—— ‘\npractical for me to take part tion was very much like Her-in it.” ",
"He referred to the con- bert’s original amendment flict between his “abolish stu- and could Just as easily have dent government” campaign, been adopted over Betinis’ and the nature of the conven-proposal without necessitat-tion designed to reorganize ing an executive order,” Gab-student government. ",
"rielson explained.",
"\nBetinis established the con-^ Senate President Bob Grif-vention by an executive orderi fin. ",
"a member of the Consti-:shortly before the semester, tutional Convention, outlined break to resolve the differ-ithe future course of the con-iences over his own constitu- stitutional convention as tional amendment and that of threefold:\nAMS President Adam Her- First, to ask Dr. Topping to bert. ",
"forward the recommendations\nFollowing Betinis’ exit, the of his Ad Hoc Committee on stunned 14 - member conven- Student Government to the tion elected Senator Brooke convention.",
"\nGabrielson, Senate rules com-1 Second, to attempt to de-mittee chairman, to preside fine the functions and areas | over subsequent convention of need of student govern-j meetings. ",
"ment:\nGabrielson described Beti- Finally, to attempt a joint nis’ move of withdrawing meeting between the conven-his original proposal, calling tion delegates and the mem-for a constitutional conven- bers of the ad hoc committee, tion, and then resigning as During his statement to the chairman as a “matter of convention, Betinis enumerat-prestige. ",
"e(j forces on campus which he\nThe “compromise conven- (Continued on Page 2)\nAssembly to Hear Justice Goldberg\nSupreme Court Associate Justice Arthur Goldberg will address a religious convocation at 2 p.m. Tuesday in Bovard Auditorium, University Chaplain John Cantelon announced yesterday.",
"\nJustice Goldberg, representing the Jewish faith, will be the second of three religious speakers this year. ",
"He will follow Father Adrian Von Kaam, a leading Catholic educator from Pittsburg's Duquesne University, who spoke last semester.",
"\nJustice Goldberg will also talk informally with students about the Peace Corps at 11 a.m. in the YWCA and will speak to the Faculty Club at noon in Town and Gowti.",
"\nHis visit is sponsored by the chaplain's office in conjunction with the Arthur S. Wolpe B'nai B’rith Hillel Memorial Fund, the B'nai\nB'rith Hillel Foundation and the Cultural Events Committee.",
"\nJustice Goldberg received a BS in law and a PhD in jurisprudence from Northwestern University.",
"\nHe was editor-in-chief of the Illinois Law Review and began private law practice in 1929.",
"\nAppointed general consul for the Congress of Industrial Organizations in 1948, Justice Goldberg served as general consul for the industrial union department of the AFL-CIO from 1955 to 1961.",
"\nIn 1961 President John F. Kennedy appointed him Secretary of Labor. ",
"He held this post until his appointment as associate justice of the Supreme Court on Aag- 29. ",
"1962.",
"\nJustice Goldberg has published numerous articles and is author of the book “AFL-CIO: Labor United.”",
"\nDr. Eleanor Metheny, physical education professor, delighted faculty members yesterday with a light but instructive speech on how college professors may quietly relieve their tensions through appropriate physical exercises.",
"\nSpeaking on the topic “Enjoy Your Tensions,” Dr. Metheny, author and world traveler, addressed an audience of approximately 150 professors and guests at the noon faculty luncheon.",
"\nDr. Metheny's demonstra-i tion of the “silent scream” technique of relieving anxieties brought more than silent laughter from her audience.",
"\nEmphasizing that the “scream” should be saved for emergencies, Dr. Metheny suggested that for smaller, more everyday frustrations, one might find the “drop your jaw” approach more convenient.",
"\nThis method, explained Dr. Metheny, involves “looking like an idiot.” ",
"She asked the\naudience to try it, producing loud laughter in the room.",
"\nFor professors suffering from insomnia. ",
"Dr. Metheny offered an original yawn producing exercise labeled the “sleep maker.”",
"\nShe also advocated that if each of her methods is followed simultaneously, one does not have enough time to think about his problems.",
"\n“I'm the only professor who puts her class to sleep on purpose,” Dr. Metheny said.",
"\nAfter Dr. Metheny's talk,' Dr. Edwin C. Robbins, president of the Faculty Center Association, announced the appointment of Dr. Tillman J.! ",
"Hall, physical education pro-: fessor, to the post of program chairman for the spring semester.",
"\nArthur J. Goldberg, associate justice of the Supreme Court, will be guest speaker at next week's meeting. ",
"Members of the faculty planning to attend are urged to make reservations early.",
"\nLINQUISTIC COMMUNICATION—Kent\nS. Cathcart, instructor in English com-1 munication, instructs a student on the\nForeign students enrolled reel-type wrhich is capable of in the English Communica- recording for approximately tion Program for Foreign Stu- one hour.",
"\ndents (ECPFS) witnessed the Movable desk tops enable opening of a new $20,000 the laboratory to be used as language laboratory yester- a regular classroom. ",
"The desk (jay tops protect the machinery\nThe laboratory, under the when not in use and form a direction of Kent S. Cath- writing surface for class-cart, instructor in English work.",
"\ncommunication, has a capaci- Each booth is also equip-ty of 24 students and is de-:ped with a counter which1 signed to assist students in tabulates the amount of tape| overcoming linguistic prob- recorded, providing easy re-lems as they learn the Eng- call of previous dialogue, lish language. “",
"This new laboratory will\nThe master-conlrol system allow the student to receive provides a two-way communi- more instruction hours in the cation between the teacher oral-aural communications. ",
"It and students, either indivi- is hoped that this will enable dually or collectively. ",
"the department to serve these\nHigh-fidelity tape and foreign students more effi-movable desk tops are two ciently,” Cathcart said, unique features of the new The language laboratory is lab. ",
"The use of high-fidelity the latest development in the tape in recording student dia-history of the English com-logues gives an accurate re- munication Program for For-production of the voice. ",
"eign Students. ",
"ECPFSC was\nThe tape is a seven-inch, started in 1958 by a faculty\ncommission in an attempt to coach foreign students in the English language so that they ma^)succeed at the university without a linguist'c handicap.",
"\nThe ECPFSC. ",
"aimed at r>-ishing student English ahili!;-rather than tearhing Engiish. ",
"covers all of the language skills. ",
"However, there is a bv-product of the program in-tercultural exchange.",
"\nThe program consists of classes in language orientation, conversation and discussion taught by a faculty of eight. ",
"Dr. Robert Kaplan, coordinator of ECPFSC, stated that the “principle purpose for the existence of this program is to serve student needs.”",
"\nApproximately 1,500 foreign students have completed courses under the auspices of the ECPFSC. ",
"There are 150 students currently enrolled in courses, Kaplan said.",
"\nuses of new equipment in the language lab. ",
"The lab is designed to aid foreign students who are learning English.",
"\n\nPAGE THREE:\nUniversity of Southern California\nPAGE FOUR:\nrour Preps Will Delight T\"V TT ^ I IT /\\ \"^^T\" ^°)an ^w'mrners Trojan Audience Soon I m / \\ I I . \\ ",
"wEll I 1%. % #",
"^l /\"% 1^1 Test Trio of Northern Powers\nVol. ",
"XVI LOS ANGELES, CALIFORNIA, THURSDAY, FEBRUARY 11,1965 No. ",
"61\nROW RUNNERS—G reek Week preliminary relays at- final competition. ",
"One weary runner takes a short nap tracted both participants and onlookers. ",
"Coeds watch before his next race. ",
"Phi Psis captured top honors yes-as their fraternity neighbors vie for a place in today's terday. ",
"KAs, Kappa Sigs and Sig Eps gained the finals.",
"\nCREEK RELAY FINALS\nFour Houses to Battle\nPhi Kappa Psi, Kappa Sigma, Kappa Alpha and Sigma Phi Epsilon fraternities will compete in the finals of the Greek Relays today at 2 p.m. on the Row.",
"\nThe four entrants in the fraternity relay finals were determined yesterday in the preliminary heats. ",
"Selection was on a time basis, with Phi Kappa Psi posting the top mark of 1:03.9.",
"\nKappa Sigma was second with 1:04.2, Kappa Alpha was a tenth of a second behind with 1:04.3 and Sigma Phi Epsilon was fourth with 1:05.0.",
"\nOther Greek Activities\nThe Greek Relays are a part of Greek Week activities, which will also feature a Grecian Ball tomorrow night from 8 to 12 midnight in the ballroom of the International Hotel.",
"\nActivity on the Row will begin this afternoon at 2 p.m. with the sorority gunnysack race on the lawn of the Alpha Delta Pi house.",
"\nThe fraternity relay finals will follow on 28th Street between University Avenue and Figueroa Street.",
"\nThe sorority suitcase relay will be run last.",
"\nWomen in the suitcase relay will be required to wear men’s clothing over their own. ",
"They must take off the men’s clothes and pack their suitcases before passing them on to their teammates.",
"\nDelta Gamma is the defending champion of the suitcase relay. ",
"Sigma Chi was the winner of last year’s fraternity relay rnce. ",
"The gunnysack race is new this year.",
"\nDefending champion Sigma Chi ran the course today in 1:05.0, but was disqualified for passing the baton too far out on the first pass in front of the KA house.",
"\nNumerous protests were lodged by the brothers of Sigma Chi, who felt that an injustice had been done to their fraternity.",
"\n“Look, this race is a big thing with us,” a Sigma Chi pleaded.",
"\n“After all,” added another, “we also dropped the baton. ",
"If we hadn’t dropped the baton, we would have the best time here. ",
"Why, that would have knocked two whole seconds off our time.”",
"\n. ",
"Despite their pleading Sigma Chi remains disqualified.",
"\nImpromptu Race\nAbout 100 row residents witnessed the preliminary relay races which were interrupted by a momentary, impromptu motorcycle race.",
"\nThe prelims were run from University Avenue to the Kappa Alpha house; the second leg was made from the KA house to Woody’s; the third leg from Woody’s to the KA house and finally from the KA house back to University Avenue.",
"\nA street dance climaxed yesterday’s Greek Week activities. ",
"The Parleys played at the event held on the Row.",
"\nTrophies for the relays will be given at the Grecian Ball Friday night.",
"\nOi\ns\n*\nDIFFICULTIES OVERCOME\nOn Nerve Relief\n$20,000 Language Lab Aids Foreign Students.",
"\nBetinis Resigns Position As Convention Chairman\nIR Panel\nQuestions\nClergymen\nBy CHUCK CONYERS\nThree representatives of campus religion last night tried to convince members of Sigma Gamma Sigma, international relations sorority, that U. S. missionaries do not forcibly impose the Ju-deo-Christian ethic upon their charges.",
"\nPanel members for the discussion in the YWCA included Charles Doak, campus Presbyterian pastor; Milton R. Hughes, director of the Southern Baptist-sponsored Baptist Student Union; and Ben L. Cohen, director of the B’nai B'rith Hillel Foundation on campus.",
"\nMission Defended\nTne principal discussion began after Rev. Mr. Hughes stated his belief that Christian mission enterprises have won friends for the United States. ",
"He added that several young nations have gained responsibility and sovereignty through mission upbringing and education.",
"\nWhen the importance of contemporary missions in foreign societies was challenged, Rev. Mr. Hughes admitted the place of such outposts has changed because of post World War II anticolonialism feelings abroad.",
"\n“Now missionaries are there as advisers while nationals do the work. ",
"The job of a missionary is now to work himself out of a job,” Rev. Mr. Hughes said.",
"\nHe was then asked if missionaries force Western tradition upon their flock.",
"\nGive Peace “They try not to force Western tradition upon people but to give them the peace of God. ",
"Missionaries teach religion but also try to teach English and reading. ",
"The missionary does not think his way is the only way as far as the cultural customs go,” he replied.",
"\nOne sorority member argued that missionaries try to bring the Judeo - Christian ethic of Western civilization to the countries rather than accepting their ethics.",
"\n“I would say that most missionaries try to share their way of thinking. ",
"They give the people the chance to accept or reject their beliefs,” he said.",
"\nMARCEL BOLOMET\n... French lecturer\nNew Study System Can Save Time\nStudents who feel pressured by a constant lack of time to accomplish all their goals might find their tension unnecessary if they adopt the study system of Marcel Bolomet, former USC scholastic advisor and French lecturer.",
"\nGamma Phis Test\nBolomet is currently trying out his system with Gamma Phi Beta sorority.",
"\nThe purpose behind his method is to help students organize their time and discover their potentials. ",
"He wants them to clearly see their goals for the future, to decide what type of person they want to be and from this step work toward successfully accomplishing their well-marked ideals.",
"\nConstruct Chart\nThe mechanics of his system begin with constructing a chart containing 24 slots on one side and the days of the week on the adjoining side. ",
"At the beginning of the week the student plots the use of the entire 168 hours in the week. ",
"He indicates thei time devoted to individual studies, social activities, conferring with friends and all other actions for 24 hours. ",
"The insertions should be written in pencil so the time allotted for one event can be interchanged with another on, that same day.",
"\nAs a result the student is able to account for each hour of his time and see how much free time he has.",
"\nAccording to Bolomet, the student becomes the master of his time rather than its slave when he has some knowledge of how his time b spent.",
"\nSudden Move Stuns Members\nBy ELLIOT ZWIEBACH\nASSC President John Betinis resigned his chairmanship of the Constitutional Convention last night.",
"\nHis unexpected resignation followed a statement in which he said “it would be hypocritical of me to chair this convention and im-'—— ‘\npractical for me to take part tion was very much like Her-in it.” ",
"He referred to the con- bert’s original amendment flict between his “abolish stu- and could Just as easily have dent government” campaign, been adopted over Betinis’ and the nature of the conven-proposal without necessitat-tion designed to reorganize ing an executive order,” Gab-student government. ",
"rielson explained.",
"\nBetinis established the con-^ Senate President Bob Grif-vention by an executive orderi fin. ",
"a member of the Consti-:shortly before the semester, tutional Convention, outlined break to resolve the differ-ithe future course of the con-iences over his own constitu- stitutional convention as tional amendment and that of threefold:\nAMS President Adam Her- First, to ask Dr. Topping to bert. ",
"forward the recommendations\nFollowing Betinis’ exit, the of his Ad Hoc Committee on stunned 14 - member conven- Student Government to the tion elected Senator Brooke convention.",
"\nGabrielson, Senate rules com-1 Second, to attempt to de-mittee chairman, to preside fine the functions and areas | over subsequent convention of need of student govern-j meetings. ",
"ment:\nGabrielson described Beti- Finally, to attempt a joint nis’ move of withdrawing meeting between the conven-his original proposal, calling tion delegates and the mem-for a constitutional conven- bers of the ad hoc committee, tion, and then resigning as During his statement to the chairman as a “matter of convention, Betinis enumerat-prestige. ",
"e(j forces on campus which he\nThe “compromise conven- (Continued on Page 2)\nAssembly to Hear Justice Goldberg\nSupreme Court Associate Justice Arthur Goldberg will address a religious convocation at 2 p.m. Tuesday in Bovard Auditorium, University Chaplain John Cantelon announced yesterday.",
"\nJustice Goldberg, representing the Jewish faith, will be the second of three religious speakers this year. ",
"He will follow Father Adrian Von Kaam, a leading Catholic educator from Pittsburg's Duquesne University, who spoke last semester.",
"\nJustice Goldberg will also talk informally with students about the Peace Corps at 11 a.m. in the YWCA and will speak to the Faculty Club at noon in Town and Gowti.",
"\nHis visit is sponsored by the chaplain's office in conjunction with the Arthur S. Wolpe B'nai B’rith Hillel Memorial Fund, the B'nai\nB'rith Hillel Foundation and the Cultural Events Committee.",
"\nJustice Goldberg received a BS in law and a PhD in jurisprudence from Northwestern University.",
"\nHe was editor-in-chief of the Illinois Law Review and began private law practice in 1929.",
"\nAppointed general consul for the Congress of Industrial Organizations in 1948, Justice Goldberg served as general consul for the industrial union department of the AFL-CIO from 1955 to 1961.",
"\nIn 1961 President John F. Kennedy appointed him Secretary of Labor. ",
"He held this post until his appointment as associate justice of the Supreme Court on Aag- 29. ",
"1962.",
"\nJustice Goldberg has published numerous articles and is author of the book “AFL-CIO: Labor United.”",
"\nDr. Eleanor Metheny, physical education professor, delighted faculty members yesterday with a light but instructive speech on how college professors may quietly relieve their tensions through appropriate physical exercises.",
"\nSpeaking on the topic “Enjoy Your Tensions,” Dr. Metheny, author and world traveler, addressed an audience of approximately 150 professors and guests at the noon faculty luncheon.",
"\nDr. Metheny's demonstra-i tion of the “silent scream” technique of relieving anxieties brought more than silent laughter from her audience.",
"\nEmphasizing that the “scream” should be saved for emergencies, Dr. Metheny suggested that for smaller, more everyday frustrations, one might find the “drop your jaw” approach more convenient.",
"\nThis method, explained Dr. Metheny, involves “looking like an idiot.” ",
"She asked the\naudience to try it, producing loud laughter in the room.",
"\nFor professors suffering from insomnia. ",
"Dr. Metheny offered an original yawn producing exercise labeled the “sleep maker.”",
"\nShe also advocated that if each of her methods is followed simultaneously, one does not have enough time to think about his problems.",
"\n“I'm the only professor who puts her class to sleep on purpose,” Dr. Metheny said.",
"\nAfter Dr. Metheny's talk,' Dr. Edwin C. Robbins, president of the Faculty Center Association, announced the appointment of Dr. Tillman J.! ",
"Hall, physical education pro-: fessor, to the post of program chairman for the spring semester.",
"\nArthur J. Goldberg, associate justice of the Supreme Court, will be guest speaker at next week's meeting. ",
"Members of the faculty planning to attend are urged to make reservations early.",
"\nLINQUISTIC COMMUNICATION—Kent\nS. Cathcart, instructor in English com-1 munication, instructs a student on the\nForeign students enrolled reel-type wrhich is capable of in the English Communica- recording for approximately tion Program for Foreign Stu- one hour.",
"\ndents (ECPFS) witnessed the Movable desk tops enable opening of a new $20,000 the laboratory to be used as language laboratory yester- a regular classroom. ",
"The desk (jay tops protect the machinery\nThe laboratory, under the when not in use and form a direction of Kent S. Cath- writing surface for class-cart, instructor in English work.",
"\ncommunication, has a capaci- Each booth is also equip-ty of 24 students and is de-:ped with a counter which1 signed to assist students in tabulates the amount of tape| overcoming linguistic prob- recorded, providing easy re-lems as they learn the Eng- call of previous dialogue, lish language. “",
"This new laboratory will\nThe master-conlrol system allow the student to receive provides a two-way communi- more instruction hours in the cation between the teacher oral-aural communications. ",
"It and students, either indivi- is hoped that this will enable dually or collectively. ",
"the department to serve these\nHigh-fidelity tape and foreign students more effi-movable desk tops are two ciently,” Cathcart said, unique features of the new The language laboratory is lab. ",
"The use of high-fidelity the latest development in the tape in recording student dia-history of the English com-logues gives an accurate re- munication Program for For-production of the voice. ",
"eign Students. ",
"ECPFSC was\nThe tape is a seven-inch, started in 1958 by a faculty\ncommission in an attempt to coach foreign students in the English language so that they ma^)succeed at the university without a linguist'c handicap.",
"\nThe ECPFSC. ",
"aimed at r>-ishing student English ahili!;-rather than tearhing Engiish. ",
"covers all of the language skills. ",
"However, there is a bv-product of the program in-tercultural exchange.",
"\nThe program consists of classes in language orientation, conversation and discussion taught by a faculty of eight. ",
"Dr. Robert Kaplan, coordinator of ECPFSC, stated that the “principle purpose for the existence of this program is to serve student needs.”",
"\nApproximately 1,500 foreign students have completed courses under the auspices of the ECPFSC. ",
"There are 150 students currently enrolled in courses, Kaplan said.",
"\nuses of new equipment in the language lab. ",
"The lab is designed to aid foreign students who are learning English."
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[
"Send to a friend\n\nLow blood pressure, known as hypotension, is often overlooked as a contributing factor for fatigue and diminished brain health. ",
"Those with a blood pressure below the recommended 120/80 are often told that their numbers are terrific with nothing to worry about. ",
"This may not be entirely accurate for some individuals. ",
"Blood pressure levels should be evaluated in context of symptoms. ",
"Fatigue, poor concentration, and feeling dizzy upon standing—or as chronic symptoms, but worse when upright—may reflect low blood pressure or hypotension. ",
"This article is a follow-up to an earlier article - Low Blood Pressure Causes Brain Atrophy.",
"\n\nLow Blood Pressure Definition\n\nMayo Clinic’s definition of low blood pressure is a blood pressure reading lower than 90 systolic or 60 diastolic. ",
"Either or both systolic and diastolic number can be low. ",
"ICD-10 definition is hypotension as “a blood pressure that is below the normal expected for an individual in a given environment. ",
"It is a considered only a problem if it causes symptoms.”",
"\n\nLow blood pressure may cause symptoms when it drops suddenly when arising as in orthostatic hypotension or it may be chronically too low which is hypotension. ",
"There may also be orthostatic intolerance which may have a different nuance than orthostatic hypotension. ",
"General symptoms for all concerns include dizziness, lightheadedness, fainting, blurred vision, nausea, fatigue, and poor concentration. ",
"Some individuals may have feel chilled more easily. ",
"If blood pressure drops dangerously low it may be related with shock which may lead to confusion, cold, clammy skin, rapid, shallow breathing, and weak and rapid pulse.",
"\n\nMany may not realize that their symptoms of brain fog and fatigue may be related to diminished blood pressure, but rather blame it on other things. ",
"If you have these symptoms and a tendency towards low blood pressure, take note.",
"\n\nLow Blood Pressure Causes\n\nFrequent or ongoing low blood pressure may be related with multiple symptoms. ",
"It may be found with pregnancy, heart problems such as valve problems, heart attack, and congestive heart failure, and endocrine disorders related with thyroid, adrenal/Addison’s disease, parathyroid, hypoglycemia, and diabetes. ",
"Addison’s disease is diagnosed when 90 percent or more of the adrenal glands have been destroyed. ",
"Autonomic insufficiency linked with Parkinson’s disease and autoimmune disorders like POTS/postural orthostatic tachycardia syndrome are linked with low blood pressure.",
"\n\nLow blood pressure may reflect acute or chronic dehydration, anemia and blood loss, severe infections and severe allergies. ",
"Medications such as diuretics, heart medications, Parkinson’s medications or levodopa, some antidepressants, and erectile dysfunction meds may cause low blood pressure. ",
"It may be related with exhaustion and severe stress.",
"\n\nLow catecholamine (dopamine, adrenaline/epinephrine and noradrenaline/norepinephrine) levels are another cause of orthostatic hypotension and low blood pressure. ",
"These compounds function as neurotransmitters and stress hormones. ",
"They are produced by the brain, inner portion of the adrenal glands/adrenal medulla and the sympathetic autonomic nervous system in response to stress.",
"\n\nThese neurochemicals help your body adjust to many different stresses including changes in body position from lying to standing. ",
"Very low or undetectable levels of noradrenaline and adrenaline are found in the blood, urine, and cerebrospinal fluid may be found in autonomic insufficiency and contribute to symptoms of chronic low blood pressure or hypotension.",
"\n\nEating large meals or high amounts of processed carbs or starches may worsen low blood pressure. ",
"Crossing your legs and low salt diets may contribute to or lower blood pressure. ",
"Heat and humidity will worsen low blood pressure, whereas cooler temperatures and climates may support vasoconstriction and blood pressure in some individuals.",
"\n\nSome nutritional supplements when taken in high doses or beyond personal need may affect individuals with sensitive blood pressure regulation or symptomatic low blood pressure. ",
"It is not that the nutrient is bad as it may be helping other things in the body. ",
"It does mean that there are other things to work on and that dosage may need to change. ",
"Observe your response and adjust usage accordingly. ",
"If fatigue levels worsen after ingestion of a nutrient, it may be its natural relaxation of blood vessels and other support is needed.",
"\n\nLow Blood Pressure Evaluation\n\nIf you are unsure if you have low blood pressure, keep track of what your levels are when you go see your health care professional. ",
"Blood pressure may also be checked at the pharmacy or grocery store, your local fire department, your chiropractor’s or physical therapist office, your parish nurse or school nurse or with your own personal monitors. ",
"Make sure the cuff fits and the arm position is correct. ",
"Keep track of your symptoms if your blood pressure was found questionable.",
"\n\nIn long-standing, complex cases where numerous causes have been ruled out and the person is still quite symptomatic, a tilt table test may be performed. ",
"This tests the autonomic nervous system response to standing and evaluates blood pressure management. ",
"This test is often used to help evaluate concerns in patients with Chronic Fatigue Syndrome/CFS, Postural Orthostatic Tachycardia Syndrome (POTS), or other disorders with autonomic dysfunction.",
"\n\nBe aware that a short tilt table test of 2-3 minutes may miss the diagnosis. ",
"A 10-20 minutes standing tilt table test may be needed to fully see blood pressure changes and diagnose POTS/Postural Orthostatic Tachycardia Syndrome, autonomic insufficiency or other disorder related with hypotension. ",
"If room temperature with the tilt table test is cool, it may potentially inadvertently affect test results as the body constricts blood vessels in response cooler room temperatures.",
"\n\nBlood tests may be used to evaluate endocrine function like the adrenals and cortisol and catecholamine levels as these neurochemicals affect blood pressure and blood vessel constriction. ",
"Too much of these compounds can lead to hypertension, but when there is too little of these compounds present, it may contribute to hypotension.",
"\n\nLow Blood Pressure Effects\n\nAging, low blood pressure and orthostatic hypotension do not mix well. ",
"Risk potential and total number of falls increase with aging. ",
"Research shows that individuals 65 years and older with orthostatic hypotension were much more likely to experience falls compared to those with normal or elevated blood pressure.",
"\n\nMood changes with depression may occur with low blood pressure. ",
"Low blood pressure, depression, and autonomic insufficiency are now recognized to precede the tremors and movement problems associated with Parkinson’s Disease. ",
"Meta-analyses show that orthostatic hypotension is an independent predictor of adverse cardiovascular events and a 36 percent increase in “all cause mortality”.",
"\n\nDiminished Blood Flow to the Brain\n\nOne concern that is often not discussed with hypotension is the decrease in blood flow to the brain. ",
"Fainting, dizziness, blurred vision, decreased concentration and brain fatigue are symptoms that reflect the brain is simply not getting enough blood flow. ",
"Significant or severe low blood pressure may affect the brain and heart, even causing damage. ",
"Research tells us that long-term low blood pressure is linked with brain atrophy more so than hypertension. ",
"Please see the article Low Blood Pressure Linked with Brain Atrophy for more information.",
"\n\nIf your brain is not getting adequate circulation from diminished blood pressure, think about how it affects you with daily tasks due to increased fatigue, loss of stamina, diminished mental clarity, lower body temperature, etc. ",
"It will be harder to follow directions or participate in conversations, balance your checkbook, to make decisions, to learn and comprehend, and make new memories.",
"\n\nIf you lay down with your feet up above your heart and your energy level, mental clarity, and overall strain in the body improves, it may not just be the rest break to the muscles that causes the improvement. ",
"If symptoms improve within a few moments, it is likely that blood flow and hemodynamics have improved. ",
"This may reflect that the brain is getting more oxygen and the heart and nervous system don’t have to work so hard to nourish the brain with low blood pressures. ",
"Measures should be taken to help support blood pressure and to identify the cause for the low blood pressure.",
"\n\nPOTS, CFS, Fibromyalgia\n\nIndividuals with complex autoimmune disorders often experience a myriad of symptoms including low or dysregulated blood pressure challenges. ",
"They look well on the outside, yet struggle to get through the day. ",
"This is often the case for individuals with Chronic Fatigue Syndrome (CFS), Fibromyalgia, POTS, Ehlers-Danlos, chronic Lyme, Mast Cell Disorders, and other chronic disorders with autonomic nervous system dysfunction and blood pressure challenges. ",
"Impaired blood flow in the brain, orthostatic intolerance, low blood pressure, autonomic nervous system changes are present in these disorders that often lead to the symptoms of brain fog, brain fatigue and cognitive challenges.",
"\n\nThose with orthostatic intolerance or who have symptoms that flare when being upright have decreased brain perfusion and may not have changes in heart rate as seen in POTS. ",
"Other studies on CFS patients note decreased blood perfusion in the brain, low blood pressure, and changes in pH in muscle tissues which reflects autonomic nervous system and vascular dysfunction.",
"\n\nCFS research published earlier in 2018 demonstrates that patients with Chronic Fatigue Syndrome have lower systolic blood pressure and more of an exaggerated rhythm throughout the day. ",
"Hypotension and orthostatic intolerance is common in CFS. ",
"Symptoms of fatigue, dizziness, poor concentration, nausea, and tremors may reflect autonomic dysregulation in the body and changes of blood flow and pressure in the brain. ",
"Researchers note that the lower the systolic blood pressure in CFS patients, the more debilitating the fatigue is. ",
"If you ask a patient with CFS and hypotension who is in tune with their body, they can tell you the same thing.",
"\n\nHelpful Hints for Blood Pressure\n\nAdequate hydration and salt intake are two fundamental ways to help keep blood volume up and reduce risk for low blood pressure. ",
"We need at least 2000 mg of salt per day for general health. ",
"For those with low blood pressure, that intake may need to markedly increase to 4000-6000 mg per day or higher. ",
"Use Celtic or Himalayan sea salt rather than processed table salt.",
"\n\nSome individuals who are very weak or have sensitive systems may need to consume smaller meals four times per day rather than three meals. ",
"Large meals can cause the blood pressure to drop. ",
"Consume quality protein, complex carbohydrates, and some good fats to keep your blood sugar steady. ",
"If your energy crashes after a large meal and it is not due to food intolerance reactions or blood sugar swings, make sure you take enough digestive enzymes to help digest your meal. ",
"Do not drink large amounts of water or iced beverages with your meals as this impedes digestion. ",
"Try to rest for a few minutes after a meal rather than going back to work. ",
"Put your feet up periodically to help blood flow to the brain and heart. ",
"Compression stockings and sometimes medications are prescribed to help debilitating hypotension.",
"\n\nLow blood pressure is nothing to ignore, yet it is often dismissed as “normal”. ",
"If you have symptoms with your low blood pressure work on ways to naturally support it. ",
"Your brain, heart and all tissues in the body rely on a healthy blood pressure in order to do their daily duties. ",
"If your car only ran at low RPMs and was sputtering, wouldn’t you want to take it in to the mechanic to get it fixed?",
"\n\nDiscover\n\n*These statements have not been evaluated by the Food and Drug Administration. ",
"These products are not intended to diagnose, treat, cure, or prevent any disease. ",
"Note: the asterisk mark following a paragraph and linking to the above FDA disclaimer applies to any or all statements in that paragraph."
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"\n\nIs AI contributing to the inequality in America? - ",
"BIackSwan\nhttp://www.machinedlearnings.com/2012/03/are-we-bad-guys.html?utm_source=twitterfeed&utm_medium=twitter\n\n======\nTyDurden\nOverpopulation & unemployment go hand-in-hand with robotics & artificial\nintelligence. ",
"Pair that with technology suppression(human race stuck on planet\nEarth), and it makes for a very toxic melting pot...\n\nWe have 100 years tops before SkyNET becomes self-aware. ",
"Just because there's\na common stigma NOT to make an intelligent A.I. doesn't mean someone won't\nrelease one into the wild out of the blue...\n\n-Tyler Durden\n\n"
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"A San Joaquin Valley school district agreed Wednesday to pay $10.5 million to a former student who suffered brain damage during a pep rally, when he was attacked and beaten while wearing the costume of a chicken -- the mascot of a rival football team.",
"\n\nThe Kern County High School District agreed to settle the lawsuit filed by Mitch Carter. ",
"The vast majority of the award will be paid by the district’s insurance carrier.",
"\n\nThe announcement of the settlement brought a swift end to the damages phase of the trial, when jurors decide how much money to award. ",
"On Monday, jurors found the school district liable for Carter’s injuries.",
"\n\nIn December 2010, Carter -- then 17 years old -- was wearing a chicken costume as part of a skit at Bakersfield High School that mocked the mascot of Clovis West High School in Clovis.",
"\n\n\nDuring the pep rally, students, including members of the varsity football team, swarmed Carter, landing kicks and blows and piling onto him, according to the complaint filed in Kern County Superior Court.",
"\n\n“He was put in the most hated, personified figure at that time: the opposing team’s mascot,” said one of his attorneys, Beverly Hills-based lawyer Nicholas Rowley. “",
"They dressed him up and had him play the fool.”",
"\n\nAfter the attack, Carter incurred a mild traumatic brain injury and suffered damage to his pituitary gland, Rowley said. ",
"Carter has suffered anxiety and depression and has faced difficulty in school, earning failing grades in his courses.",
"\n\n“It changed his whole personality,” Rowley said. “",
"The boy that left that morning to go to school never came back home.”",
"\n\n\nMedical bills have totaled about $103,000 thus far, and future medical care is estimated to cost about $5 million, the attorney added.",
"\n\nMidway through the skit but before the public beating, Carter had tried to stop his performance. ",
"After two students hit and knocked him over, he told a school administrator that he would not continue wearing the chicken costume, Rowley said.",
"\n\nThe administrator scolded Carter, telling him he’d have to pay the $75 cost to rent the chicken costume if he did not continue the skit, the attorney said.",
"\n\nCarter’s attorneys noted that the student wasn’t the first person to be assaulted while wearing a costume at a pep rally.",
"\n\n\nIn 2005, a Bakersfield High School teacher wore a costume of an opposing team’s mascot and ended up with a torn rotator cuff, five broken ribs and back injuries.",
"\n\nThe lawsuit contended that the school neglected to quickly intervene after students mobbed Carter. ",
"Rowley claimed that the school created an environment that, in effect, permitted violence.",
"\n\nAttorneys for the school district countered that Carter volunteered to wear the chicken costume for the skit, which was performed without school approval.",
"\n\n“The reason things got out of hand is because Bret Mitchell Carter took it upon himself to create a skit unapproved that was poorly planned, ill advised and caused things to go south,” defense attorney Michael Kellar told the jurors, according to the Bakersfield Californian.",
"\n\n\n“Mr. Carter’s motivated by greed for money to tell absurd stories in an effort to justify how this went down.”",
"\n\nIn a statement released to the Bakersfield Californian on Wednesday, the school district said it understood why its insurance carrier opted to reach a settlement.",
"\n\n“The district is going to take this opportunity to evaluate its standards, policies and practices to ensure that every student is educated in a safe and secure environment -- because every parent and student should expect nothing less.”",
"\n\nRalph Wegis, the Bakersfield-based attorney who has represented Carter since the beginning of the case, said the jury’s decision and the sizable settlement pointed to more systemic problems facing the school district.",
"\n\n\n“Things need to be changed,” Wegis said. “",
"People need to be protected from going through what this family has gone through.”",
"\n\nAfter the settlement was announced, Carter joined his mother and attorney for a news conference and said the sizable award did not make him feel vindicated.",
"\n\n“I just wanted people to hear my story,” Carter said. “",
"I would trade everything just to have a full functioning brain.”",
"\n\nCarter said he hoped to finish college and attend law school in the future, adding: “I want to be able to stand up for people that need their voices heard.”",
"\n\n\nFor more news in California, follow @MattHjourno.",
"\n\nALSO\n\nHang glider dies after crashing into tree in the San Bernardino National Forest\n\n\nWoman who drove with pedestrian in windshield is charged with vehicular manslaughter while drunk\n\nYouTube star charged with filing false police report after saying he was beaten near WeHo gay club\n\nUPDATES:\n\n8:50 p.m.: This article was updated throughout.",
"\n\n\nThis article was originally published at 5:15 p.m."
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[
"1. ",
"Introduction\n===============\n\nIt has been over 40 years since biological markers were first introduced as a way to detect and manage thyroid cancer \\[[@B1-cancers-02-00885],[@B2-cancers-02-00885]\\]. ",
"As the understanding of the molecular pathogenesis of thyroid cancer evolved, the concept of thyroid biomarkers changed. ",
"Thus, biomarkers, also known as molecular markers, biological markers, or tumor markers, have become useful not only for detecting thyroid cancer early, but also for detecting recurrent and persistent disease and for predicting the effectiveness of surgical removal, radioiodine ablation, and chemotherapy \\[[@B3-cancers-02-00885]\\].",
"\n\nIn theory, biomarkers should tip the scales in favor of early detection. ",
"According to the most recent statistics in 2009, the incidence of thyroid cancer accounts for approximately 37,200 cases, yet causes only 0.29% of all cancer deaths \\[[@B4-cancers-02-00885]\\]. ",
"Although death attributed to thyroid cancer is uncommon, because of improvements in imaging techniques, the number of thyroid cancers detected has risen steadily over the past decade \\[[@B5-cancers-02-00885]\\]. ",
"Thyroid cancer is now the sixth most common cancer in women and the second most common cancer in women under 40 years of age \\[[@B4-cancers-02-00885],[@B6-cancers-02-00885]\\].",
"\n\nFine needle aspiration (FNA) cytology remains the procedure of choice for preoperatively establishing the diagnosis of a thyroid nodule. ",
"Despite the benefits of FNA cytology for diagnosing papillary, medullary, and anaplastic thyroid cancer, it is not helpful in determining whether follicular or Hurthle cell thyroid tumors are benign or malignant. ",
"Also some FNA cytology results suggest, but do not definitively diagnose, papillary thyroid cancer. ",
"Consequently, the management of the indeterminate thyroid follicular nodule and the suggestive but non-diagnostic papillary nodule continues to be suboptimal. ",
"The idea of identifying a single genetic mutation or cell surface marker that would distinguish a benign thyroid follicular nodule from that of a thyroid carcinoma or clarify the diagnosis of papillary thyroid carcinoma is still elusive. ",
"This is because biomarkers can encompass a spectrum of a thyroid follicle cell's physiological state, *en route* to its transformation from a benign to a malignant state \\[[@B3-cancers-02-00885]\\]. ",
"The recent multistep model of thyroid follicular carcinogenesis, based upon Vogelstein's model for colon cancer \\[[@B7-cancers-02-00885]\\], is an excellent model to study the role of biomarkers to improve thyroid cancer detection. ",
"This model also provides a firm basis for the selection of patients who can be safely observed or who need lobectomy or total thyroidectomy.",
"\n\nMolecular techniques as diagnostic tools are rapidly changing. ",
"Two of the earliest biomarkers discovered in thyroid cancer stemmed from the seminal work that established serum assays for measuring serum thyroglobulin and calcitonin levels \\[[@B8-cancers-02-00885],[@B9-cancers-02-00885]\\]. ",
"Their discovery helped revolutionize the management of patients with thyroid cancer in terms of earlier diagnosis and more sensitive follow-up. ",
"Biomarker studies have expanded to include genetic mutations and molecular changes to detect some of the earliest events in thyroid tumorigenesis. ",
"As technology improved, high-throughput genomic and proteomic assays have become the latest methods for identifying a multitude of biomarkers to reflect a molecular profile signature for each tumor type at any given stage \\[[@B3-cancers-02-00885]\\]. ",
"This article reviews the literature on thyroid cancer biomarker research as it has evolved over time, beginning with serum-based biomarkers, followed by mutation-based biomarkers, and ending with the more current genomic and proteomic-based biomarker profiles. ",
"Each of the following sections is a synopsis of the most clinically significant thyroid cancer biomarkers to date. ",
"These sections are not meant to be exhaustive or all encompassing lists. ",
"Instead, they are a starting point for understanding each type of biomarker and its utility in the field of thyroid cancer research.",
"\n\n2. ",
"Serum-Based Biomarkers\n=========================\n\nSerum biomarkers represent the first generation of thyroid biomarkers. ",
"Ideally, a serum biomarker is one that is highly sensitive and specific, can establish diagnostic certainty, and can be easily measured \\[[@B10-cancers-02-00885]\\]. ",
"This definition has remained fairly consistent over several decades, despite the introduction of complex bioinformatics systems that are being used as analytical tools to identify new biomarkers. ",
"Herberman \\[[@B11-cancers-02-00885]\\] proposed five criteria for a biomarker to be useful in cancer detection: (1) measurement should be simple, reproducible, easily available and cost effective; (2) should detect a quantitative difference so that it can distinguish those with or without disease; (3) should have high sensitivity; (4) should be able to monitor for recurrence of disease, and (5) should have high specificity.",
"\n\n2.1. ",
"Calcitonin\n---------------\n\nCalcitonin, an anti-hypercalcemic hormone secreted by the parafollicular C-cells of the thyroid, fulfills the above-mentioned criteria as a serum-based marker for medullary thyroid cancer (MTC) \\[[@B12-cancers-02-00885]\\]. ",
"In conjunction with FNA, calcitonin is a biomarker that helps optimize the sensitivity and specificity of diagnosing MTC. ",
"However, it is not a foolproof test, and calcitonin measurements have differed depending on the assays used \\[[@B13-cancers-02-00885]\\]. ",
"Radioimmunoassays were initially used to detect calcitonin in the circulation. ",
"Because these assays used polyclonal antibodies, several different monomers of calcitonin could be detected, making the test less specific \\[[@B14-cancers-02-00885]\\]. ",
"However, studies have demonstrated that both immunoradiometric assays and immunochemiluminescent assays, which are based on monoclonal antibodies, can eliminate variation and improve reliability and sensitivity in the measurement of calcitonin \\[[@B15-cancers-02-00885],[@B16-cancers-02-00885]\\]. ",
"In addition, increased calcitonin levels are not exclusive to MTC, as they have been reported in other conditions such as C-cell hyperplasia, thyroid nodules of follicular cell origin, increasing age, increased body-mass index, cigarette smoking, breast feeding and small cell carcinoma of the lung \\[[@B15-cancers-02-00885],[@B17-cancers-02-00885]\\]. ",
"However, hypercalcitonemia in the above-mentioned conditions occur with a prevalence of less than 5% \\[[@B15-cancers-02-00885]\\].",
"\n\nThe role of calcitonin as a screening marker for MTC has shifted from a \"mass-screening\" paradigm to use for specific situations. ",
"For instance, there are two types of medullary thyroid cancer: a familial (\\~25%) type and a sporadic (\\~75%) type. ",
"The level of calcitonin in MTC is associated with an increasing tumor burden as evidenced by a large tumor size along with the presence of lymph node or distant metastasis \\[[@B18-cancers-02-00885],[@B19-cancers-02-00885]\\]. ",
"Some rare patients with poorly differentiated or metastatic MTC have relatively low calcitonin levels due to cellular heterogeneity and their tumors stain less well for calcitonin and more for carcinoembryonic antigen (CEA). ",
"These patients have more aggressive disease \\[[@B20-cancers-02-00885]\\]. ",
"Overall, calcitonin is more sensitive for documenting recurrent tumor but CEA levels are better predictors of tumor aggressiveness. ",
"Provocative stimulation tests using calcium or pentagastrin increase the sensitivity of calcitonin. ",
"Provocative stimulation of calcitonin, has in general been replaced by genetic testing using *RET* mutations because the latter is more sensitive and specific \\[[@B21-cancers-02-00885],[@B22-cancers-02-00885]\\].",
"\n\n2.2. ",
"Thyroglobulin\n------------------\n\nSerum thyroglobulin is a glycoprotein that is synthesized by thyroid follicular cells and is the precursor molecule for the production of thyroxine (T4) and triiodothyronine (T3) \\[[@B23-cancers-02-00885]\\]. ",
"Thyroglobulin is made by normal, hyperplastic and neoplastic thyroid tissue. ",
"Thyroglobulin is a valuable serum marker for detecting recurrent or persistent well-differentiated thyroid cancer of follicular cell origin, as there should be no thyroglobulin present after a total thyroidectomy unless residual thyroid tissue is present \\[[@B24-cancers-02-00885],[@B25-cancers-02-00885]\\]. ",
"Although mainly localized in the colloid, thyroglobulin can be resorbed into the peripheral circulation, allowing it to be detected in the patient's serum. ",
"Van Herle initially developed an assay that measured serum thyroglobulin using a double radioimmunoassay technique \\[[@B8-cancers-02-00885]\\]. ",
"As is true for the serum detection of calcitonin, advancements are continually being made that improve both the sensitivity and specificity of serum thyroglobulin detection \\[[@B24-cancers-02-00885]\\]. ",
"However, problems still exist with the interpretation of these assays, given the presence of certain conditions. ",
"For example, the presence of anti-thyroglobulin antibodies interfere with blood thyroglobulin levels often with inappropriately low and rarely falsely high thyroglobulin levels \\[[@B26-cancers-02-00885],[@B27-cancers-02-00885]\\]. ",
"Thyroglobulin assays are not as reliable when patients are on thyroid hormone suppressive therapy. ",
"In addition, there are many benign thyroid conditions (e.g., thyroiditis, thyrotoxicosis, benign adenoma and iodine deficiency) that may cause a false-positive reading, as reflected by increased thyroglobulin levels \\[[@B28-cancers-02-00885]\\]. ",
"For thyroglobulin to serve as a reliable and sensitive marker for recurrent or persistent differentiated thyroid cancer, two conditions must be met. ",
"First, the patient must undergo total (or near total) thyroidectomy and then receive radioiodine ablation of the remaining thyroid remnant so as to assure that any subsequent thyroglobulin measurements are due to recurrent thyroid cancer and not persistent normal thyroid tissue. ",
"Second, to improve the sensitivity of measured serum thyroglobulin, thyroid stimulating hormone (TSH) stimulation must occur either through thyroid hormone withdrawal or by administering recombinant human TSH (rhTSH) \\[[@B23-cancers-02-00885]\\]. ",
"Unfortunately, even if all the above-mentioned criteria are met, thyroglobulin levels although generally very helpful in documenting tumor recurrence, may still rarely be undetectable in the serum when there is known residual disease \\[[@B24-cancers-02-00885]\\]. ",
"This is most likely to occur in patients with elevated antithyroglobulin antibodies.",
"\n\nThe serum-based biomarkers, calcitonin and thyroglobulin, provided the impetus for future research in the discovery of biomarkers in thyroid cancer. ",
"Though relatively useful as initial tests, calcitonin and thyroglobulin have several limitations. ",
"More recently, molecular studies using reverse transcriptase polymerase chain reaction (RT-PCR) have been used to measure tissue-tumor specific messenger RNA levels in the circulation. ",
"Both thyroglobulin (Tg) mRNA and TSH receptor mRNA have been extensively studied. ",
"Initially, the sensitivity and specificity of both were questioned because they were routinely found in the blood of healthy subjects. ",
"Some of the explanations for this included ectopic transcription of thyroglobulin mRNA in lymphocytes and alternative splicing of thyroglobulin mRNA in thyroid follicular cells \\[[@B29-cancers-02-00885],[@B30-cancers-02-00885]\\]. ",
"Other researchers have found that primer selection was an integral step in determining that Tg mRNA and TSHr mRNA are highly sensitive and specific markers for detecting thyroid cancer recurrence in patients who are on thyroid hormone suppressive therapy or who have circulating antithyroglobulin antibodies \\[[@B25-cancers-02-00885],[@B31-cancers-02-00885]\\].",
"\n\nThe first generation of biomarkers, the serum-based biomarkers, has helped improve the management of patients with thyroid cancer, but further advances are necessary. ",
"As the understanding of thyroid carcinogenesis continues to unfold, it is becoming clear that the single serum biomarker approach is too simplistic.",
"\n\n3. ",
"Mutation Based Biomarkers\n============================\n\nThe next step in the evolution of thyroid cancer biomarkers was the study of genetic mutations in thyroid tumors. ",
"Genetic alterations in thyroid tumors can be divided into two categories, inheritable (germline) mutations, and sporadic (somatic) mutations. ",
"Investigations into the inheritable and sporadic mutations in thyroid cancer have proceeded in parallel with one another. ",
"Although many gene mutations have been studied, only one inheritable genetic mutation and five to eight sporadic mutations are currently of significance ([Table 1](#cancers-02-00885-t001){ref-type=\"table\"}). ",
"The single known inheritable gene mutation associated with thyroid cancer is a point mutation in the *RET* proto-oncogene that causes medullary thyroid cancer. ",
"The association of a genetic mutation with medullary thyroid cancer was first hypothesized in the late 1980s, but was not specifically identified until 1993 \\[[@B32-cancers-02-00885]\\]. ",
"The first sporadic mutation identified in thyroid cancer was described in 1987 and involved a genetic defect in the RAS protein family \\[[@B33-cancers-02-00885]\\]. ",
"In 1990, somatic *RET/PTC* translocations were identified in papillary thyroid cancer. ",
"In 1992, *P53* mutations in anaplastic thyroid cancer and *NTRK1* mutations in papillary thyroid cancer were identified \\[[@B34-cancers-02-00885],[@B35-cancers-02-00885],[@B36-cancers-02-00885]\\]. ",
"There was a subsequent lull in the discovery of genetic mutations in thyroid cancer until the year 2000, when *PAX8/PPARgamma* translocations were found in follicular thyroid cancers \\[[@B37-cancers-02-00885]\\]. ",
"This was followed by the discovery of *BRAF* mutations, first in melanoma, then in papillary thyroid cancer in 2003. *",
"BRAF* mutations are the most common somatic mutations in papillary thyroid cancer and are possibly one of the most significant genetic mutation findings in thyroid cancer research \\[[@B38-cancers-02-00885]\\]. ",
"Several investigators have continued to search for the genetic mutation responsible for familial non-medullary thyroid cancer, but the responsible genetic mutation has not yet been identified.",
"\n\ncancers-02-00885-t001_Table 1\n\n###### \n\nFrequency of Sporadic Mutations in Thyroid Cancer by Phenotype.",
"\n\n Mutation Adenoma PTC FTC Anaplastic Ref.",
"\n --------------------------------------------------------------- --------------- ----- --------- ------------ -----------------------------\n BRAF 0% 44% \\<1% 24% \\[[@B39-cancers-02-00885]\\]\n RET/PTC Controversial 35% 0% 0% \\[[@B40-cancers-02-00885]\\]\n RAS 13% 10% 40% 22% \\[[@B41-cancers-02-00885]\\]\n PAX8/PPARδ 11% 0% 36% 0% \\[[@B42-cancers-02-00885]\\]\n P53 0% 1% 1% 55% \\[[@B43-cancers-02-00885]\\]\n NTRK1 Unknown 12% Unknown Unknown \\[[@B44-cancers-02-00885]\\]\n PTC: papillary thyroid cancer, FTC: follicular thyroid cancer \n\nAs the understanding of thyroid tumor formation advanced, the focus of mutation-based biomarkers shifted from single genetic mutations to molecular signatures and panels of multiple mutations. ",
"This shift in approach happened because investigators realized that the progression from a normal cell to a tumor is a complex process that likely involves multiple genetic and possibly epigenetic events, making identification of a single mutation responsible for sporadic thyroid cancer unlikely. ",
"Instead, a spectrum of many different mutations might be responsible for sporadic thyroid cancer. ",
"The discovery of single genetic mutations remains important for understanding cancer formation, but single mutations are not practical clinical biomarkers for thyroid cancer. ",
"Panels of DNA mutations are being explored as thyroid cancer biomarkers. ",
"Another area of interest is in microsatellite instability, which can be used as a biomarker for cancer development.",
"\n\n3.1. ",
"BRAF\n---------\n\nBRAF is part of the RAF family (A, B, and C) of serine/threonine kinases. ",
"These proteins relay signals from membrane-bound receptors to downstream regulators of the MAPK pathway that ultimately control the expression of several genes responsible for cell proliferation, differentiation, and apoptosis ([Figure 1](#cancers-02-00885-f001){ref-type=\"fig\"}). ",
"Most *BRAF* mutations are caused by a single point mutation that replaces valine for glutamic acid at position 600, and is designated *BRAF*^V600E^. The *BRAF*^V600E^ mutation is currently the most common genetic mutation found in papillary thyroid cancer (29--83%) \\[[@B39-cancers-02-00885]\\]. ",
"It is also frequently found in the tall-cell variant of papillary thyroid cancer, poorly differentiated thyroid cancer, and undifferentiated thyroid cancer of papillary origin. ",
"It has rarely been found in follicular thyroid neoplasms \\[[@B45-cancers-02-00885]\\]. ",
"Another activating *BRAF* mutation, *BRAF*^K601E^, has been found in thyroid adenomas and the follicular variant of papillary thyroid cancer \\[[@B46-cancers-02-00885],[@B47-cancers-02-00885]\\]. ",
"Some, but not all, investigators have found that activating *BRAF* point mutations confer a high rate of transformation to malignancy, and have been associated with later age at presentation, extra-tumoral extension, lymph node metastasis, and a higher rate of recurrence \\[[@B45-cancers-02-00885],[@B48-cancers-02-00885]\\]. ",
"A recent meta-analysis of 1168 patients confirmed an association between the *BRAF*^V600E^ mutation and higher clinical staging as well as increased extratumoral invasion \\[[@B49-cancers-02-00885],[@B50-cancers-02-00885]\\]. ",
"As a biomarker, *BRAF* is almost exclusively found in papillary thyroid carcinomas, and can be used as a marker for this tumor type. *",
"BRAF* is also thought to confer a poorer prognosis compared to sporadic thyroid tumors that do not have the mutation, but for this to be significant in clinical decision-making, more prospective studies need to be done.",
"\n\n3.2. ",
"RET/PTC Rearrangements (Sporadic Papillary Thyroid Cancer)\n---------------------------------------------------------------\n\n*RET* mutations that cause sporadic papillary thyroid cancer are rearrangements rather than point mutations like the ones found in familial medullary thyroid cancer. *",
"RET* gene gain-of-function happens when the *RET* gene is rearranged and connected to a new promoter from a gene that is constitutively expressed in thyroid follicular cells. ",
"This leads to uncontrolled downstream signaling and activation of the MAPK pathway. ",
"At least 15 separate genes are known to rearrange with the *RET* gene, causing mutations that are responsible for papillary thyroid cancer. ",
"In fact, these mutations are almost exclusively found in papillary thyroid cancer. ",
"The *RET/PTC* 1 and 3 rearrangements are by far the most common. ",
"A high frequency of *RET/PTC* rearrangements have been found in papillary thyroid cancers after radiation exposure, both from nuclear accidents, such as Chernobyl, and from exposure to external beam radiation therapy. *",
"RET/PTC* rearrangements are also common in childhood papillary thyroid cancer. ",
"A high frequency of *RET/PTC* rearrangements have also been identified in papillary microcarcinomas, suggesting that they are an early event in the formation of papillary thyroid cancer. ",
"The *RET/PTC* rearrangements are important for understanding how sporadic thyroid cancer occurs, but are not useful as clinical biomarkers.",
"\n\n{#cancers-02-00885-f001}\n\n3.3. ",
"RET Point Mutations (Familial Medullary Thyroid Cancer)\n------------------------------------------------------------\n\nThe *RET* gene, first identified as a proto-oncogene in 1985 \\[[@B51-cancers-02-00885]\\], encodes the RET receptor: a plasma membrane bound tyrosine-kinase, which is expressed in neuroendocrine and neural cells. ",
"This means that the *RET* gene product is expressed in normal thyroid parafollicular or c-cells, but not in normal thyroid follicular cells. ",
"Multiple endocrine neoplasia type 2A (MEN 2A) was assigned to chromosome 10 by linkage analysis in 1987, when the location of the *RET* gene was still unknown \\[[@B52-cancers-02-00885]\\]. ",
"The following year, the nucleotide sequence of the *RET* gene was determined and in 1989, the *RET* gene was mapped to chromosome 10q11.2 \\[[@B53-cancers-02-00885],[@B54-cancers-02-00885]\\]. ",
"In 1991, a linkage analysis of MEN 2A patients narrowed the linkage to chromosome 10q11.2, which at that time was known to be the location of the *RET* gene as well \\[[@B55-cancers-02-00885]\\]. ",
"This finding was confirmed in 1993, the same year that specific germline mutations of the *RET* gene were found in families with MEN 2A, which definitively established the association of *RET* point mutations on chromosome 10q11.2 with the development of medullary thyroid cancer \\[[@B32-cancers-02-00885],[@B56-cancers-02-00885],[@B57-cancers-02-00885]\\]. ",
"These mutations were also linked to MEN 2B patients and to families with familial medullary thyroid cancer who develop medullary thyroid cancer without other endocrinopathies.",
"\n\nPoint mutations of the *RET* gene that cause medullary thyroid cancer result in a gain of function of the RET receptor. ",
"Loss-of-function mutations of the *RET* gene do not cause thyroid cancer, but instead cause Hirchsprung's disease and congenital megacolon. ",
"Multiple individual *RET* gene point mutations have been identified and have led to the genotype-phenotype correlation of *RET* mutations and the stratification of hereditary medullary thyroid cancer into three risk groups (Levels 1, 2, and 3). ",
"These levels correlate with the genotype as well as with the age of onset and the level of aggressiveness of thyroid cancer, with Level 1 mutations being the least aggressive and Level 3 mutations being the most aggressive. ",
"Because the penetrance of thyroid cancer is nearly 100% in patients with *RET* point mutations, prophylactic thyroidectomy is now a standard of care in all patients with these mutations. ",
"Current recommendations call for prophylactic thyroidectomy between the ages of five and 10 for individuals with Level 1 mutations, at no later than five years of age for Level 2 mutations, and at diagnosis or no later than six months of age for Level 3 mutations \\[[@B58-cancers-02-00885],[@B59-cancers-02-00885]\\]. ",
"Earlier operation is indicated for these patients if they are not serum calcitonin-negative and thyroid node negative by ultrasound. ",
"Genetic testing, because it is more sensitive and specific, has replaced calcitonin monitoring to detect which patients are susceptible to developing familial medullary thyroid cancer \\[[@B60-cancers-02-00885]\\]. ",
"If the serum calcitonin level is increased or there are thyroid nodules by ultrasound then surgery is indicated. ",
"The hereditary *RET* point mutations are the most specific biomarkers in clinical use today for diagnosing patients who will develop medullary thyroid cancer. ",
"No other currently used thyroid cancer biomarker is as sensitive or specific.",
"\n\n3.4. ",
"RAS\n--------\n\nThe products of the *RAS* gene family (H, N, and K) are a group of intracellular proteins that relay signals from activated membrane-bound receptors to downstream kinases of the MAPK pathways that regulate cell proliferation, differentiation, and cell survival ([Figure 1](#cancers-02-00885-f001){ref-type=\"fig\"}). ",
"Up to 30% of human tumors are thought to have *RAS* mutations. ",
"Activating point mutations in the *RAS* genes produce constitutively active RAS proteins that cause uncontrolled activation of their downstream cellular control pathways, leading to aberrant cell proliferation and differentiation.",
"\n\nIn thyroid tumors, the most frequent *RAS* mutations are found in the *N-RAS* gene, followed by *H-RAS*, and least frequently, *K-RAS*. *",
"RAS* mutations are found in a wide variety of thyroid tumors including follicular adenomas, follicular carcinomas, papillary carcinomas, poorly differentiated carcinomas, and undifferentiated carcinomas. ",
"They are more frequently associated with follicular tumors than papillary tumors. ",
"Because *RAS* mutations are found in the entire spectrum of thyroid cancers, and with increasing frequency as tumors become more undifferentiated, *RAS* mutations have been suggested to be a biomarker for a more aggressive form of thyroid cancer \\[[@B61-cancers-02-00885]\\]. ",
"However, *RAS* mutations cannot be used to reliably differentiate between benign and malignant tumors. *",
"RAS* mutations do seem to confer an increased risk of uncontrolled proliferation, which predisposes the cells to accumulation of other genetic defects and eventual tumor formation. ",
"As a biomarker, this means that a benign follicular tumor with a *RAS* mutation may have a higher likelihood of subsequently transforming into a follicular carcinoma, but the exact risk is not known.",
"\n\n3.5. ",
"PAX8/PPARgamma\n-------------------\n\nPPARgamma is a nuclear receptor involved in cell cycle control and apoptosis. ",
"PAX8 is a transcription factor involved in the development and regulation of thyroid-specific genes. ",
"An abnormal translocation causes a fusion of the *PAX8* promoter with the *PPARgamma* gene, forming the mutated PAX8/PPARgamma fusion protein (PPFP), which causes inactivation of the wild-type PPARgama gene. *",
"PAX8/PPARgamma* translocations are found predominantly in follicular thyroid carcinomas. ",
"However, they have also been found, with a much lower frequency, in follicular adenomas and papillary carcinomas. *",
"PAX8/PPARgamma* mutations are typically associated with follicular thyroid cancers that present at an earlier age and with a high frequency of vascular invasion. ",
"This predisposition to early vascular invasion means that few tumors with these mutations are found in the pre-malignant state, which is why few follicular adenomas are found with the mutation. ",
"As a biomarker, a *PAX8/PPARgamma* rearrangement is a strong indicator that a tumor is a follicular carcinoma with an early propensity for vascular invasion, but it is not sensitive or specific enough to be used on its own to reliably distinguish between benign and malignant follicular neoplasms. ",
"However, if a *PAX8/PPARgamma* mutation is found in a follicular neoplasm, there is a higher likelihood that the lesion is or will progress to a follicular cancer \\[[@B62-cancers-02-00885]\\].",
"\n\n3.6. ",
"P53 Tumor Suppressor\n-------------------------\n\nP53 is an important tumor suppressor that regulates cell cycle arrest and apoptosis. ",
"When a cell is damaged, p53 is responsible, via a cascade of downstream regulators, for either halting the cell cycle to allow DNA repair, or for initiating apoptosis in severely damaged cells. ",
"In these ways, p53 acts to prevent damaged cells from converting into tumors. ",
"Mutations that cause inactivation of the *P53* gene may be found in up to 50% of all human malignancies, making it the most common mutation associated with human cancer \\[[@B63-cancers-02-00885]\\]. *",
"P53* mutations are common in poorly and undifferentiated thyroid cancers and in thyroid cancer cell lines, but are rare in well-differentiated thyroid cancers. ",
"This pattern of distribution suggests that an initial inciting event destabilizes the thyroid cell DNA, causing thyroid cancer formation, and after that initial event, *P53* mutations cause the thyroid cancer to progress to an undifferentiated state \\[[@B64-cancers-02-00885]\\]. ",
"This theory helps to explain the high prevalence of *P53* mutations in anaplastic thyroid cancer compared to well-differentiated thyroid cancer. ",
"There is also evidence that p53 inactivation caused by events other than mutations may play a role in well-differentiated thyroid tumors \\[[@B65-cancers-02-00885],[@B66-cancers-02-00885]\\]. ",
"Because of their high incidence in undifferentiated thyroid cancer, the presence of *P53* mutations may be predictive of a highly aggressive thyroid cancer, but on its own, sensitivity is too low to be used as a reliable clinical biomarker.",
"\n\n3.7. ",
"NTRK1\n----------\n\nThe neurotrophic tyrosine kinase type 1 (*NTRK1*) gene encodes a tyrosine kinase cell surface receptor known as NTRK1 that binds its ligand nerve growth factor (NGF). ",
"After binding to NGF, the NTRK1 receptor activates the RAS proteins, which in turn are responsible for signaling via the MAPK pathway and controlling cell proliferation, differentiation, and apoptosis ([Figure 1](#cancers-02-00885-f001){ref-type=\"fig\"}) \\[[@B44-cancers-02-00885]\\]. ",
"In the mid 1990s, three genes, tropomyosin 3 (TPM3), translocated promoter region (TPR), and TRK fused gene (TFG), were found to rearrange with *NTRK1* to form translocations that are associated with papillary thyroid cancer \\[[@B36-cancers-02-00885],[@B44-cancers-02-00885]\\]. ",
"The incidence of *NTRK1* translocations is exclusive to papillary thyroid cancer, but with a much lower frequency (5--15%) than *RET* translocations or *BRAF* point mutations. ",
"The few studies that have been done on the prognosis of *NTRK1* translocations seem to show that they have a worse prognosis than *RET* translocations \\[[@B67-cancers-02-00885]\\]. ",
"However, data on NRTK1 incidence and prognosis is limited, making *NTRK1* a poor clinical biomarker at this time.",
"\n\n3.8. ",
"DNA Mutation Panels\n------------------------\n\nPapillary and follicular thyroid cancers are hypothesized to develop via distinct mutation pathways, with more than 70% of papillary thyroid cancers caused by mutations of the RET/RAS/BRAF/MAPK pathway, and more than 80% of follicular thyroid cancers caused by mutations of the *RAS* genes or *PAX8/PPARgamma* translocations \\[[@B68-cancers-02-00885]\\]. ",
"In most thyroid cancers, these mutations are mutually exclusive events, meaning only one of these mutations is found in any particular cancer \\[[@B38-cancers-02-00885]\\]. ",
"When these mutations are used as independent biomarkers their sensitivity and specificity are too low to be clinically relevant. ",
"But, because almost 70--80% of thyroid cancers should have at least one of these mutations, a panel of all the mutations may be able to improve the diagnostic accuracy of thyroid tumor FNA cytology. ",
"In 2009, a prospective study was completed of 470 thyroid nodule FNAs using a diagnostic panel of DNA mutation testing of all these genes \\[[@B68-cancers-02-00885]\\]. ",
"This mutation-based diagnostic panel found that if any one of these mutations (*RET*, *RAS*, *BRAF*, *MAPK*, *PAX8/PPARgamma*) were detected in an FNA sample, there was a 97% chance that the nodule was malignant. ",
"The panel also demonstrated 100% sensitivity and 100% specificity in diagnosing FNA samples of \"indeterminate\" cytology as either benign or malignant. ",
"However, the sensitivity of the panel decreased to 75% for FNA samples classified as \"follicular neoplasm\", and 60% for FNA samples classified as \"suspicious for malignancy\". ",
"Up to 30% of thyroid cancers will have no currently known mutations, thus further decreasing the utility of current DNA mutation panels. ",
"Signatures from gene expression profiles will eventually be used to construct new DNA mutation panels for FNA-based diagnosis of thyroid nodules.",
"\n\n3.9. ",
"Microsatellite Instability\n-------------------------------\n\nMicrosatellites are simple DNA sequences of two to five nucleotide base pairs that are repeated multiple times. ",
"These repeated sequences are found scattered throughout the genome, and are a normal part of the genetic makeup. ",
"Microsatellites are useful as highly specific DNA markers in genetic analysis because they are highly polymorphic and normal variation occurs between individuals. ",
"In 1993, it was discovered that variations in the length of microsatellites (microsatellite instability) at individual loci could be used as a marker of DNA mutation and instability in colorectal cancer \\[[@B69-cancers-02-00885]\\]. ",
"Microsatellite instability was eventually determined to occur secondary to mutations of DNA mismatch repair genes. ",
"The defective mismatch repair genes cause the microsatellites to be replicated incorrectly. ",
"More importantly, these defective repair genes allow accumulation of multiple somatic point mutations that eventually lead to tumor formation. ",
"In this way, microsatellite instability can be used as a genetic signature that identifies cells at risk for becoming malignant, and can possibly allow for early intervention. ",
"Since the first study demonstrating microsatellite instability in thyroid lesions in 1997 \\[[@B70-cancers-02-00885]\\], small panels of microsatellites have been studied in thyroid cancer, with varying outcomes. ",
"Microsatellite instability has been demonstrated in both papillary and follicular thyroid tumors, as well as in benign thyroid adenomas \\[[@B71-cancers-02-00885],[@B72-cancers-02-00885]\\]. ",
"However, others have been unable to find any microsatellite instability in thyroid tumors \\[[@B73-cancers-02-00885]\\]. ",
"One problem with studying microsatellite instability is deciding which microsatellites to include in the analysis. ",
"High-throughput studies of the entire genome that would generate a genetic signature have not been done, but are needed to better determine the clinical utility of microsatellite instability in thyroid cancer. ",
"Currently, microsatellite instability cannot be used as a clinical biomarker for thyroid cancer because its frequency in various types of thyroid tumors is still debatable.",
"\n\n4. ",
"Epigenetic Biomarkers\n========================\n\nEpigenetics is defined as the study of heritable changes in gene expression that occur independent of changes in the primary DNA sequence. ",
"Mechanisms of epigenetic regulation include DNA methylation, histone protein modification, nucleosome positioning, and microRNA silencing. ",
"In thyroid cancer, DNA methylation, histone modifications, and microRNA silencing have all been studied, but there is minimal data on nucleosome positioning. ",
"All these mechanisms regulate DNA by silencing the expression of genes when they are activated. ",
"Defects in these mechanisms lead to aberrant gene expression and both have been implicated in multiple human cancers. ",
"One of the most exciting aspects of epigenetic regulation is that unlike genetic mutations, these processes are easily reversible with various therapeutic agents. ",
"Because of this, markers of epigenetic causes of thyroid cancer have been investigated. ",
"As with the other thyroid cancer biomarkers, work has proceeded from initially identifying these changes in thyroid cancer, to identifying epigenetic \"signatures\" that can predict cancer formation before the cancer becomes clinically apparent so that treatment can begin at an early stage.",
"\n\n4.1. ",
"Hypermethylation\n---------------------\n\nDNA methylation is an epigenetic regulatory mechanism involved in silencing gene expression that is particularly important in normal embryogenesis. ",
"It occurs by adding a methyl group to the cytosine residue of a CpG dinucleotide. ",
"Regions of DNA that contain multiple copies of CpG dinucleotides are termed CpG islands and are usually located at the 5' end of gene promoters. ",
"Gene silencing after methylation of a CpG island occurs by either blocking the binding of transcription factors to the promoter region or by recruitment of methyl-binding DNA transcription repressors to the promoter. ",
"Aberrant methylation, or hypermethylation, of tumor suppressor genes has been identified in many human tumors including thyroid tumors \\[[@B74-cancers-02-00885]\\]. ",
"Hypermethylation of multiple genes has been identified in association with the PIK3/AKT pathway in follicular thyroid cancers, and of the MAPK pathway in papillary thyroid cancers \\[[@B75-cancers-02-00885],[@B76-cancers-02-00885]\\]. ",
"Hypermethylation has also been identified in benign thyroid tumors, though to a lesser extent than in thyroid carcinomas. ",
"Investigations have also linked *BRAF* mutations to hypermethylation, which may be associated with increased production of vascular endothelial growth factor in *BRAF*-positive thyroid tumors \\[[@B77-cancers-02-00885]\\]. ",
"Other genes identified as hypermethylated in thyroid cancer include the *TSH* receptor, *NIS*, *PCNA*, *CDKN2A*, *FGFR2*, and *RASSF1A*. ",
"High-throughput technology has shown that patterns of methylation that correlate with tumor formation and progression produce a genetic signature called the CpG island methylator phenotype (CIMP). ",
"The CIMP signature confers a high degree of genetic instability and has been identified as an early marker of progression from benign to malignant neoplasia in colon cancer \\[[@B78-cancers-02-00885]\\]. ",
"A CIMP signature has not been identified in thyroid cancers, but is the next step in the evolution of hypermethylation as a thyroid cancer biomarker.",
"\n\n4.2. ",
"MicroRNA\n-------------\n\nMicroRNAs, also known as miRNAs or miRs, were discovered in 1993, and since then, more than 700 human miRNAs have been catalogued \\[[@B79-cancers-02-00885],[@B80-cancers-02-00885]\\]. ",
"They are single-stranded RNAs of 20--25 nucleotides that down-regulate gene expression at the post-transcriptional level via translational repression, mRNA cleavage, and mRNA decay. ",
"They can act as either oncogenes or tumor suppressors and have been identified in the regulation of a wide variety of functions including hematopoiesis, cell proliferation, cell survival and apoptosis \\[[@B81-cancers-02-00885]\\]. ",
"A single miRNA can regulate the expression of hundreds of different genes. ",
"The first study of miRNA expression in thyroid cancer was in 2005, and found 17 miRNAs that were over expressed and six miRNAs that were under expressed \\[[@B82-cancers-02-00885]\\]. ",
"Most studies on thyroid cancer miRNA expression are in papillary thyroid cancer and have consistently shown upregulation of miRNAs 146b, 221, 222, 224, 155, and 181b. ",
"Fewer studies have been done on follicular thyroid cancers, but upregulation has been identified in miRNAs 155, 187, 221, 222, and 224 \\[[@B83-cancers-02-00885]\\]. ",
"As is true for genomics, miRNA \"signatures\" are beginning to be identified, which may prove useful for diagnosing thyroid cancer from FNA samples. ",
"A few small prospective trials using FNA samples have established miRNA as a possible diagnostic tool, but larger clinical trials are needed to confirm the utility of miRNAs as a clinical biomarker \\[[@B84-cancers-02-00885]\\].",
"\n\n5. ",
"Genomics\n===========\n\nThe term genomics was first used in 1987 and refers to the study of the structure and organization of a genome of a particular organism. ",
"This includes the mapping and sequencing of the genome, as well as the analysis of the information gained from mapping and sequencing in the context of their biological significance and biomedical application \\[[@B85-cancers-02-00885]\\]. ",
"Technology is expanding the study of genomics and our understanding of the genome at an almost exponential pace. ",
"A full review of genomics, even in the context of thyroid cancer, is beyond the scope of this review. ",
"The following general overview is intended as a starting point for understanding the important findings in the field of genomics as they relate to thyroid cancer.",
"\n\nInitially, the field of genomics was focused on mapping the human genome and understanding the effects of single genes in a system. ",
"The sequencing of the human genome, along with the development of high throughput technologies, have shifted the focus of the field of genomics to gene expression profiling and the quest to identify genetic signatures of disease. ",
"Many different gene expression profiling technologies are currently in use, including cDNA microarrays, oligonucleotide arrays, and Serial Analysis of Gene Expression (SAGE) \\[[@B86-cancers-02-00885],[@B87-cancers-02-00885],[@B88-cancers-02-00885]\\]. ",
"These technologies allow the simultaneous study and comparison of the expression of thousands of genes in varying conditions. ",
"The utility of this approach is in recognizing patterns of gene expression to gain insight into biological processes rather than focusing on individual gene expression. ",
"These technologies are being used in cancer research to elucidate the mechanisms of tumor formation to identify a patient's risk of developing cancer, to predict prognosis once cancer has developed, and ultimately to design targeted therapies for cancer treatment. ",
"The hope is that gene expression profiling will someday lead to personalized medicine, in which a DNA signature unique to each patient can be developed and allow tailoring of treatment specific to that patient's needs.",
"\n\nOne of the first gene expression profiles done in thyroid cancer was on a group of 8 papillary thyroid cancers in 2001 \\[[@B89-cancers-02-00885]\\]. ",
"This study identified a papillary thyroid cancer gene signature that consisted of more than 220 differentially expressed genes, many of which were previously identified as being differentially expressed in papillary thyroid cancer, and many of which were not. ",
"In 2005, a DNA microarray analysis on papillary thyroid cancer identified genetic signatures that distinctly correlate with mutations in *BRAF*, *RAS*, and *RET/PTC* papillary thyroid cancers \\[[@B90-cancers-02-00885]\\]. ",
"These genetic signatures support the theory that sporadic papillary thyroid cancers originate via mutations in the RET/RAS/BRAF/MAPK pathway, and could also predict tumors with these mutations with high accuracy. ",
"This study also identified specific genetic signatures that distinguish classic papillary thyroid cancer from the tall cell and follicular variants.",
"\n\nOther gene expression profile studies have identified distinct genetic signatures that correlate with the *PAX8/PPARgamma* translocation in follicular thyroid cancer, and in *RET/PTC* rearrangements in radiation-induced papillary thyroid cancer, and can help predict an increased risk of invasion in papillary thyroid cancer \\[[@B91-cancers-02-00885],[@B92-cancers-02-00885],[@B93-cancers-02-00885]\\]. ",
"Multiple gene expression profiling studies on thyroid cancer have been completed in a relatively short period of time. ",
"This is reflected in a 2006 meta-analysis of 21 gene expression profile studies \\[[@B94-cancers-02-00885]\\]. ",
"The meta-analysis highlights both the strengths and weaknesses of high throughput gene expression profiling. ",
"The analysis identified a small group of genes that were consistently differentially expressed in multiple studies. ",
"Within this group were a subset of genes known to be involved with thyroid cancer, and a subset of genes that had previously not been identified in relation to thyroid cancer. ",
"However, several genes identified as being involved in thyroid cancer in individual studies were not consistently found across multiple studies. ",
"This highlights the fact that gene expression profiling is still an evolving field with many different and complicated technologies. ",
"Because these studies generate such large amounts of data, complex mathematical formulas and statistical analyses are required. ",
"Currently, the analysis of high-throughput gene expression profile studies has not been standardized, which might be partially responsible for the occasional contradictory findings associated with these studies. ",
"Even though these studies have already produced many exciting findings regarding the development and biology of thyroid cancer formation, there is currently not enough concordance of these genetic signatures to use them as biomarkers for diagnosis or prognosis of patients with these tumors.",
"\n\n6. ",
"Proteomics\n=============\n\nProteomics is defined as the study of protein structure and function. ",
"The term was first introduced as an analogy to \"genomics,\" but in this case referring to the entire protein spectrum \\[[@B95-cancers-02-00885]\\]. ",
"Before high-throughput proteomics was developed, protein expression was studied by immunohistochemistry, serving as an adjunct to FNA cytopathology, hoping to improve the diagnostic accuracy in indeterminate thyroid nodule FNA samples. ",
"Immunohistochemical markers have been used to determine the molecular phenotype of these indeterminate lesions.",
"\n\n6.1. ",
"Immunohistochemistry\n-------------------------\n\nSeveral immunohistochemical markers representing different components of the cell, such as the membrane, the cytoplasm, or the nucleus, have been studied in thyroid neoplasms \\[[@B96-cancers-02-00885]\\]. ",
"Some of the antibodies that have been examined include galectin-3, Hector Battifora mesothelial cell antibody (HBME-1), cytokeratin-19, *RET*, TTF-1, hTERT, telomerase, p27 and p53 to name a few ([Table 2](#cancers-02-00885-t002){ref-type=\"table\"}). ",
"Two markers that have been extensively studied are galectin-3 and HMBE-1. ",
"When each of these antibodies was individually examined, variable results were obtained. ",
"Some studies found that these markers could differentiate between benign and malignant thyroid lesions and that the protein expression varied as a function of the tumor stage of papillary thyroid cancer \\[[@B97-cancers-02-00885],[@B98-cancers-02-00885],[@B99-cancers-02-00885],[@B100-cancers-02-00885],[@B101-cancers-02-00885]\\]. ",
"Although these markers supported the diagnosis for the classical type of papillary thyroid cancer, they did not help for its histological variants. ",
"On the other hand, when these markers were combined (e.g., galectin-3 + HBME-1; HBME-1 + CK-19; galectin-3 + HBME-1 + CK-19), several investigators reported improved sensitivity and specificity for the detection of cancer seen on tissue or FNA samples \\[[@B101-cancers-02-00885],[@B102-cancers-02-00885],[@B103-cancers-02-00885],[@B104-cancers-02-00885]\\]. ",
"Consequently, these results suggest that panels or patterns of these and other markers may represent the future of analyzing protein expression to improve the diagnostic certainty of the indeterminate thyroid nodular lesion.",
"\n\ncancers-02-00885-t002_Table 2\n\n###### \n\nSignificant IHC Markers in Differentiated Thyroid Cancer.",
"\n\n Marker Expression in thyroid cancer Marker Expression in thyroid cancer\n ------------ ------------------------------ ------------ ------------------------------\n Galectin-3 Upregulated HER4 Upregulated\n CK19 Upregulated TG Downregulated\n VEGF Downregulated MIB-1 Upregulated\n Aurora-A Upregulated Caveolin Upregulated\n p16 Upregulated Aurora-C Upregulated\n AR Upregulated S100 Upregulated\n HBME-1 Upregulated MRAS Downregulated\n Bcl-2 Downregulated c-kit Downregulated\n Cyclin-D1 Upregulated HER3 Upregulated\n CAV-1 Upregulated RET Upregulated\n Cyclin-E Upregulated AMF-R Upregulated\n E-CAD Downregulated MLH1 Downregulated\n Clusterin Upregulated AAT Upregulated\n CR Upregulated TTF-1 Upregulated\n IGFBP5 Upregulated PGI Upregulated\n P21 Upregulated HSP-27 Downregulated\n IGFBP2 Upregulated Syntrophin Upregulated\n CTNNB1 Upregulated \n\n**Bold = most discriminatory markers**; IHC = immunohistochemical. ",
"With kind permission from Springer Science+Business Media: *Annals of Surgical Oncology*, Molecular Phenotyping of Thyroid Tumors Identifies a Marker Panel for Differentiated Thyroid Cancer Diagnosis, Vol. ",
"15, 2008, 2811--2826, Wiseman SM *et al*., ",
"Table 5, © 2008 The Society of Surgical Oncology, Inc.\n\n6.2. ",
"High-Throughput Proteomics\n-------------------------------\n\nThe study of biomarkers has now expanded to include the analysis of the proteome, which investigates protein expression and modification, along with change in protein activity and protein localization \\[[@B3-cancers-02-00885]\\]. ",
"The information gained from the proteome reflects not only the expressed genomic profile of the cell, but also takes into account post-translational changes that are not detected at the mRNA level, as well as protein expression at different time points along a cell's spectrum of neoplastic transformation \\[[@B3-cancers-02-00885],[@B95-cancers-02-00885],[@B105-cancers-02-00885]\\]. ",
"The advantage of proteomics is the ability to detect biomarkers from the patient's serum or plasma. ",
"The idea that a tumor is capable of \"leaking\" proteins into the circulation ultimately allows potential serum biomarkers to be identified \\[[@B106-cancers-02-00885]\\].",
"\n\nThe study of proteomics uses high-throughput technologies, which combines multidimensional separation systems based on mass spectrometry analysis and ProteinChip technology (Ciphergen Biosystems, USA) to improve sensitivity and specificity of a complex mixture of proteins and peptides from either tissue or serum \\[[@B107-cancers-02-00885],[@B108-cancers-02-00885]\\]. ",
"Currently there are two proteomic methods for identifying biomarkers. ",
"The first method uses mass spectrometry (MS) to identify protein patterns by determining the mass:charge ratio and amino-acid sequence of proteins within a given specimen. ",
"When MS is combined with an ionization technique, such as matrix-assisted laser desorption ionization-time of flight (MALDI-TOF), surface-enhanced laser desorption/ionization-time of flight (SELDI-TOF) or electrospray ionization, it improves the sensitivity and rapid detection of cancer-specific biomarkers and proteomic-profile patterns \\[[@B3-cancers-02-00885],[@B108-cancers-02-00885],[@B109-cancers-02-00885]\\]. ",
"The second method uses the technique of proteolytic digestion by exposing the protein samples to high efficiency liquid chromatography along with tandem MS analysis. ",
"This technique improves both protein identification and abundance, which translates into detecting and identifying larger amounts of peptides from complex protein mixtures \\[[@B109-cancers-02-00885],[@B110-cancers-02-00885]\\]. ",
"Once MS has established a protein profile pattern, this data is read by artificial intelligence-based bioinformatics systems that serve as diagnosticians and help discriminate between normal, benign, pre-malignant, or malignant disease from a patient's serum or tissue \\[[@B108-cancers-02-00885]\\].",
"\n\nThe first study that established a proteomic profile of benign and malignant human thyroid tissue was reported in 2002 \\[[@B111-cancers-02-00885]\\]. ",
"In that study, cathepsin B was found to be up-regulated in the neoplastic tissues of follicular adenoma, follicular carcinoma, and papillary carcinoma when compared to non-neoplastic thyroid tissue. ",
"Unfortunately, cathepsin B could not distinguish between follicular adenoma and follicular carcinoma. ",
"The study also detected the up-regulation of two proteins within papillary thyroid cancer: ATP synthase D chain (ATPQ) and prohibitin (PHB) \\[[@B111-cancers-02-00885]\\]. ",
"The clinical and therapeutic usefulness of these findings are still unknown.",
"\n\nTwo subsequent studies using two-dimensional difference gel electrophoresis (2D-DIGE) and MALDI-TOF mass spectrometry attempted to delineate a set of thyroid biomarkers through protein analysis. ",
"Brown *et al.* ",
"\\[[@B112-cancers-02-00885]\\] compared the protein expression profiles of tissue from papillary thyroid cancer (PTC) specimens with matched normal thyroid tissue from the same patients and normal thyroid tissue from benign follicular adenomas. ",
"Using statistical validation software they discovered three novel biomarkers for PTC: (1) S100A6 (an isoform of S100 protein); (2) peroxiredoxin 2, and (3) heat shock protein 70 (HSP70), all three of which showed more than a two-fold difference in protein expression. ",
"In addition, they also confirmed the overexpression of known PTC biomarkers as identified by mRNA expression, which included galectin-3, cytokeratin-19 and cathepsin-B. Interestingly, they found two distinct forms of cathepsin-B in PTC, one form that was under expressed and another form (more acidic) that was over-expressed. ",
"The authors noted that cathepsin-B is involved in thyroglobulin expression and it is well known that serum thyroglobulin is a marker for disease recurrence. ",
"The differences seen in the different forms of cathepsin-B protein expression reflect the need for further study of these post-translational changes to validate these proteins as thyroid cancer biomarkers \\[[@B112-cancers-02-00885]\\]. ",
"The second study, conducted by Netea-Maier *et al.* ",
"\\[[@B113-cancers-02-00885]\\], examined protein abundance differences between thyroid follicular adenomas and follicular carcinomas. ",
"They discovered a statistically significant difference in protein abundance among 43 proteins between follicular thyroid carcinoma and follicular adenoma. ",
"Based upon previous reports they decided to examine three proteins (HSP gp96, PDI A3 and calreticulin) in greater detail and subsequently performed immunohistochemical analysis on a different set of paraffin-embedded tissue blocks for validation purposes \\[[@B113-cancers-02-00885]\\]. ",
"These three proteins (HSP gp96, PDI A3 and calreticulin) are involved in protein folding and consequently a subset (HSP gp96 and calreticulin) may play a role in anti-cancer immunogenicity \\[[@B114-cancers-02-00885],[@B115-cancers-02-00885]\\]. ",
"Immunohistochemical analysis revealed that calreticulin was the best single marker for distinguishing between follicular adenoma and follicular thyroid carcinoma, and all three markers together showed a high sensitivity at predicting widely invasive follicular thyroid carcinoma \\[[@B113-cancers-02-00885]\\]. ",
"Because the discovery proteomics and the immunohistochemical validation studies proved to be accurate for distinguishing between follicular adenoma and follicular thyroid carcinoma for these three markers, prospective studies are now needed to prove the clinical utility of this approach.",
"\n\nTo expand upon the study of proteomics, Giusti *et al.* ",
"\\[[@B116-cancers-02-00885],[@B117-cancers-02-00885]\\] was the first group to analyze the fluid obtained from fine needle aspirates (FNA) of thyroid cancer. ",
"They used a combination of 2D-GE and MALDI-TOF-MS analysis to acquire a proteomic profile of a variety of classes of proteins from an FNA thyroid biopsy. ",
"In a follow-up study, they then examined the proteomic profiles of FNA samples from papillary thyroid tumors (comparing the classical and tall cell variants) as well as control FNA specimens from the opposite (unaffected) thyroid lobe. ",
"They discovered 17 protein spots that could reliably distinguish between papillary thyroid cancer and normal thyroid tissue \\[[@B117-cancers-02-00885]\\]. ",
"Some of these proteins had been previously linked to thyroid cancer by other methods \\[[@B111-cancers-02-00885],[@B112-cancers-02-00885],[@B118-cancers-02-00885],[@B119-cancers-02-00885]\\]. ",
"The proteomic analysis also showed a difference in protein expressed between the classical variant of PTC and the tall cell variant of PTC. ",
"In particular, ferritin heavy chain \\[FHC\\], peroxiredoxin 1 \\[PRX1\\] and 6-phosphogluconate dehydrogenase \\[6-PDGH\\] were exclusively upregulated in the tall cell variant of papillary thyroid cancer \\[[@B117-cancers-02-00885]\\].",
"\n\nThe most recent proteomic studies on thyroid cancer utilized SELDI-TOF mass spectrometry to distinguish between PTC and benign thyroid disease based upon serum protein profiles \\[[@B107-cancers-02-00885],[@B120-cancers-02-00885]\\]. ",
"The major advantage of the SELDI-TOF technique is that it uses minute amounts of serum, coupled to bioinformatics systems that allow for high-throughput technology. ",
"According to Wulfkuhle \\[[@B108-cancers-02-00885]\\], proteomic patterns or fingerprints, instead of protein identities, can generate discernable biomarker patterns. ",
"According to Wang \\[[@B107-cancers-02-00885]\\], different protein expression factors can distinguish among four distinct categories: (1) PTC *vs.* normal thyroid tissue; (2) PTC *vs.* benign thyroid nodules; (3) various pathological stages of PTC; and (4) different pathological types of thyroid cancer. ",
"Thus, several biomarker patterns could be identified as a distinguishing feature within each category. ",
"Similarly, when three different groups were examined: (1) benign nodular disease *vs*. ",
"PTC; (2) benign nodular disease *vs.* controls; and (3) PTC *vs.* controls, a set of protein biomarkers distinguished benign nodular disease from PTC, based on different proteomic expression patterns, with a high degree of sensitivity and specificity (85.7%, 100%, respectively) \\[[@B120-cancers-02-00885]\\]. ",
"Fan *et al.* ",
"\\[[@B121-cancers-02-00885]\\] utilized several proteomic techniques to identify and validate their findings. ",
"The proteomic profiles of serum samples from patients diagnosed with PTC and non-cancer controls were analyzed. ",
"SELDI-TOF-MS technology was used to screen for potential candidate protein patterns and high pressure liquid chromatography, tandem MS and ProteinChip immunoassays were used to purify, identify and confirm the protein biomarkers, respectively. ",
"They identified a set of three protein biomarkers (haptoglobin alpha-1 chain, apolipoprotein C-I and apolipoprotein C-III) capable of differentiating between PTC and non-cancer controls. ",
"Another study used tissue from three patients diagnosed with PTC \\[[@B122-cancers-02-00885]\\]. ",
"Data analysis from three different protein chips identified approximately 63 protein biomarkers that could potentially distinguish between PTC and normal thyroid tissue from the same patients. ",
"The drawback of this study was that they failed to determine the identities of these 63 protein biomarkers (via SELDI-TOF-MS) and only three samples of PTC were examined. ",
"However, expanding this technique to a larger group of samples with different thyroid pathologies may help to identify a panel of protein biomarkers to help distinguish benign from malignant thyroid neoplasms.",
"\n\nMore recent proteomic research in thyroid cancer has focused on discovering biomarkers with the potential to identify therapeutic targets. ",
"Neutrophil gelatinase-associated lipocalin (NGAL) is a protein involved in the inflammatory and immune response system and is expressed in the FRO thyroid cancer cell line (a human anaplastic thyroid cancer cell line). ",
"Through 2D-GE and MALDI-TOF analysis, NGAL was identified as a mediator of NF-κB, a protein responsible for oncogenic activity in poorly differentiated thyroid cancer cell lines \\[[@B123-cancers-02-00885]\\]. ",
"In addition, NGAL may serve as a potential therapeutic target due to its regulatory activity of iron. ",
"NGAL is responsible for transporting iron from the extracellular space into the cell. ",
"Increased intracellular iron levels have been shown to contribute to excess DNA synthesis via multiple transcription factors leading to tumor development \\[[@B124-cancers-02-00885],[@B125-cancers-02-00885],[@B126-cancers-02-00885]\\]. ",
"This information may help identify therapeutic targets against NGAL and its role in iron uptake because iron chelators are already in clinical use today.",
"\n\nIn 2009, a glycoproteomic profile of biomarkers was obtained by using liquid chromatography-tandem mass spectrometry to identify cell surface and secreted glycoproteins \\[[@B127-cancers-02-00885]\\]. ",
"Both well-differentiated (FTC-133, XTC-1, TPC-1) and undifferentiated (ARO, DRO-1) thyroid cancer cell lines were studied, and an average of 150 glycoproteins were identified for each cell line examined. ",
"Most of the glycoproteins were either cell surface proteins or secretory proteins. ",
"Interestingly, a subset of five cell membrane glycoproteins (vasorin, NCAM-1, trophoblast glycoprotein, discoidin and integrin alpha-5 chain) were only found in the well-differentiated thyroid cancer cell lines as compared to the undifferentiated anaplastic cell lines \\[[@B127-cancers-02-00885]\\]. ",
"This information holds potential for developing therapeutic strategies at the level of cell surface, integral membrane, or secretory glycoproteins, but validation studies are still needed.",
"\n\nA 2009 study reported the presence of *RET* oncogene mutations in human medullary thyroid cancer cell lines (TT and MZ-CRC-1) based on proteomic analysis and antibody-based validation techniques \\[[@B128-cancers-02-00885]\\]. ",
"The study compared expression profiles of downstream signaling elements (e.g., *RAS*-mitogen-activated protein kinases \\[MAPKs\\], phosphatidylinositol 3-kinase \\[PI3K\\], c-jun N-terminal kinase \\[JNK\\]) from *RET*-MEN2A and *RET*-MEN2B expressing cells. ",
"There were 41 phosphotyrosine proteins detected downstream following *RET* activation from two different medullary thyroid cancer cell lines and the EGFR pathway was over-expressed. ",
"It is hoped that this information might provide new targets for anti-cancer therapies. ",
"The analysis of proteomics coupled with the analysis of gene array expression and epigenetic changes is unraveling the oncogenic pathways for the development of targeted therapeutics, as seen in breast, prostate and colorectal cancer \\[[@B129-cancers-02-00885]\\].",
"\n\n7. ",
"Conclusions\n==============\n\nThe study and identification of biomarkers in thyroid cancer has a long history. ",
"From the beginning, the goal was to find markers that can identify benign and malignant thyroid tumors and to predict the behavior of these thyroid cancers. ",
"The evolution of thyroid biomarker research reflects the standard thinking of the time and has changed in parallel with the advancement of technology. ",
"Although thyroid cancer is one of the least deadly forms of cancer, research in the field has remained on the cutting edge of science and technology, but better diagnostic tests and predictors of tumor aggressiveness are necessary. \"",
"Finding a cure\" for thyroid cancer should remain a strong impetus for continued research to identify new biomarkers and therapeutics for this disease.",
"\n\nThis review highlights the fact that even though a significant amount of progress has been made in cancer research, we are only at the beginning of fully understanding how normal cells progress to tumors. ",
"This is why many thyroid cancer biomarkers have been identified, but few have made it into routine clinical practice. ",
"Calcitonin and thyroglobulin are markers for disease recurrence after surgical resection, and the *RET* point mutations are used to predict susceptibility and prognosis of a patient with familial medullary thyroid cancer. ",
"None of the other biomarkers reviewed in this study are currently in routine clinical use, but are being used as building blocks for the future to improve treatment.",
"\n\nBiomarker research continues to advance. ",
"Serum-based markers are moving from proteins secreted by tumors to detecting minute amounts of genetic material being shed by the tumor into the circulation. ",
"DNA mutations are still being studied and discovered, and have progressed from single mutation discoveries to the synthesis of \"mutation pathways\" and panels of DNA mutations as a single \"signature\". ",
"Epigenetic regulation has become a part of biomarker research, and will increase in use as a biomarker as our understanding of these processes improves. ",
"High-throughput technologies have advanced our ability to study the human genome, but much work in the field still needs to be done in the validation and quality control of the outcome data. ",
"These technologies have also greatly expanded our ability to study the proteome, going from the study of single proteins to identifying proteomic signatures of thousands of expressed proteins. ",
"We have also begun to see a crossover of the different disciplines. ",
"Proteomics is being applied to serum-based studies, genomics is helping to identify new DNA mutations, and epigenetic regulation has changed our understanding of how DNA regulation takes place.",
"\n\nThyroid biomarker investigations were once the realm of a single scientist working on elucidating the mechanisms and utility of a single biomarker. ",
"We are moving into an era of biomarker research that requires a multi-disciplinary team of biologists, geneticists, statisticians, oncologists, surgeons, and even engineers. ",
"This team has the task of understanding and interpreting data from new technologies and incorporating these findings into our understanding of tumor formation. ",
"It may be that a combination of serum based biomarkers, epigenetics, genomics, and proteomics will be required to produce reliable and clinically helpful biomarkers. ",
"It could also be that ideas and mechanisms that have not yet been discovered will be the key to thyroid biomarkers. ",
"New areas of study like metabolomics, the study of the end products of cellular processes, and the kinome, the full complement of human protein kinases, have not been studied in thyroid cancer. ",
"Thyroid biomarker discovery remains an exciting field of thyroid cancer research, and will remain on the cutting edge of technology.",
"\n\nThe authors would like to thank the Friends of Endocrine Surgery for their support as well as the Cedars Cancer Institute of the McGill University Health Centre, Montreal, Quebec, Canada (EJM is the recipient of the Henry R. Shibata Fellowship Award). ",
"The authors would also like to thank Pamela Derish from the UCSF Department of Surgery Publication Office for her editorial assistance.",
"\n\n[^1]: RHG and EJM contributed equally to this work.",
"\n"
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"You don’t have to be a full-fledged Trekkie to be familiar with the famous “Vulcan blessing” from Star Trek — “Live long and prosper” — or with the splay-fingered\n\n“Vulcan salute”\n\nthat is generally used when this saying is spoken.",
"\n\nIf you’re a fan of the original Star Trek television series, you’re undoubtedly know the episode that introduced the Vulcan blessing and salute.",
"\n\nIt’s titled “Amok Time” and today is its anniversary.",
"\n\nIt was first aired on September 15, 1967 (as Episode 1 of Season 2).",
"\n\nI remember watching “Amok Time” that night in ‘67 as a teenager and working to make my fingers split apart in proper Vulcan fashion.",
"\n\nToday, like many Trek fans, I consider it one of the best episodes of the original series.",
"\n\nThe script for “Amok Time” was written by the legendary science fiction author Theodore Sturgeon .",
"\n\nIt’s one of three scripts he wrote for Star Trek. ",
"Sturgeon also penned the script for the humorous “Shore Leave” episode from Season 1 and a script titled “Joy Machine” that was never produced.",
"\n\nIn addition to being the first Trek episode to feature the Vulcan blessing and salute, “Amok Time” is the only episode of the original Trek series that includes scenes set on Vulcan, the home planet of Mr. Spock (actor Leonard Nimoy).",
"\n\nMany key elements of Vulcan culture created by this episode were used throughout the rest of the original series — and in the following Trek spin-off TV series and movies.",
"\n\nThe most memorable initial use of the Vulcan blessing and salute comes near the end of the “Amok Time” episode.",
"\n\nAs Spock prepares to leave the planet, he says to the female Vulcan leader, T’Pau: “Live long, T’Pau, and prosper.”",
"\n\nT’Pau responds: “Live long and prosper, Spock.”",
"\n\nAs they speak, they give each other the Vulcan salute.",
"\n\nAccording to Nimoy, he invented the hand gesture himself and adapted it from one used by Jewish priests while giving a blessing.",
"\n\n* * * * * * * * * *\n\nComments? ",
"Corrections? ",
"Post them on the Famous Quotations Facebook group .",
"\n\nFurther reading…"
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"Fox News has issued an apology to its viewers and Hillary Clinton after a guest compared the former secretary of state to “herpes” during a broadcast on Thursday.",
"\n\nFox News Guest Compares Hillary Clinton To ‘Herpes That Won’t Go Away’\n\nMORE\n\nFox News Guest Compares Hillary Clinton To ‘Herpes That Won’t Go Away’\n\nFox News has issued an apology to its viewers and Hillary Clinton after a guest compared the former secretary of state to “herpes” during a broadcast on Thursday.",
"\n\nFox News host Rick Leventhal who appeared visibly taken aback, issued the an on-air apology and cut off his guest, Anna Paulina, immediately after she called Hillary Clinton “herpes” who just won’t “go away” Full Article Here: https://www.buddylist.co/posts/fox-news-guest-compares-hillary-clinton-to-herpes-that-wont-go-away\n\n✅Please support the Tip Jar here: https://www.seekingthetruth.info/tipjar/\n\n👕☕Buy a T-Shirt or Coffee Mug https://teespring.com/stores/seekingthetruth\n\n☑️ https://www.patreon.com/truthseeker\n\n😎 Join us on Buddylist. ",
"A true free speech social network. ",
"https://www.buddylist.co\n\n🤑My Bitcoin Wallet Address: 17S5o2Thyqcw9TqtFyQAQdAtjSwJJ95rcE\n\n💻50% Off Coupon for Life Torguard VPN Promo code: truth101\n\n💻Torguard Offshore No Logs VPN Services: https://goo.gl/iBDGTj\n\nFor Push Notifications please Subscribe on my website. ",
"Go there and click the bell and you will get notified with every new video.",
"\n\nhttps://www.seekingthetruth.info\n\nJoin me on my new Social Network called Buddylist to discuss more. ",
"https://www.buddylist.co/share/Qu_na2NZKNS9FWro?utm_source=manual\n\nBefore and After the 2016 Election: https://www.youtube.com/watch?v=1z6VfcvMVig\n\n#SeekingTheTruth\n\nMy YouTube Channel: https://www.youtube.com/seekingthetruth101\n\nFind me on\n\nSteemit: https://steemit.com/@truthseeker101\n\nBitchute: https://www.bitchute.com/channel/seekingthetruth/\n\nBuddylist: https://www.buddylist.co/members/837095\n\nDLive: https://dlive.io/@truthseeker101\n\nMinds: https://www.minds.com/SeekingTheTruth101\n\nCopyright Disclaimer Under Section 107 of the Copyright Act 1976, allowance is made for \"fair use\" for purposes such as criticism, comment, news reporting, teaching, scholarship, and research. ",
"Fair use is a use permitted.",
"\n\n\"Fair Use\" guidelines: www.copyri.."
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"The MRI apparatus irradiates a high frequency wave on a body under examination (hereafter referred to as the subject) placed in a uniform static magnetic field to excite nuclear spins inside the subject, receives MR signals generated by the nuclear spins, and renders an image of the subject (imaging). ",
"Irradiation of the high frequency wave and reception of the MR signals are conducted by a resonance type RF coil.",
"\nGenerally, in imaging by the MRI apparatus, an MR signal of hydrogen nuclide (1H) is used. ",
"However, these days, there is multinuclear imaging whereby MR signals from plural kinds of nuclides, such as fluorine (19F), phosphorus (31P), and sodium (23Na), in addition to hydrogen nucleus, are detected and are put into images. ",
"The multinuclear imaging is used in the case where an in-body distribution of a component nuclide contained inside the body other than hydrogen is intended to be grasped, in the case where a contrast medium having fluoride is intended to be visualized, and the like.",
"\nIn the multinuclear imaging, when MR signals from plural kinds of nuclides are detected with a resonance type RF coil, the resonance frequencies of the RF coil must be tuned in to frequencies of the MR signals of the respective nuclides. ",
"For example, in the case where an MRI apparatus of 3T imaging two kinds of nuclides, hydrogen and fluoride, the resonance frequency of the RF coil is tuned in to 127.8 MHz for hydrogen and to 120.3 MHz for fluoride, respectively.",
"\nFor this reason, the multinuclear imaging uses the following two methods: a plurality of RF coils having different resonance frequencies that are tuned in to respective frequencies of the MR signals of a plurality of nuclides that are intended to be detected; and a single RF coil capable of changing the resonance frequency in response to the nuclide to be detected. ",
"The RF coil that is used in imaging in which two kinds of nuclides are considered as objects to be detected, which is a main stream in the multinuclear imaging, and that is capable of changing the resonance frequency includes: (1) A coil utilizing a fact that a birdcage type RF coil has a plurality of resonance modes (e.g., see Gerald B. Matson et al., “",
"A Practical Double-Tuned 1H/31 P Quadrature Birdcage Headcoil Optimized for 31P Operation,” Magnetic Resonance in Medicine, pp. ",
"173-182, 1999); (2) A coil changing its resonance frequency using a variable capacitor; (3) A coil changing its resonance frequency by an arrangement of a capacitor and an inductor (a double tunable RF coil); (4) A coil changing its resonance frequency by varying the distance between an RF shield and the RF coil (e.g., see Dongfeng Lu and Peter M. Joseph, “A Technique of Double Resonant Operation of F-19 and H-1 Quadrature Birdcage Coils, ” Abstracts of Society of Magnetic Resonance in Medicine, p. 531, 1990, and JP-A-Hei7(1995)-155307); etc."
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"AMD Ryzen Threadripper Unboxing and Installation\n\nPosted by Nathan Kirsch| Thu, Aug 03, 2017 - 4:05 PM\n\nWe wanted to post a good video showing our unboxing of the AMD Ryzen AMD Threadripper processor, but our attempts at a good unrehearsed video without proper lighting was a fail. ",
"So, we apologize to all the millennials as our unboxing coverage is mostly going to be images and text that you actually have to read.",
"\n\nWe were expecting a board and processor like normal, but we ended up receiving two large boxes that tipped the scales at 55 pounds! ",
"This was not going to be your average board and processor! ",
"One boxed included a large pelican case with the AMD Ryzen Threadripper logo and a plaque that custom engraved for the site. ",
"The other box included an ASUS ROG Zenith Extreme X399 motherboard ($549.99), Thermaltake Flow Riing 360 TT Premium Edition CPU liquid cooler ($168.99), two G.Skill TridentZ RGB DDR4 16GB 3200MHz memory kits ($207.37 each), Samsung 960 Pro PCIe NVMe SSD ($279.99) and a Thermaltake Toughpower Grand 1200W power supply ($247.87). ",
"This is nearly $1,650 of hardware that is needed to get an AMD Ryzen Threadripper system up and running! ",
"You still need the storage drive, graphics card/s, mass storage drive, keyboard, mouse, display and of course the operating system of choice.",
"\n\nInside the other box we found three AMD Ryzen Threadripper processors! ",
"We had the Ryzen Threadripper 1950X 16-core, 32-thread processor ($999) and then the Ryzen Threadripper 1920X 12-core, 24-thread processor ($799). ",
"Both of these processors are launching on August 10th, so we while we can show you the processors we can’t show you any performance numbers other than Cinebench. ",
"The third and final processor was actually in a display case with some custom engraving to the site. ",
"The opening of the pelican case was pretty neat as AMD placed lighting behind the processors and the light coming through the packaging looked great.",
"\n\nHere is the AMD Ryzen Threadripper processor that we were given for display on our bookshelf of goodies. ",
"We pulled off the AMD Phenom II Black Edition TWKR edition and AMD FX belt buckle to show off some swag from years past.",
"\n\nIf you’ve ever owned an AMD Athlon X64, Phenom, Phenom II or a processor along those lines you’ll appreciate the image above as it just goes to show how large this new processor is. ",
"The AMD Ryzen Threadripper processors have 4,094 pads on the bottom of the processor that line up with the pins on the AMD TR4 socket.",
"\n\nHere is a look at the bottom of the AMD Ryzen Threadripper processor next to AMD FX Processor belt buckle. ",
"The scorpion is on there as that was the codename of the platform for AMD Bulldozer.",
"\n\nOpening up the AMD Ryzen Threadripper processor is an experience unlike any other we’ve had before with a processor. ",
"You can tell AMD put a ton of time and effort into making this a fun user experience! ",
"You first have to rip off the cardboard that is looped around the packaging. ",
"You then remove the packing halves. ",
"In the lower section you’ll find the warranty details, case badge sticker, T20 Torx tool that is set for proper torque and a water cooling mount for Asetek water coolers. ",
"You then need to take apart the center section that holds the processor. ",
"Once a tension spring is removed you can slide the processor out. ",
"Note that the processor resides in a bright orange carrier frame and it needs to remain in this orange frame!",
"\n\nNow that the AMD Ryzen Threadripper processor is taken apart you can install it into the motherboard. ",
"We have nine images in the gallery above that visually show you the installation process into the ASUS ROG Zenith Extreme X399 motherboard. ",
"We had no issues installing the processor and having to screw things down instead of flipping levers is no big deal. ",
"This method takes a little longer, but it feels like you are doing something important and is a rewarding experience. ",
"The correct torque setting is 1.6nm (14.2 in-lbs) on all three screws, but luckily the included torque wrench is set for that. ",
"We covered the installation processor in a news post last week, so we won’t go into super detail on it.",
"\n\nOnce you get the processor all mounted up you can put on your CPU cooler of choice. ",
"Right now there are 25 liquid and air cooling options and we’ve listed them all below. ",
"You may notice that the head or block of the thermal solution does not completely cover the integrated heat spreader (also known as processor lid) of the Ryzen Threadripper. ",
"This is acceptable because base still covers both silicon chips under the Threadripper heat spreader. ",
"To ensure best performance, please ensure the thermal paste applied to the head spreads evenly and completely across the surface of the head to use the full contact area. ",
"More coolers are being designed for this platform, but these will all do for now!"
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"DNA a atacat cu contestaţie în anulare decizia Curţii de Apel Alba Iulia prin care a fost anulată, în 9 februarie, condamnarea de trei ani de închisoare cu suspendare pentru abuz în serviciu în cazul fostului secretar general al CJ Hunedoara, Dan Daniel.",
"\n\nPurtătorul de cuvânt al Curţii de Apel Alba Iulia, Cosmin Muntean, a declarat, joi, corespondentului MEDIAFAX, că instanţa ”a respins ca nefondată” contestaţia în anulare depusă de DNA.",
"\n\nProcurorul şef al DNA, Laura Codruţa Kovesi, a declarat, în februarie, că DNA va ataca cu o contestaţie în anulare decizia Curţii de Apel Alba prin care a fost anulată o condamnare.",
"\n\n„După apariţia deciziei am analizat motivarea, am decis să o atacăm cu contestaţie în anulare. ",
"Este o decizie izolată. ",
"Am studiat practica judiciară şi este o soluţie izolată. ",
"În cursul anului trecut, după decizia CCR, au fost multe soluţii de condamnare pentru abuz, peste 60 de persoane au fost condamnate. ",
"CCR nu a dezincriminat infracţiunea de abuz în serviciu. ",
"Nu avem de a face cu o dezincriminare. ",
"Atacăm cu o contestaţie în anulare. ",
"Dacă vom găsi şi alte decizii, vom sesiza procurorul general cu un recurs în interesul legii. ",
"Pentru motivul invocat în contestaţia la executare, pentru aceleaşi motive, în situaţii similare, contestaţiile la executare au fost respinse”, a declarat procurorul şef al DNA.",
"\n\nÎn data de 9 februarie, Curtea de Apel Alba Iulia a anulat condamnarea de trei ani de închisoare cu suspendare pentru abuz în serviciu în cazul fostului secretar general al Consiliului Judeţean Hunedoara, Dan Daniel. ",
"Judecătorul a considerat că în cauza pe care a judecat-o a apărut „o dezincriminare a faptei” ca urmare a deciziei CCR. ",
"Mai exact, decizia a fost dispusă ca urmare a unei măsuri extraordinare de atac, contestaţia la executare, făcută după ce procesul în apel s-a terminat fiind dispusă o decizie definitivă. ",
"Ulterior sentinţei finale, Daniel Dan a contestat decizia, invocând decizia CCR.",
"\n\nAstfel, DNA a folosit o altă cale extraordinară de atac, contestaţia în anulare care se poate face, printre alte situaţii, „când împotriva unei persoane s-au pronunţat două hotărâri definitive pentru aceeaşi faptă.”",
"\n\nJudecătorii de la Alba au explicat decizia susţinând că a intervenit o modificare a elementelor constitutive ale infracţiunii, ca urmare a deciziei CCR privind abuzul în serviciu.",
"\n\n„Având în vedere conţinutul infracţiunii prevăzute de art.297 alin.1 C.pen., ",
"Curtea de Apel Alba Iulia constată că, urmare a deciziei de neconstituţionalitate menţionate, practic, a intervenit o modificare a elementelor constitutive ale acestei infracţiuni, sub aspectul laturii obiective, fiind limitat domeniul de aplicare al acestei infracţiuni prin excluderea tuturor cazurilor în care norma a cărei încălcare este imputată subiectului activ al infracţiunii nu este o lege, o ordonanţă sau o ordonanţă de urgenţă. (..) ",
"Nu se poate considera (aşa cum a apreciat prima instanţă) că decizia nr.405/2016 este doar o decizie de interpretare, instanţa constituţională admiţând prin decizia menţionată o excepţie de neconstituţionalitate”, se arată în motivarea instanţei.",
"\n\nDan Daniel a fost condamnat în primă instanţă în 2015 într-un dosar care a vizat infracţiuni de serviciu comise la organizarea concursului pentru postul de director al Direcţiei Generale de Asistenţă Socială şi Protecţia Copilului Hunedoara."
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"There aren't many Pokemon games outside the Nintendo handheld consoles, and I didn't like Pokemon Go. ",
"But now there appears to be a change in strategy at Nintendo. ",
"The first Pokemon game for the Switch, Pokemon Quest, was just released, and several more Pokemon games, some more similar to the classics, have been announced.",
"\n\nI tried Pokemon Quest, and the first thing I saw was the start screen that said \"Tap to start\". ",
"Tap? ",
"It turned out that was an omen: Pokemon Quest plays horribly with a JoyCon controller, it is designed for touch screens. ",
"However the Switch isn't a great mobile handheld console, due to very limited battery life. ",
"So my first thought was \"why don't they bring this out for my iPad instead\". ",
"It turns out that this is exactly what they are going to do end of this month. ",
"I wonder if I can link my progress from the Switch to the iOS.",
"\n\nPokemon Quest is very much designed like a mobile game: It is Free2Play and has the usual Gacha collection mechanic, as well as energy running out after a number of expeditions. ",
"However the game isn't overly pushy with its monetization, and can be played reasonably well without paying anything. ",
"The real time combat requires some getting used to, I prefer the classic turn-based combat. ",
"But I'll play this until the \"Pokemon Let's Go\" games come out, which are supposed to bring back the gameplay of Pokemon Yellow.",
"\n\nI think the Switch has been improving batteries in newer models...I picked one up a few months back, and on a recent trip observed it ran for five hours of intense gameplay without issue while on a trip before needing a recharge."
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"Hepatocyte growth factor protects hepatoblastoma cells from chemotherapy-induced apoptosis by AKT activation.",
"\nHepatocyte growth factor/scatter factor (HGF) is a ubiquitously expressed molecule that elicits pleiotropic functions on epithelial cells, including mitogenic, motogenic, differentiating, angiogenic and morphogenic effects. ",
"In hepatoblastoma (HB), post-operative residual tumor growth and tumor recurrences are often associated with markedly elevated serum levels of HGF, suggesting a link between this molecule and tumor malignancy. ",
"Here, we demonstrate that HGF has no impact on overall cell viability and proliferation of HB cells, although signal transduction occurs downstream of HGF, such as c-Met phosphorylation, activation of phosphoinositide 3-kinase (PI3K)/AKT and mitogen-activated protein kinase (MAPK)/ERK-1/2 signaling. ",
"Instead of being mitogenic, HGF confers anti-apoptotic properties upon serum starvation and moreover protects HB cells against strong apoptotic inducers such as cisplatin and camptothecin, thereby contributing to chemotherapeutic resistance. ",
"This effect is mainly dependent on the PI3K/AKT signaling pathway, since inhibition by wortmannin resulted in abrogation of HGF-mediated survival, whereas inhibition of the MAPK pathway had no effect. ",
"Together, these findings highlight the importance of HGF in tumor cell survival and suggest that HGF and its cognate receptor c-Met should be considered as a candidate for combined therapeutic strategies of advanced pediatric liver tumors."
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"How My Best Friend Rania Crashed a Party and Saved the World\n\nBy Ada Hoffmann\n\nIllustration by Bryan Prindiville\n\nWhatever You Say I Am by Bryan Prindiville\n\nRania Mehanna was my best friend, so when I bumped into her crying in the stairwell at lunch hour, I was stupid enough to rush right to her. ",
"She freaked out and shook her head, trying to brush her eyes off all clumsy. ",
"Rania didn’t cry much, and she didn’t have the hang of it.",
"\n\nShe hadn’t picked the spot at random, of course. ",
"She’d crumpled up in one of the blind spots where the school security cameras don’t quite see. ",
"She didn’t want the world to know she was crying. ",
"World Savers don’t cry alone in corners. ",
"I had to be careful not to blow her cover.",
"\n\nEveryone knows this stuff. ",
"But when your best friend is hunched over sobbing, basic stuff slips your mind.",
"\n\nI stopped, then grinned and hopped over the rest of the way, like I was just happy to see her. ",
"Rania glared at me, but she stopped trying to jam her fists into her eyes.",
"\n\n“Hey,” I said.",
"\n\n“Hey, Emma.”",
"\n\n“You want to be alone?”",
"\n\nShe hesitated. “",
"No.”",
"\n\nI slid down beside her. “",
"What’s up?”",
"\n\n“Nothing.” ",
"She shook her head.",
"\n\n“‘Kay, then. ",
"I got us movie tickets. ",
"Two seats. ",
"Rania Mehanna and Emma Cruz for Catgirls vs. Pterodactyls. ",
"Opening night. ",
"I mean, if you still want to go.”",
"\n\nShe sighed. “",
"Yeah, I guess.”",
"\n\n“With Suman Bachchan as the pterodactyl trainer.”",
"\n\n“Yeah whatever.”",
"\n\nIt wasn’t like Rania not to squeal over Suman Bachchan. ",
"She scowled even deeper when I said his name. ",
"Boy trouble, then. ",
"I lowered my voice. “",
"It’s Jacob, isn’t it?”",
"\n\nRania buried her face in her hands. “",
"He stood me up again. ",
"That’s all. ",
"I should be used to it by now.”",
"\n\n“Bastard,” I said.",
"\n\n“Like you’d know.”",
"\n\nI hated when she needled me about that. ",
"I never got into the “guys” thing. ",
"Or the “girls” thing. ",
"Dating wasn’t my scene. ",
"But I knew what it meant to care about people. ",
"I had friends I’d take a bullet for — Rania first. ",
"Even when she was giving me the death glare from Mars.",
"\n\nRania snorted and shoved me. “",
"That’s not a date with me. ",
"That’s a date with Suman Bachchan’s cheekbones.”",
"\n\nI shoved back. “",
"Then Stephen Hawking: Space Pirate. ",
"Same analogy.”",
"\n\n“Dude. ",
"I’m allowed to stand you up if the movie sucks.”",
"\n\n“Filmgate gave it 8 out of 10.”",
"\n\n“Filmgate is full of Numbers Fiends. ",
"They’ll give 8 out of 10 to any pile of crap with a space pirate.”",
"\n\n“So who says it sucks?”",
"\n\n“Moviedock.” ",
"Which was as full of World Savers as Filmgate was full of Numbers Fiends. “",
"No female characters in the whole movie. ",
"Not even a waitress or something. ",
"Just a bunch of white dudes with speech synthesizers blowing things up.”",
"\n\n“But you don’t have a problem with all the bikinis in Catgirls?”",
"\n\n“Yeah, well.” ",
"She waved me off. “",
"Suman Bachchan. ",
"Shut up.” ",
"Now she was smiling, at least. “",
"Did you hear the Infallible Cloud changed his label? ",
"He’s not a full World Saver anymore, just a World Saver slash Pleasure Seeker. ",
"Can you believe it?”",
"\n\nI shrugged. “",
"I don’t know the guy, except for his cheekbones.”",
"\n\n“Yeah. ",
"Stupid raters.” ",
"She pulled her knees up to her chest. “",
"Stupid everyone.”",
"\n\n“Rania,” I said, “how long has it been since you and Jacob talked?”",
"\n\nRania plopped her head down in her hands. “",
"A month now. ",
"I mean, I text him. ",
"I post on his wall. ",
"Nothing stalkery. ",
"But he’s all ‘Oh hey, I’m out someplace, talk to you later.’ ",
"I ask him to come and see me, but he ignores it.”",
"\n\n“So go see him anyway. ",
"He’s got GPS, doesn’t he?”",
"\n\n“You know why I can’t go see him.”",
"\n\nI did. ",
"Jacob was a Pleasure Seeker, all jokes and persuasion with nothing on the inside. ",
"Which was fine, as long as he and Rania were happy. ",
"But his dad had a new job and the family had moved to Brightside: one of those prissy gated communities in the suburbs, full of Upward Movers and Monocle Men. ",
"World Savers did not go to parties in Brightside. ",
"Rania couldn’t go to him without losing some serious personality points. ",
"And Jacob wouldn’t go to her.",
"\n\n“So don’t go see him. ",
"Set your AllBook status to Single. ",
"It’s not like he deserves any better.”",
"\n\nShe snapped back up straight. “",
"World Savers do not break up online. ",
"That’s a sign of poor empathy and lack of respect for your partner, and if you don’t even respect the person you’re dating, how can you—”\n\nI raised a hand. “",
"Stop. ",
"Respect goes both ways, remember? ",
"Right now you don’t owe him any. ",
"Besides, everybody’s got little marks on their record. ",
"Even the Pope.”",
"\n\n“Easy for you to say. ",
"You’re a Relator. ",
"You can make friends in any cesspool of privilege you like.”",
"\n\n“Sorry,” I said. “",
"It sucks. ",
"You’re right.”",
"\n\nShe closed her eyes and breathed in and out.",
"\n\n“I wish I was a hacker,” she said. “",
"I wish I could just go see him, cuss him out, break things off, and then erase it all from the system and still be a World Saver and get into pre-med at Harvard. ",
"I’d do that.”",
"\n\nI wanted to tell Rania that even the rudest breakup in the world wouldn’t stop her from being a World Saver. ",
"She was the real thing: single-handedly raising thousands of dollars for World Hacks one month, volunteering around the world in another, and keeping her grades up high the whole time. ",
"She didn’t quit: her only problem was that she couldn’t set out to fix her personal life with the same zeal as fixing everything else.",
"\n\nBut Rania had her eye on some picky colleges. ",
"The kids who got in had records spotless like you wouldn’t believe. ",
"These were kids who never went through a Delinquent or Self Destructor phase, not even something semi-respectable like Beta Girl. ",
"They’d been World Savers since they were two years old and their parents posted about how fast they’d learned the alphabet. ",
"I had a theory those kids weren’t the actual best World Savers, but they looked great on paper. ",
"By those standards, one mark on Rania’s record was a pretty big deal.",
"\n\nI knew those problems went way deeper than what I could deal with. ",
"I knew I should sit tight, let Rania cry on my shoulder, validate her feelings. ",
"But all of a sudden I had a bright idea.",
"\n\n“Do you want it that bad, Rania? ‘",
"Cause I’ve got a lot of Numbers Fiends on my friends list.”",
"\n\nParker Austin[Upward Mover] wrote\nNot according to her profile. ",
"Dude, you haven’t met her at all this month. ",
"Love you to prove me wrong, but twenty bucks says you’re keeping her around for the personality points.",
"Yesterday at 4:25 pm\n\nI didn’t know a thing about sneaking past the Infallible Cloud. ",
"But I had two thousand people on my friends list, give or take. ",
"Maybe forty of them were Numbers Fiends, and twenty of those were local. ",
"AllBook would log anything we said online, so I had some face-to-face schmoozing to do.",
"\n\nI had plans that evening already, of course. ",
"Bowling night with my friends at yearbook committee. ",
"So I typed out a bunch of heartfelt apologies and cancelled those. ",
"Then I got to trolling for Numbers Fiends. ",
"Gita Jha, my favourite, was out of town, and the next two had their schedules bricked right up, but I got a hit with number four: Deborah Hendrix, a frizzy-haired girl I played video games with every couple of months. ",
"Numbers Fiends aren’t picky: a quick query if she wanted to hang out tonight, and I had her.",
"\n\nDeborah had the kind of house that smelled like old pizza boxes and chips, with her parents holed up in home offices all night, but she led me all perky through the mess to the den. ",
"In here, there was enough clear floor to move around. ",
"Three of the walls were plastered with wide-format screens, and a patched-up maroon sofa leaned against the fourth. ",
"Deborah took out an old-fashioned handheld remote.",
"\n\n“The arm controls are broken,” she apologized, pointing at the sofa’s raggedy arm. “",
"You know how it is. ",
"I got a new platformer, though. ",
"With alien battles! ",
"You still like those?”",
"\n\n“Sure.”",
"\n\nWe clicked our way through the login and into the new game. ",
"Pictures popped up on the walls: a maze of golden walkways, gears, and ramps over a steaming green sea. ",
"We ran in place, and our avatars on the big central screen ran through the maze—or the maze moved around them. ",
"We jumped from one walkway to another, grabbed power-ups, sometimes pushed each other into the water to see what would happen. (",
"Answer: You die.) ",
"We dodged the flocks of flying 8 balls. ",
"On the first big stairway, with a mean-looking alien at the top, I moved one of my arms wrong and the stairs folded up into a slide. ",
"My avatar fell all the way down and landed in a green bubble-blowing machine, which swelled me up and popped me into nonexistence.",
"\n\nDeborah giggled. “",
"Oh, man! ",
"I missed that — I never died that way. ",
"Try it again.”",
"\n\nI held my arms out straight and tried to do the biggest steps I could up the staircase, but I couldn’t get it right. ",
"By the fourth bubble-popping death we were both hunched over laughing on the floor.",
"\n\n“You suck at this,” said Deborah.",
"\n\n“I’ll try again when the room stops spinning.” ",
"The alien on the wall fidgeted, and a return to last save point? ",
"window popped up, but I ignored them. “",
"Hey, while I was here, I wanted to ask your help with something.”",
"\n\n“Yeah, sure.” ",
"Deborah waved me aside, still giggling. “",
"You’ve got to get to the top of the stairs, though. ",
"The alien battles are awesome. ",
"They’ve got all these tentacles and you have to — Wait, what do you need help with?” ",
"She scowled. “",
"I am so tired of people using me as a math tutor. ",
"I like you because you spend time with me, not my textbooks. ",
"If this visit is about math, I swear—”\n\n“No, no. ",
"Not math. ",
"Do you know Rania Mehanna?”",
"\n\n“Sorta.”",
"\n\n“She and I want to crash a party.”",
"\n\nI already knew it had to be a party. ",
"Showing up at his house would be creepy, and his parents would see us.",
"\n\nDeborah raised a bushy eyebrow. “",
"A party? ",
"And you asked me?”",
"\n\n“It’s not just any party. ",
"It’s in Brightside. ",
"If the Infallible Cloud gets word Rania went to a party in Brightside, it’ll lower her World Saver rating. ",
"She’s extra worried about her rating. ",
"Wants to get into Harvard, you know. ",
"But her boyfriend’s in Brightside and she needs to talk to him. ",
"So we need to get to the party, do what needs doing, and get out again, without anyone knowing. ",
"And for that, we need expert help. ",
"You in?”",
"\n\nDeborah scowled. ",
"She didn’t have the best dating life either. ",
"Mostly she’d gone for the boys — and a few girls — who asked for her help with their math homework, despite the way she complained about them, and they tended to dump her in the spring when math class ended. “",
"Why’s Rania dating someone in Brightside?”",
"\n\n“I don’t know. ",
"Animal magnetism. ",
"Four months ago they did Model UN and she babbled nonstop about the way he swaggered around pretending to be the Russian Federation. ",
"Then all of a sudden they were dating. ",
"He didn’t used to be in Brightside, you see.”",
"\n\n“Oh.”",
"\n\n“But now he’s standing her up all the time and he won’t change his status back to Single.”",
"\n\n“Oh, and she can’t change it herself without a face-to-face meeting. ",
"I’ve heard of that.” ",
"Deborah tugged on her hair, thinking it over. “",
"Yeah, I guess it’s worse than not dating. ",
"You sure you have to go into Brightside to find him? ",
"He never goes out to the movies or something?”",
"\n\n“Yeah, he goes to the movies. ",
"At the Brightside Cinema. ",
"They’ve got everything in there, trust me. ",
"Logic says he’ll run off on vacation at some point, but that could be months, and even then he might not go anywhere Rania can reach him. ",
"He might head straight to the airport and jet off to Tuvalu. ",
"So to get to him, we’ve got to go in.”",
"\n\nDeborah thought it over. “",
"Okay,” she said at last. “",
"I’m in. ",
"On one condition.”",
"\n\n“Yeah?”",
"\n\nShe grinned and sat up straight, which swung her avatar’s head right into an oncoming 8 ball. “",
"I want to go to the party.”",
"\n\n--\n\nCelebrity Sues Over “Fraudulent” Personality Rating\n\nThe Infallible Cloud is not so infallible according to Sadie Clare Wolfe, star of Electric Mermaid and Babes of the Apocalypse. ",
"When her personality rating changed from Upward Mover to Beta Girl, Wolfe sued the six major rating consortiums, calling the decision “fraudulent” and claiming that the change would cost her hundreds of thousands of dollars in lost fan revenue.",
"\n\nThe rating consortiums stand firmly behind their product, saying the change to Beta Girl was simply an honest algorithmic assessment of her behaviour. ",
"But Wolfe and her legal counsel claim the models behind personality ratings are subtly flawed. ",
"Two other B-list celebrities, Sophie Xu and Dimitra Panagiotopoulos, have filed suit on similar grounds.",
"\n\nThe furore has led some to speculate that Suman Bachchan, the heartthrob star of Catgirls vs. Pterodactyls, may also speak up. ",
"Bachchan, noted for his philanthropy and charity advocacy but also a regular at Hollywood parties, was recently reclassified as a World Saver / Pleasure Seeker, rather than simply a World Saver.",
"\n\nAt press time, Bachchan’s agent had not responded to inquiries.",
"\n\n--\n\nRania’s house was the opposite of Deborah’s: it looked like something out of a furniture ad. ",
"Cozy, mind you, with cushions and blankets over the armchairs, and it smelled like cinnamon and honey from the kitchen. ",
"But Rania’s mom had a fit if anyone so much as got crumbs on the floor.",
"\n\nDeborah was already sprawled out on the carpet by the time I got there. ",
"Rania was perched on the edge of the couch. ",
"She looked worried that I hadn’t found someone better. ",
"I had to introduce them and convince Deborah to sit on a chair like a normal person. ",
"Then we got down to business.",
"\n\n“No. ",
"Trust me, it does GPS even if you turn it off. ",
"Leave it here. ",
"Same with whatever else you’ve got. ",
"Tablets, book reader, music, even your ID. ",
"And no ultra-good clothes. ",
"Secondhand is best. ",
"Nothing with an RFID.”",
"\n\nRania nodded slow.",
"\n\n“That’ll stop the Infallible Cloud from tracking you. ",
"Next thing we have to do is block face recognition.”",
"\n\nI raised an eyebrow. “",
"They’ve got face recognition at the door?”",
"\n\nDeborah sighed like she was talking to idiots. “",
"People are taking pictures at this party, right? ",
"And posting them? ",
"It’s not a Luddite party? ",
"Even if you stay out of trouble, you’re going to be in the background of someone’s photo. ",
"Soon as they post it, AllBook runs face recognition, auto-tags you, and links to your account. ",
"In case someone goes ‘Who was that girl?’ ",
"and wants to friend you. ",
"So we need to give you a face AllBook can’t identify. ",
"Got any interesting makeup?”",
"\n\nRania made a face. “",
"Can’t I just wear big sunglasses, or…”\n\n“Obvious disguise attempt is obvious. ",
"No. ",
"You want something that screws up the algorithms, but doesn’t look like you’re trying to screw the algorithms. ",
"Best thing is dressing you up like a World Hating Poet. ",
"They’re all drawing spiderwebs under their eyes, right?”",
"\n\n“Yeah.”",
"\n\n“So you wear your darkest, laciest clothes. ",
"I’ve got a lace trim hat you can borrow. ",
"And Emma paints you with the biggest, most asymmetrical spiderweb ever. ",
"Darken the cheekbones and nose, brighten it up above the eyes, and with any luck AllBook gets so confused it can’t tag you.”",
"\n\n“But I’m not a World Hating Poet,” said Rania.",
"”I can’t have that on my record.”",
"\n\n“My parents’ car.” ",
"My mom was a doctor; we weren’t Upward Movers, but we had enough money to park in their street, and taking the bus was too risky. “",
"I drive. ",
"We go in, find Jacob, Rania takes him aside, I stand lookout, and you enjoy the party. ",
"Everyone’s happy, right?”",
"\n\n“Right,” Deborah said fast, and Rania nodded, but that worry in her eyes was getting worse.",
"\n\nI chewed my lip. “",
"You want moral support?”",
"\n\n“And a witness. ",
"So he can’t blow it off.”",
"\n\n“Okay. ",
"Deborah, would you stand lookout?”",
"\n\nShe sat up straight. “",
"I wanted to see the party.”",
"\n\n“You can still see it. ",
"We’ll even wait around in the car when we’re done if you want extra time. ",
"You just need to stick with us for a little while and let us know about anything suspicious.”",
"\n\nDeborah raised her chin. “",
"Sure. ",
"I’ll do the social-skills stuff. ",
"See if I can’t.”",
"\n\n--\n\nSuman Bachchan [World Saver / Pleasure Seeker] wroteWore an old suit for Catgirls vs. Pterodactyls opening night instead of buying new. ",
"Donated the difference. ",
"Still look GREAT! ",
"Dunno why everyone’s still talking about the Pleasure Seeker label. ",
"Just a label, guys.",
"Yesterday at 12:31 pm | Comment | Like 1542 people like this\n\n--\n\nThe party was ridiculous fancy. ",
"Deborah’s gaming system was space-cool by my standards, but this was from another planet. ",
"Not just screens on the walls, but 3D projections outward and inward all over, making the whole place an ultra-real projected desert full of postcard dunes. ",
"The resolution was so good you could see through the walls, into the distance, where the dunes went on forever and ever. ",
"We would have all bumped into the walls if someone hadn’t projected pictures of railings on them.",
"\n\nLots of the guests were wearing imitation desert clothes to match. ",
"Girls with Jell-O shots tripped past us in headscarves and belly-dancer pants, and pale blond Monocle Men fiddled with their keffiyehs.",
"\n\nRania wrinkled her nose. ",
"Orientalism, she mouthed when I caught her eye, and I nodded. ",
"Ew. ",
"Rania’s mom wore a headscarf for real, and most of these people in fake hijab would’ve whispered and called her a terrorist outside this party. ",
"Too bad it would have wrecked our disguise if she said anything. ",
"I guess this was why World Savers didn’t go to these parties.",
"\n\nDeborah stared around at everything making fish faces.",
"\n\n“Ohmygod,” she whispered. “",
"Look, look, this is top-of-the-line early adopter stuff. ",
"Musta cost more than my parents’ car. ",
"And what’s th—”\n\nA robot camel the size of a Golden Retriever slammed into her. ",
"I winced. ",
"How much money did you have to waste to get custom robots just for one party? ",
"But Deborah made a wibbly face, like it was a kitten, and threw her arms around it. “",
"Ohmygod, Emma, look! ",
"It’s so realistic. ",
"Look how they did the joints, you can hardly tell it’s motors under there and not muscles. ",
"The fur, even. ",
"You’re not looking.”",
"\n\n“A little. ",
"Seen a few of them in Jacob’s AllBook photos. ",
"Never in person.”",
"\n\nWe sauntered around, avoiding the photo-ops, and Rania found a redheaded boy browsing the retro-cool paper bookshelf.",
"\n\n“Hey,” she said. “",
"You seen Jacob?”",
"\n\n“Yeah.” ",
"He pointed. “",
"Kitchen.”",
"\n\nIn the kitchen Deborah started bug-eyed grinning again. ",
"The place was like a posh cooking show, all sterile metallic lines and big spaces. ",
"A hot desert sun beat down above the oven, and a cool oasis powered the sink. ",
"A steel screen door led out to the back porch. ",
"Jacob and like twenty other people stood around talking beside it.",
"\n\nJacob had the most ridiculous outfit in the whole party. ",
"Not just a keffiyeh, but a whole costume from that godawful remake of The Sheik. ",
"Trying too hard. ",
"That would get him Beta Girl if he kept at it, but not Upward Mover.",
"\n\nRania started at him, then paused. “",
"If I go get him, people will realize it’s me. ",
"Is that going to be…?”",
"\n\n“Yeah, I’ll do it. ",
"You two wait outside.”",
"\n\nI sauntered in and spent a couple minutes joining the crowd, laughing at the jokes, pointing out movie references. ",
"Easy peasy. ",
"Jacob noticed me and frowned, but it looked like he didn’t want to make a scene, either.",
"\n\n“Hey, Jacob?” ",
"I asked after we’d exchanged a few not-so-bon mots.",
"\n\n“Yeah?”",
"\n\nI did a flirty head tilt. “",
"Wanted to talk to you outside for a bit. ",
"You mind?”",
"\n\nI never flirt. ",
"Jacob knew me well enough to know. ",
"But Jacob never didn’t flirt, and the others in the crowd grinned, poking him to follow me.",
"\n\nHe sighed. “‘",
"Kay, sure, whatever. ‘",
"Scuse me, guys.”",
"\n\nI led him out the screen door. ",
"There was a bigass back yard, the kind with a pool and a porch and a big dog-walking lawn full of cute stands of trees. ",
"He got pale when he saw Rania and Deborah. “",
"What is this, some kind of girl-mob?”",
"\n\n“Those trees.” ",
"I pointed. “",
"Over there.”",
"\n\n“You’re ganging up on me?”",
"\n\n“I’m ganging up on you,” Rania said. “",
"Emma is here as a witness. ",
"And Deborah… is here.”",
"\n\n“Standing lookout,” said Deborah.",
"\n\n“Why are you dressed like that?” ",
"said Jacob.",
"\n\n“Long story,” I said. “",
"Let’s just get to the trees.”",
"\n\nHe gave me a nervous look, but he let Rania and me frog-march him into the trees, away from the splashing, chlorine-smelling pool crowd.",
"\n\n“First thing,” said Rania once we were out of sight. “",
"I’m not going to be seen talking to you in that.”",
"\n\nJacob couldn’t have taken off the whole costume without stripping, but he took off the headdress, at least. “",
"Nag, nag, nag. ",
"This is why I don’t call you more often. ",
"You’re so busy being a World Saver you can’t have fun.”",
"\n\n“Um,” he said. “",
"I mean, it’s not that simple. ",
"I was going to call you, it’s just that things keep coming up.”",
"\n\n“Then you do want to see me?”",
"\n\n“Um,” said Jacob.",
"\n\n“You see, the last six times you said you wanted to see me, we set a date and you never showed up. ",
"It kind of gives the impression you don’t. ",
"Isn’t that right, Emma?”",
"\n\nI shrugged. ",
"I knew a script when I heard one. “",
"Sounds right to me.”",
"\n\nJacob scowled. “",
"Okay, maybe I don’t.”",
"\n\n“Then we’re breaking up?” ",
"said Rania.",
"\n\nJacob paused.",
"\n\n“Look, you don’t know how it is for me. ",
"Everyone here’s been an Upward Mover for years. ",
"I’ve got to work quadruple to fit in. ",
"At least if I’ve got a girlfriend, that gives me something to talk about.”",
"\n\nI raised my eyebrows. “",
"You talk to them about Rania? ",
"There’s nothing to talk about, at the rate you two have been going out. ",
"Unless you’re making things up.”",
"\n\nI wondered if he was making things up. ",
"It was the sort of thing he’d do, and it might cost Rania more personality points than she thought. ",
"Boys like Jacob made up weird sex stories, or even crime stories, to get attention. ",
"That kind of gossip might change her label for real.",
"\n\n“Please,” said Jacob, ignoring me. “",
"If you find someone else, then sure, that’s cool, but why not keep our status at ‘Dating’ till then? ",
"For show? ",
"It would help my personality rating. ",
"And I know how you feel about your personality rating. ",
"I know you understand. ",
"I need this.” ",
"He reached out and rested a hand on her shoulder. “",
"Save the world for me a little, Rania, sweetie. ",
"Just for a while.”",
"\n\nRania wavered. ",
"She bit her lip.",
"\n\nI looked around quick, trying to squint through the tree needles and assess the danger. ",
"Clumps of people all over the backyard, and a big splash-war in the pool, but nobody dangerous close. ",
"Not even Deborah.",
"\n\nShe’d wandered off. ",
"I thought I saw her frizzy head in the distance, talking to someone. ",
"Flirting, to judge from the body language.",
"\n\nDuh. ",
"I should have seen it coming. ",
"Deborah might not go to another high school party ever. ",
"How could I expect her to stay in the trees, missing it all?",
"\n\n“I don’t know,” Rania said. “",
"Maybe…”\n\nI wanted to run and drag Deborah back, but Rania needed me here more. ",
"Her weak spot was weaker than I thought.",
"\n\n“You think it’s saving the world,” I said, “helping you swan around in those clothes? ",
"You think the world will improve one bit because you, Jacob Harrington, were an Upward Mover?”",
"\n\n“Well, sure,” said Jacob.",
"\n\nRania rallied. ",
"She raised her head a bit higher. “",
"And you think I owe that to you? ",
"You want me to give up my personality rating, my chances at a good college and a World Saver job, and most of my chances at finding someone better than you — all so, what, you can impress people at racist parties?”",
"\n\nJacob raised his voice. “",
"You think you can call me racist?”",
"\n\n“I think I can tell you,” Rania said, raising hers to match, “that World Saving has as much to do with this as pigs do with the International Linear Collider. ",
"And that you are no longer my boyfriend, as of now. ",
"You can put that on AllBook!”",
"\n\n“Because why?” ",
"He was shouting now. “",
"Because you’re on your high horse about the clothes at this party? ",
"Who even invited you? ",
"I didn’t!”",
"\n\n“I don’t have to explain anything to you!”",
"\n\n“Guys.” ",
"I was starting to freak out. ",
"Yelling like this was a good way to get noticed fast. “",
"Guys, can we quiet—”\n\nJacob shouted past me. “",
"It’s my love life, and you owe me an explanation! ",
"This is the worst thing about World Savers. ",
"You prance around deciding who’s worthy of help and who’s not, and you think you’re so good. ",
"But when an ordinary guy needs a little bit of slack—”\n\nRania wasn’t looking at me either. “",
"Don’t you tell me what World Savers are like. ",
"You’ve never been a quarter of the way to World Saver in your life!”",
"\n\n“And damned glad that I—”\n\nThere was a quick rustle in the branches, then a click and a flash.",
"\n\nI turned. ",
"A whole gaggle of Upward Movers in imitation hijab stood there giggling, snapping pictures with their phones.",
"\n\nRania turned the color of feta cheese.",
"\n\n“I’m going home,” she said in a strangled voice, and she rushed out of the trees.",
"\n\nI didn’t chase Rania down right away. ",
"She didn’t like to talk when she was that upset, and she wasn’t going to get far without a wallet or bus pass. ",
"Instead I pushed past the Upward Movers and ran to Deborah.",
"\n\nShe was flirting, all right. ",
"At least the trollop who’d distracted her was well-dressed. ",
"No costume, just a smart vest, boots, and slacks, with boyish bowl hair.",
"\n\n“Deborah,” I said through my teeth.",
"\n\nShe startled. “",
"Oh, hi, Emma. ",
"This is Caroline. ",
"We were just — I mean, is everything all—”\n\n“A horde of Upward Movers walked in on Rania and Jacob and me while you were gone. ",
"So no, everything is not all right. ",
"Thanks for asking.”",
"\n\nDeborah flushed bright. ",
"She knew what she’d done.",
"\n\n“Walked in?” ",
"said Caroline. “",
"What were they doing in there?”",
"\n\n“Breaking up,” Deborah moaned, burying her face in her hands.",
"\n\n“Why on earth would you need to break up with nobody watching? ",
"For that matter, if you needed to break up with nobody watching, why on earth would you try to do it at a party?”",
"\n\n“Long story. ",
"C’mon, Deborah.” ",
"I tugged Deborah’s wrist. “",
"We’ve got to take her home. ",
"You can look Caroline up on AllBook later.”",
"\n\n“But—”\n\nCaroline waved a vague hand. “",
"It’s all right. ",
"I’ll probably look you up before you even get home. ",
"Have fun.”",
"\n\n“Bye!” ",
"Deborah called as we hurried away.",
"\n\nWe found Rania pacing the sidewalk. ",
"It was a well-lit street, no traffic, but it looked like she’d remembered she didn’t have a bus pass or taxi money, and it hadn’t improved her mood.",
"\n\n“I’m sorry,” Deborah said. “",
"I’m really sorry!”",
"\n\n“Don’t talk to me,” Rania snapped. ",
"She had this apocalyptic redness under her eyes.",
"\n\n“I’ll drive you home now,” I said. “",
"I’ve got the keys. ",
"It’s going to be okay.”",
"\n\n“Don’t. ",
"Talk. ",
"To. ",
"Me.”",
"\n\nBut she got in the car.",
"\n\nThe ride was hot and silent until we dropped off Deborah. ",
"After that, Rania started throwing words like bricks.",
"\n\n“My life,” she said, “is ruined.”",
"\n\n“Maybe not.” ",
"I’d been trying to figure out what to say ever since starting the engine. “",
"Maybe you lost a couple of points. ",
"Maybe not going would have been worse, in the long run. ",
"Maybe his points would have rubbed off on yours.”",
"\n\n“I should have just changed my status to Single. ",
"I shouldn’t have let you talk me into this.”",
"\n\n“You did a good thing, Rania. ",
"You confronted someone who’d been hurting you. ",
"It would have worked if not for Deborah and the shouting. ",
"You were brave.”",
"\n\nI stared at her. ",
"She was my best friend. ",
"She had the biggest, most tempestuous heart. ",
"But she was no World Hating Poet. ",
"She didn’t do the “leaving forever!” ",
"thing lightly.",
"\n\nShe slammed the car door and stalked off.",
"\n\n--\n\nDeborah Hendrix[Numbers Fiend] wrote\nARGH. ",
"No wonder I never get invited to parties, I screw EVERYTHING up.",
"Yesterday at 11:01 pm | Comment | Like\n\n--\n\nCaroline Ward[Monocle Man / Numbers Fiend] wrote\nNonsense. ",
"138 people tonight by last count, incl. ",
"crashers. ",
"Each on average participates in dozens of activities. ",
"Hardly any of these screwed up, most not by you. ",
"Also enjoyed yr company. ",
"Thinking of mtg again at more intellectual venue. ",
"Shame about angry friends though. ",
"What happened there?Yesterday at 11:15 pm\n\nCaroline Ward[Monocle Man / Numbers Fiend] wrote\nAh. ",
"Received email. ",
"Thinking over reply. ",
"Possible I can find a way to help…Today at 12:02 am\n\n--\n\nSchool sucked. ",
"I mean, I had plenty to do: I passed notes in science class, giggled with the Beta Girls by my locker, organized up a storm with yearbook committee (with constant apologies for missing bowling night), and kicked butt at volleyball practice. ",
"But I kept seeing Rania out of the corner of my eye, slumped over her desk, puffy-eyed. ",
"Glaring when she caught my eye.",
"\n\nIt wasn’t all my fault, but she was hurting, and it had been my idea. ",
"I had to make it up to her. ",
"And groveling empty-handed wasn’t going to work. ",
"World Savers are about results.",
"\n\nI knew I had twenty-four hours: the Infallible Cloud’s average latency. ",
"The pictures of Rania at the party — yelling, red-faced, with a World Hating Poet spiderweb on her face — had gone up last night. ",
"The makeup must have worked, because AllBook didn’t automatic tag her. ",
"But Jacob sure as hell knew who she was, and he’d tagged the picture in the wee hours of the morning.",
"\n\n“What’s on your mind?” ",
"said my friend Isabella while we toweled off after volleyball. “",
"You’ve been out of it.”",
"\n\nIsabella wrinkled her nose. “",
"So it’s, what, one of those World Saver spats? ",
"She yelling at you?”",
"\n\n“No, she unfriended me.”",
"\n\n“Yikes.”",
"\n\nWhat could I do? ",
"I wasn’t Deborah, and even Deborah couldn’t hack a system like AllBook. ",
"I was a Relator. ",
"I just talked to people.",
"\n\nSo.",
"\n\nI was a Relator. ",
"I had two thousand AllBook friends. ",
"How many of them could I talk to in an evening?",
"\n\n--\n\nEmma Cruz[Relator] wrote\nMy friend Rania Mehanna is a World Saver. ",
"Top of every social science class. ",
"You should hear her talk about her fundraising for World Hacks or the volunteer trip she took to India. ",
"She’s ambitious. ",
"Wants to change things.",
"\n\nWe need World Savers because things need changing. ",
"Always will. ",
"We build the best systems we can, but one way or another we’ll always need better.",
"\n\nRania’s ex-boyfriend, Jacob, used the system against her. ",
"He wanted all the social points of a girlfriend and none of the responsibility. ",
"So when he moved to Brightside, an Upward Mover community, he stayed there. ",
"Rania couldn’t visit him without risking her World Saver points, and he refused to visit her. ",
"If she dumped him online, that would have risked her points too: let’s face it, breaking up via text is still a dick move.",
"\n\nSo with a couple of friends, Rania snuck into Brightside and stood up to him. ",
"She had Numbers Fiend advice for the sneaking. ",
"Turned out that wasn’t enough. ",
"Now Rania’s despondent because the pictures are up online and her World Saver label’s in danger.",
"\n\nBut World Saving shouldn’t just mean doing the right volunteer work and having the right opinions. ",
"World Saving is risky. ",
"Rania Mehanna had to choose between fixing her personal life and saving her reputation. ",
"That choice is itself unjust. ",
"But when it was reputation or justice, Rania picked justice.",
"\n\nThis is the principle World Saving works on, big or small. ",
"It’s not Big Oil or the rainforest, but this is what it looks like.",
"\n\nRania thinks the Infallible Cloud doesn’t agree. ",
"But the Infallible Cloud’s supposed to reflect what people think. ",
"Which is why I need you all to click “Like”, and repeat this yourself:\n\nRania Mehanna is a World Saver.",
"\nSpread the word.",
"\n\nToday at 4:36 pm | Comment | Like 1663 people like this\n\n--\n\nThe next day I couldn’t even get to my locker past the crowd of people saying:\n\n“Oh, wow, your post!”",
"\n\n“I wish I had the balls to post something like that.”",
"\n\n“Go Rania!”",
"\n\nJust like in the comments. ",
"So far, so good. ",
"But Rania pushed past them all and howled, “Emmmaaaa!”",
"\n\nShe hugged me and everyone crowded in.",
"\n\n“It was the least I could do.”",
"\n\n“No, no. ",
"Emma. ",
"When was the last time you looked at your AllBook feed?”",
"\n\nShe took a minute to get the words out, and by then other people chimed in.",
"\n\n“Oh. ",
"I think I saw this too.”",
"\n\n“Saw what?”",
"\n\n“Let her tell it, dude.”",
"\n\nRania took a huge breath and shrieked, “Suman Bachchan!”",
"\n\nAt which point everybody started cheering and talking so fast that it took like ten minutes to get out my phone and check AllBook. ",
"I thought I was going to lose Rania in all the high-fiving. ",
"But there it was:\n\n--\n\nSuman Bachchan [World Saver / Pleasure Seeker] wrote\nEveryone keeps asking why I’m not up in arms about the label change. ",
"Maybe I’ve got other things to worry about? ",
"Just maybe?",
"\nOn that note, a friend linked me to this.",
"\nGo read it, guys. ",
"She says it better than I could.",
"\nRania Mehanna is a World Saver.",
"Today at 8:19 am | Comment | Like 4324 people like this\n\n--\n\nI stared at my phone. ",
"I like to think I’m a supportive friend. ",
"But the first thing I thought was, “Oh, God. ",
"I’m going to be swamped.”",
"\n\nThen I started laughing.",
"\n\n“How the heck? ",
"How did that happen?”",
"\n\nRania made her way back to me. “",
"That stupid Monocle-Man-slash-Numbers-Fiend who was talking to Deborah. ",
"Her uncle knows Suman Bachchan’s dad. ",
"Can you believe it?”",
"\n\nI hugged her again.",
"\n\n“So we’re still friends?”",
"\n\n“Total friends. ",
"And we’re still on for Catgirls vs. Pterodactyls.”",
"\n\n“Awesome.”",
"\n\nShe hooked her arm in mine, and we marched to science class. ",
"Rania was what Rania was, and there was a lot of world left to save.",
"\n\nLike this:\n\nAda Hoffmann is a mild-mannered computer scientist by day and a writer by night. \"",
"How My Best Friend Rania Crashed a Party and Saved the World\" is her first foray into hard science fiction. ",
"Previous work has appeared in Strange Horizons, AE, Shimmer, and elsewhere. ",
"She also blogs about autism in speculative fiction. ",
"Find her at ada-hoffman.com or on Twitter at @xasymptote.",
"\n\nBryan Prindiville is currently an Art Director for Catholic Relief Services (CRS) where he has also worked as a designer and illustrator since late 2000. ",
"In his free time he has had a hand in a number of webcomics including Bassetville and Hello with Cheese. ",
"Traditionally published work can be found in Rum and Runestones, Tee Morris' All a Twitter, and others. ",
"Less traditionally he can be found as a member of the live art entertainment show Super Art Fight. ",
"More information and work are available at his sketch blog, bryanprindiville.com.",
"\n\nSTEP RIGHT UP! ",
"GET YER CLOWNS HERE!",
"\n\nClowns: The Unlikely Coulrophobia Remix is hot off the presses! ",
"So get your hot, pressed clowns today! ",
"Or if you prefer your clowns cold-pressed, never fear. ",
"A clown is nothing if not adaptable."
] | {
"pile_set_name": "Pile-CC"
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"Living Like More Than A Conqueror – Part 2\n\nWe have been given the ability to choose to be more than a conqueror, and at times we don’t have to apologize for it.",
"\n\nRomans 8:37 reiterates the fact that we are more than conquerors through Christ who loved us. ",
"We have to live continually with the awareness that we have been predestined to win and conquer. ",
"However, our positional authority to win lies solely on our faith. ",
"Matthew 21:22 says – whatever things you ask in prayer, believing, then you shall have it. ",
"Believing as a conqueror is the way to live.",
"\n\nAs we renew our minds with the promises of God in His Word, then we gradually begin to build the mindset of a conqueror – who is capable of doing what other people cannot do.",
"\n\nGideon was an example of a champion with a renewed mindset. ",
"God called him to reduce the crowd of soldiers he had to fight the Midianites. ",
"A bunch of soldiers who were predominantly laden with fear. ",
"So he sent about 22,000 men back home who were not fit for the war as champions.",
"\n\nThen of the few that were remaining, Gideon still had to thin down the\n\ncompany to a handful of 300 men – with the strength of champions. ",
"And with the 300 men, against thousands from the enemy’s camp – they brought victory to the camp of Israel. ",
"All they did was chant “the sword of the Lord and of Gideon,” and the enemy’s camp fled into oblivion. ",
"God picks the illogical of men and substances to bring Him victory and all of the glory.",
"\n\nGod is, however, looking for people who will not argue with His methods and would just execute His divine strategy to the letter – to bring in the victory just to the glory of His name.",
"\n\nJoshua was also a leader, champion and conqueror who used the words of his mouth by faith to stop the sun and the moon, and used them to his advantage in positional authority as a conqueror.",
"\n\nThe Creator of the sun and the moon therefore listened to Joshua for a win!",
"\n\nThe only set-back champions have is fear. ",
"When they have courage, then they become conquerors.",
"\n\n…excerpts of Pastor Dawn’s sermon as penned by the stylus of a ready scribe! (",
"Ps. ",
"45:1)"
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"China to Triple Nuclear Power Capacity by 2026\n\nThe Three Mile Island Nuclear Plant is seen in the early morning hours March 28, 2011 in Middletown, Pennsylvania. (",
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"\n\n\"The pace of nuclear power projects in recent years is not in accordance with this target, and it's necessary to make sure the nuclear industry is back on track,\" He Yu, China General Nuclear Power's chairman, said.",
"\n\nAnalytics firm BMI Research noted that if the project is realized, China will overtake the United States with the most nuclear power. ",
"The country is allocating $570 billion to build more than 60 nuclear power plants in the next 10 years.",
"\n\nAside from that, China also plans to produce 58 gigawatts of nuclear power into operation by the end of the decade, doubling its current 27 gigawatts capacity. ",
"It also plans to add more and bring 150 gigawatts by 2030. ",
"And by 2050, it plans to more than double its nuclear power capacity to 350 gigawatts, allocating over a trillion dollars in nuclear investment.",
"\n\nChina's nuclear ambitions have faced repeated delays, including a suspension of approval for three years from 2011 as it performed safety procedures following Japan's Fukushima disaster. ",
"If production of third-generation nuclear reactors are not sped up, China would likely fail to meet its plan to decrease emissions.",
"\n\nThe US is currently expecting to produce 100 gigawatts of nuclear power in 2030. ",
"A gigawatt of power could provide energy to about 700,000 homes.",
"\n\nAs of now, China has 30 nuclear reactors, accounting for 2.5 percent of its electricity supply. ",
"It plans to build 24 more and increase the construction of a large commercial scale reprocessing plant.",
"\n\nMeanwhile, the world's nuclear capacity is expected to increase by 60 percent by 2040."
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" COURT OF APPEALS\n SECOND DISTRICT OF TEXAS\n FORT WORTH\n\n NOS. ",
"02-15-00093-CV\n 02-15-00094-CV\n 02-15-00095-CV\n\n\nIN THE MATTER OF S.L.T.\n\n\n ------------\n\n FROM COUNTY COURT AT LAW NO. ",
"1 OF DENTON COUNTY\n TRIAL COURT NOS. ",
"JV-2014-00246, DH-2014-0616, JV-2014-00509\n\n ------------\n\n MEMORANDUM OPINION1 AND JUDGMENT\n ------------\n\n We have considered “Appellant’s Motion For Voluntary Dismissal.” ",
"It is the\n\ncourt’s opinion that the motion should be granted; therefore, we dismiss the\n\nabove appeals. ",
"See Tex. ",
"R. App. ",
"P. 42.1(a)(1), 43.2(f).",
"\n\n Costs of the appeals shall be paid by appellant, for which let execution\n\nissue. ",
"See Tex. ",
"R. App. ",
"P. 42.1(d).",
"\n\n PER CURIAM\n\nPANEL: GARDNER, WALKER, and MEIER, JJ.",
"\n\nDELIVERED: April 9, 2015\n 1\n See Tex. ",
"R. App. ",
"P. 47.4.",
"\n\f"
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"Integrated care for people with long-term mental and physical health conditions in low-income and middle-income countries.",
"\nIntegrated care is defined as health services that are managed and delivered such that people receive a continuum of health promotion, disease prevention, diagnosis, treatment, disease management, rehabilitation, and palliative care services, coordinated across the different levels and sites of care within and beyond the health sector and, according to their needs, throughout the life course. ",
"In this Review, we describe the most relevant concepts and models of integrated care for people with chronic (or recurring) mental illness and comorbid physical health conditions, provide a conceptual overview and a narrative review of the strength of the evidence base for these models in high-income countries and in low-income and middle-income countries, and identify opportunities to test the feasibility and effects of such integrated care models. ",
"We discuss the rationale for integrating care for people with mental disorders into chronic care; the models of integrated care; the evidence of the effects of integrating care in high-income countries and in low-income and middle-income countries; the key organisational challenges to implementing integrated chronic care in low-income and middle-income countries; and the practical steps to realising a vision of integrated care in the future."
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"WAKEFIELD, Mass. - The Women’s Hockey East Association and the College of the Holy Cross announced today that Holy Cross has accepted an invitation to join the conference as its 10th member school. ",
"The Crusaders will begin play as a member of Women’s Hockey East in the 2018-19 season with games being played on the Holy Cross campus in the newly renovated Hart Center at the Luth Athletic Complex.",
"\n\n“It is always exciting when we are able to grow our membership ranks and particularly so with an institution of such unique prestige and character as Holy Cross,” said Hockey East Commissioner Joe Bertagna. “",
"We look forward to joining forces with the Crusaders to bring the Hockey East brand to Central Massachusetts and its great hockey fans.”",
"\n\nWomen’s Hockey East began as a six-member conference with Boston College, the University of Connecticut, the University of Maine, the University of New Hampshire, Northeastern University, and Providence College competing in the inaugural 2002-03 season. ",
"The conference has added a total of three teams on two occasions prior to today; Boston University and Vermont joined the league in 2005, while Merrimack College became the most recent addition in 2015.",
"\n\nBoth the Terriers and Warriors joined Women’s Hockey East as first-time varsity programs. ",
"Boston University has gone on to win five WHEA Tournament championships and appear in two NCAA title games, while Merrimack reached the playoffs in 2016-17 in just its second season in the conference.",
"\n\nTake a look at our accomplishments from this season!#RiseTogetherhttps://t.co/Gb1B2Szqe2 — Holy Cross W. Hockey (@HCrossWHockey) March 2, 2017\n\n“This is a historic day for the College of the Holy Cross and for our women’s ice hockey program,” said Holy Cross Director of Athletics Nathan Pine. “",
"I’d like to thank Commissioner Bertagna and the Women’s Hockey East membership for their confidence in Holy Cross and for this wonderful opportunity to join their ranks. ",
"We are committed to competing for championships in Hockey East, and I’m confident all of our constituencies will be excited for this important move.”",
"\n\nHoly Cross first fielded a varsity women’s ice hockey team in 1999-2000, and has won five ECAC East/New England Hockey Conference Open Tournament titles throughout its history – in 2009, 2010, 2011, 2015, and 2016. ",
"The Crusaders have posted a winning record in each of the last 11 seasons, including a combined mark of 46-8 over the past two years (22-5 in 2016-17; 24-3 in 2015-16).",
"\n\n“This is a great day for our Holy Cross women’s ice hockey family,” said Holy Cross head coach Peter Van Buskirk. “",
"The leadership on our campus has shown a great commitment to the future of our program, and everyone from our current student-athletes to our dedicated alumni base is looking forward to our new home in Hockey East.”"
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"\nThe value of repeat hepatectomy for intrahepatic recurrence after curative resection of neuroendocrine liver metastasis (NELM) remains unclear. ",
"The aim of this retrospective cohort study was to determine the significance of repeat hepatectomy for recurrent NELM. ",
"Patients who underwent hepatectomy for NELM between 1994 and 2016 were identified. ",
"The indications for a first hepatectomy were adequate liver remnant volume and no extrahepatic metastasis. ",
"The diagnosis of recurrent NELM was based on radiographic examinations. ",
"The indications for a repeat hepatectomy were the same as those for the first hepatectomy. ",
"Clinicopathologic factors, short-term survival, and long-term survival were investigated using clinical records. ",
"Forty-four patients enrolled in this study. ",
"Thirty-three patients among them underwent a curative hepatectomy, and 28 of them developed recurrence. ",
"Of them, 16 patients underwent a repeat hepatectomy. ",
"The overall survival of the repeat hepatectomy cohort (n = 16) was significantly better than that of the no repeat hepatectomy cohort (n = 12) (P < .001). ",
"The progression free survival after the first hepatectomy (n = 44) and that after repeat hepatectomy (n = 16) were similar (P = .546). ",
"No significant difference was seen between the frequency of major complications (Clavien-Dindo score ≥ 3a) after the first and repeat hepatectomy (P = .279). ",
"No repeat hepatectomy (hazard ratio [HR] 5.0, P = .036) was identified as an independent predictive factor of a poor outcome among the recurrent cohort, along with the presence of multiple nodules (HR 26.2, P = .008) and a CA19-9 level ≥40 U/mL (HR 11.2, P = .012). ",
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"How to Choose a Reliable Online Blackjack Casino\n\nOne of the interesting things about the internet gambling is that you are offered the chance to choose from a wide array of online casinos. ",
"A significant majority of these casinos offer players (potential players) the chance to try themselves out in blackjack games. ",
"What this means, is that you need to very careful when selecting your casino; you need to consider the features of the casino as well as other important information on the casino.",
"\n\nCheck out our blackjack guide below, as it can be very useful for you when choosing the casino that will suit your needs.",
"\n\nReputation Is Everything\n\nOf course you want to hurriedly register and start playing, but a few minutes to check through the casino you are registering with will not take your game away. ",
"One very important factor to consider is the reputation of the casino. ",
"Now, the question you have on your mind is most likely – how will I know if the casino is reputable? ",
"Not so hard, simply check their licenses and look at the sort of gaming authority license they have. ",
"A truly reputable company will definitely list their its licenses on its website. ",
"This way you can check and see whether they can render to you the type of service you consider to be “the best online blackjack experience.”",
"\n\nAre They a Newbie-Friendly Platform?",
"\n\nWhether you run a business or you are learning a skill, whatever you find yourself doing, you started from somewhere. ",
"There will always be newbies in every aspect of life. ",
"As you are looking to start your online blackjack playing experience, it is highly recommended you start with a casino company that appreciates newcomers. ",
"The best online casinos have formed the habit of giving newcomers the chance of using play money to test their platform and see if their gaming software suits them or not.",
"\n\nYou will always find casinos offering special bonuses exclusively for new players. ",
"They offer various incentives that will entice new players to their platforms, from welcome bonuses to deposit bonuses and lots more, so be sure to get yourself the best deal available.",
"\n\nWhat Payment Methods Are Included?",
"\n\nPlayers who love the online blackjack game can make their payments through several payment platforms. ",
"Ensure you check the available payment options the platform you want to choose allows, and be sure you can use those platforms before choosing them. ",
"It is very advisable that you go with a platform that allows you to make instant deposits and withdrawals, so as to facilitate smooth conduction of transactions between you and your casino of your choice.",
"\n\nWhat Game Variations Are Available?",
"\n\nAfter you have taking time to peruse over the technical capabilities of the platform, what you need to consider next is the variety of games the platform has on offer for players. ",
"As earlier mentioned that there are several blackjack modes, it will be best if you register with a casino that offers you enough playing options, so that when you try them, you will know the one(s) that suits you the best.",
"\n\nWhile we are still on the topic, bear in mind that you should adjust your strategy based on the variation of the game. ",
"The Blackjack chart can be a great source of help for whichever version you pick, as it gives you a fair amount of tactical options you can employ for the game."
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"Bait and Switch\n\nGoodfella: Jack Nicholson (left, with Matt Damon) chews the scenery as a mob boss in The Departed.",
"\n\nNo studio director was a greater hero to the Hong Kong new wave than Martin Scorsese. ",
"John Woo dedicated The Killer to him; Wong Kar-wai modeled his first feature, As Tears Go By, after Mean Streets; Taxi Driver's rain-slicked slo-mo urban stylistics worked their way into countless lesser HK films. ",
"With The Departed, Scorsese returns the compliment striving to bring it all back home by remaking the strongest Asian action film of the past few years.",
"\n\nAndrew Lau and Andy Mak's 2002 Infernal Affairs has a scenario so excellent it's amazing it didn't appear until movies entered their second century (and readers are invited to let me know if it had): A powerful mob boss grooms a follower from childhood to join the police even as the police plant an undercover agent within the boss's crime family. ",
"Each organization realizes that it has been infiltrated, but it's not until the movie approaches its climax and after each mole has been enjoined to, in effect, find himself that the deeply buried parallel informants figure out each other's identity.",
"\n\nIts key dramatic roles assumed mainly by cell phones, Infernal Affairs is one of those rare movies in which the premise is the star; Scorsese, however, has necessarily packed his remake with names. ",
"Matt Damon plays the rogue cop (Andy Lau's role in the original), with Leonardo DiCaprio as his undercover counterpart (embodied with soulful vulnerability by Tony Leung). ",
"Towering over both youngsters, Jack Nicholson has the meaty and here vastly inflated role of the patriarchal crime boss. ",
"Eric Tsang stole Infernal Affairs with his high-spirited moonfaced malevolence; Nicholson is handed the keys to the kingdom in his first scene.",
"\n\nThe Departed is a wildly commercial project, but let no one imagine it a work for hire. ",
"The opening burst of rock 'n' roll mayhem flashback street fight scored to the Rolling Stones is instant Scorsese. ",
"Transposing the action to the mean streets of South Boston, the filmmaker gets to refresh his stock in trade: The foul-mouthed protagonists are cops rather than wiseguys and, as the gangsters are Irish, the colorful ethnic invective can encompass Italians. ",
"The Deuce is here the Combat Zone; the resident, porn-haunted madman is a crime boss; the John Ford movie that's quoted isn't The Searchers but, inevitably, The Informer (it's shown on TV). ",
"Catholic guilt, however, is a constant.",
"\n\nInfernal Affairs was surprisingly cool and effectively restrained for HK action, but Scorsese raises the temperature with every ultra-violent interaction. ",
"The surplus of belligerence and slur reach near-Tarantino-esque levels appropriate as he's staking a claim to QT's turf. ",
"Screenwriter William Monahan (responsible for last year's Ridley Scott crusades spectacle Kingdom of Heaven) provides some choice insults: \"Hey, go save a kitten in a tree, you fuckin' homos,\" Damon taunts his rivals in a police-fireman football game. ",
"No opportunity is missed to call a priest a pederast. (",
"In addition to a flair for the schoolyard bon mot, Monahan's major contribution is combining the spies' two love interests into one very, very nervous shrink played by Vera Farmiga thus creating the possibility for yet another betrayal.)",
"\n\nThe film craft which is to say, the camera placement and Thelma Schoonmaker's slam-bang editing is first-rate, at least initially. ",
"Running two and a half hours, The Departed is 50 minutes longer than the original; the set pieces are more baroque than suspenseful; and the texture overwhelms the premise. (",
"By the time of the not-even-climactic mondo bloodbath car-crash shootout, the story's a sodden mess.) ",
"Farmiga's role notwithstanding, psychology is primitive: Damon's rogue cop starts to crack under the strain of his charade as DiCaprio's volatile, Valium-gobbling fake mobster grows increasingly anxious. (",
"DiCaprio's character is never sufficiently motivated, but his stolid non-performance is ultimately more affecting than Damon's one-note interpretation.) ",
"In a movie characterized by broad accents and broader characterizations, Mark Wahlberg has the distinction of delivering the least nuanced, most sympathetic (and funniest) turn, as the mouthiest cop on the force.",
"\n\nOf course, the Damon and DiCaprio characters are supposed to be acting. ",
"Not so Nicholson's, but never to be ignored, the star skews crazier and crazier, appearing with stage blood up to his elbows, playing pointless drunk scenes, pulling whimsical rat faces, reprising his satanic role in The Witches of Eastwick after a night at the opera. (",
"For all the sulfur, Ray Winstone is far scarier as the mobster's second in command.) ",
"Scorsese has a long history of burdening films with unpleasant and even atrocious central performances, and Nicholson seems bent on twirling the mustache off Daniel Day-Lewis' heavy in Gangs of New York a role that really belonged to producer Harvey Weinstein.",
"\n\nNeither a debacle nor a bore, The Departed works but only up to a point, and never emotionally even if the director does contrive to supply his version of a happy ending. \"",
"I don't want to be a product of my environment,\" Nicholson boasts at the onset. \"",
"I want my environment to be a product of me.\" ",
"Yeah, yeah, and that's the problem. ",
"Overwrought as The Departed may be, it's nothing that wouldn't have been cured by losing Jack (and maybe half an hour). ",
"Too bad the bottom line meant Scorsese had to sell that hambone Mephistopheles his soul."
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"Bill Gates recently bought the rights to a series of lectures by legendary Caltech physicist Richard Feynman. ",
"The former Microsoft head’s purchase shows that the cultural and scientific legacy of Feynman remains strong even 21 years after his death.",
"\n\nThe lectures, given in 1964 as part of Cornell University’s Messenger Lecture Series, were filmed by the BBC, who had retained the rights since. ",
"Gates purchased the lectures for an undisclosed amount.",
"\n\nBut what would the former Microsoft head want with the copyright to lectures by the revered physicist? ",
"In a recent interview with the CERN Bulletin, Gates said that his only plan is to make the footage freely available to the public.",
"\n\nAdd to that Gates’ reverence for Feynman, and it makes sense. ",
"The lectures are only the latest addition to Gates’ personal collection of Feynman-related material, which includes original manuscripts of some of Feynman’s best known work. ",
"Read more\n\nRadiolab has started a new season and you owe it to yourself to go check out their newest show on Stochasticity.",
"\n\n“Stochasticity, is just a wonderfully slippery and smarty-pants word for randomness. ",
"How big a role does randomness play in our lives? ",
"Do we live in a world of magic and meaning or … is it all just chance and happenstance? ",
"To tackle this question, we look at the role chance and randomness play in sports, lottery tickets, and even the cells in our own body. ",
"Along the way, we talk to a woman suddenly consumed by a frenzied gambling addiction, two friends whose meeting seems purely providential, and some very noisy bacteria.”"
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"Incidence and risk factors of toxoplasmosis in a cohort of human immunodeficiency virus-infected patients: 1988-1995. ",
"HEMOCO and SEROCO Study Groups.",
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"It increased from 0.7 per 100 person-years in 1988 to 2.1 per 100 person-years in 1992, as a result of the increasing prevalence of patients with CD4 cell counts below 200/microL. It decreased thereafter to 0.2 per 100 person-years in 1995, while the proportion of patients receiving specific prophylaxis was increasing. ",
"A Toxoplasma antibody titer of >150 IU/mL was an important predictor of toxoplasmosis (adjusted relative risk [aRR], 3.6 [95% confidence interval, 2.1-6.0]), independent of a CD4+ cell count of <200/microL (aRR, 20.8) and specific prophylaxis (aRR, 0.2 [0.1-0.3]). ",
"The median CD4+ cell count was 389/microL at the time the antibody titer was first noted to be >150 IU/mL, while the median CD4 cell count at onset of toxoplasmosis was 58/microL. Thus, disease was diagnosed 10 days to 74 months after the rise in Toxoplasma antibody titers. ",
"While the risk factors for development of toxoplasmosis remain incompletely defined, the importance of specific prophylaxis for patients with low CD4 cell counts and high Toxoplasma antibody titers is supported by these findings."
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"Estación Rancagua\n\nEstación Rancagua, is a railway station of the Empresa de los Ferrocarriles del Estado in Rancagua, Chile. ",
"It is the main railway station in the Libertador General Bernardo O'Higgins Region. ",
"Estación Rancagua is part of the Red Sur EFE, the TerraSur inter-city service, and the Metrotrén commuter service stops here.",
"\n\nLines and trains \nThe following lines and trains pass through or terminate at Estación Rancagua:\n\nRed Sur EFE\nTerraSur inter-city service (Alameda - Estación Chillán)\nMetrotrén commuter service (Alameda - Estación San Fernando)\n\nAdjacent stations\n\nExternal links \n Empresa de los Ferrocarriles del Estado\n Metrotrén\n Terrasur\n\nRancagua\nCategory:Buildings and structures in O'Higgins Region\nCategory:Transport in O'Higgins Region"
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"A Study of Pell Grants in Alabama\n\nBelow you will find two reports pertaining to the impact of Pell Grants in the state of\nAlabama. ",
"The first report, A Study of Pell Grants in Alabama, is a narrative describing the impact of\nPell Grants on students at Alabama’s public universities and colleges over the last several years. ",
"The\nreport then goes on to discuss how recent restrictions placed on Pell eligibility will impact students in the\nfuture. ",
"The initial report is 20 pages long.",
"\n\nThe second part of the report, The Impact of Pell Grants at Alabama Public Universities and\nTwo-Year Colleges: Technical Report, provides a breakdown, institution by institution (in alphabetical order by\nfour-year and two-year institutions), of the findings. ",
"A word of caution: the Technical Report is approximately\n130 pages long. ",
"You may want to be selective in printing the second attachment.",
"\n\nBelow you will find links to the two reports. ",
"Earlier versions of these reports contained errors. ",
"Please discard\nany version of these two reports in which the date November 26, 2012 does not appear on either the footer of\nA Study of Pell Grants in Alabama or on the cover page of The Impact of Pell Grants at Alabama Public\nUniversities and Two-Year Colleges: Technical Report."
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"Gavdari-ye Mohammad Qoli Rusta\n\nGavdari-ye Mohammad Qoli Rusta (, also Romanized as Gāvdārī-ye Moḩammad Qolī Rūstā) is a village in Sefidar Rural District, Khafr District, Jahrom County, Fars Province, Iran. ",
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"awesome build..... what did u use to illuminate ur bubble shifter...... i have that same one in my RA43 celica\n\ni made it. ",
"i use a juice bottle cap and glued in LED strip shooting upward. ",
"and hard wired it to a lighter adaptor. ",
"make sure you buy the LED strip(ebay) with same voltage as the lighter adaptor. ",
"i cut up my old cellphone lighter charger.",
"\n\nawesome build..... what did u use to illuminate ur bubble shifter...... i have that same one in my RA43 celica\n\ni made it. ",
"i use a juice bottle cap and glued in LED strip shooting upward. ",
"and hard wired it to a lighter adaptor. ",
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"Objectives were to assess associations between overnight HR, lying patterns and RFI, and between acute stress HR and RFI. ",
"Heifer calves (n=107; 408±28 days of age, 341±42.2 kg) and yearling heifers (n=36; 604±92 days of age, 539±52.2 kg) were exposed to a performance test to determine productive performance. ",
"Overnight HR (electrode based) and lying patterns (accelerometer based) were monitored on a subgroup of heifer calves (n=40; 20 lowest RFI; 20 highest RFI). ",
"In the 10-min acute stress assessment, all heifers were individually exposed to the opening and closing of an umbrella and HR before (HRBEF), in response to (HRMAX), after (HRAFT) and change (HRCHG; HRAFT-HRBEF) as a result of exposure were determined. ",
"Using polynomial regression, rate of HR decrease pre-exposure (β 1) and rates of HR increase (β 2) and decrease (β 3, β 4) post-exposure were determined. ",
"Heifer calves in the overnight assessment were classified into equal RFI groups (low RFI; high RFI) and HR means were treated as repeated measures and compared using multiple regression. ",
"In the acute stress assessment, heifers were classified within cattle category into equal RFI groups (low RFI; high RFI) and means and polynomial regression parameters were compared using multiple regression. ",
"Low-RFI heifer calves had a lower overnight HR (69.2 v. 72.6 BPM), similar HR change from lying to standing intervals (8.9 v. 9.2 BPM) and similar time lying (61.1% v. 64.5%) compared with high-RFI heifer calves. ",
"Low-RFI heifer calves had a higher absolute HRMAX (162.9 v. 145.7 BPM) and β 2 (-0.34 v. -0.20) than high-RFI heifer calves. ",
"Low-RFI yearling heifers had similar acute stress HR means and a lower β 1 (0.003 v. 0.006) than high-RFI yearling heifers. ",
"Overnight HR and acute stress HR are potential indicators of RFI in heifer calves. ",
"However, acute stress HR results varied in yearling heifers, suggesting previous handling experience and/or age influence stress response. ",
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"-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en.",
"\n\n-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en.",
"\n\n-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en.",
"\n\nOnce there was loving couple travelling in a bus in a mountainous area. ",
"They decided to get down at some place.",
"\n\nAfter the couple got down at some place the bus moved on. ",
"As the bus moved on, a huge rock fell on the bus from the mountain and crushed the bus to crumbs. ",
"Everybody on board was killed. ",
"The couple upon seeing that, said, \"*We wish we were on that bus*\" Why do u think they said that ?",
"\n\nScroll down for answer...\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\n....\n\nANSWER :\n\n*If they had remained on the bus instead of deciding to get down, the resulting time delay could have been avoided and the rock would have fallen after the bus had passed ..!!!*",
"\n\n*Think positive in life always and look for opportunities when u can help others..\n\n....Many times in life, the opposite of Success is not Failure, its\n\nQuitting. ",
"Winners never quit, quitters never Win....*\n\n\"Win as if you were used to it, lose as if you enjoyed it for a change...\"\n\n-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en.",
"\n\n-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en.",
"\n\n-- You received this message because you are subscribed to the Google Groups \"Samsongroup\" group. ",
"To post to this group, send email to samsongroup@googlegroups.com. ",
"To unsubscribe from this group, send email to samsongroup+unsubscribe@googlegroups.com. ",
"For more options, visit this group at http://groups.google.com/group/samsongroup?hl=en."
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"I had to block another family member and unapologetically shared with another, who is willing to listen. ",
"To be rejected by family hurts, especially parent and children, Since I came out as gay, I have thought several times of taking my own life, the rejection cuts deep into my heart and soul, but knowing Christ and believing in an awesome God has been a better option.",
"\n\nI am truly not worried as the inclusive christian ministry with House of Rainbow brings me in contact with many people who are same gender loving, thousands of people around the world just like me. ",
"Above all I have encountered God with unimaginable love.",
"\n\nTo be able, on a daily basis tell gay people God love us just the way we are is a humbling experience and I thank God for calling me.",
"\n\nGAY means God Adores You, God Accepts You, God Affirms You.",
"\n\nJeremiah 1:5 Before I formed you in the womb I knew you, and before you were born I consecrated you; I appointed you a prophet to the nations.’",
"\n\nPsalm 139:14 I praise you because I am fearfully and wonderfully made; your works are wonderful, I know that full well. ",
"I am not ashamed to be gay, I regret not knowing better before I got married to a woman in 1991, thank God I was divorced by 1994.There are many millions of stories only if we pay attention. ",
"I thought I will share this, I apologise if this is uncomfortable. ",
"But I needed to share. ",
"God bless you. ",
"Jide Macaulay"
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"Critical role of miR-125b in lipogenesis by targeting stearoyl-CoA desaturase-1 (SCD-1).",
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"Stearoyl-CoA desaturase-1 (SCD-1), a key enzyme in lipogenesis, catalyzes the conversion of SFA to MUFA, and inhibition of its activity impairs lipid synthesis. ",
"As posttranscriptional regulators, microRNAs are involved in many pathways of lipid metabolism; however, their effect on SCD-1 has not been reported. ",
"In this study, miR-125b was identified as a potential regulator of SCD-1 using bioinformatics analysis. ",
"Here, we validated SCD-1 as the target of miR-125b using a dual luciferase assay. ",
"During adipogenesis, a synthetic mimic or inhibitor was used to overexpress or reduce the expression of miR-125b in porcine adipocytes. ",
"Overexpression of miR-125b reduced the accumulation of lipid droplets and triglycerides concentration and repressed SCD-1 protein expression and MUFA composition. ",
"The inhibitor had the reverse effect. ",
"Small interfering RNA against tested in adipocytes further proved the direct correlation between miR-125b and SCD-1. ",
"Moreover, in vivo experiments in mice showed that injection of miR-125b expression vector decreased the hepatic triglycerides concentration relative to saline. ",
"This study indicated that miR-125b regulates lipogenesis by targeting SCD-1; therefore, miR-125b might be applied in therapy of lipid metabolism disorders."
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"Q:\n\nMatrix Averaging of duplicate Rows\n\nI've got a cvs file with 2 columns (see below Matrix 1 for example). ",
" I would like to create a program to average the second column of the matrix for all the duplicate numbers in the first column. ",
" So for instance in the matrix below there are two rows of \"2\" in the first column. ",
" Those rows would be averaged into one column of ((356+456)/2 = 406) etc. ",
" So the final matrix would like the Matrix 2 at the bottom. ",
" Any ideas on how to do this? ",
" \nMatrix 1\nmat1 <- structure(c(1, 2, 2, 3, 4, 4, 4, 5, 234, 356, 456, 745, 568, \n 998, 876, 895), .Dim = c(8L, 2L))\nmat1\n [,1] [,2]\n[1,] 1 234\n[2,] 2 356\n[3,] 2 456\n[4,] 3 745\n[5,] 4 568\n[6,] 4 998\n[7,] 4 876\n[8,] 5 895\n\nMatrix 2\nmat2 <- structure(c(1, 2, 3, 4, 5, 234, 406, 745, 814, 895), .Dim = c(5L, 2L))\nmat2\n [,1] [,2]\n[1,] 1 234\n[2,] 2 406\n[3,] 3 745\n[4,] 4 814\n[5,] 5 895\n\nA:\n\nWhat about\nas.matrix(aggregate(mat1[,2],by = list(mat1[,1]),FUN = mean))\n\n"
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"Question:\n\nWhat is a snap switch?",
"\n\nSnap switches are a kind of switch designed to be activated with a specific type of action, either snapping to the left or right or by a spring operated snapping mechanism. ",
"In order to activate the circuit (or deactivate it depending on how the snap switches are installed) a specific type of force and motion is required. ",
"Snap switches are often used in panels for door locks, motor control, timers and other functions. ",
"They are designed so that accidental activation of the circuit is much less likely due to the motion required to activate the unit.",
"\n\nWith an ordinary push button or switch, accidental contact may cause the circuit to activate; with snap switches the accidental contact doesn't equal unintended results unless the force and direction of the contact are equivalent to intentional activation. ",
"There are three kinds of general-use snap switches; one type made specifically for AC, another combining AC and DC capabilities.",
"\n\nEver wonder how you could make your life better and more fun? ",
"So have tens of millions of fans who have turned to LifeTips for answers over the past decade. ",
"We keep the tips, advice, books, podcasts and writing services flowing, so you can keep your life and business growing in the right direction. ",
"Upward!"
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"AD ASTRA\n\nPG-13 | 2 hours 3 minutes • Advance Tickets\n\nClosed Captioning, Descriptive Audio\n\nReading Cinemas\nManville with TITAN LUXE\n\nAbout\n\nAstronaut Roy McBride (Brad Pitt) travels to the outer edges of the solar system to find his missing father and unravel a mystery that threatens the survival of our planet. ",
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"Miami (AFP) - A major glacier in northeastern Greenland is rapidly crumbling into the Atlantic Ocean and experts warned on Thursday the breakup will likely raise global sea level by 18 inches (a half meter).",
"\n\nThe glacier, known as Zachariae Isstrom, \"entered a phase of accelerated retreat in 2012\" tripling the pace of melting. ",
"It is now losing mass at a rate of five billion tons per year, said the study in the journal Science.",
"\n\n\"The shape and dynamics of Zachariae Isstrom have changed dramatically over the last few years,\" said lead author Jeremie Mouginot, an assistant researcher in the Department of Earth System Science at the University of California, Irvine.",
"\n\nThe glacier is dumping \"high volumes of icebergs into the ocean, which will result in rising sea levels for decades to come,\" he said.",
"\n\nThe findings are based on 40 years of satellite data from global space agencies as well as aerial surveys monitoring the shape, size and position of glacial ice over time.",
"\n\nWarmer ocean water is eroding the glacier from below, and increasing air temperatures are melting it from above.",
"\n\n\"Zachariae Isstrom is being hit from above and below,\" said senior author Eric Rignot, professor of earth system science at UCI.",
"\n\n\"The top of the glacier is melting away as a result of decades of steadily increasing air temperatures, while its underside is compromised by currents carrying warmer ocean water, and the glacier is now breaking away into bits and pieces and retreating into deeper ground.\"",
"\n\nAnother large glacier near Zachariae Isstrom -- known as Nioghalvfjerdsfjorden -- is also melting but not as quickly because it is protected by an inland hill.",
"\n\nThe study said the two glaciers \"make up 12 percent of the Greenland ice sheet and would boost global sea levels by more than 39 inches if they fully collapsed.\""
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[
"Background {#Sec1}\n==========\n\nOver the last years, the development of novel antibody-drug conjugates (ADCs) has been a trending topic in cancer therapy \\[[@CR1]\\]. ",
"This technology uses an antibody to deliver a cytotoxic drug selectively to a tumor cell population by targeting tumor-associated receptors. ",
"There are several reasons driving the development of antibody-drug conjugates for cancer treatment. ",
"One feature is that a humanized or human antibody bears very high specificity and affinity towards its cancer specific antigen \\[[@CR2]\\]. ",
"Another feature is that some of the most potent non-targeted chemotherapeutics such as maytansine have extensive side effects in patients \\[[@CR3]\\], which intuitively could be reduced by targeting the drugs to their intended site of action. ",
"However, the transition of antibody-drug conjugates into the clinic would benefit greatly from non-invasive precision predictors that could potentially allow for patient stratification and early evaluation of effectiveness. ",
"Patient stratification according to the expression of membrane-bound antigens typically requires invasive biopsy via needle aspiration of multiple tissue regions to overcome tumor heterogeneity. ",
"In addition, decisions regarding patient management are often made by using archived biopsy results that do not necessarily reflect antigen expression at the time of treatment.",
"\n\nMirvetuximab soravtansine (IMGN853) is an antibody-drug immunoconjugate that consists of a humanized monoclonal antibody (M9346A) targeting folate receptor alpha (FRα)-positive cancer cells \\[[@CR4]\\] attached to a highly potent cytotoxic maytansinoid, DM4 (Fig.",
" [1](#Fig1){ref-type=\"fig\"}) \\[[@CR5]\\]. ",
"Although IMGN853 has been evaluated in a phase 1 expansion clinical study \\[[@CR6], [@CR7]\\] and is currently being evaluated in the Forward1 (NCT02631876) phase 3 clinical study, it is noteworthy that IMGN853 is not the only folate receptor targeting therapeutic approach currently evaluated in clinical trials \\[[@CR8], [@CR9]\\]. ",
"Nonetheless, the antibody, with its high specificity and affinity towards FRα, has limited distribution in normal human tissue \\[[@CR10], [@CR11]\\] and mainly targets the overexpression in epithelial ovarian cancer and non-small cell lung cancer \\[[@CR12], [@CR13]\\]. ",
"After endocytosis, the ADC releases its toxic payload inside the cell via lysosomal degradation of the antibody and disulfide reduction of the sulfo-SPDB linker. ",
"Following binding of DM4 to the microtubules, the targeted cancer cell undergoes mitotic arrest and cell death.",
"Fig. ",
"1Concept of an antibody-based companion diagnostic for cancer therapy using positron emission tomography (PET) imaging. ",
"The humanized antibody, M9346A, functions as targeting vector for antibody-drug-conjugate as well as companion diagnostic\n\nIn order to improve the drug development process of novel therapeutic agents, it is becoming more and more important to concurrently develop companion diagnostics \\[[@CR14]--[@CR16]\\]. ",
"With this in mind, the aim of this study was to develop and evaluate an ^89^Zr-labeled antibody derived from the FRα-targeting antibody, M9346A, as an immuno-positron emission tomography (immuno-PET) companion diagnostic agent for the antibody-drug conjugate IMGN853 that is based on the same humanized antibody (Fig.",
" [1](#Fig1){ref-type=\"fig\"}). ",
"In order to develop a M9346A-based PET imaging agent, we asked the following questions: (1) Is it possible to achieve the immunoreactivity after modification with desferrioxamine (DFO) as a chelator and radiolabeling with Zr-89? (",
"2) Is this antibody-based PET imaging agent able to delineate ovarian cancer xenografts? (",
"3) Does the pharmacokinetics of the radiopharmaceutical match with the pharmacokinetic profile of the antibody-drug-conjugate? (",
"4) Can we predict antibody-drug conjugate success using the novel companion diagnostic?",
"\n\nMethods {#Sec2}\n=======\n\nMaterials {#Sec3}\n---------\n\nUnless otherwise stated, all chemicals and solvents were used without further purification. ",
"Water used for this study was ultrapure (\\> 18.2 MΩcm^−1^ at 25 °C). ",
"Phosphate-buffered saline (PBS) as well as cell growth medium was purchased from the Media Preparation Facility at Memorial Sloan Kettering Cancer Center (MSK) (New York, NY). ",
"Humanized monoclonal antibody recognizing FRα and M9346A, as well as antibody-drug-conjugate IMGN853 were provided by ImmunoGen, Inc. (Waltham, MA) and further purified via a PD10 desalting column (GE Healthcare). ",
"Concentrations of solutions containing antibody were determined by using a NanoDrop™ 2000 spectrophotometer from Thermo Fisher Scientific (Waltham, MA). ",
"The bifunctional chelator *p*-isothiocyanatobenzyl-desferrioxamine (DFO-Bz-NCS) was purchased from Macrocycles (Plano, TX). ",
"^89^Zr-oxalate was acquired from 3D Imaging, LLC (Maumelle, AR). ",
"HPLC reactions (1.0 mL/min, phosphate-buffered saline) were performed on a Shimadzu UFLC HPLC system equipped with a DGU-20A degasser, a SPD-M20A UV detector, a LC-20AB pump system, and a CBM-20A communication BUS module using a size exclusion column (GE Superdex™ 200, 10/300 GL).",
"\n\nFunctionalization and radiolabeling of antibody M9346A {#Sec4}\n------------------------------------------------------\n\nThe humanized antibody, M9346A, was functionalized with the bifunctional chelator *p*-isothiocyanatobenzyl-desferrioxamine (DFO-Bz-NCS) with a 1:3.5 antibody:DFO ratio, as previously described \\[[@CR17], [@CR18]\\]. ",
"Specifically, a solution of previously purified antibody M9346A (500 μL, 2.27 mg) in PBS was adjusted to pH = 8.5 with 1 M Na~2~CO~3~ (1.0 μL). ",
"Then, a solution of DFO-Bz-NCS (4.2 mM, 4.0 μL) in dimethylsulfoxide was slowly added and the reaction mixture was incubated at 37 °C for 90 min. ",
"Afterwards, the DFO-modified antibody was purified using a PD-10 desalting column and isolated in PBS solution for in vitro and in vivo studies.",
"\n\nA solution of ^89^Zr-oxalate in 1 M oxalic acid (5.0 μL, 85.1 MBq, 2.3 mCi) was added to phosphate-buffered saline (100 μL), and a 1 M sodium carbonate solution (4.8 μL) was used to adjust the pH to 7.1--7.4. ",
"Then, a solution of DFO-M9346A (500 μL, 500 μg) was added to the neutralized Zr-89 solution and the reaction mixture was incubated at 37 °C for 1 h. After purification using PD-10 desalting columns (GE Healthcare), \\[^89^Zr\\]Zr-DFO-M9346A (85.7 ± 5.7% isolated radiochemical yield and 3.08 ± 0.43 mCi/mg specific activity) was isolated in a PBS solution (2.0 mL) and radiochemical purity (98.0 ± 0.7%) was determined through radio-instant thin/layer chromatography. ",
"Then, the resulting ^89^Zr-radiolabeled antibody solution was diluted with PBS to the desired volume for in vitro and in vivo studies.",
"\n\nCancer cell lines {#Sec5}\n-----------------\n\nWe chose four cancer cell lines with variable FRα expression levels to correlate imaging findings. ",
"The HeLa-derived cancer cell line KB (high expression), the ovarian cancer cell line OV90 (low/medium expression), the human lung cancer cell line H2110 (low/medium), and the human lung cancer cell line A549 (low/no expression) were purchased from ATCC (Manassas, VA). ",
"No further cell line authentication was conducted, and the cell lines were expended by passaging 2--3 times, aliquoted, and frozen in liquid nitrogen. ",
"For use in in vitro as well as in vivo experiments, the cell lines were grown in medium recommended by American Type Culture Collection (ATCC) and passaged regularly at 70--80% confluence every 3--4 days. ",
"All cell lines were cultured at 37 °C and 5% carbon dioxide.",
"\n\nCharacterization and in vitro experiments {#Sec6}\n-----------------------------------------\n\n\\[^89^Zr\\]Zr-DFO-M9346A was investigated for stability by incubating the radioligand in 1% bovine serum albumin at 37 °C for 3 days. ",
"Therefore, radiochemical purity of \\[^89^Zr\\]Zr-DFO-M9346A was tested at various time points via radio-instant thin layer chromatography (Eckert & Ziegler, Germany) with ethylendiaminetetraacetic acid (50 mM) as mobile phase.",
"\n\nA Lindmo assay \\[[@CR19]\\] was performed to validate the integrity of the ^89^Zr-radiolabeled antibody M9346A. In detail, serial dilutions of KB cells were incubated with one concentration of \\[^89^Zr\\]Zr-DFO-M9346A in Eppendorf vials (1.5 mL) on a shaking rack. ",
"After incubation at room temperature for 1 h, cells were washed with ice-cold PBS and the retained radioactivity on cells was measured using a WIZARD^2^ automatic γ-counter from PerkinElmer.",
"\n\nFurthermore, an in vitro binding assay was performed with cell lines expressing different levels of FRα, as previously described \\[[@CR20]\\]. ",
"One day prior to the in vitro experiment, the cells lines KB, OV90, and H2110, as well as A549 (each 1.0 × 10^6^ cells per well, \\> 90% viability for each cell line) were seeded into 6-well plates containing growth medium (2.0 mL) and incubated at 37 °C to form subconfluent cell monolayers. ",
"Then, growth medium was removed from the 6-well plates, cells were washed with PBS, and 900 μL of growth medium was added. ",
"After 1 h of incubation at 37 °C, a solution of \\[^89^Zr\\]Zr-DFO-M9346A (0.5 μCi, 100 ng) in PBS (100 μL) was added to the wells. ",
"For blocking studies, cells were pre-incubated with humanized antibody M9346A (10 μg) 5 min prior to the addition of ^89^Zr-radiolabeled antibody (0.5 μCi, 100 ng). ",
"To determine the number of cells at the time of the experiment, a separate set (*n* = 3) of wells for each cell line and experiment was analyzed by detaching the cells with trypsin and counted immediately using a Vi-cell XR cell viability analyzer (Beckman Coulter). ",
"After 1 h post-incubation at 37 °C, the supernatant of each well was collected together with washing solution of ice-cold PBS. ",
"Then, cells were lysed with sodium hydroxide solution (1 M, 1.0 mL) for 5 min at room temperature. ",
"Finally, each cell suspension was collected and all radioactive samples were measured using a WIZARD^2^ automatic γ-counter from PerkinElmer (Waltham, MA). ",
"Receptor-specific uptake was determined by correlating cell-bound activity relative to non-bound activity in the supernatant and displayed as a percentage of applied activity per 5.0 × 10^5^ cells.",
"\n\nAnimals {#Sec7}\n-------\n\nAll in vivo studies and procedures were performed in accordance with an approved protocol from the Institutional Animal Care and Use Committee at MSK. ",
"All in vivo experiments were carried out in female, athymic nude mice (Envigo; outbread; 6--8 weeks, 20--25 g). ",
"Subcutaneous KB and OV90 xenografts (150 ± 20 mm^3^) were established as previously described \\[[@CR5], [@CR20]\\]. ",
"In detail, suspended KB cells (1.5 × 10^6^, viability \\> 93%) or suspended OV90 cells (1.0 × 10^7^, viability: \\> 95%) in a solution containing a 1:1 mixture of Matrigel (Becton Dickinson, Bedford, MA) and cell culture media (no FBS for OV90 xenografts) were subcutaneously inoculated on the right shoulder of anesthetized mice (1.5--2.0% isoflurane (Baxter Healthcare) in medical air (2 L/min)). ",
"Prior in vivo studies, KB tumors were grown for 9--10 days post-implantation and OV90 tumors were grown for 21 days.",
"\n\nIn vivo PET/CT imaging and biodistribution {#Sec8}\n------------------------------------------\n\nFor in vivo studies, PET images were recorded using a small-animal Inveon® PET/CT system from Siemens (Knoxville, TN) and mice were anesthetized with 1.5--2.0% isoflurane at 2.0 L/min flow of medical air. ",
"PET images were analyzed using AsiPro VM™ software (Concorde Microsystems) and Inveon research workplace 4.1 software (Siemens Healthcare). ",
"For biodistribution studies, mice were euthanized at pre-determined time points through asphyxiation with carbon dioxide and organs of interest were collected, weighed, and counted using a WIZARD^2^ automatic γ-counter from PerkinElmer. ",
"The percentage of tracer uptake stated as percentage injected activity per gram of tissue (%IA/g) was calculated as the activity associated with tissue per organ weight per actual injected dose, decay corrected to the start time of counting. ",
"KB tumor-bearing mice (*n* = 10) were injected with \\[^89^Zr\\]Zr-DFO-M9346A (7.75 ± 0.25 MBq, 209.0 ± 6.7 μCi, 50 μg) in PBS (200 μL). ",
"PET images were acquired at 4 h, 24 h, 48 h, and 72 h post-injection, and biodistribution studies were performed at 24 h and 72 h (each cohort, *n* = 5). ",
"OV90 tumor-bearing mice (*n* = 6) were injected with \\[^89^Zr\\]Zr-DFO-M9346A (2.02 ± 0.08 MBq, 48.4 ± 2.0 μCi, 50 μg) in PBS (200 μL). ",
"PET images were acquired at 24 h and 48 h post-injection, and biodistribution studies were performed at 24 h and 48 h post-injection of radioligand. ",
"Each tumor-bearing cohort exhibited expected bone uptake upon release of Zr-89 from the chelator desferrioxamine (DFO) of about 5--10 %IA/g.\n\nRadiolabeling of ADC {#Sec9}\n--------------------\n\nAll iodine-131 radiolabeling reactions were performed in pre-coated Iodogen (1,3,4,6-tetrachloro-3α,6α-diphenyl glycoluril) tubes (100 μg per tube). ",
"For instance, an aqueous solution of Na^131^I (37.0 MBq, 1.0 mCi) was added to a phosphate buffered saline solution of IMGN853 (250 μg, 45 μL) and the tube containing the reaction mixture was gently agitated at 1-min intervals at room temperature over a period of 10 min. ",
"After purification using PD-10 desalting columns, \\[^131^I\\]-IMGN853 (32.1 MBq, 0.87 mCi) was isolated in 87% radiochemical yield and high radiochemical purity (99.9%).",
"\n\nBlood half-life studies {#Sec10}\n-----------------------\n\nFemale athymic nude mice (6--8 weeks, *n* = 5) were injected intravenously with \\[^89^Zr\\]Zr-DFO-M9346A (2.3 ± 0.2 MBq, 62.6 ± 4.6 μCi, 25 μg) or \\[^131^I\\]-IMGN853 (2.7 ± 0.1 MBq, 73.4 ± 2.6 μCi, 25 μg). ",
"At predetermined time points (0.5 h, 1.0 h, 3.0 h, 6.0 h, 24 h, 48 h, and 72 h), a sample of blood was obtained from the great saphenous vein of each animal in a heparin-coated capillary glass tube and the weights of the collected blood samples were obtained by using an analytical balance Toledo XS105 from Mettler. ",
"The radioactivity of the blood samples was recorded with a WIZARD^2^ automatic γ-counter from PerkinElmer. ",
"The residual radiotracer, expressed as a percentage injected dose per gram (%IA/g), was calculated as the activity present in the blood weighed per actual injected dose, decay corrected to the time of counting.",
"\n\nBiodistribution studies {#Sec11}\n-----------------------\n\nFor biodistribution studies, OV90 tumor-bearing nude mice (6--8 weeks, *n* = 5) were injected intravenously with \\[^131^I\\]-IMGN853 (1.42 ± 0.02 MBq, 38.3 ± 0.4 μCi) in PBS (200 μL). ",
"At 48 h post-injection, mice were euthanized and organs of interest were collected, weighed, and counted with a WIZARD^2^ automatic γ-counter from PerkinElmer.",
"\n\nIn vivo therapy studies {#Sec12}\n-----------------------\n\nTumor volumes for all mice were measured via manual caliper measurements of the longest dimension (*x*), shortest dimension (*y*), and height (z), assuming an ellipsoid shape (*V* = (3.1415/6) × (x × y × z) \\[mm^3^\\]). ",
"During tumor measurements, weight for all mice was recorded, and after initial tumor measurements, mice were randomized into three cohorts (*n* = 3--5) per cohort ensuring all cohorts had tumor volumes of 150--200 mm^3^. One day after initial tumor volume measurements, mice in the therapy cohort A were co-administered with a solution of ADC (1.25 mg/kg, 31.2 μg) and companion imaging agent \\[^89^Zr\\]Zr-DFO-M9346A (3.34 ± 0.04 MBq, 90.3 ± 1.1 μCi, 25 μg) in PBS (200 μL). ",
"Cohort B was injected with the companion imaging agent \\[^89^Zr\\]Zr-DFO-M9346A (3.68 ± 0.07 MBq, 99.5 ± 1.8 μCi, 25 μg). ",
"Cohort C was injected with PBS (200 μL). ",
"Two days after administration, mice of cohort A and cohort B were anesthetized with 1.5--2.0% isoflurane at 2.0 L/min flow of medical air and PET/CT imaging was accomplished over 10 min using small-animal Inveon® PET/CT system from Siemens (Knoxville, TN). ",
"Then, tumor volumes were determined via caliper measurement every 3 to 4 days up to an endpoint volume of \\> 1000 mm^3^. Additionally, all mice were assessed twice per week throughout the study for outward signs of toxicity and decreasing body weight.",
"\n\nResults {#Sec13}\n=======\n\nDevelopment of \\[^89^Zr\\]Zr-DFO-M9346A {#Sec14}\n--------------------------------------\n\nModification of the humanized monoclonal antibody (mAb), M9346A, with the bifunctional chelator DFO-Bn-NCS was accomplished using a previously reported method (Fig.",
" [2a](#Fig2){ref-type=\"fig\"}, Additional file [1](#MOESM1){ref-type=\"media\"}: Figure S1) \\[[@CR17], [@CR18], [@CR21]\\]. ",
"First, desferrioxamine was coupled to lysine-NH~2~ groups of the parent antibody (1:3.5 antibody: DFO ratio) at pH = 8.5 over 90 min in phosphate buffered saline, followed by purification using PD-10 desalting columns. ",
"Subsequently, radiolabeling of the modified mAb with neutralized \\[^89^Zr\\]Zr-oxalic acid solution proceeded in PBS at 37 °C over 60 min, followed by purification using PD-10 desalting columns. ",
"DFO-M9346A was radiolabeled reliably with good specific activity (3.08 ± 0.43 mCi/mg) and radiochemical purity (98.0 ± 0.7%). ",
"Lindmo assays of the resulting radiolabeled antibody construct confirmed that the immunoreactivity of the construct was established through binding of the antibody to FRα in vitro (83.4 ± 3.5%, Additional file [2](#MOESM2){ref-type=\"media\"}: Figure S2). ",
"Stability of \\[^89^Zr\\]Zr-DFO-M9346A was examined in bovine serum albumin at 37 °C over a period of 72 h and showed that more than 95% of the radioligand remained intact (Additional file [3](#MOESM3){ref-type=\"media\"}: Figure S3). ",
"Further characterization of \\[^89^Zr\\]Zr-DFO-M9346A included an in vitro cell uptake assay using cancer cell lines with various expression levels of FRα (Fig.",
" [2](#Fig2){ref-type=\"fig\"}c). ",
"We observed highly specific cell-associated uptake and retention in KB, OV90, and H2110 after 1 h of incubation at 37 °C (43.9 ± 0.9%, 17.6 ± 0.5%, and 17.3 ± 0.8%, per 500,000 cells respectively), corresponding to their folate receptor expression levels \\[[@CR5]\\]. ",
"The cancer cell line A549 with very low expression levels of FRα displayed negligible uptake (0.4 ± 0.1%).Fig. ",
"2Synthesis and characterization of \\[^89^Zr\\]Zr-DFO-M9346A. **a** Conjugation of *p*-isothiocyanatobenzyl-desferrioxamine (DFO-Bz-NCS) to humanized antibody M9346A in basic aqueous conditions at 37 °C for 90 min and ^89^Zr-radiolabeling of DFO-M9346A at 37 °C for 60 min. **",
"b** After purification of reaction mixture using a PD-10 desalting column, quality control of \\[^89^Zr\\]Zr-DFO-M9346A was performed using radio-instant thin layer chromatography in a solution of EDTA (50 mM). **",
"c** In vitro uptake studies of either \\[^89^Zr\\]Zr-DFO-M9346A (100 ng) or a mixture of \\[^89^Zr\\]Zr-DFO-M9346A (100 ng) and M9346A (10 μg) using cancer cell lines KB, OV90, H2110, and A549 with various expression levels of FRα. ",
"\\*\\**P* \\< 0.01; \\*\\*\\**P* \\< 0.001. ",
"Error bars represent the SD\n\nIn vivo and ex vivo experiments with \\[^89^Zr\\]Zr-DFO-M9346A in subcutaneous epithelial cancer xenografts {#Sec15}\n---------------------------------------------------------------------------------------------------------\n\nWe first asked whether \\[^89^Zr\\]Zr-DFO-M9346A can target a tumor in vivo known to be high in expression of FRα (Fig.",
" [3a](#Fig3){ref-type=\"fig\"} and Additional file [4](#MOESM4){ref-type=\"media\"}: Figure S4). ",
"After tail vein injection of the radioligand, small-animal PET imaging (4 h, 24 h, 48 h, and 72 h) and biodistribution studies (24 h and 72 h) were conducted using KB tumor-bearing mice (female, athymic, *n* = 10). ",
"Serial PET imaging showed clear delineation of the tumor already after 24 h with low uptake in normal tissue. ",
"The maximum intensity projections (MIPs) indicated that blood-pool and background activity cleared over time, leading to improved tumor-to-background ratios at 48 h post-injection. ",
"Ex vivo biodistribution data corroborated the PET data, as high tumor localization of \\[^89^Zr\\]Zr-DFO-M9346A at 24 h (30.1 ± 1.9 %IA/g, *n* = 5) was observed and tumor uptake increased over time out to 72 h (45.8 ± 29.0 %IA/g, *n* = 5) post-injection, suggesting an optimal time point for future PET imaging studies without increasing bone uptake over time (Additional file [5](#MOESM5){ref-type=\"media\"}: Figure S5).Fig. ",
"3In vivo studies of \\[^89^Zr\\]Zr-DFO-M9346A: **a** PET images acquired at 24 h and 48 h post-injection and biodistribution studies performed at 24 h and 72 h with \\[^89^Zr\\]Zr-DFO-M9346A intravenously administered in KB tumor bearing mice. **",
"b** PET images acquired at 24 h and 48 h post-injection and biodistribution studies performed at 24 h and 72 h with \\[^89^Zr\\]Zr-DFO-M9346A intravenously administered in tumor bearing mice\n\nAfter the first proof-of-concept experiment using KB cells, we set out to validate \\[^89^Zr\\]Zr-DFO-M9346A in a more clinically relevant cancer model using the ovarian cancer cell line OV90. ",
"To this end, \\[^89^Zr\\]Zr-DFO-M9346A was intravenously injected into female athymic nude mice bearing a subcutaneous OV90 xenograft on the right shoulder. ",
"PET imaging and ex vivo biodistribution after 24 h and 48 h confirmed efficient retention of the radioligand in the tumor (24.2 ± 6.3 %IA/g, *n* = 6) with very limited background uptake (Fig.",
" [3b](#Fig3){ref-type=\"fig\"} and Additional file [6](#MOESM6){ref-type=\"media\"}: Figure S6).",
"\n\nPharmacokinetic head-to-head comparison between IMGN853 and \\[^89^Zr\\]Zr-DFO-M9346A {#Sec16}\n-----------------------------------------------------------------------------------\n\nAfter successful characterization of the antibody-based PET imaging agent showing very promising performance in vivo, we next set out to validate that the antibody-drug conjugate and companion diagnostic show nearly identical pharmacokinetic profiles. ",
"In order to achieve this goal, we performed a head-to-head comparison between both antibody constructs. ",
"In order to do that, we radiolabeled the antibody-drug conjugate, IMGN853, through direct halogenation with I-131 (Fig.",
" [4a](#Fig4){ref-type=\"fig\"}). ",
"\\[^131^I\\]-IMGN853 was isolated in good isolated radiochemical yields (86.7 ± 13.5%) and high radiochemical purity (99.3 ± 0.7%). ",
"Based on previous in vitro studies with \\[^89^Zr\\]Zr-DFO-M9346A, we tested whether the radiolabeled ADC retains its ability to bind to FRα and performed an in vitro uptake with using KB cells (Fig.",
" [4b](#Fig4){ref-type=\"fig\"}). ",
"Then, we went ahead and compared the drugs' pharmacokinetic behavior by evaluating each tracer's blood half-life in vivo. ",
"Figure [4c](#Fig4){ref-type=\"fig\"} shows the correlation of tracer (%IA/g) in the blood at predetermined time points. ",
"The blood half-life for both radioligands was determined through serial bleeds in healthy mice, and the obtained data points were compared to each other. ",
"We found a good correlation between \\[^131^I\\]-IMGN853 and \\[^89^Zr\\]Zr-DFO-M9346A (*R*^2^ = 0.9736). ",
"Ex vivo biodistribution data revealed very similar biodistribution pattern in all organs compared to the companion diagnostic with slightly lower tumor uptake of \\[^131^I\\]-IMGN853 of 17.3 ± 5.2 %IA/g (Additional file [7](#MOESM7){ref-type=\"media\"}: Figure S7).Fig. ",
"4Pharmacokinetic head-to-head comparison between antibody-drug-conjugate, IMGN853 and companion diagnostic, \\[^89^Zr\\]Zr-DFO-M9346A. **a** Introduction of the radionuclide I-131 by direct halogenation of the antibody in the presence of Iodogen as chemical oxidant yielding radiolabeled antibody-drug-conjugate \\[^131^I\\]-IMGN853 (32.2 MBq, 0.87 mCi, 250 μg) in high radiochemical purity (\\> 98%). **",
"b** In vitro characterization of \\[^131^I\\]-IMGN853. ",
"Incubation of KB cells with radiolabeled ADC or with a mixture of radiolabeled ADC (100 ng) and M9346A (10 μg). **",
"c** Comparison of blood half-lives of \\[^131^I\\]-IMGN853 and \\[^89^Zr\\]Zr-DFO-M9346A (dashed line indicates the 95% confidence band)\n\nLeveraging PET imaging with \\[^89^Zr\\]Zr-DFO-M9346A during therapy with IMGN853 {#Sec17}\n-------------------------------------------------------------------------------\n\nFinally, we determined whether \\[^89^Zr\\]Zr-DFO-M9346A could be used as a companion diagnostic during ADC therapy to predict therapy outcome. ",
"For these experiments (Fig.",
" [5a](#Fig5){ref-type=\"fig\"}), mice bearing OV90 tumor xenografts were injected with an imaging dose of \\[^89^Zr\\]Zr-DFO-M9346A (90.3 ± 1.1 μCi, 25 μg) as well as with a therapy dose of IMGN853 (1.25 mg/kg, 31.2 μg). ",
"PET images were acquired 2 days post-administration, and tumor volume of each mouse was tracked over several days until the end of the study. ",
"In general, administration of \\[^89^Zr\\]Zr-DFO-M9346A during ADC treatment with IMGN853 confirmed good tumoral uptake with high tumor-to-tissue contrast (Fig.",
" [5b](#Fig5){ref-type=\"fig\"}). ",
"After quantification of the acquired PET images, we observed differences in tumoral uptake with the lowest uptake of 42.4 %IA/g and the highest of 61.6 %IA/g. Following the tumor volume of individual mice revealed a differential response in ADC treatment (Fig.",
" [5c](#Fig5){ref-type=\"fig\"}). ",
"At 6 weeks post-treatment, two mice of the therapy cohort receiving IMGN853 (1.25 mg/kg) responded by inhibited tumor growth, whereas three mice showed similar tumor growth curves as the cohorts injected with \\[^89^Zr\\]Zr-DFO-M9346A (positive control) and PBS (negative control). ",
"No weight losses were observed in any cohort of the therapy study (Additional file [8](#MOESM8){ref-type=\"media\"}: Figure S8). ",
"Correlation between tumoral uptake (%IA/g) of \\[^89^Zr\\]Zr-DFO-M9346A and therapy outcome shows that in this cohort, mice with uptake greater than 50 %IA/g appeared to be responders and mice with lower than 50 %IA/g were non-responders (Fig.",
" [5d](#Fig5){ref-type=\"fig\"}). ",
"The cutoff level at 50 %IA/g was arbitrarily set and corresponds to the average tumor uptake of \\[^89^Zr\\]Zr-DFO-M9346A in this cohort.",
"Fig. ",
"5Therapy study with antibody-drug-conjugate, IMGN853, and companion diagnostic, \\[^89^Zr\\]Zr-DFO-M9346A, using OV90 xenografts. **",
"a** Overall design of the therapy study including inoculation of cancer cells, injection of ADC and companion diagnostic, PET imaging after 2 days, and continuously measuring tumor growth. **",
"b** PET images of mice injected with \\[^89^Zr\\]Zr-DFO-M9346A (90.3 ± 1.1 μCi, 25 μg) and IMGN853 (1.25 mg/kg, 31.2 μg) and quantification through ROI analysis of coronal slices. **",
"c** Plot of the individual tumor volume post treatment (days). **",
"d** Correlation of tracer uptake (%IA/g) and therapy outcome\n\nDiscussion {#Sec18}\n==========\n\nOver the last decades, companion diagnostic agents have become more and more important in the development process of targeted drugs, particularly in oncology \\[[@CR14]--[@CR16], [@CR22]\\]. ",
"Especially, nuclear medicine-based companion diagnostics targeting cancer-specific membrane bound receptors have received considerable attention \\[[@CR23]--[@CR26]\\]. ",
"In this study, we report a nuclear medicine-based companion diagnostic, \\[^89^Zr\\]Zr-DFO-M9346A, for the ADC mirvetuximab soravtansine (IMGN853). ",
"Currently, patient selections in clinical trials of IMGN853 are based on immunohistochemical (IHC) assessment of archival tumor tissue. ",
"Expression levels of FRα are not contemporaneous, and both intra- and inter-lesion tissue sampling is limited, making a non-invasive whole body imaging approach more favorable.",
"\n\nFollowing published protocols \\[[@CR17], [@CR18]\\], we were able to modify and radiolabel the FRα-targeting antibody M9346A in good radiochemical yields (88 ± 5%) and high-specific activities (3.08 ± 0.43 mCi/mg). ",
"\\[^89^Zr\\]Zr-DFO-M9346A was highly stable in human plasma (\\> 97% at 24 h). ",
"It also displayed relatively high immunoreactivity (\\> 80%) indicating conservation of the ability to bind to FRα in vitro. ",
"Furthermore, we observed high uptake in KB, OV90, and H2110 cells (43.9 ± 0.9%, 17.6 ± 0.5%, and 17.3 ± 0.8%, per 500,000 cells respectively). ",
"Co-incubation of OV90 and H2110 with a 100-fold excess of antibody M9346A further corroborates specific uptake. ",
"In case of the KB cancer cell line, incomplete blocking may be a result of the high abundance of FRα per cell in comparison to the other FRα-positive cancer cell lines \\[[@CR27]\\]. ",
"Nevertheless, reduction in uptake was statistically significant (*p* \\< 0.001). ",
"Binding to A549 was negligible (\\< 1%), since FRα expression levels are reported to be low in comparison with other cancer lines.",
"\n\nEncouraged by the successful in vitro characterization of \\[^89^Zr\\]Zr-DFO-M9346A, we next sought to test the companion diagnostic in tumor-bearing mice. ",
"Initially, we injected the PET imaging agent in KB tumor bearing mice. ",
"Already 24 h after tracer administration, we observed good delineation of the tumor. ",
"According to the maximum-intensity-projection, slow blood clearance of the antibody construct caused accumulation in the heart region. ",
"However, qualitatively improved PET images were observed at 48 h and 72 h post-injection. ",
"Biodistribution data supported the results of the PET imaging studies, observing good anticipated tumor-to-blood ratios of 5.9 and tumor-to-bone ratios of 5.4 at 72 h post-administration of \\[^89^Zr\\]Zr-DFO-M9346A. In an OV90 tumor-bearing PET imaging study, we observed similar uptake patterns with good delineation of tumor and high imaging contrast. ",
"Based on the biodistribution studies, we reached slightly lower uptake of the companion diagnostic, which is likely due to the lower expression level of FRα in OV90 cells compared to KB cells.",
"\n\nWe next wanted to set stage for being able to predict ADC therapy success using our new companion diagnostic. ",
"Both antibody-based constructs had similar pharmacokinetic profiles, similar to what we showed with a nanoparticle construct earlier \\[[@CR28]\\]. ",
"Therefore, we were able to directly radiolabel the ADC with iodine-131. ",
"A standard in vitro uptake assay showed that binding as well as specificity of the antibody-based construct towards FRα was conserved, indicating minor modifications of the antibody. ",
"A direct comparison of the blood half-life of each radiolabeled antibody construct, \\[^131^I\\]-IMGN853 and \\[^89^Zr\\]Zr-DFO-M9346A, successfully indicated nearly identical pharmacokinetic behaviors in vivo. ",
"Radioiodination of antibodies with Iodogen is a standard method, suggesting that it is very unlikely that labeling conditions will lead to degradation of IMGN853. ",
"Since the antibody exhibits a more than 20-fold higher molecular weight in comparison with the added modifications (3--4 DM4 per antibody and 3--4 chelator per antibody), we expected minor alterations in the pharmacokinetics of each construct. ",
"However, other physiochemical properties such as polarity and charge of a molecule can influence drug pharmacokinetics as well.",
"\n\nWe tested the ability of \\[^89^Zr\\]Zr-DFO-M9346A to predict FRα-targeted therapy with IMGN853 in vivo. ",
"The penultimate goal of this study was to distinguish between responders and non-responders at the beginning of a therapeutic intervention---which might be an additional first step to improve the interpretation of current and future clinical trials using IMGN853. ",
"Two days post-co-administration of \\[^89^Zr\\]Zr-DFO-M9346A and IMGN853 in OV90 tumor-bearing mice, we were able to delineate OV90 tumors, in which we observed heterogeneity of companion diagnostic, presumably dictated not only by target expression, but also tumor physiology \\[[@CR29]\\]. ",
"By measuring the tumor volume of each animal over time, we were able to establish a threshold of tracer uptake at 50 %IA/g. A higher value indicated that the animal responded to the therapy, whereas a lower value indicated a non-responder.",
"\n\nConclusion {#Sec19}\n==========\n\nIn summary, we have developed an antibody-based companion diagnostic measuring FRα expression in ovarian cancer during ADC therapy. ",
"\\[^89^Zr\\]Zr-DFO-M9346A was found to be straightforward to radiolabel with Zr-89 after conjugation of desferrioxamine to the antibody. ",
"Our PET imaging agent showed excellent in vivo performance delineating FRα-positive tumors with high tumor-to-background ratios. ",
"Successful in vivo correlation of the pharmacokinetics between companion diagnostic and the radiolabeled ADC allowed us to perform therapy studies allowing the precision prediction of responders and non-responders in small animal studies. ",
"In humans, ADC sensitivity likely depends on FRα expression. ",
"Intuitively, one of the next steps should be to use this imaging technology in more advanced and heterogeneous mouse models of ovarian cancer. ",
"We are confident that \\[^89^Zr\\]Zr-DFO-M9346A could ultimately be a suitable companion diagnostic for IMGN853 cancer therapy, quantitatively assessing the folate receptor expression levels non-invasively in patients.",
"\n\nAdditional file\n===============\n\n {#Sec20}\n\nAdditional file 1:**Figure S1.** ",
"Size exclusion chromatograms of (**a**) M9346A and (**b**) DFO-M9346A. (PDF 279 kb) Additional file 2:**Figure S2.** ",
"Lindmo immunoreactivity assay for ^89^Zr-labeled DFO-M9346A. (PDF 280 kb) Additional file 3:**Figure S3.** ",
"Stability study in human plasma of \\[^89^Zr\\]Zr-DFO-M9346A. (PDF 280 kb) Additional file 4:**Figure S4.** ",
"Serial PET imaging at 4 h, 24 h, 48 h, and 72 h post-administration of \\[^89^Zr\\]Zr-DFO-M9346A in KB tumor bearing mice. (",
"PDF 335 kb) Additional file 5:**Figure S5.** ",
"Ex vivo biodistribution study at 24 h and 72 h post-injection of \\[^89^Zr\\]Zr-DFO-M9346A in KB tumor bearing mice. (",
"PDF 335 kb) Additional file 6:**Figure S6.** ",
"Ex vivo biodistribution study at 24 h and 48 h post-injection of \\[^89^Zr\\]Zr-DFO-M9346A in OV90 tumor bearing mice. (",
"PDF 335 kb) Additional file 7:**Figure S7.** ",
"Side-by-side comparison of the ex vivo biodistribution at 48 h post-injection of \\[^89^Zr\\]Zr-DFO-M9346A and \\[^131^I\\]-IMGN853 in OV90 tumor bearing mice. (",
"PDF 102 kb) Additional file 8:**Figure S8.** ",
"Weights of individual mouse over time during treatment. (",
"PDF 280 kb)\n\nADC\n\n: Antibody-drug conjugate\n\nCT\n\n: Computed tomography\n\nDFO\n\n: Deferoxamine\n\nFRα\n\n: Folate receptor alpha\n\nHPLC\n\n: High-performance liquid chromatography\n\nmAb\n\n: Humanized monoclonal antibody\n\nMSKCC\n\n: Memorial Sloan Kettering Cancer Center\n\nPBS\n\n: Phosphate buffered saline\n\nPET\n\n: Positron emission tomography\n\nWe thank the staff members of the Small Animal Imaging Core Facility as well as the Radiochemistry and Molecular Imaging Probe Core Facility at Memorial Sloan Kettering Cancer Center. ",
"The authors especially thank Brian Zeglis for helpful discussions.",
"\n\nFunding {#FPar1}\n=======\n\nThis work was supported by National Institutes of Health (NIH) grants R01 CA204441 (T.R.) and P30 CA008748. ",
"The authors further thank ImmunoGen, Inc. for generous support.",
"\n\nAvailability of data and materials {#FPar2}\n==================================\n\nPlease contact the author for data requests.",
"\n\nCB, JH, JFP, JL, NVKP, JAK, and TR designed the experiments. ",
"CB, JH, KG, and AS conducted the experiments. ",
"CB and TR analyzed the data. ",
"CB and TR wrote the paper. ",
"All authors read and approved the final manuscript.",
"\n\nEthics approval {#FPar3}\n===============\n\nAll in vivo studies and procedures were performed in accordance with an approved protocol from the Institutional Animal Care and Use Committee at Memorial Sloan Kettering Cancer Center (MSK).",
"\n\nConsent for publication {#FPar4}\n=======================\n\nNot applicable.",
"\n\nCompeting interests {#FPar5}\n===================\n\nJose F. Ponte is an employee of ImmunoGen, Inc. The authors declare that they have no competing interests.",
"\n\nPublisher's Note {#FPar6}\n================\n\nSpringer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.",
"\n"
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"Case: 13-3053 Document: 13 Page: 1 Filed: 03/11/2013\n\n\n\n\n NOTE: This order is nonprecedential.",
"\n\n United States Court of Appeals\n for the Federal Circuit\n __________________________\n\n AVON JARRETT SPEARMAN,\n Petitioner,\n\n v.\n MERIT SYSTEMS PROTECTION BOARD,\n Respondent.",
"\n __________________________\n\n 2013-3053\n __________________________\n\n Petition for review of the Merit Systems Protection\n Board in case no. ",
"DC0752120306-I-1.",
"\n __________________________\n\n ON MOTION\n __________________________\n\n ORDER\n\n The Department of the Treasury moves to recaption to\n name the Merit Systems Protection Board as respondent,\n and for a 21-day extension of time for the Board to file its\n response brief.",
"\n\n Pursuant to 5 U.S.C. § 7703(a)(2), the Board is desig-\n nated as the respondent when the Board's decision con-\n cerns the procedure or jurisdiction of the Board. ",
"The\n employing agency is designated as the respondent when\n\fCase: 13-3053 Document: 13 Page: 2 Filed: 03/11/2013\n\n\n\n\n AVON SPEARMAN V. MSPB 2\n\n\n the Board reaches the merits of the underlying case. ",
"Here,\n the Board dismissed Spearman’s appeal for lack of jurisdic-\n tion. ",
"Thus, the Board is the proper respondent in this\n petition for review.",
"\n Accordingly,\n IT IS ORDERED THAT:\n\n The motions are granted. ",
"The revised official caption\n is reflected above. ",
"The Board should calculate its brief\n due date from the date of filing of this order.",
"\n\n FOR THE COURT\n\n\n /s/ Jan Horbaly\n Jan Horbaly\n Clerk\n s21\n\f"
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"Taiwan PMI\n\nTaiwan: Manufacturing PMI falls to over one-year low\n\nNovember 3, 2014\n\nThe manufacturing Purchasing Managers’ Index (PMI), elaborated by HSBC and Markit, fell from 53.3 in September to 52.0 in October. ",
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"Q:\n\nUse same string format for multiple items in Python 3\n\nAs an example, I have the following numbers assigned to different variables:\na = 1.23981321; b = 34; c = 9.567123\n\nBefore I print these variables, I format them to 4 decimal places:\n'{:.4f} - {:.4f} - {:.4f}'.format(a, b, c)\n\nWhich prints the following:\n'1.2398 - 34.0000 - 9.5671'\n\nInstead of assigning :.4f to each placeholder { }, is there a way to declare :.4f once to format all of the values?",
"\n\nA:\n\na = 1.23981321\nb = 34\nc = 9.567123\n\nprint('{:.4f} - {:.4f} - {:.4f}'.format(a, b, c)) # Original\n\nprint('{a:{f}} - {b:{f}} - {c:{f}}'.format(a=a, b=b, c=c, f='.4f')) # New\n\nIt's easier to do if you use keyword arguments so you can have {a}, as opposed to positional arguments. ",
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"Q:\n\nAdding a wiki post of third party iOS tools\n\nEarlier this week, there was a question posted of third party libraries that people use for iOS. ",
" It was closed, but had lots of responses. ",
"I think there should be a wiki post that contains all of the ones recommended here to point new users too.",
"\nLink to Question here\n\nA:\n\nFirst off, we've found that list-style questions don't work well in the format of this site. ",
"They generally devolve into piles of dozens, if not hundreds, of answers that aren't curated well. ",
"People won't read the whole list of pages of answers, so they just keep posting duplicates and the whole thing ends up being a mess. ",
"That's why that question was closed.",
"\nSecond, this question has been posed many times in the past. ",
"Two of the ones I can find right now are\n\n\"Open source iOS components? ",
"Reusable views, controllers, buttons, table cells, etc?\"",
"\n\"Useful iPhone Libraries\"\n\nDo we need yet another one? ",
"I suppose we could link to the above from the actual iphone and ios tag wikis, to point people at these canonical questions.",
"\nStack Overflow really isn't designed for these kinds of lists. ",
"Thankfully, there are many other places designed to host such a list, like Cocoa Controls or Verious, among others.",
"\n\n"
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[
"Introduction {#s1}\n============\n\nDuring mitosis, chromosomes are bound to microtubules emanating from both poles of the mitotic spindle via sister-kinetochores and aligned on the metaphase plate precisely in the middle of the spindle. ",
"The equatorial position of the metaphase plate is a distinctive feature of metazoan, plant, and many fungal cells. ",
"A centered metaphase plate is established even in asymmetrically dividing cells, where daughter cells of unequal size are obtained by an asymmetric positioning of the spindle prior to anaphase (e.g., in *Caenorhabditis elegans* embryos) or an asymmetric elongation of the spindle in anaphase (e.g., in *Drosophila melanogaster* embryonic neuroblasts \\[[@bib16]; [@bib39]\\]). ",
"However, the reason why the metaphase plate is located in the middle of the spindle is not known. ",
"One hypothesis is that the centered position facilitates the synchronous arrival of chromosomes at spindle poles during anaphase to prevent chromosomes from being caught on the wrong side of the cytokinetic furrow ([@bib30]; [@bib11]). ",
"Elegant work in meiotic praying mantis cells demonstrated that the equatorial positioning of the metaphase plate is not a mere consequence of bipolar kinetochore--microtubule attachments, as trivalent sex-chromosome align in the middle of the spindle, even though trivalent attachment does not favor an equatorial position ([@bib30]). ",
"Moreover, previous studies in *Chlamydomonas rheinhardtii* and *C. elegans* showed that an asymmetry in centriole numbers at spindle poles led to an asymmetric metaphase plate position, even though chromosomes established bipolar attachments ([@bib12]; [@bib17]). ",
"While in algae, longer half-spindles were associated with the pole containing fewer centrioles, in nematodes, longer half-spindles emanated from the pole containing more centrioles. ",
"However, whether cells react to asymmetrically located metaphase plates and the long-term consequences of this asymmetry are not known. ",
"Here, we investigated these questions in human tissue culture cells. ",
"We find that cells correct metaphase plate position before anaphase onset, we demonstrate that a centered metaphase plate position relies on the spindle assembly checkpoint (SAC) to provide sufficient time for this correction mechanisms, and we show that a failure to correct plate position leads to asymmetric cell divisions.",
"\n\nResults {#s2}\n=======\n\nCells center the metaphase plate position before anaphase onset {#s2-1}\n---------------------------------------------------------------\n\nTo monitor the relative position of the metaphase plate in the spindle over time, we recorded by time-lapse imaging HeLa cells stably expressing eGFP-centrin1 (centriole marker) and eGFP-CENPA (kinetochore marker) and automatically tracked centrosomes and the metaphase plate using an in-house developed software ([@bib14]; [@bib42]). ",
"Metaphase or late prometaphase cells were recorded over a short period of 5 min in 3D at a resolution of 7.5 s under conditions of low phototoxicity compatible with anaphase entry ([@bib14]). ",
"By plotting the ratio R of the half-spindle lengths of metaphase cells at the onset of our recordings (first three time points), we found a broad distribution centered around median R = 0.98, which represents nearly equal half-spindle lengths. ",
"When analyzing the subset of cells that entered anaphase during our recordings 30 s before anaphase, we found a sharp R distribution in the middle of the spindle (median R = 1.02; [Figure 1A](#fig1){ref-type=\"fig\"}): less than 10% of the R values were smaller than 0.85 or larger than 1.15 at anaphase onset, while in the metaphase population over 24.2% were outside of these boundaries. ",
"This suggested a centering mechanism for the metaphase plate as cells progressed towards anaphase. ",
"To test this hypothesis, we aimed to create asymmetric spindles by generating cells with an asymmetric centriole distribution, using small interfering (si)RNAs against Sas-6, a protein required for centriole duplication ([@bib23]). ",
"This procedure was used on a set of HeLa eGFP-centrin cells that co-expressed either eGFP-CENPA, α-tubulin-mRFP (spindle marker), or Histone H2B-mRFP (chromosome marker). ",
"Every wild-type mitotic cell contains four centrioles: one oldest (grandmother) centriole, one older (mother) centriole, and their two respective daughter centrioles ([@bib31]), which all have different eGFP-centrin signal intensities ([@bib20]). ",
"A 24-hr Sas-6 depletion led to a mix of cells with two centrioles per pole, one centriole per pole, or one pole with one centriole and the other pole with two centrioles (called from here on 2:2, 1:1, or 2:1 cells; [Figure 1B,C](#fig1){ref-type=\"fig\"}). ",
"Our intensity measurements revealed that in 2:1 cells it was most often the oldest centriole that gave rise to a daughter centriole, probably due to limiting levels of Sas-6 (data not shown). ",
"Tracking of HeLa eGFP-centrin1/CENPA cells indicated that the distribution of half-spindle ratios during metaphase was broad in 2:2 or 1:1 cells, but that on average the plate was located in the middle of the cell (median R = 1.04 (2:2) and 1.03 (1:1); note that 2:2 cells served as control transfection for all subsequent experiments; to be consistent in 2:2 or 1:1 cells, R was calculated as the length of the half-spindle associated to the grandmother centriole divided by the opposite half-spindle length, while in 2:1 cells, R represents the ratio of the half-spindle associated to the 2-centriole pole over the opposite half-spindle length; [Figure 1D](#fig1){ref-type=\"fig\"}). ",
"As in wild-type cells, the range of R had narrowed by the time cells were about to enter anaphase, consistent with a centering process ([Figure 1E](#fig1){ref-type=\"fig\"}). ",
"In 2:1 cells, we found two half-spindles of unequal length, with a shorter half-spindle associated with the pole containing one centriole (median R = 1.12, p \\< 0.0001 in Mann--Whitney U test compared to symmetric distribution; R \\> 1.15 in 42.9% of the cells; [Figure 1D](#fig1){ref-type=\"fig\"}). ",
"This asymmetry was corrected before anaphase onset (median R = 1.03; [Figure 1F](#fig1){ref-type=\"fig\"}), consistent with the existence of a centering process. ",
"Since our videos were much shorter than the overall duration of metaphase, we were unable to directly visualize the centering process. ",
"To circumvent this difficulty, we recorded longer time series of 2:1 cells, monitoring them for 15 min at 30-s intervals. ",
"By plotting R over time in 15 cells that spend at least 7 min in metaphase before entering anaphase, we could directly observe how cells centered the plate position over time, while reducing the variability in metaphase plate position ([Figure 1G,H](#fig1){ref-type=\"fig\"}).10.7554/eLife.05124.003Figure 1.Cells center the position of the metaphase plate before anaphase onset.(**A**) Distribution of spindle ratio R in metaphase cells in wild-type eGFP-centrin1/CENPA HeLa cells during metaphase in general (black curve) or 30 s before anaphase onset (red curve). ",
"The spindle ratio R was calculated by dividing the half-spindle length L1 associated with the grandmother centriole (brightest eGFP-centrin1) by the other half-spindle length L2 (for all cell numbers in all experiments see [Table 1](#tbl1){ref-type=\"table\"}). (**",
"B**) Depletion of Sas-6 but not control depletion leads to the gradual loss of centrioles after 24, 48, or 72 hr, resulting in a mixed population of cells with different number of centrioles as indicated. ",
"Centrioles were visualized based on images of eGFP-centrin1/mRFP-α-tubulin cells as shown in **C** (n = 50 cells per experiment, in 3 experiments, error bars indicate s.e.m.). (**",
"C**) Immunofluorescence images of eGFP-centrin1 (green)/mRFP-α-tubulin (red) cells stained with DAPI (blue) with different centriole configurations as indicated. ",
"Scale bar indicates 2 μm. (**",
"D**) Distribution of spindle ratio R in metaphase cells in *siSas-6*-transfected eGFP-centrin1/CENPA cells with different centriole configurations (2:2, 2:1, or 1:1). ",
"The spindle ratio was calculated by dividing the half-spindle length L1 associated with the grandmother centriole (2:2 and 1:1 cells) or the half-spindle length associated with 2 centrioles (2:1 cells) by the other half-spindle length L2. ",
"The spindle ratio of 2:1 cells was significantly different from the ratios seen in 2:2 or 1:1 cells (T-test with Welch\\'s correction, 2:1 \\> 2:2, p = 0.018). (**",
"E**, **F**) Distribution of spindle ratio R in Sas-6-depleted eGFP-centrin1/CENPA 2:2 (**E**) or 2:1 (**F**) cells in metaphase and before anaphase onset. ",
"2:1 cells have a significantly more asymmetric plate position in metaphase when compared to cells just before anaphase (Mann--Whitney U test, p = 7.16 × 10^−7^). (**",
"G**) Plot of half-spindle ratio R over time in 15 individual eGFP-centrin1/CENPA 2:1 cells that entered anaphase during live-cell imaging (black lines). ",
"The timeline was synchronized to anaphase onset (t = 0); the red curve indicates the median of R, and red surface the 95% confidence interval. ",
"Note how median R approaches 1 over time and how its variability decreases. (**",
"H**) Time-lapse images of a eGFP-centrin1/CENPA 2:1 cell as analyzed in **G**. ",
"Half-spindle ratio R and time before anaphase are indicated for each frame. ",
"Number of centrioles was determined in IMARIS in 3D (see 3D-insets in green), \\* denotes the pole with 1 centriole.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.003](10.7554/eLife.05124.003)10.7554/eLife.05124.004Table 1.Number of cells in every experiment**DOI:** [http://dx.doi.org/10.7554/eLife.05124.004](10.7554/eLife.05124.004)ConditionN° of cellsWT40WT at anaphase onset42WT + MPS1-IN19*SiSas-6* 2:2 cells41*SiSas-6* 2:2 cells at anaphase onset29*SiSas-6* 2:2 cells + MPS1-IN19*SiSas-6* 2:1 cells59*siSas-6* 2:1 cells + MG13218*SiSas-6* 2:1 cells at anaphase onset33*SiSas-6* 2:1 cells in long term videos14*SiSas-6* 2:1 cells + MPS1-IN26*SiSas-6* 1:1 cells36*SiKif2a + siMCAK*20*SiKif2a + siMCAK* at anaphase onset41*SiKif2a + siMCAK + siSas-6* 2:1 cells29*SiKif2a + siMCAK + siSas-6* 2:1 cells at anaphase onset24*SiSas-6* 2:1 cells + ZM110*SiSas-6 2:2 + DMSO*15*SiSas-6 2:2* + taxol11*SiSas-6 2:1 + DMSO*16*SiSas-6 2:1* + taxol15Centriole laser-ablation (2:1)11Control laser-ablation8Centriole laser-ablation (2:1) + Mps1 inhibitor6Control laser-ablation + Mps1 inhibitor6\n\nThe SAC provides cells with sufficient time to center the metaphase plate {#s2-2}\n-------------------------------------------------------------------------\n\nTo investigate how cells center the metaphase plate, we analyzed the timing of 2:1 cells in comparison to 2:2 or 1:1 cells by monitoring either HeLa eGFP-centrin2/H2B-mRFP or HeLa eGFP-centrin1/mRFP-α-tubulin cells. ",
"A 2:2, 1:1, or a 2:1 centriole configuration affected neither the timing of bipolar spindle assembly nor the time it took to align all chromosomes on the metaphase plate ([Figure 2A,B](#fig2){ref-type=\"fig\"}). ",
"Anaphase onset, however, was delayed by 12 min ([Figure 2C](#fig2){ref-type=\"fig\"}, p = 0.003 in Mann--Whitney U test) in 2:1 cells, when compared to 2:2 or wild-type cells, resulting in 2:1 cells spending twice the amount of time in metaphase ([Figure 2D](#fig2){ref-type=\"fig\"}, p = 0.015 in Mann--Whitney U test). ",
"Anaphase time in 2:1 cells was also delayed in eGFP-centrin1/mRFP-α-tubulin cells 2:1 cells ([Figure 2E](#fig2){ref-type=\"fig\"}), indicating that it is a robust phenomenon (p = 0.003 in Mann--Whitney U test). ",
"In contrast, anaphase was not delayed in 1:1 cells, indicating that the longer metaphase timing seen in 2:1 cells was not caused by the loss of daughter centrioles, but the consequence of centriole asymmetry ([Figure 2C,E](#fig2){ref-type=\"fig\"}). ",
"Anaphase onset is under the control of the SAC, which inhibits the anaphase-promoting complex/cyclosome and delays anaphase onset if kinetochores are not properly attached to spindle microtubules ([@bib18]; [@bib8]). ",
"To test whether the observed metaphase delay depended on the SAC, we co-depleted the SAC protein Mad2 or inhibited the SAC kinase Mps1 in Sas-6-depleted cells ([Figure 2---figure supplement 1](#fig2s1){ref-type=\"fig\"}). ",
"In the absence of Mad2, 2:2, 2:1, 1:1, or control-treated cells had the same anaphase timing, indicating that the anaphase delay in 2:1 cells depends on the SAC ([Figure 2F](#fig2){ref-type=\"fig\"}). ",
"The same result could be found in Mps1-inhibited cells ([Figure 2---figure supplement 2](#fig2s2){ref-type=\"fig\"}). ",
"To test whether metaphase plate position is corrected as a result of the additional time provided by the SAC, we prolonged metaphase in *siSas-6*-treated cells by 1 hr by adding the proteasome inhibitor MG132 and found a symmetric plate position in 2:1 cells (median R = 0.98; [Figure 2G](#fig2){ref-type=\"fig\"}). ",
"In contrast, when we abrogated the SAC in metaphase cells by adding an inhibitor of the SAC kinase Mps1, MPS1-IN ([@bib21]), 2:1 cells showed a bimodal distribution: 53% entered anaphase with symmetric spindles, with a median R centered around 1, however, the other 47% had asymmetric spindles with a distribution centered around R = 1.15 ([Figure 2H](#fig2){ref-type=\"fig\"}, Mann--Whitney U test, p = 0.032 when compared to untreated 2:1 cells at anaphase onset). ",
"We conclude that the centering of the metaphase plate is not a mechanical consequence of cells preparing for anaphase, but that it depends on the SAC, which provides 2:1 cells with sufficient time to build a symmetric spindle. ",
"Mps1 inhibition in metaphase also led to a higher rate of segregation errors in 2:1 than in 2:2 or wild-type cells; this increase was reproducible, but not statistically significant ([Figure 2I,J](#fig2){ref-type=\"fig\"}).10.7554/eLife.05124.005Figure 2.The SAC delays anaphase in cells with asymmetric spindles allowing the centering of the metaphase plate.(**A**) Boxplots of the spindle assembly time (NBD until bipolar spindle formation) in wild-type and Sas-6-depleted 2:2, 1:1, or 2:1 eGFP-centrin1/mRFP−α-tubulin cells. ",
"Numbers indicate the median value, n = 49--59 cells in 2--6 experiments. (**",
"B**--**D**) Boxplots for the time between NBD and metaphase (**B**); the time between NBD and anaphase onset (**C**); and the time between metaphase and anaphase (**D**) in wild-type and Sas-6-depleted 2:2, 1:1, or 2:1 eGFP1-centrin2/H2B-mRFP cells. \\* ",
"indicates statistically significant difference in **C** (Mann--Whitney U test, p = 0.003), and **D** (Mann--Whitney U test, p = 0.015), n = 36--100 cells in 6--13 experiments. (**",
"E**) Boxplot for the time between NBD and anaphase B in wild-type and Sas-6-depleted 2:2, 1:1, or 2:1 eGFP1-centrin1/ mRFP−α-tubulin cells. \\* ",
"indicates statistically significant difference (Mann--Whitney U test, p = 0.003). (**",
"F**) Boxplots for the time between NBD and anaphase onset in Mad2-depleted or Mad-2/Sas-6-depleted 2:2, 1:1 or 2:1 eGFP1-centrin2/H2B-mRFP cells. ",
"2:1 cells are not delayed (Mann--Whitney U test, p = 0.836). (**",
"G**) Distribution of spindle ratios R in 2:1 eGFP-centrin1/CENPA cells treated with DMSO or MG132. ",
"For cell numbers see [Table 1](#tbl1){ref-type=\"table\"}. (**",
"H**) Distribution of spindle ratio R in Sas-6-depleted eGFP-centrin1/CENPA 2:1 cells in metaphase, at anaphase onset, or at anaphase onset when treated in metaphase with the Mps1 inhibitor MPS1-IN-1. ",
"Values for metaphase and anaphase without Mps1-IN treatment were taken from [Figure 1E](#fig1){ref-type=\"fig\"} for comparison. ",
"MPS1-IN treated 2:1 anaphase cells are significantly more asymmetric (Mann--Whitney U test, p = 0.032). (**",
"I**) Quantification of chromosome segregation errors in eGFP-centrin1/CENPA cells under the indicated conditions (n = 17--30 cells; N = 4--13 experiments). ",
"Error bars indicate s.e.m. (**",
"J**) Illustrative live-cell imaging stills of eGFP-centrin1/CENPA cells in anaphase with (right panel) or without (left panel) chromosome segregation errors. ",
"SAC, spindle assembly checkpoint.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.005](10.7554/eLife.05124.005)10.7554/eLife.05124.006Figure 2---figure supplement 1.Validation of Sas-6 and Mad2 co-depletion.(**A**, **B**) Immunoblots of wild-type, *siControl*-, *siSas-6*, or *siSas-6/siMad2* treated eGFP1-centrin2/H2B-mRFP HeLa cells probed with (**A**) Sas-6 and α-tubulin antibodies or (**B**) with Mad2 and α-tubulin antibodies.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.006](10.7554/eLife.05124.006)10.7554/eLife.05124.007Figure 2---figure supplement 2.Mps1 inhibition suppresses the anaphase delay in 2:1 cells.",
"Boxplots for the time between NBD and anaphase onset in wild-type, *siSas-6-*treated 2:2, 1:1, or 2:1 eGFP1-centrin2/H2B-mRFP cells treated with an Mps1 inhibitor. ",
"Numbers indicate average anaphase times. ",
"Note that 2:1 cells are not delayed when compared to 2:2 cells (Mann--Whitney U test, p = 0.9673).**DOI:** [http://dx.doi.org/10.7554/eLife.05124.007](10.7554/eLife.05124.007)\n\nAn asymmetric distribution of centrioles leads to an imbalance of microtubule stability {#s2-3}\n---------------------------------------------------------------------------------------\n\nTo understand the mechanism behind metaphase plate centering, we investigated how the loss of a daughter centriole affects the spindle poles and why it may affect the position of the metaphase plate. ",
"We first tested whether loss of centrioles reduces the polar ejection force ([@bib35]), as a possible explanation for the shorter half-spindle attached to the pole with fewer centrioles. ",
"Wild-type eGFP-centrin1/mRFP−α-tubulin cells were compared with cells treated with RNAis against Sas-6 or the chromokinesin Kid, the main driving force of the polar ejection force ([@bib44]). ",
"Cells were subjected to a half-hour treatment with the Eg5 inhibitor monastrol to obtain monopolar spindles ([@bib25]), fixed and stained for HEC1 to label kinetochores. ",
"The average distance between kinetochores and centrosomes in monopolar spindles reflects the polar ejection force ([@bib44]), as seen by a reduced distance in Kid-depleted cells ([Figure 3A](#fig3){ref-type=\"fig\"}). ",
"Loss of 2 daughter centrioles in Sas-6-depleted cells (one in each pole), however, did not reduce this distance, indicating that daughter centriole loss does not reduce the polar ejection force ([Figure 3A](#fig3){ref-type=\"fig\"}). ",
"In *C. elegans*, half-spindle size has been linked to the abundance of TPX-2, an activator of Aurora-A, a critical regulator of centrosome maturation ([@bib31]; [@bib12]); however, by quantitative immunofluorescence we found no significant differences for TPX-2 or for phosphorylated Aurora-A (the active form of Aurora-A) between the two spindle poles in Sas-6-depleted 2:1 cells ([Figure 3B](#fig3){ref-type=\"fig\"}). ",
"Next, we quantified the levels of centrobin, which stabilizes microtubule minus-ends, since it localizes only to daughter centrioles ([@bib46]; [@bib15]). ",
"Consistent with the daughter-specific localization, centrobin was only present on the pole with two centrioles in 2:1 cells; in contrast, α-tubulin density in each half-spindle and the abundance of the centrosome proteins γ-tubulin, pericentrin, ninein, and p150^glued^ (a marker for the dynein/dynactin motor complex) on the two spindle poles in 2:1 cells were as symmetric as in wild-type or 2:2 cells ([Figure 3C--F](#fig3){ref-type=\"fig\"}). ",
"Since centrobin depletion leads to a SAC-dependent mitotic arrest due to unstable microtubule minus-ends and unstable kinetochore--microtubules ([@bib15]), we hypothesized that the asymmetric localization of centrobin in 2:1 cells may lead to a difference in microtubule stability between the two poles. ",
"To test this, we subjected 2:2, 2:1, and 1:1 eGFP-centrin1/CENPA cells to a 7-min cold treatment on ice to depolymerize the astral microtubules and stained the remaining cold-resistant kinetochore--microtubules ([@bib37]). ",
"The stability of kinetochore--microtubules varied from cell to cell, but in 75% of the 2:2 or 1:1 cells the microtubule minus-ends showed the same stability at both spindle poles ([Figure 3G,H](#fig3){ref-type=\"fig\"}). ",
"In contrast, in 2:1 cells, an unequal stability of the minus-ends was visible in over 50% of the cells, and in most cases it was the pole with 2 centrioles that had more stable minus-ends ([Figure 3G,H](#fig3){ref-type=\"fig\"}). ",
"This difference in kinetochore--microtubule minus-end stability suggested that microtubule minus-ends might depolymerize faster in the absence of a daughter centriole, which would explain the shorter half-spindles associated with the 1-centriole pole. ",
"We also noted in our live-cell imaging experiments that \\>70% of the spindles in 2:1 eGFP-centrin1/mRFP-α-tubulin cells (but not 2:2 or 1:1 cells) were rotating, a phenomenon that could reflect a difference in minus-end stability of the two astral microtubule populations, resulting in an imbalance of cortical forces in the two half-spindles ([Figure 3I](#fig3){ref-type=\"fig\"} and [Videos 1--3](#video1 video2 video3){ref-type=\"other\"}). ",
"To confirm this hypothesis, we first tested whether a 1-hr MG132 treatment, which leads to symmetric spindles in 2:1 cells, is accompanied by an equalization in microtubule stability at the two spindle poles. ",
"While in control-treated cells, we saw an unequal stability of the minus-ends in 49 ± 4% of the 2:1 cells, this was only the case in 22 ± 6% of the MG132-treated 2:1 cells (p = 0.0275 in unpaired t-test; [Figure 3J](#fig3){ref-type=\"fig\"}). ",
"In contrast, in 2:2 cells, MG132 treatment did not affect the percentage of cells with unequal minus-end stability (14% each; [Figure 3J](#fig3){ref-type=\"fig\"}). ",
"Next, we tested whether the asymmetry of the spindle and the spindle rotation phenotype could be suppressed by increasing microtubule stability. ",
"We therefore co-depleted the microtubule-depolymerases KIF2a and MCAK, a condition known to increase microtubule stability ([@bib9]). ",
"KIF2a/MCAK depletion per se did not affect the position of the metaphase plate (median R = 1.01) or the ratio of cells with rotating spindles ([Figure 3K,L](#fig3){ref-type=\"fig\"}). ",
"However, when KIF2a/MCAK was co-depleted in 2:1 cells, the spindle asymmetry was statistically less pronounced (median spindle ratio R = 1.08 and 19.2% R \\> 1.15 compared to median R = 1.12 and 42.9% R \\> 1.15 in Sas-6 deplete cells, p = 0.039 Mann--Whitney test), and spindles did not rotate ([Figure 3L,M](#fig3){ref-type=\"fig\"}). ",
"To corroborate these results, we also stabilized microtubules by applying a brief, 30 min, 10 nM taxol treatment to Sas-6-depleted cells. ",
"While control (DMSO)-treated 2:1 cells were still asymmetric (median R = 1.23 and 56.3% R \\> 1.15), taxol-treated cells were much closer to symmetry (median R = 1.02 and 13.3% R \\> 1.15, p = 0.0062; [Figure 3N](#fig3){ref-type=\"fig\"}). ",
"We conclude that loss of a single centriole leads to a difference in minus-end microtubule stability between the two spindle poles, that this difference plays a large role in the asymmetric positioning of the metaphase plate, and that cells reduce this difference as they center the metaphase plate before anaphase onset.10.7554/eLife.05124.008Figure 3.2:1 cells have half-spindles with different microtubule stability and fail to mature kinetochore--microtubule attachments.(**A**) eGFP-centrin1 cells (green) were treated with monastrol and stained for HEC1 (red; right panel) to calculate the distance between kinetochores and the closest centriole. ",
"The left panel shows the distribution of centriole-HEC1 distances from n =16−28 cells, \\>1000 kinetochores. ",
"Scale bar indicates 2 μm. (**",
"B**, **C**) The difference in centrosomal levels of TPX-2 and phospho-Aurora-A (**B**), and centrobin between each spindle pole was quantified in HeLa eGFP-centrin1 by immunofluorescence using the indicated formula, and plotted as boxplots for each centriole configuration; n = 12--63 cells. ",
"Scale bar indicates 2 μm. (**",
"D**) Representative image of wild-type and *siSas-6*-treated 2:1 eGFP-centrin1 (green)/mRFP−α-tubulin (white) cells stained with anti-centrobin sera (red). ",
"Insets show magnified centrioles. ",
"Scale bar indicates 2 μm. (**",
"E**) Quantification of the difference in α-tubulin (red signal) levels between the two half-spindles, according to the formula shown in the box. ",
"Results were plotted in the left panel using a boxplot. ",
"n = 19--25 cells, N = 2 experiments. ",
"Scale bar indicates 2 μm. (**",
"F**) The difference in centrosomal levels of γ-tubulin, pericentrin, ninein and p150^glued^ between each spindle pole was quantified as in (**B**) and plotted as boxplots for each centriole configuration; n = 18--68 cells. (**",
"G**) Immunofluorescence images of 2:2 and 2:1 *siSas-6* eGFP-centrin1/CENPA (green) cells treated for 7 min with ice-cold medium and stained with anti-α-tubulin sera (magenta). ",
"Subsetted images are maximum intensity projections of 10 stacks (z = 0.2 μm) around centrioles. ",
"Scale bar indicates 2 μm. (**",
"H**) Quantification of kinetochore--microtubule minus-end stability at poles. ",
"Bar graph indicates percentage of cells that have asymmetric levels of kinetochore--microtubule minus-ends at the poles after cold-treatment; n = 26--49 cells; \\* indicates that within the 2:1 cell population the minus-end stability was significantly higher at the pole with 2 centriole (p = 0.00082 exact binomial test). (**",
"I**) Quantification of spindle rotation in control- and Sas-6-depleted 2:2, 2:1, or 1:1 eGFP-centrin1 (green)/mRFP−α-tubulin (red) cells based on time-lapse images as shown in the right panels. ",
"Times indicate minutes after NBD. ",
"Scale bar indicates 5 μm. ",
"A spindle was counted as rotating if it had turned by more than 90° in X/Y. n = 32--122 cells in 2--6 experiments. \\* ",
"indicates significant difference; Fisher\\'s exact test p = 8.39e-09. (**",
"J**) Quantification of kinetochore--microtubule stability at poles. ",
"Bar graph indicates percentage of cells that have more stable kinetochore--microtubule minus-ends either at the pole with the grandmother centriole (2:2) cells or at the 2-centriole pole (2:1 cells); n = 20--40 cells in N = 3 independent experiments; \\* indicates that the MG132 treatment significantly reduced the percentage of cells with more stable minus-ends at the 2-centriole pole (p = 0.0275 in unpaired t-test). (****",
"K****) Distribution of spindle ratios R in wild-type and siKIF2a/MCAK-treated HeLa eGFP-centrin1/CENPA cells in metaphase. ",
"For cell number see [Table 1](#tbl1){ref-type=\"table\"}. (****",
"L****) Quantification of spindle rotation in wild type, Sas-6-depleted 2:1, KIF2a/MCAK-depleted, or KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin2/H2B-mRFP cells, n = 18--37 cells in 1--3 experiments. \\* ",
"indicates significant difference; p = 0.024 in Fisher\\'s exact test. ",
"See also [Videos 1--3](#video1 video2 video3){ref-type=\"other\"}. (**",
"M**) Distribution of spindle ratios R in Sas-6-depleted 2:1 and KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin1/CENPA cells in metaphase. ",
"For cell numbers see [Table 1](#tbl1){ref-type=\"table\"}. (**",
"N**) Distribution of spindle ratios R in metaphase in Sas-6-depleted 2:2 and 2:1 cells treated either with DMSO or 10 nM taxol. ",
"For cell numbers see [Table 1](#tbl1){ref-type=\"table\"}.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.008](10.7554/eLife.05124.008)Video 1.Sas-6-depleted 2:2 HeLa cell expressing eGFP-centrin1 (centriole marker; green) and mRFP-α-tubulin (microtubules; red) in mitosis.",
"Time is indicated in minutes.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.009](10.7554/eLife.05124.009)10.7554/eLife.05124.009Video 2.Sas-6-depleted 2:1 HeLa cell expressing eGFP-centrin1 (centriole marker; green) and mRFP-α-tubulin (microtubules; red) in mitosis.",
"Time is indicated in minutes. ",
"Note the spindle rotation movements.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.010](10.7554/eLife.05124.010)10.7554/eLife.05124.010Video 3.Sas-6-depleted 1:1 HeLa cell expressing eGFP-centrin1 (centriole marker; green) and mRFP-α-tubulin (microtubules; red) in mitosis.",
"Time is indicated in minutes.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.011](10.7554/eLife.05124.011)10.7554/eLife.05124.011\n\n2:1 cells form amphitelic kinetochore--microtubule attachments that take longer to stabilize {#s2-4}\n--------------------------------------------------------------------------------------------\n\nIn the next step, we investigated whether the difference in minus-end stability in 2:1 cells translated into kinetochore--microtubule attachment defects sensed by the SAC. ",
"The SAC responds to unattached kinetochores or to sister-kinetochores with insufficient tension (measured as the distance between the two sister-kinetochores) that become transiently detached due to the kinase activity of Aurora-B ([@bib8]). ",
"Depletion of centrobin on both spindle poles leads to unstable kinetochore--microtubules and reduced inter-kinetochore distances that result in a permanent mitotic arrest ([@bib15]). ",
"When we tracked kinetochores in 2:1 eGFP-centrin1/CENPA cells, however, we found no difference in inter-kinetochore distances or the oscillatory sister-kinetochore movements along the spindle axis ([Figure 4A,B](#fig4){ref-type=\"fig\"}; [@bib14]). ",
"Since 42.9% of the 2:1 cells have asymmetric spindles, we restricted our analysis to only include cells that have a spindle ratio R \\> 1.15, but again found no change in inter-kinetochore distances ([Figure 4A](#fig4){ref-type=\"fig\"}). ",
"As SAC satisfaction is also linked to intra-kinetochore stretching ([@bib24]; [@bib41]), we further measured the intra-kinetochore distance between CENPA and the outer kinetochore protein HEC1 and found no change in 2:1 cells compared to 2:2 or 1:1 cells ([Figure 4C](#fig4){ref-type=\"fig\"}); the observed distance of 110 nm is consistent with previous studies ([@bib43]). ",
"Overall, this indicated that sister-kinetochores in 2:1 cells formed bipolar attachments with mechanical behaviors that were indistinguishable from normal cells. ",
"Since a single unattached kinetochore is sufficient to elicit a SAC response ([@bib36]; [@bib3]), we next investigated whether 2:1 cells have rare unattached kinetochores, by measuring the proportion of metaphase cells with Mad2-positive kinetochores (marker for unattached kinetochores). ",
"When compared to 2:2, 1:1, or wild-type cells, we found a small but non-significant increase in the proportion of Mad2-positive 2:1 cells ([Figure 4D](#fig4){ref-type=\"fig\"}), which could point to a minor or transient attachment defect caused by the imbalance of microtubule stability within the spindles of 2:1 cells. ",
"Such imbalance could lead to kinetochores that are not fully attached, that is, not bound by the full complement of microtubules (up to 25 microtubules per kinetochore in vertebrate cells; [@bib34]). ",
"To detect immature attachments, metaphase eGFPcentrin1/CENPA cells were stained with antibodies against SKAP, a marker for kinetochores fully bound by stable microtubules ([@bib38]), and analyzed for the presence of SKAP on kinetochores: cells in which more than 30% of the kinetochore-pairs were SKAP-negative were considered to have partially unstable kinetochore--microtubules ([Figure 4E,F](#fig4){ref-type=\"fig\"}). ",
"Such cells were rare in wild-type, 2:2, or 1:1 cells (\\<10%), but formed a substantial proportion of 2:1 cells (22%; [Figure 4F](#fig4){ref-type=\"fig\"}). ",
"These SKAP-negative kinetochores were distributed in a symmetric manner (were present both on the 1- or 2-centriole side; data not shown), and they all disappeared if 2:1 cells were briefly treated with 10 nmol taxol ([Figure 4F](#fig4){ref-type=\"fig\"}). ",
"Moreover a 1-hr treatment with the proteasome inhibitor MG132 also abolished the subpopulation of 2:1 cells with SKAP-negative kinetochores ([Figure 4F](#fig4){ref-type=\"fig\"}). ",
"Overall this suggested that in 2:1 cells, sister-kinetochores form amphitelic microtubule attachments, but that differences in minus-end stability delay stabilization of kinetochore--microtubule attachments in both half-spindles, as reflected by higher levels of SKAP-negative kinetochores.10.7554/eLife.05124.012Figure 4.2:1 cells have immature kinetochore--microtubule attachments.(**A**, **B**) Analysis of inter-kinetochore distances and sister-kinetochore oscillations in wild-type, Sas-6-depleted 2:2, 1:1, 2:1, or the subset of 2:1 eGFP1-centrin1/CENPA metaphase cells with an asymmetric plate position based on our in-house kinetochore tracking assay ([@bib14]), n = 620--889 kinetochores in 36--48 cells. ",
"The distribution of inter-kinetochore distances (CENPA to CENPA distance) is shown in **A** (no significant difference, t-test, p = 0.99), and the autocorrelation of the sister-kinetochore movements in **B**. ",
"The first minima of the autocorrelation curve indicate the half-period of the chromosome oscillations, and their depth the regularity of the oscillations. (**",
"C**) Distribution of intra-kinetochore distances in wild-type and Sas-6-depleted 2:2, 1:1, or 2:1 eGFP1-centrin1/CENPA metaphase cells. ",
"Cells were stained with antibodies against the N-terminus of HEC1. ",
"Using the tracking assay, we determined for each sister-kinetochore pair the CENPA--CENPA and the HEC1-HEC1 distances, and calculated the CENPA-HEC1 distances by halving the difference, n = 701--790 kinetochores in 26--30 cells in 3 experiments (no significant difference, Mann--Whitney test, 2:1 vs 2:2, p = 0.203). (**",
"D**) Quantification of Mad2-positive kinetochores in wild-type or Sas-6-depleted 2:2, 1:1, or 2:1 cells. ",
"eGFP-centrin1 (green) metaphase cells were stained with anti-Mad2 (green), and CREST sera (magenta; left panel) and the number of Mad2-positive kinetochores quantified in the right panel (n = 50--102 cells in 2 (wt) or 8 (*siSas-6*) experiments; no significant difference was found; Fisher\\'s exact test, p = 0.17). ",
"Scale bar indicates 2 μm. (**",
"E**, **F**) Quantification of SKAP-negative kinetochores in wild-type, *siSas-6*-depleted 2:2, 2:1, 2:1 + DMSO, 2:1, 2:1 + MG132, and 2:1 + taxol-treated eGFP-centrin1/CENPA cells. ",
"Cells were stained with antibodies against the kinetochore protein SKAP (magenta), as shown in **E** (maximum-intensity projection of 8 stacks \\[z = 0.3 μm\\]). ",
"Using the eGFP-CENPA (green) signal, we quantified the number of sister-kinetochore pairs with at least one SKAP-negative kinetochore (as shown in inlets). ",
"Quantification in **F** shows the percentage of cells where more than 30% of the sister-pairs were SKAP-negative. ",
"Fisher\\'s exact test for 2:1 \\> 2:2, p = 0.0013. ",
"n = 46--72 cells in N = 4--7 experiments. ",
"Scale bar indicates 2 μm. (**",
"G**) Example of an eGFP-centrin1/CENPA (green) cell in which a single daughter centriole was ablated (white arrow indicates the location of the laser pulse). ",
"Cells were fixed and stained with anti-centrin sera (magenta) to confirm the loss of a centriole, as opposed to the mere bleaching of eGFP-centrin1. ",
"Scale bar indicates 5 μm. (**",
"H**) Plot of half-spindle ratio R over time in 11 single eGFP-centrin1/CENPA cells in which a single centriole was ablated. ",
"The time point of laser ablation is t = 0. ",
"The thick red curve indicates the median of R of laser ablated 2:1 cells (\\* denotes when median R is asymmetric \\[p \\< 0.01\\]), the thick black curve indicates the median R distribution of 8 control-ablated cells. (**",
"I**) Average inter-kinetochore distances in eGFP-centrin1/CENPA cell before or after a single daughter centriole was ablated as determined by the kinetochore tracking assay. ",
"Error bars indicate s.e.m. ",
"n = 11 cells, 2 time points before and after ablation and on average 20 kinetochores per cell. \\* ",
"denotes a statistically significant difference (p = 0.001 in two-tailed paired t-test).**DOI:** [http://dx.doi.org/10.7554/eLife.05124.012](10.7554/eLife.05124.012)\n\nSince our results contrasted with recent hypotheses that suggest that correct force generation at kinetochores is locally regulated and does not require a direct connection between kinetochores and centrosomes ([@bib40]), we aimed to exclude a possible off-target effect of Sas-6 depletion on kinetochore--microtubule attachments. ",
"For this purpose, we used laser-microsurgery to ablate one daughter centriole in cells that had already reached metaphase. ",
"As in Sas-6-depleted cells, 2:1 cells generated by microsurgery resulted in an asymmetric plate location, as, on average, R increased from 1.06 to 1.16 after laser-ablation ([Figure 4G,H](#fig4){ref-type=\"fig\"} and [Video 4](#video4){ref-type=\"other\"}). ",
"These cells displayed a reduced inter-kinetochore distance (mean of 1.03 μm before ablation to 0.95 μm after the ablation, p = 0.001 in paired two-tailed t-test), which led to a prolonged metaphase arrest ([Figure 4I](#fig4){ref-type=\"fig\"}); in contrast, a control laser pulse in the vicinity of the spindle pole did not affect half-spindle lengths or inter-kinetochore distances (data not shown). ",
"This confirmed that loss of a single daughter centriole can directly affect force generation at sister-kinetochores, consistent with our findings that Sas-6-depleted 2:1 cells show defects in the quality of kinetochore--microtubule attachments. ",
"We also conclude that an acute laser-ablation of a daughter centriole in metaphase has a more severe effect on the forces acting on kinetochore than Sas-6 depletion. ",
"Possible explanations for this difference could be that Sas-6-depleted 2:1 cells might have more time to adapt to the lack of a missing centriole as they progressively build up the spindle, or that the laser-ablation destroys not just a centriole but also part of enzymatic activities in the vicinity of the centriole, such as minus-end depolymerases, which in normal cells are known to exert a pulling force on kinetochore-fibers ([@bib29]).Video 4.Laser-ablated 2:1 HeLa cell expressing eGFP-centrin1 (centriole marker) and eGFP-CENPA (kinetochore marker) in metaphase.",
"Note the asymmetric metaphase plate position after the ablation of a single centriole.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.013](10.7554/eLife.05124.013)10.7554/eLife.05124.013\n\nStabilization of kinetochore--microtubules suppresses the SAC-dependent delay in 2:1 cells {#s2-5}\n------------------------------------------------------------------------------------------\n\nTo confirm the presence of partially destabilized kinetochore--microtubule attachments in *siSas-6*-treated 2:1 cells and test whether they are the cause of the SAC response, we used the Aurora-B inhibitor ZM1. ",
"Aurora-B inhibition does not overcome a SAC-dependent mitotic arrest caused by unattached kinetochores (nocodazole treatment), but overcomes a mitotic arrest caused by insufficient tension at sister-kinetochores in monopolar spindles (monastrol treatment), as Aurora-B inhibition stabilizes kinetochore--microtubules and prevents loss of kinetochore--microtubule attachment ([Figure 5A](#fig5){ref-type=\"fig\"}; [@bib7]; [@bib22]). ",
"When we acutely inhibited Aurora-B in metaphase 2:1 cells, cells entered anaphase with asymmetric spindles (median R = 1.18, Mann--Whitney U test, p = 7.57 × 10^−6^ when compared to anaphase onset without Aurora-B inhibitor), indicating that a stabilization of kinetochore--microtubules satisfies the SAC in 2:1 cells ([Figure 5B](#fig5){ref-type=\"fig\"}). ",
"To confirm these findings in an independent manner, we also tested whether depletion of the microtubule depolymerases KIF2a/MCAK would allow 2:1 cells to enter anaphase with asymmetric spindles. ",
"Depletion of KIF2a/MCAK per se did not affect plate position at anaphase onset (median R = 1.03); however, it allowed 2:1 cells to enter anaphase with asymmetric spindles (median R 1.08, p \\< 0.0001 compared to symmetric distribution in Mann--Whitney U test; [Figure 5C](#fig5){ref-type=\"fig\"}). ",
"KIF2a/MCAK depletion also rescued the anaphase timing difference in 2:1 cells compared to 2:2 cells and led to the correct loading of SKAP on kinetochores ([Figure 5D,E](#fig5){ref-type=\"fig\"}; note that KIF2a/MCAK depletion alone led to a small anaphase delay when compared to wild-type cells, as previously reported \\[[@bib10]\\]). ",
"This showed that co-depletion of KIF2a/MCAK overrides the SAC response in 2:1 cells, leaving them no time to center the metaphase plate, indicating that unstable kinetochore--microtubules cause the SAC-dependent anaphase delay. ",
"KIF2a/MCAK co-depletion, however, did not suppress a mitotic arrest caused by the presence of insufficient tension (monastrol treatment) or unattached kinetochores (nocodazole treatment; [Figure 5A](#fig5){ref-type=\"fig\"}). ",
"Since the spindle checkpoint response has been shown to be subject to off-target effects ([@bib13]; [@bib45]), we repeated these experiments with an alternative set of KIF2a and MCAK siRNAs. ",
"These experiments validated our initial findings, confirming that KIF2a/MCAK suppress the anaphase delay seen in Sas-6-depleted 2:1 cells ([Figure 5---figure supplement 1A](#fig5s1){ref-type=\"fig\"}). ",
"Validation of Sas-6, KIF2a, and MCAK depletion siRNA treatments by immunoblotting also showed that in the triple MCAK/KIF2a/Sas-6 depletion, KIF2a was only partially depleted ([Figure 5---figure supplement 1B](#fig5s1){ref-type=\"fig\"}). ",
"We therefore tested whether the depletion of MCAK alone in 2:1 cells is sufficient to suppress the mitotic delay. ",
"As this was not the case, we conclude that depletion of both MT-depolymerases is necessary to overcome the instability of kinetochore--microtubules in 2:1 cells ([Figure 5---figure supplement 1C](#fig5s1){ref-type=\"fig\"}).10.7554/eLife.05124.014Figure 5.Depleting KIF2a and MCAK overcomes the SAC response in 2:1 cells.(**A**) Mitotic index of untreated, ZM-1-treated, MPS1-IN-treated, control-depleted, or KIF2a/MCAK-depleted cells treated for 16 hr with nocodazole (unattached kinetochores) or monastrol (lack of tension), n ≥ 400 cells in 3--4 experiments, error bars indicate s.e.m. \\* ",
"ZM1 and MPS1-IN overcome a monastrol arrest (t-test p \\< 0.0001), and MPS1-IN overcomes a nocodazole arrest (t-test p = 0.0044). (**",
"B**) Distribution of spindle ratio R in Sas-6-depleted eGFP-centrin1/CENPA 2:1 cells at anaphase onset treated with or without the Aurora-B inhibitor ZM1. ",
"Data from [Figure 1E](#fig1){ref-type=\"fig\"} without Aurora-B inhibition are shown for comparison. ",
"Aurora-B inhibition allows cells to enter anaphase with asymmetric spindles (n = 12 cells; Mann--Whitney U test, p = 7.57 × 10^−6^). (**",
"C**) Distribution of spindle ratios R in Sas-6-depleted 2:1, KIF2a/MCAK-depleted, and KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin1/CENPA cells in metaphase or at anaphase onset. (**",
"D**) Boxplots of anaphase timing of wild-type, KIF2a/MCAK-depleted, or KIF2a/MCAK/Sas-6 depleted 2:2 and 2:1 eGFP1-centrin2/H2B-mRFP cells. ",
"n = 23--61 cells in 1--3 experiments. (**",
"E**) Quantification of SKAP-negative kinetochores as in [Figure 4F](#fig4){ref-type=\"fig\"} in Sas-6-depleted 2:1 and KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin1/CENPA cells. ",
"SAC, spindle assembly checkpoint.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.014](10.7554/eLife.05124.014)10.7554/eLife.05124.015Figure 5---figure supplement 1.Validation of Sas-6, KIF2a, and MCAK co-depletion.(**A**) Boxplots of anaphase timing of *siControl*, *siSas-6* 2:2, *siSas-6* 2:1, *siCtrl/KIF2a/MCAK*, *siKIF2a/MCAK/Sas-6* 2:2, and *siKIF2a/MCAK/Sas-6* 2:1 eGFP1-centrin1/CENPA cells. ",
"n = 14--78 cells in 3 independent experiments. ",
"Note that *siSas-6* 2:1 cells are delayed compared to siSas-6 2:2 cells (p \\< 0.00001 in Mann--Whitney test), but that *siKIF2a/MCAK/Sas-6* 2:1 are not delayed compared to *siKIF2a/MCAK/Sas-6* 2:2 cells. (**",
"B**) Immunoblots of eGFP1-centrin2/H2B-mRFP cells treated with the indicated siRNA and probed with anti-Sas6, anti-MCAK, anti-KIF2A and anti-α-tubulin (loading control) antibodies. ",
"The relative ratio with the α-tubulin signal is quantified for each condition on the right panels. ",
"Note the KIF2a siRNA only led to a partial depletion. (**",
"C**) Boxplots of anaphase timing of *siControl*, *siCtrl/MCAK, siMCAK/Sas-6* 2:2, *siMCAK/Sas-6* 2:1 eGFP1-centrin1/CENPA cells. ",
"n = 9--169 cells in 3 independent experiments.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.015](10.7554/eLife.05124.015)\n\nAnaphase entry with asymmetric spindles leads to asymmetric cell division {#s2-6}\n-------------------------------------------------------------------------\n\nSince the mere depletion of KIF2a/MCAK allowed 2:1 cells to enter anaphase with asymmetric spindles, we could investigate the functional importance of a centered metaphase plate position beyond anaphase onset (Aurora-B inhibition could not be used, since it blocks anaphase B and cytokinesis \\[[@bib7]\\]). ",
"We found that KIF2a/MCAK/Sas-6-depleted 2:1 cells had slightly higher rates of chromosome segregation errors (statistically insignificant difference, p = 0.4713, Fischer\\'s exact test) when compared to KIF2a/MCAK-depleted cells ([Figure 6A](#fig6){ref-type=\"fig\"}). ",
"Strikingly KIF2a/MCAK-depleted 2:1 cells also underwent asymmetric cell divisions, yielding two daughter cells of different volumes ([Figure 6B](#fig6){ref-type=\"fig\"} and [Videos 5, 6](#video5 video6){ref-type=\"other\"}): while the two daughter cell volumes never differed by more than 20% in wild-type or KIF2a/MCAK depleted cells, 50% of KIF2a/MCAK/Sas-6-depleted 2:1 cells yielded one daughter cell that was at least 20% larger than its sister progeny ([Figure 6C](#fig6){ref-type=\"fig\"}). ",
"This suggested that an asymmetric metaphase plate position leads to asymmetric cell division. ",
"A recent study, however, demonstrated that asymmetric position of the entire spindle can also lead to asymmetric cell divisions in HeLa cells ([@bib19]). ",
"To discriminate between the two possibilities, we quantified the position of the spindle center in relationship to the cell center at anaphase onset, which was often not centered in the middle of the cell at anaphase onset, as has been previously reported ([Figure 6D](#fig6){ref-type=\"fig\"}; [@bib4]; [@bib19]). ",
"The extent to which the spindle centers were offset was the same in KIF2a/MCAK-depleted and KIF2a/MCAK-depleted 2:1 cells, indicating that changes in spindle position were not at the origin of the asymmetric cell divisions seen in KIF2a/MCAK-depleted 2:1 cells. ",
"To independently confirm our hypothesis that an asymmetric metaphase plate position in anaphase leads to an asymmetric cell division, we acutely abrogated the spindle checkpoint with an Mps1 inhibitor in control-depleted cells, Sas-6-depleted 2:2 cells, Sas-6-depleted 2:1 cells in late prometaphase/early metaphase (as visualized by live-cell imaging, where 43% of the cells have a metaphase plate not in the middle of the spindle, see [Figure 1F](#fig1){ref-type=\"fig\"}) or Sas-6-depleted 2:1 cells that were in late metaphase, which have symmetric spindles. ",
"Mps1 inhibition did not give rise to asymmetric cell division in control-depleted cells, 2:2 cells, or 2:1 cells treated in late metaphase; in contrast, in late prometaphase/early metaphase 2:1 cells with an asymmetric plate position, Mps1 inhibition resulted in an asymmetric cell division in 45% of the cases ([Figure 6E](#fig6){ref-type=\"fig\"}). ",
"The same result was also seen after acute Mps1 inhibition in laser-ablated cells: cells that were treated with a laser pulse in the cytoplasm divided in a symmetric manner; in contrast, 67% of the cells in which a single centriole was laser-ablated divided in an asymmetric manner when treated with an Mps1 inhibitor ([Figure 6F](#fig6){ref-type=\"fig\"} and supplementary [Videos 7, 8](#video7 video8){ref-type=\"video\"}). ",
"We conclude that entry into anaphase with an asymmetric position of the metaphase plate results in an asymmetric cell division.10.7554/eLife.05124.016Figure 6.An asymmetric plate position at anaphase onset leads to segregation errors and asymmetric cell division.(**A**) Quantification of chromosome segregation errors in KIF2a/MCAK-depleted and KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin1/eGFP-CENPA cells based on time-lapse images. ",
"n = 12--21 cells in 2--4 experiments. ",
"Error bars indicate s.e.m. (**",
"B**, **C**) Wild-type, KIF2a/MCAK-depleted or KIF2a/MCAK/Sas-6-depleted 2:1 eGFP-centrin1/CENPA cells were recorded by time-lapse imaging using the eGFP-centrin1 signal to count centrioles and phase contrast to detect the cell membrane as shown in **B** for a siKIF2a/MCAK/Sas-6 2:1 cell (scale bar = 5 μm). ",
"Phase contrast images were used to quantify the ratio of the two daughter cell sizes, which was plotted as a histogram in **C**. ",
"Half the Kif2a/MCAK/Sas-6-depleted 2:1 cells had a ratio of over 1.2, a ratio never observed in other conditions (t-test with Welch\\'s correction between KIF2a/MCAK and KIF2a/MCAK/Sas-6 2:1, p = 3.1e-05; n = 20--45 cells in 2--4 experiments). (**",
"D**) Quantification of spindle center position in relation to cell center in KIF2a/MCAK-depleted or KIF2a/MCAK/Sas-6 2:1 eGFP-centrin1/CENPA cells at anaphase onset. ",
"n = 20--45 cells in 2--4 experiments. (**",
"E**) Wild-type HeLa eGFP-centrin1/eGFP-CENPA cells, 2:2 cells, 2:1 cells in late prometaphase (still 1--2 chromosomes not perfectly aligned on the plate), or 2:1 cells in late metaphase (plate perfectly in the middle) were treated with an Mps1 inhibitor and recorded by time-lapse imaging using phase contrast to detect the cell membrane as shown in **B**. ",
"Shown is the ratio of the two daughter cell sizes; 45% of the 2:1 cells treated in late prometaphase/early metaphase had a ratio of over 1.2, a ratio never observed in other conditions (t-test with Welch\\'s correction between 2:1 cells in late prometaphase and 2:1 cells in late metaphase, p = 0.0141; n = 11--17 cells in 3 experiments). ",
"See also [Videos 5, 6](#video5 video6){ref-type=\"other\"}. (**",
"F**) HeLa eGFP-centrin1/CENPA cells were treated with a laser pulse in the cytoplasm (control) or ablation of a centriole (2:1 cells) and acutely treated with an Mps1 inhibitor to force cells into anaphase. ",
"Shown is the ratio of the two daughter cell sizes. ",
"Note that 67% of the 2:1 cells treated in late prometaphase/early metaphase had a ratio of over 1.2, a ratio never observed in other conditions (t-test with Welch\\'s correction between control and 2:1 cells p = 0.0451; n = 6). ",
"See also [Videos 7, 8](#video7 video8){ref-type=\"other\"}.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.016](10.7554/eLife.05124.016)Video 5.KIF2a/MCAK-depleted HeLa cell expressing eGFP-centrin1 (centriole marker; green) and eGFP-CENPA (kinetochore marker; green) entering anaphase and recorded with phase contrast microscopy.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.017](10.7554/eLife.05124.017)10.7554/eLife.05124.017Video 6.KIF2a/MCAK/Sas-6-depleted 2:1 HeLa cell expressing eGFP-centrin1 (centriole marker; green) and eGFP-CENPA (kinetochore marker; green) entering anaphase and recorded with phase contrast microscopy.",
"Note the asymmetric cell division.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.018](10.7554/eLife.05124.018)10.7554/eLife.05124.018Video 7.Laser-ablated control (ablation in the cytoplasm) HeLa cell expressing eGFP-centrin1 (centriole marker) and eGFP-CENPA (kinetochore marker) treated in metaphase with an Mps1 inhibitor.",
"Shown is the GFP-fluorescence channel (left) and the DIC channel (right). ",
"Note how the cell divides in a symmetric manner.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.019](10.7554/eLife.05124.019)10.7554/eLife.05124.019Video 8.Laser-ablated 2:1 HeLa cell expressing eGFP-centrin1 (centriole marker) and eGFP-CENPA (kinetochore marker) treated in metaphase with an Mps1 inhibitor.",
"Shown is the GFP-fluorescence channel (left) and the DIC channel (right). ",
"Note how the cell divides in an asymmetric manner.**DOI:** [http://dx.doi.org/10.7554/eLife.05124.020](10.7554/eLife.05124.020)10.7554/eLife.05124.020\n\nDiscussion {#s3}\n==========\n\nHere, we show that an equatorial position of the metaphase plate in the middle of the spindle is necessary for symmetric cell divisions and demonstrate that cells actively center the metaphase plate before anaphase onset. ",
"Metaphase plate centering requires the SAC, which provides cells with enough time to correct metaphase plate position. ",
"The SAC responds to subtle defects in kinetochore--microtubule stability that arise in cells with an asymmetric plate position and an imbalance of centrioles, implying that the SAC is more sensitive than previously assumed.",
"\n\nRecent studies have shown that proper positioning of the spindle ensures symmetric cell divisions, and that, deviations from a symmetric position are corrected by dynein-dependent cortical forces and membrane elongation during anaphase ([@bib19]). ",
"Here, we find that this external cortical correction mechanism in anaphase is complemented in metaphase by an internal centering mechanism that ensures a symmetric position of the metaphase plate within the spindle with the help of the SAC. ",
"This centering mechanism is particularly visible in cells with an asymmetric distribution of centrioles (2:1 cells), but it also acts in wild-type cells, indicating that it is active in every cell division. ",
"The centering of the metaphase plate is not just a consequence of establishing stable bipolar attachments at kinetochores, since KIF2a/MCAK/Sas-6-depleted 2:1 cells fail to center the plate, despite having reached stable bipolar attachments that satisfy the SAC; it is an active correction process, which in part depends on the regulation of microtubule dynamics, but whose precise molecular mechanisms will need to be uncovered.",
"\n\nKIF2a/MCAK-depleted 2:1 cells fail to center the metaphase plate before anaphase and divide asymmetrically. ",
"These cells have no defect in spindle positioning when compared to the symmetrically-dividing KIF2a/MCAK-depleted cells, implying that the asymmetric position of the metaphase plate is the source of asymmetric cell division. ",
"This hypothesis is confirmed by our analysis of 2:1 cells treated with an Mps1 inhibitor: only 2:1 cells treated in late prometaphase/early metaphase, which still have asymmetrically positioned metaphase plates, give rise to asymmetric cell division, whereas the same 2:1 cell population treated in late metaphase, which have a centered metaphase plate position, carry out symmetric cell divisions. ",
"We thus postulate that a symmetric metaphase plate position is essential for symmetric cell divisions, explaining why it is conserved in all metazoans, plants, and many fungi. ",
"Control of this parameter is essential, since differences in cell size have been linked to cell fate ([@bib19]). ",
"Metaphase plate position may also play a crucial role in asymmetric cell divisions that depend on asymmetric spindles in anaphase, such as in embryonic *D. melanogaster* neuroblasts. ",
"To form asymmetric spindles in a controlled and stereotypical manner, cells need an internal reference in space: breaking an existing symmetry, that is, a symmetric metaphase plate position, provides such a reference point. ",
"This is consistent with the progression of embryonic fly neuroblasts, which first align the metaphase plate in the middle of the spindle, before undergoing an asymmetric elongation of the spindle in anaphase. ",
"Our results also shed light on the mechanisms controlling the position of the cytokinetic furrow. ",
"Original studies in sand dollar eggs showed that the position of the centrosomes is a key determinant of the cytokinetic furrow position ([@bib33]); later studies in *C. elegans* found that a second signal emanating from the spindle midzone also contributes to the positioning of the cytokinetic furrow ([@bib5]; [@bib1]). ",
"A role for chromosomes was, however, discarded in these two organisms, since midzone formation and cytokinesis did not require them. ",
"In contrast, in human cells, chromosomes stabilize microtubules of the midzone and thus favor the formation of a cytokinetic furrow ([@bib2]). ",
"Here, we show that 2:1 cells only misplace the cytokinetic furrow in the presence of an asymmetric plate position in metaphase, implying that the position of the metaphase plate plays a crucial fine-tuning role in the positioning of the cytokinetic furrow. ",
"Future studies will have to test whether the metaphase plate acts via the microtubules of the midzone, or as recently postulated, by influencing the cortical populations of Anillin and Myosin in anaphase in a Ran-GTP-dependent manner ([@bib19]).",
"\n\nThe depletion of KIF2a/MCAK satisfies the SAC in 2:1 cells, but not in cells with unattached or tension-free kinetochore-microtubule attachments, indicating that the SAC responds to kinetochore--microtubule attachments defects less severe than lack of attachment or a tension defect. ",
"What might be these defects? ",
"Kinetochores in 2:1 cells bind a sufficient number of microtubules to form amphitelic attachments and stretch the two sister-kinetochores apart, but a number of kinetochores do not bind the full complement of stable microtubules required for SKAP loading. ",
"It is established that the SAC responds to detached kinetochores and is satisfied when kinetochores have bound the full set of microtubules. ",
"Based on our results, we postulate that the SAC also responds if a kinetochore is only bound by a fraction of the full set of microtubules. ",
"Such a lack of full occupancy would delay anaphase onset, such as in 2:1 cells; it would also explain the tendency for a higher rate of chromosome segregation errors in 2:1 cells treated with an Mps1 inhibitor or KIF2a/MCAK siRNAs. ",
"Stabilizing those kinetochore--microtubules by inhibiting Aurora-B or depleting KIF2a/MCAK establishes full binding of microtubules, SKAP loading, and satisfies the SAC, allowing cells to enter anaphase. ",
"This suggests a SAC that is more sensitive than a checkpoint that only senses detached kinetochores or kinetochores that become detached due to a tension defect. ",
"A SAC that detects such minor defects in kinetochore--microtubule occupancy caused by an imbalance of microtubule stability within the spindle would be able to indirectly probe for plate positioning, giving cells time to correct this imbalance and ensure a symmetric metaphase plate position. ",
"Such graded response to microtubule occupancy within a kinetochore complements studies showing that the SAC acts in a graded manner when it comes to the number of unattached kinetochores ([@bib3]; [@bib6]).",
"\n\nMaterials and methods {#s4}\n=====================\n\nCell culture, siRNA, and drug treatments {#s4-1}\n----------------------------------------\n\nHeLa cells were grown in Dulbecco\\'s modified medium containing 10% Fetal Calf Serum (FCS), 100 U/ml penicillin, 100 mg/ml streptomycin, at 37°C with 5% CO~2~ in a humidified incubator. ",
"HeLa eGFP-centrin1/eGFP-CENPA, eGFP-Centrin1/mRFP-α-tubulin, and eGFP-Centrin2/H2B-mRFP (kind gift of U Kutay, ETH) cells were further maintained in 250 ng/ml puromycin and 250 μg/ml G418. ",
"Live-cell imaging experiments were performed at 37°C in Lab-Tek II chambers (Thermo Fischer, Switzerland) with Leibovitz L-15 medium containing 10% FCS. ",
"SiRNA oligonucleotides (Invitrogen and Thermo Fisher, Switzerland) against control (Scrambled), Sas-6, Mad2, Kid1, KIF2a, and MCAK were transfected using Oligofectamine (Invitrogen; [@bib28]; [@bib10]; [@bib23]; [@bib44]; [@bib27]). ",
"Mad2 and Kid1 depletion had been previously validated in our laboratory ([@bib28]; [@bib44]), Sas-6 depletion was validated by counting centrioles in all experiments; Mad2 and Sas-6 co-depletion was additionally validated by immunoblotting ([Figure 2---figure supplement 1](#fig2s1){ref-type=\"fig\"}); KIF2a/MCAK depletion in Sas-6-depleted cells was validated by immunoblotting ([Figure 2---figure supplement 1](#fig2s1){ref-type=\"fig\"}). ",
"To exclude off-target effects, KIF2a and MCAK were also depleted with an alternative set of pooled siRNAs (ON-TARGETplus Human KIF2C (11,004) and ON-TARGETplus Human KIF2a (3796) siRNA---SMARTpools; GE Healthcare, Switzerland). ",
"For drug treatments, cells were treated with 100 μM monastrol for 3 hr, 1 μM MG132 for 1 hr, or 10 nM taxol (all Sigma-Aldrich, Switzerland) for 15 min before fixation for immunofluorescence or live-cell imaging. ",
"The Mps1 inhibitors MPS1-IN-1 (10 μM; kind gift of NS Gray; \\[[@bib21]\\]) or Reversine (10 μM, Sigma--Aldrich), or the Aurora-B inhibitor ZM1 (2 μM, Tocris, United Kingdom) were added to metaphase cells during live-cell imaging. ",
"To determine the response to spindle poisons, cells were treated for 16 hr with combinations of 100 ng/ml nocodazole, 100 μM monastrol, 10 μM MPS1-IN-1 or 2 μM ZM1, and the percentage of mitotic cells determined by phase contrast microscopy.",
"\n\nImmunofluorescence {#s4-2}\n------------------\n\nCells were fixed with methanol at −20°C for 6 min, or with 20 mM PIPES(pH 6.8), 10 mM EGTA, 1 mM MgCl~2~, 0.2% Triton X-100, 4% formaldehyde for 7 min at room temperature. ",
"For the cold-stable assay, cells were incubated in cold medium whilst placed on ice for 7 min. ",
"The following primary antibodies were used: rabbit anti-Mad2 (1:1000; Bethyl); mouse anti-α-tubulin (1:10,000) and rabbit anti-γ-tubulin (1:2000; both Sigma--Aldrich); mouse anti-HEC1 (1:1000), mouse anti-pericentrin (1:2000, kind gift of U Kutay), rabbit anti-ninein (1:500), mouse anti-TPX-2 (1:250), and mouse anti-centrobin (1:1000; all Abcam, United Kingdom); mouse anti-p150^Glued^ (1:500; Becton Dickinson, Switzerland); rabbit anti-phospho-Aurora-A (1:1000; Cell signalling, Danvers, MA); rabbit anti-centrin (1:1000) and affinity-purified rabbit anti-SKAP (1 mg/ml; \\[[@bib38]\\]; both gifts of I Cheeseman). ",
"Cross-adsorbed secondary antibodies were used (Invitrogen). ",
"Three-dimensional image stacks of mitotic cells were acquired in 0.2-μm steps using a 100× NA 1.4 objective on an Olympus DeltaVision microscope (GE Healthcare) equipped with a DAPI/FITC/TRITC/CY5 filter set (Chroma, Bellow Falls, VT) and a CoolSNAP HQ camera (Roper Scientific, Tucson, AZ). ",
"For quantitative measurements, 3D image stacks were deconvolved with SoftWorx (GE Healthcare) and quantified with SoftWorx, Imaris (Bitplane, Switzerland) or ImageJ. Images were mounted as figures using Adobe Illustrator. ",
"Kinetochore protein intensities were measured as a ratio to the CREST signal as described ([@bib26]). ",
"To monitor the polar ejection force, the distance between centrosomes and kinetochores was measured as described ([@bib44]).",
"\n\nLive-cell imaging {#s4-3}\n-----------------\n\nFor mitotic timing experiments, cells were recorded every 3 or 4 min as three-dimensional image stacks (12 × 1 μm steps using a 60× 1.4 NA objective, or 7 × 2 μm stacks using a 40× 1.3 NA objective) on an Olympus DeltaVision microscope equipped with a GFP/mRFP filter set (Chroma) and a CoolSNAP HQ camera. ",
"To monitor cell contours, cells were illuminated with white light and recorded by phase-contrast microscopy. ",
"Time-lapse videos were visualized in Softworx to quantify mitotic timing and to detect rotating spindles.",
"\n\nKinetochore tracking, metaphase plate position, spindle positioning, intra-kinetochore distance, and SKAP assays {#s4-4}\n----------------------------------------------------------------------------------------------------------------\n\nFor kinetochore tracking, plate width and plate position experiments, fluorescence time-lapse imaging of metaphase HeLa eGFP--centrin1/eGFP-CENPA cells was recorded with a 100× 1.4 NA objective on an Olympus DeltaVision microscope. ",
"35 Z-sections 0.5 μm apart were acquired with a sampling rate of 7.5 s for a total duration of 5 min. ",
"Three-dimensional image stacks were deconvolved with SoftWorx and subjected to the kinetochore tracking assay analysis run in MATLAB (The Math Works, Inc, Natick, MA), to asses inter-kinetochore distances and kinetochore oscillations ([@bib14]). ",
"The tracking assay was also used to quantify the length of the two half-spindles: the tracking assay estimates the metaphase plate by fitting a plane to the calculated kinetochore positions; metaphase plate position relative to the spindle poles was calculated using a custom MATLAB function that detects centrioles and calculates plate position as the intersection of the fitted plane with the spindle axis. ",
"The earliest time point data of each cell imaged was used for plate position and inter-kinetochore distance analysis to ensure that data come from early metaphase cells. ",
"To measure plate position at anaphase and to better visualize the centering mechanisms, we used a temporal resolution of 30 s and applied our combined kinetochore and centrosome tracking analysis. ",
"Videos were manually screened for the presence of chromosome segregation errors. ",
"To determine spindle positions within cells, we used the centrosome positions to determine the center of the spindle (equidistant to both centrosomes) and compared it to the cell center, which was determined using phase contrast images (point on the spindle axis that is equidistant to both cell cortexes). ",
"To measure intra-kinetochore distances and SKAP signals, HeLa eGFP--centrin1/eGFP-CENPA cells were fixed and stained with anti-HEC1 and anti-SKAP antibodies, respectively. ",
"Three-dimensional image stacks of fixed cells were subjected to the kinetochore tracking assay for sister--kinetochore pair identification. ",
"Imaris was used in conjunction with a custom MATLAB function ([Source Code 1](#SD1-data){ref-type=\"supplementary-material\"}) to measure the HEC1 and SKAP signals of the detected sister-kinetochores.",
"\n\nLaser ablation experiments {#s4-5}\n--------------------------\n\nCentriole ablations were carried out by 2--4 series (10 Hz repetition rate) of second-harmonic, single-mode, 532-nm pulses of an Nd:YAG laser (ULTRA-CFR TEM00 Nd:YAG from Big Sky Laser, Quantel, United Kingdom). ",
"The pulse width was 8 ns and the pulse energy used was 1.5--2 μJ. A more detailed description of the laser-microsurgery unit can be found in ([@bib32]). ",
"Imaging and laser focusing was performed using a 100× 1.4 NA plan-Apochromatic DIC objective on a Nikon TE2000U inverted microscope equipped with a Yokogawa CSU-X1 spinning-disk confocal head and an iXon^EM^+ Electron Multiplying CCD camera.",
"\n\nStatistical methods {#s4-6}\n-------------------\n\nStatistical analyses were performed in R 2.15.0. ",
"Unpaired t-tests with Welch\\'s correction and Mann--Whitney U tests (against 2:2 cells) were carried out to check for the statistical significance of normal and non-normal distributed data, respectively. ",
"Count data were analyzed using the Fisher\\'s Exact test. ",
"Levene\\'s test of equality of variance was used to check for equal variances, using the lawstat package (α = 0.05). ",
"Graphs were plotted in R using the ggplot2 package and mounted in Adobe Illustrator.",
"\n\nFunding Information\n===================\n\nThis paper was supported by the following grants:\n\n- http://dx.doi.org/10.13039/501100004362Schweizerische Herzstiftung (Swiss Heart Foundation) 3100A0-120728 to Patrick Meraldi.",
"\n\n- http://dx.doi.org/10.13039/501100003006Eidgenössische Technische Hochschule Zürich (Federal Institute of Technology Zurich) to Patrick Meraldi.",
"\n\n- Université de Genève to Patrick Meraldi.",
"\n\n- http://dx.doi.org/10.13039/501100001706Louis-Jeantet Foundation (Fondation Louis-Jeantet) to Patrick Meraldi.",
"\n\n- http://dx.doi.org/10.13039/501100003043EMBO to Chia Huei Tan.",
"\n\n- http://dx.doi.org/10.13039/501100001645Boehringer Ingelheim Fonds (BIF) to Ivana Gasic.",
"\n\n- http://dx.doi.org/10.13039/501100000781European Research Council (ERC) to Helder Maiato.",
"\n\nWe thank the LMC of the ETH Zurich for microscopy support, U Kutay for the HeLa H2B-mRFP/eGFP-centrin2 cell line (ETHZ, CH), I Cheeseman for the SKAP and centrin antibodies (MIT, USA), Nathaniel Gray for the Mps1 inhibitor (Harvard, USA), and Ed Harry (Univ. ",
"of Warwick, UK) for adapting the tracking code to Nikon files. ",
"We thank J Pines (Univ. ",
"of Cambridge, UK), Andrew McAinsh (Univ. ",
"of Warwick, UK), Monica Gotta (Univ. ",
"of Geneva, CH), and the Meraldi lab members for critical discussions. ",
"PM was funded by an SNF-project grant, the Swiss Cancer league, the ETH Zurich, the University of Geneva and the Louis-Jeantet Foundation, CHT was supported by a short-term EMBO fellowship and IG by a Böhringer Ingelheim fellowship. ",
"CHT and IG are part of the MLS PhD School. ",
"HM is funded by the seventh framework program grant PRECISE from the European Research Council.",
"\n\nAdditional information {#s5}\n======================\n\nThe authors declare that no competing interests exist.",
"\n\nCHT, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article.",
"\n\nIG, Acquisition of data, Analysis and interpretation of data.",
"\n\nSPH-R, Acquisition of data, Analysis and interpretation of data.",
"\n\nDD, Acquisition of data, Analysis and interpretation of data.",
"\n\nMB, Acquisition of data, Analysis and interpretation of data.",
"\n\nHM, Acquisition of data, Analysis and interpretation of data.",
"\n\nPM, Conception and design, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article.",
"\n\nAdditional files {#s6}\n================\n\n10.7554/eLife.05124.021\n\n###### \n\nCustom built software in Matlab.",
"\n\n**DOI:** [http://dx.doi.org/10.7554/eLife.05124.021](10.7554/eLife.05124.021)\n\n10.7554/eLife.05124.022\n\nDecision letter\n\nPines\n\nJon\n\nReviewing editor\n\nThe Gurdon Institute\n\n,\n\nUnited Kingdom\n\neLife posts the editorial decision letter and author response on a selection of the published articles (subject to the approval of the authors). ",
"An edited version of the letter sent to the authors after peer review is shown, indicating the substantive concerns or comments; minor concerns are not usually shown. ",
"Reviewers have the opportunity to discuss the decision before the letter is sent (see [review process](http://elifesciences.org/review-process)). ",
"Similarly, the author response typically shows only responses to the major concerns raised by the reviewers.",
"\n\nThank you for sending your work entitled \"The equatorial position of the metaphase plate ensures symmetric cell divisions\" for consideration at *eLife*. ",
"Your article has been favorably evaluated by Tony Hunter (Senior editor), a Reviewing editor, and three reviewers.",
"\n\nThe Reviewing editor and the other reviewers discussed their comments before we reached this decision, and the Reviewing editor has assembled the following comments to help you prepare a revised submission.",
"\n\nThere is a general consensus that this paper is interesting and worthy of publication in *eLife* but a number of concerns must first be addressed. ",
"In particular, the authors should:\n\n1\\) Show how active the centering mechanism is with more time-lapse images. ",
"The authors should also show an R vs. time graph for all their individual traces.",
"\n\n2\\) Provide stronger arguments that the asymmetry of the plate in 2:1 cells is due to differences in minus end stability.",
"\n\n3\\) Improve the evidence that SKAP loading is a read-out for stability. ",
"The authors should also show data to clarify that SKAP loading is specifically reduced on the 1-centriole side, instead of sometimes being reduced on the 1-centriole side, and sometimes on the 2-centriole side.",
"\n\n4\\) Provide more data in support of the reproducibility and robustness of the laser ablation assay, and explain the change in spindle length after centriole ablation.",
"\n\n5\\) Back-up their claim that an asymmetric plate causes an asymmetric division.",
"\n\n6\\) Validate their RNAi and rescue experiments.",
"\n\nIn addition to these concerns the authors should comment on possible differences between laser ablation and Sas-6 depletion, and more clearly explain how they compute asymmetry. ",
"The authors should also make it explicit that the delay in mitosis observed in cells whose metaphase plate is off-centre is a consequence of an imbalance in microtubule forces resulting from kinetochore-microtubule occupancy that is monitored by the SAC.",
"\n\n10.7554/eLife.05124.023\n\nAuthor response\n\n*1) Show how active the centering mechanism is with more time-lapse images. ",
"The authors should also show an R vs. time graph for all their individual traces*.",
"\n\nThe R-values presented in the original manuscript were calculated using an automated tracking assay for kinetochores (GFP-CENP-A) and centrosomes (GFP-centrin1) recorded with a 7.5 seconds resolution. ",
"We knew from previous studies that this temporal resolution was necessary to catch record and analyze chromosome movements in metaphase (Jaqaman et al., ",
"J. Cell Biol. ",
"2010). ",
"To avoid photo-toxicity we were however forced to work with low intensities and to limit our movies to 5 minutes. ",
"This meant that only a subset (10-15%) of recorded cells entered anaphase within those 5 minutes, and that it is therefore not possible to visualize the full scale of the plate positioning correction, which often lasts more than 5 minutes, in such movies. ",
"Our representation of R is therefore built on a population analysis of cells either early in metaphase or just before anaphase. ",
"This population analysis shows that just before anaphase onset, cells have on average a metaphase plate that is located more precisely in the middle of the spindle.",
"\n\nTo improve the manuscript we first explain this fact in the text (subsection \"Cells center the metaphase plate position before anaphase onset\"); second, to better document the metaphase plate centering process we also recorded 15 minutes movies of 2:1 cells using a temporal resolution of 30s. ",
"We show in the novel [Figure 1G and 1H](#fig1){ref-type=\"fig\"} the R-values over time for 15 single cells, as well as the median R and the 95% confidence interval for the cell population as they approach anaphase. ",
"These experiments directly visualize and validate the existence of a centering mechanism, providing a clearer representation to the reader.",
"\n\n*2) Provide stronger arguments that the asymmetry of the plate in 2:1 cells is due to differences in minus end stability*.",
"\n\nFirst, we would like to point out that our data showed that depletion of KIF2a/MCAK only partially corrects the asymmetry of the metaphase plate position in 2:1 cells. ",
"This suggests that the difference in microtubule stability is an important factor in the asymmetry of the plate, but certainly not the only one. ",
"We have now made this fact clearer in our manuscript (subsection \"An asymmetric distribution of centrioles leads to an imbalance of microtubule stability\").",
"\n\nSecond, to further support our hypothesis of a differential minus-end stability that causes the asymmetric plate positioning in 2:1 cells, we measured with a cold-stable assay minus-end stability in 2:1 cells after a 1 hour MG132 treatment. ",
"Since MG132 treatment leads to a symmetric position of the metaphase plate, our hypothesis predicted an attenuated difference in minus end stability between the 1- and 2-centriole pole. ",
"This is exactly what we found, as now shown in the novel [Figure 3J](#fig3){ref-type=\"fig\"}.",
"\n\nTo more directly test our hypothesis we also tested whether a brief treatment with taxol, a drug that stabilizes microtubules, would correct metaphase plate position, as seen after KIF2a/MCAK depletion. ",
"As shown in the novel [Figure 3N](#fig3){ref-type=\"fig\"}, taxol treatment largely corrects the metaphase plate position, validating our hypothesis that the asymmetric plate position is caused by differences in microtubule stability.",
"\n\n*3) Improve the evidence that SKAP loading is a read-out for stability. ",
"The authors should also show data to clarify that SKAP loading is specifically reduced on the 1-centriole side, instead of sometimes being reduced on the 1-centriole side, and sometimes on the 2-centriole side*.",
"\n\nOur interpretation of SKAP as a marker for microtubule stability is not our claim, but a conclusion of Schmidt et al., ",
"JCB 2010, which showed that SKAP levels are inversely proportional to Aurora-B levels at kinetochores, which is itself inversely proportional to microtubule stability. ",
"This model is supported by our experiments showing that KIF2a/MCAK depletion in 2:1 cells restores normal SKAP levels at kinetochore, despite the presence of a partially asymmetric plate position. ",
"Since KIF2a and MCAK are both microtubule depolymerases that destabilize kinetochore-microtubules we concluded that SKAP is a read-out for microtubule stability. ",
"To fully validate this hypothesis we now in addition briefly treated 2:1 cells with 10 nM taxol and stained for SKAP. ",
"We found that 100% of the kinetochores were SKAP-positive, confirming that it is a read-out for k-fiber stability (novel [Figure 4F](#fig4){ref-type=\"fig\"}).",
"\n\nWith regard to the second question, we find that SKAP is not systemically absent on the 1-centriole-side or the 2-centriole-side, but that the frequency of the absence is the same for both sides. ",
"We do not think that this invalidates our model, as the SKAP read-out just indicates a reduced stability and higher dynamics of kinetochore-microtubules on both sides of the sister-kinetochores, consistent with the fact that these kinetochores are over time moving and correcting their position towards the middle of the spindle. ",
"This is now pointed out in our manuscript (subsection \"2:1 cells form amphitelic kinetochore-microtubule attachments that take longer to stabilize\")\n\n*4) Provide more data in support of the reproducibility and robustness of the laser ablation assay, and explain the change in spindle length after centriole ablation*.",
"\n\nTo provide a better overview of our laser ablation experiment, we now show the individual traces of R over time (before and after ablation) for 11 cells, in which a single centriole was ablated, and we plot the median of this cell population (novel [Figure 4H](#fig4){ref-type=\"fig\"}). ",
"As a negative control we also plot the median R over time for control-ablated cells (laser pulse in the cytoplasm). [",
"Figure 4H](#fig4){ref-type=\"fig\"} shows how R increases after destruction of a single centriole, leading to an asymmetric position of the metaphase plate, consistent with our Sas-6 depletion experiments. ",
"Moreover, we now provide a novel Supplementary movie 4, visualizing how the ablation of a single centriole creates an asymmetric position of the metaphase plate.",
"\n\nSince such laser-ablation experiments are technically very challenging, and not easy to combine with a second assay, we used mainly Sas-6 depletion to investigate how the removal of a single centriole affects spindle length, showing how it leads to an imbalance of microtubule dynamics (see also point 2 for the additional experiments).",
"\n\n*5) Back-up their claim that an asymmetric plate causes an asymmetric division*.",
"\n\nOur claim that an asymmetric plate position at anaphase onset was supported by two independent experiments: both KIF2a/MCAK depletion and Mps1 inhibition allow 2:1 cells to enter anaphase with asymmetric spindles resulting in an asymmetric cell division. ",
"Importantly, when Mps1 was inhibited in late metaphase cells, when the plate has reached a symmetric plate position, we only observed symmetric cell division. ",
"Since these cells are, apart from the plate position, identical to the 2:1 cells that divide asymmetrically, we concluded that the plate position is a key determinant for the (a)symmetry of cell division.",
"\n\nTo fully confirm this claim, we have now performed a second back-up experiment, in which we created 2:1 cells by laser-ablation (to create an asymmetric plate position), before forcing cells into anaphase by abrogating the spindle assembly checkpoint with an Mps1 inhibitor. ",
"We found that 4 out of 6 laser-ablated 2:1 cells underwent an asymmetric cell division (ratio between daughter cell size of more than 1.2, a ratio never seen in normal cells; this study but also [@bib19]). ",
"In contrast 6 out of 6 control-ablated cells had a ratio smaller than 1.08. (",
"p-value in unpaired t-test with Welch correction p = 0.0451; new [Figure 6F](#fig6){ref-type=\"fig\"}, and Supplementary movies 6 and 7). ",
"Despite the low numbers -- laser-ablation experiments are technically very challenging -- these experiments validate our initial claim.",
"\n\n*6) Validate their RNAi and rescue experiments*.",
"\n\nWhile our initial manuscript was based mainly on functional validations (e.g. number of centrioles for Sas-6 depletion), we have now validated in addition every key siRNA (Mad2, Sas-6, KIF2a and MCAK) by immunoblotting ([Figure 2--figure supplement 1](#fig2s1){ref-type=\"fig\"}; [Figure 5--figure supplement 1](#fig5s1){ref-type=\"fig\"}).",
"\n\nMoreover, the reviewers were worried about potential siRNA off-target effects, reason for which they asked for siRNA rescue experiments. ",
"Since such rescue experiments are technically extremely challenging when analyzing double or triple siRNAs and very difficult to analyze in a conclusive manner, we have instead performed a number of complementary experiments for each siRNA or provide additional explanations.",
"\n\nSas-6 siRNA: one possibility is that Sas-6 siRNA could affect the position of the metaphase plate, microtubule stability and SKAP loading on kinetochores and/or lead to a spindle checkpoint dependent anaphase delay as an off-target effect. ",
"However, this is not possible, since such an off-target effect would also be present in 1:1 cells that have been subjected to the same siRNA. ",
"1:1 cells show, however, none of the phenotypes seen in 2:1 cells, thus excluding any off target effect. ",
"Moreover, we confirm the effects of Sas-6 depletion on spindle positioning and asymmetric cell division using laser-ablation, as an alternative tool to create 2:1 cells (e.g. new [Figure 6F](#fig6){ref-type=\"fig\"}).",
"\n\nMad2 siRNA: this siRNA is used as a known tool to inactivate the spindle checkpoint ([Figure 2F](#fig2){ref-type=\"fig\"}). ",
"Instead of performing a rescue experiment, which is very difficult since mitotic timing is sensitive to minor changes in Mad2 levels, we repeated the Sas-6 depletion in the presence of an Mps1 inhibitor to inhibit the spindle assembly checkpoint in an independent manner. ",
"These experiments validated our Mad2 depletion experiments ([Figure 2--figure supplement 2](#fig2s2){ref-type=\"fig\"}).",
"\n\nKid siRNA: this siRNA was used as a positive control for a disruption of the polar ejection force, based on previous literature. ",
"It has been previously validated in our and other laboratories (Wandke et al., ",
"JCB 2012), and we functionally validate it as a positive control in [Figure 3A](#fig3){ref-type=\"fig\"}. ",
"In contrast, we find that Sas-6 depletion has no effect on the polar ejection force. ",
"Therefore, we cannot test for an off-target effect.",
"\n\nKIF2a/MCAK depletion: this double depletion has been validated in multiple publications (in particular from the Duane Compton laboratory), and the phenotype we report is consistent with the published literature. ",
"The novel aspect of our manuscript comes from co-depletion of Sas-6 siRNA in 2:1 cells. ",
"While we appreciate that a siRNA rescue experiment would be informative, we believe that a triple siRNA rescue experiment is nearly impossible, particularly since the overexpression of either of these microtubule depolymerases is toxic. ",
"To nevertheless exclude potential off-target effects, we repeated our experiments with an alternative set of KIF2a and MCAK siRNAs. ",
"As shown in [Figure 5--figure supplement 1](#fig5s1){ref-type=\"fig\"}, this alternative set of KIF2a/MCAK siRNAs rescued the mitotic delay seen in 2:1 cells, confirming our experiments with the original KIF2a/MCAK siRNAs. ",
"Moreover, we also validated in the same Figure the new siRNA treatment by immunoblotting. ",
"Since this experiment indicated that both the old and the new siRNA against KIF2a only led to a partial depletion of KIF2a (50%), we further tested whether depletion of MCAK alone would be sufficient to overcome the mitotic arrest seen in 2:1 cells ([Figure 5--figure supplement 1C](#fig5s1){ref-type=\"fig\"}). ",
"Since this is not the case, we conclude that KIF2a co-depletion is essential to overcome the mitotic delay.",
"\n\nFinally, we emphasize that other claims associated with KIF2A/MCAK co- depletion are fully backed up by independent experiments:\n\nOur claim that stabilization of kinetochore microtubules in 2:1 cells overcomes the spindle assembly checkpoint delay and allows anaphase entry with an asymmetric plate position is confirmed by the Aurora-B inhibition experiment;\n\nOur claim that 2:1/KIF2A/MCAK-depleted cells that enter anaphase with asymmetrically positioned metaphase plate undergo an asymmetric cell division is validated by our Mps1 inhibition experiments performed on Sas-6 depleted and on laser-ablated 2:1 cells.",
"\n\n*In addition to these concerns the authors should comment on possible differences between laser ablation and Sas-6 depletion, and more clearly explain how they compute asymmetry. ",
"The authors should also make it explicit that the delay in mitosis observed in cells whose metaphase plate is off-centre is a consequence of an imbalance in microtubule forces resulting from kinetochore-microtubule occupancy that is monitored by the SAC*.",
"\n\nWe envisage two potential reasons for the stronger phenotype seen in 2:1 cells obtained through laser ablation as compared to Sas-6 depletion. ",
"First, since Sas-6 depleted cells lose a centriole in interphase, they might be better able to partially compensate for this loss as they gradually build up the mitotic spindle. ",
"Second, it is likely that the laser pulse on a mitotic spindle pole not only impairs the centriole, but that it also disrupts biochemical activities associated with the mitotic spindle pole, such as microtubule-depolymerases, which are able to exert a pulling force on k-fibers (Meunier et al., ",
"Nat. ",
"Cell Biol, 2011). ",
"Both potential explanations are now mentioned in the text (subsection \"2:1 cells form amphitelic kinetochore-microtubule attachments that take longer to stabilize\").",
"\n\nWe now also explain in the text more explicitly, how we calculate R (subsection \"Cells center the metaphase plate position before anaphase onset\").",
"\n\nFinally, we now more explicitly link the occupancy of kinetochores with the activation of the spindle assembly checkpoint in 2:1 cells (see Discussion, fourth paragraph).",
"\n\n[^1]: These authors contributed equally to this work.",
"\n"
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[
"/*\n * Licensed to the Apache Software Foundation (ASF) under one or more\n * contributor license agreements. ",
" See the NOTICE file distributed with\n * this work for additional information regarding copyright ownership.",
"\n * The ASF licenses this file to You under the Apache License, Version 2.0\n * (the \"License\"); you may not use this file except in compliance with\n * the License. ",
" You may obtain a copy of the License at\n *\n * http://www.apache.org/licenses/LICENSE-2.0\n *\n * Unless required by applicable law or agreed to in writing, software\n * distributed under the License is distributed on an \"AS IS\" BASIS,\n * WITHOUT WARRANTIES OR CONDITIONS OF ANY KIND, either express or implied.",
"\n * See the License for the specific language governing permissions and\n * limitations under the License.",
"\n */\n\npackage org.apache.spark.sql.execution.streaming\n\nimport org.json4s.",
"DefaultFormats\nimport org.json4s.jackson.",
"Serialization\n\nimport org.apache.spark.sql.sources.v2\n\ncase class RateStreamOffset(partitionToValueAndRunTimeMs: Map[Int, ValueRunTimeMsPair])\n extends v2.reader.streaming.",
"Offset {\n implicit val defaultFormats: DefaultFormats = DefaultFormats\n override val json = Serialization.write(partitionToValueAndRunTimeMs)\n}\n\n\ncase class ValueRunTimeMsPair(value: Long, runTimeMs: Long)\n"
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[
"Long time no see right? ",
"We are happy to announce that XDK2MAM Phase 1 compromised to the EDF is now finished. ",
"This last six months were really exciting as we combined the development of various open source packages to connect the Bosch XDK110 sensors node with the Tangle and traveled as much as we could to conduct “hands on” Workshops and introduce IOTA’s Ledger and Masked Authenticated Messaging as a solution to decentralize information produced by this great device.",
"\n\nWe dictated workshops in several cities of Argentina, Uruguay, Brazil and Venezuela and there is more coming! ",
"Our next workshop will be conducted in Berlin! ",
"Yes, XDK2MAM is visiting Germany and we already have plans to meet the guys behind the XDK110 at Reutlingen, Daniel Trauth and his amazing IILA and, of course, people interested on IOTA in Berlin, home of the IF.",
"\n\nXDK2MAM Phase 1: XDK110-Tangle connectivity\n\nThe first part of XDK2MAM project aimed to provide documented open source code to connect the XDK110 with the Tangle, allowing people prototyping IoT with this powerful tool to have a way to safety store the data produced. ",
"Over the last six months we have provided with code to connect this device and the Tangle via:\n\nHTTP (Code+Guide / Videotutorial)\n\n(Code+Guide / Videotutorial) MQTT (Code+Guide / Videotutorial)\n\n(Code+Guide / Videotutorial) BLE (Code+Guide / Videotutorial)\n\n(Code+Guide / Videotutorial) UDP (Code+Guide / Videotutorial)\n\n(Code+Guide / Videotutorial) USB (Code+Guide / Videotutorial)\n\n(Code+Guide / Videotutorial) IOTA Data Marketplace PoC (Code+Guide / Videotutorial)\n\nDuring this six months we also provided open source code to connect the Ruuvitag beacon to the Tangle. ",
"Despite this was out of the milestones compromised to the EDF, we thought this little device was worth a little deviation.",
"\n\nXDK2MAM Phase 1: Dashboard application\n\nAll the above methods provided several means to send data from the XDK110 to the Tangle but, in all cases, we ended up with json datasets flooding the console. ",
"This is why, our project phase 1 last milestone was to develop what we call XDK2MAM Dashboard. ",
"This application was needed to:"
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[
"---\ndate: 2017-10-10T22:35:49-04:00\ntitle: \"Flexbox\"\nseo_title: \"Flexbox | CSS | GOUP\"\nsubheader:\n greeting: CSS - Style a Website\n description: This course covers the basics of styling a website using CSS. ",
"Work your way through the videos/articles and I'll teach you everything you need to know to style a basic website!",
"\ndescription: This tutorial covers how to use flexbox in CSS.",
"\nauthor: GOUP\nimage: flexbox.png\nvideo: 6FedyF0F2i8\nurl: /web-development/css/flexbox/\nweight: 13\n---\n\n## Code\n\n{{< code lang=\"html\" file=\"site.html\">}}\n<!-- ",
"classes 'box', 'box2' and 'container' (not shown) are used for styling the elements -->\n<div class=\"flex-container container\">\n <div class=\"box flex-item\"><h1>1</h1></div>\n <div class=\"box2 flex-item\"><h1>2</h1></div>\n <div class=\"box flex-item\"><h1>3</h1></div>\n <div style=\"align-self:center;\" class=\"box\"><h1>4</h1></div>\n <div class=\"box flex-item\"><h1>5</h1></div>\n</div>\n{{< /code >}}\n\n\n{{< code lang=\"css\" file=\"style.css\">}}\n.flex-container {\n display: flex;\n flex-direction: row;\n justify-content: space-around;\n flex-wrap: wrap;\n align-items: center;\n align-content: space-between;\n}\n{{< /code >}}"
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"Chrysoexorista ochracea\n\nChrysoexorista ochracea is a species of bristle fly in the family Tachinidae. ",
"It is found in North America.",
"\n\nReferences\n\nFurther reading\n\n \n \n\nCategory:Exoristinae\nCategory:Articles created by Qbugbot\nCategory:Insects described in 1890"
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[
"// Copyright (c) 2017, 2020, Oracle and/or its affiliates. ",
"All rights reserved.",
"\n// Licensed under the Mozilla Public License v2.0\n\npackage oci\n\nimport (\n\t\"context\"\n\t\"strconv\"\n\n\t\"github.com/hashicorp/terraform/helper/schema\"\n\toci_core \"github.com/oracle/oci-go-sdk/v25/core\"\n)\n\nfunc init() {\n\tRegisterDatasource(\"oci_core_boot_volumes\", CoreBootVolumesDataSource())\n}\n\nfunc CoreBootVolumesDataSource() *schema.",
"Resource {\n\treturn &schema.",
"Resource{\n\t\tRead: readCoreBootVolumes,\n\t\tSchema: map[string]*schema.",
"Schema{\n\t\t\t\"filter\": dataSourceFiltersSchema(),\n\t\t\t\"availability_domain\": {\n\t\t\t\tType: schema.",
"TypeString,\n\t\t\t\tRequired: true,\n\t\t\t},\n\t\t\t\"compartment_id\": {\n\t\t\t\tType: schema.",
"TypeString,\n\t\t\t\tRequired: true,\n\t\t\t},\n\t\t\t\"volume_group_id\": {\n\t\t\t\tType: schema.",
"TypeString,\n\t\t\t\tOptional: true,\n\t\t\t},\n\t\t\t\"boot_volumes\": {\n\t\t\t\tType: schema.",
"TypeList,\n\t\t\t\tComputed: true,\n\t\t\t\tElem: GetDataSourceItemSchema(CoreBootVolumeResource()),\n\t\t\t},\n\t\t},\n\t}\n}\n\nfunc readCoreBootVolumes(d *schema.",
"ResourceData, m interface{}) error {\n\tsync := &CoreBootVolumesDataSourceCrud{}\n\tsync.",
"D = d\n\tsync.",
"Client = m.(*OracleClients).blockstorageClient()\n\n\treturn ReadResource(sync)\n}\n\ntype CoreBootVolumesDataSourceCrud struct {\n\tD *schema.",
"ResourceData\n\tClient *oci_core.",
"BlockstorageClient\n\tRes *oci_core.",
"ListBootVolumesResponse\n}\n\nfunc (s *CoreBootVolumesDataSourceCrud) VoidState() {\n\ts.D.SetId(\"\")\n}\n\nfunc (s *CoreBootVolumesDataSourceCrud) Get() error {\n\trequest := oci_core.",
"ListBootVolumesRequest{}\n\n\tif availabilityDomain, ok := s.D.GetOkExists(\"availability_domain\"); ok {\n\t\ttmp := availabilityDomain.(string)\n\t\trequest.",
"AvailabilityDomain = &tmp\n\t}\n\n\tif compartmentId, ok := s.D.GetOkExists(\"compartment_id\"); ok {\n\t\ttmp := compartmentId.(string)\n\t\trequest.",
"CompartmentId = &tmp\n\t}\n\n\tif volumeGroupId, ok := s.D.GetOkExists(\"volume_group_id\"); ok {\n\t\ttmp := volumeGroupId.(string)\n\t\trequest.",
"VolumeGroupId = &tmp\n\t}\n\n\trequest.",
"RequestMetadata.",
"RetryPolicy = getRetryPolicy(false, \"core\")\n\n\tresponse, err := s.Client.",
"ListBootVolumes(context.",
"Background(), request)\n\tif err !",
"= nil {\n\t\treturn err\n\t}\n\n\ts.Res = &response\n\trequest.",
"Page = s.Res.",
"OpcNextPage\n\n\tfor request.",
"Page !",
"= nil {\n\t\tlistResponse, err := s.Client.",
"ListBootVolumes(context.",
"Background(), request)\n\t\tif err !",
"= nil {\n\t\t\treturn err\n\t\t}\n\n\t\ts.Res.",
"Items = append(s.",
"Res.",
"Items, listResponse.",
"Items...)\n\t\trequest.",
"Page = listResponse.",
"OpcNextPage\n\t}\n\n\treturn nil\n}\n\nfunc (s *CoreBootVolumesDataSourceCrud) SetData() error {\n\tif s.Res == nil {\n\t\treturn nil\n\t}\n\n\ts.D.SetId(GenerateDataSourceID())\n\tresources := []map[string]interface{}{}\n\n\tfor _, r := range s.Res.",
"Items {\n\t\tbootVolume := map[string]interface{}{\n\t\t\t\"availability_domain\": *r.AvailabilityDomain,\n\t\t\t\"compartment_id\": *r.CompartmentId,\n\t\t}\n\n\t\tif r.AutoTunedVpusPerGB !",
"= nil {\n\t\t\tbootVolume[\"auto_tuned_vpus_per_gb\"] = strconv.",
"FormatInt(*r.",
"AutoTunedVpusPerGB, 10)\n\t\t}\n\n\t\tif r.DefinedTags !",
"= nil {\n\t\t\tbootVolume[\"defined_tags\"] = definedTagsToMap(r.",
"DefinedTags)\n\t\t}\n\n\t\tif r.DisplayName !",
"= nil {\n\t\t\tbootVolume[\"display_name\"] = *r.DisplayName\n\t\t}\n\n\t\tbootVolume[\"freeform_tags\"] = r.FreeformTags\n\n\t\tif r.Id !",
"= nil {\n\t\t\tbootVolume[\"id\"] = *r.Id\n\t\t}\n\n\t\tif r.ImageId !",
"= nil {\n\t\t\tbootVolume[\"image_id\"] = *r.ImageId\n\t\t}\n\n\t\tif r.IsAutoTuneEnabled !",
"= nil {\n\t\t\tbootVolume[\"is_auto_tune_enabled\"] = *r.IsAutoTuneEnabled\n\t\t}\n\n\t\tif r.IsHydrated !",
"= nil {\n\t\t\tbootVolume[\"is_hydrated\"] = *r.IsHydrated\n\t\t}\n\n\t\tif r.KmsKeyId !",
"= nil {\n\t\t\tbootVolume[\"kms_key_id\"] = *r.KmsKeyId\n\t\t}\n\n\t\tif r.SizeInGBs !",
"= nil {\n\t\t\tbootVolume[\"size_in_gbs\"] = strconv.",
"FormatInt(*r.",
"SizeInGBs, 10)\n\t\t}\n\n\t\tif r.SizeInMBs !",
"= nil {\n\t\t\tbootVolume[\"size_in_mbs\"] = strconv.",
"FormatInt(*r.",
"SizeInMBs, 10)\n\t\t}\n\n\t\tif r.SourceDetails !",
"= nil {\n\t\t\tsourceDetailsArray := []interface{}{}\n\t\t\tif sourceDetailsMap := BootVolumeSourceDetailsToMap(&r.",
"SourceDetails); sourceDetailsMap !",
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"Thymoquinone protects rat liver after partial hepatectomy under ischaemia/reperfusion through oxidative stress and endoplasmic reticulum stress prevention.",
"\nIschaemia reperfusion (I/R) is associated with liver injury and impaired regeneration during partial hepatectomy (PH). ",
"The aim of this study was to investigate the effect of thymoquinone (TQ), the active compound of essential oil obtained from Nigella sativa seeds, on rat liver after PH. ",
"Male Wistar rats were divided equally into four groups (n = 6) receiving an oral administration of either vehicle solution (sham and PH groups) or TQ at 30 mg/kg (TQ and TQ + PH groups) for 10 consecutive days. ",
"Then, rats underwent PH (70%) with 60 minutes of ischaemia followed by 24 hours of reperfusion (PH and TQ + PH groups). ",
"Alanine aminotransferase (ALT) activity and histopathological damage were determined. ",
"Also, antioxidant parameters, liver regeneration index, hepatic adenosine triphosphate (ATP) content, endoplasmic reticulum (ER) stress and apoptosis were assessed. ",
"In response to PH under I/R, liver damage was significantly alleviated by TQ treatment as evidenced by the decrease in ALT activity (P < .01) and histological findings (P < .001). ",
"In parallel, TQ preconditioning increased hepatic antioxidant capacities. ",
"Moreover, TQ improved mitochondrial function (ATP, P < .05), attenuated ER stress parameters and repressed the expression of apoptotic effectors. ",
"Taken together, our results suggest that TQ preconditioning could be an effective strategy to reduce liver injury after PH under I/R. The protective effects were mediated by the increase of antioxidant capacities and the decrease of ER stress and apoptosis."
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"Background {#Sec1}\n==========\n\nExcess intake of a high-fat diet or Western diet is generally considered unhealthy because such a diet can cause many metabolic disorders such as obesity, diabetes and atherosclerosis \\[[@CR1]--[@CR3]\\]. ",
"Although this notion has been widely accepted, the underlying mechanisms responsible for these pathophysiological consequences are still not fully understood. ",
"Gut microbiota has received more attention for its roles in both physiological and pathophysiological processes \\[[@CR4]--[@CR7]\\]. ",
"It has been shown that many metabolic diseases are related with the changes in gut microbiota. ",
"Laboratory mice are frequently used for the study of human diseases but most strains of wild-type mice do not show any metabolic disorders on a normal chow diet. ",
"However, if WT mice were fed with a diet containing 21% fat along with 0.15% cholesterol, which is often terminated as a Western diet, they show elevated plasma lipids but they do not show obvious lesions of atherosclerosis. ",
"However, if mice are genetically deficient in apolipoprotein E (apoE) such as apoE knock-out (KO) mice, they are sensitive to a Western diet and rapidly develop hyperlipidemia and atherosclerosis. ",
"ApoE is a component of all lipoproteins except for low density lipoproteins (LDL) and functions as a ligand for LDL receptors and LDL-related proteins (LRP) on the liver thereby playing an important role in the catabolism of the remnants of chylomicrons and very low-density lipoproteins. ",
"The genetic deficiency of apoE gene leads to the accumulation of cholesterol-rich remnants in plasma in both humans and animals \\[[@CR8]--[@CR12]\\]. ",
"Therefore, Western diet-fed apoE KO mice are widely used to study many facets of human hyperlipidemia and atherosclerosis \\[[@CR13]--[@CR16]\\].",
"\n\nGiven the fact that the gut microbiota plays an important role in many metabolic diseases such as obesity, diabetes, metabolic syndrome \\[[@CR17]--[@CR20]\\] and atherosclerosis \\[[@CR21]--[@CR24]\\], we envisioned that the gut microbiota may be altered in apoE KO mice when challenged with a Western diet. ",
"Driven by the popularization of high throughput sequencing-based approaches, many meta-genomic studies were carried out to characterize the profile changes of the gut microbiome in atherosclerotic individuals \\[[@CR22], [@CR23]\\] and facilitated the comprehensive understanding of the interaction between the gut microbiota and host physiology. ",
"However, as the most commonly used atherogenic mouse model, the gut microbiota of apoE KO mice compared with WT mice has not been clarified and their relationship with metabolic and pathophysiological consequences remain unclear. ",
"In the current study, we investigated the gut microbiota profiling of apoE KO mice fed a chow and western diet compared with WT mice. ",
"Our data revealed that apoE KO mice exhibited unique features of the gut microbiota diversity and compositions and these features may provide novel insight into understanding the relationship between gut microbiota and atherosclerosis.",
"\n\nMethods {#Sec2}\n=======\n\nAnimals {#Sec3}\n-------\n\nIn this study, six-week-old male C57BL/6 J mice (wild type, WT) and apoE KO were used and divided into four groups: WT mice feed on normal chow diet (WT-CD), WT mice feed on western diet (WT-HC), apoE^−/−^ mice feed on normal chow diet (apoE-CD), and apoE KO mice feed on Western diet (apoE-HC) (*n* = 7\\~ 8). ",
"The composition of normal chow diet is based on the NIH feeds standard (NIH-07, NSN-8710-00-509-7915). ",
"The Western diet contains 21% fat and 0.15% cholesterol. ",
"The diets were provided by Beijing Keao Xieli Feed Co.,Ltd. ",
"All animals were provided ad libitum access to food and water and housed under the conventional 12 h: 12 h light/dark cycle for 12 weeks. ",
"The plasma total cholesterol (TC), Triglyceride (TG) and high-density lipoprotein cholesterol (HDL-C) levels were measured at 0, 4, 8, 12 weeks from the beginning of experiment. ",
"At the end of the experiment, all mice were sacrificed by a sodium pentobarbital overdose. ",
"The heart and artery tissue and intestinal contents were collected for further processing. ",
"Animal care and experimental procedures were approved by the Animal Care and Use Committee of Xi'an Jiaotong University.",
"\n\nPlasma lipid measurement {#Sec4}\n------------------------\n\nMice were fasted for 8 h before collecting the blood samples from tail veins. ",
"Then blood samples were centrifuged at 3000 g for 15 min and supernatants were collected. ",
"The plasma levels of TC, TG, and HDL-C were measured with commercially available kits from Biosino Bio-technology and Science (Beijing) according to the manufacturer's protocols.",
"\n\nAnalysis of atherosclerotic lesions {#Sec5}\n-----------------------------------\n\nMice were euthanized by a sodium pentobarbital overdose and flushed with saline at physiological pressure, then perfused with 4% paraformaldehyde through the left ventricle. ",
"The heart and aorta were dissected, removed and fixed with parafonnaldehyde. ",
"Aortas were open longitudinally, stained with Oil Red O and digitally scanned. ",
"En face lesion area was measured with WinROOF 6.5 (Mitani Co., Fukui, Japan). ",
"To quantify atherosclerosis at the aortic root, cross-cryosections were prepared according to our previous report \\[[@CR25]\\]. ",
"In brief, part of each heart between the lower tips of the right and left atria were embedded in optimal cutting temperature compound and the aortic root was sectioned (8 μm) serially. ",
"Corresponding sections on separate slides were treated with Oil Red O staining and analyzed by WinROOF 6.5.",
"\n\nImmunohistochemistry analysis {#Sec6}\n-----------------------------\n\nFor immunohistochemistry, corresponding sections were incubated with 3% hydrogen peroxide for 30 min and blocked with 2% horse serum for 2 h. Sections were then stained with anti-MOMA2 (Cat.",
"No.ab-33,451, abcam) or anti-α-SMA (AA132, Beyotime Biotech Inc) at 4 °C for overnight. ",
"The sections were then incubated with a secondary antibody against rat IgG (Thermo Scientific) for 1 h at room temperature. ",
"The sections were visualized with a DAB substrate kit (Vector Laboratories). ",
"Finally, image processing was carried out using a Leica DMRE microscope equipped with Spot digital image analysis software and camera.",
"\n\nGut bacterial DNA extraction and sequencing {#Sec7}\n-------------------------------------------\n\nAbout each of 0.5 g cecal sample was used for DNA extraction with QIAamp Fast DNA Stool Mini kit (Qiagen) according to the manufacturer's protocols. ",
"The extracted DNA was amplified with primers targeting V4-V5 hypervariable region of bacterial 16S rRNA genes (forward: 5-CCAGCAGCYGCGGTAAN-3; reverse: 5-CCGTCAATTCNTTTRAGT-3). ",
"PCR were then performed under the following condition: denaturation (95 °C, 3 mins), amplification (25 cycles), annealing (55 °C, 30 s), and elongation (72 °C, 30 s), and final elongation (72 °C, 5 mins). ",
"Sequencing was performed on the platform of Illumina HiSeq 2000. ",
"16S rRNA sequence were explored by the package of Quantitative Insights Into Microbial Ecology (QIIME, 1.9.0) \\[[@CR26]\\]. ",
"To avoid GC bias and low-quality nucleotides, the raw sequences were filtered with the following criteria: minimum Phred score, 20; minimum number of high-quality calls, 0.75; maximum number of consecutive low-quality base calls, 3. ",
"Trimmed sequences were then assembled \\[[@CR27]\\].",
"\n\nGut microbiota diversity and composition analysis {#Sec8}\n-------------------------------------------------\n\nSequence with 97% similarity were clustered into one operational taxonomic unit by UCLUST \\[[@CR28]\\]. ",
"The closed-reference taxonomy was assigned to genus level against the Greengenes database \\[[@CR29]\\] with 0.5 confidence threshold. ",
"Sequence occurred less than three times were removed from further analysis. ",
"Estimation of Shannon-Weiner index based on 16S rRNA gene were performed to assess the gut microbiota diversity with an average sequence number of 2,4000 reads per sample. ",
"Weighted uniFrac distance \\[[@CR30], [@CR31]\\] were used for the principle coordinate analysis.",
"\n\nIdentification of gut microbiota features that discriminate groups {#Sec9}\n------------------------------------------------------------------\n\nLinear Discriminant Analysis Effect Size (LEfSe) were performed to identify the bacterial communities that differentiate different groups of mice \\[[@CR32]\\]. ",
"Gut bacterial richness was used to calculate the effect size on either dietary or apoE deficiency. ",
"The threshold on logarithmic Linear Discriminant Analysis (LDA) score were set as 2 (default). ",
"One-way ANOVA was used to analyze differences between groups and Tukey's multiple comparison test was applied to correct for multiple tests.",
"\n\nStatistics {#Sec10}\n----------\n\nThe TG, TC and HLD-C levels were represented as mean ± SEM. ",
"The richness of dominant gut bacteria was presented by average and relative abundance with more than 0.5% were considered as dominate bacterial communities. ",
"Statistical analyses were performed by one-way ANOVA followed by Student's t-test. *",
"P* \\< 0.05 was considered as statistics significant. ",
"Pearson's correlation was calculated for each pairwise combination of each gut bacterial family versus atherosclerosis lesion area ratio using Graphpad Prism 6.",
"\n\nResults {#Sec11}\n=======\n\nDyslipidemia and atherosclerosis in apoE KO mice {#Sec12}\n------------------------------------------------\n\nOn a normal chow die[t]{.ul}, apoE KO mice had significantly higher levels of plasma TC but lower levels of HDL-C than WT mice but TG levels were unchanged (Fig.",
" [1a](#Fig1){ref-type=\"fig\"}, [b](#Fig1){ref-type=\"fig\"} and [c](#Fig1){ref-type=\"fig\"}). ",
"On a Western diet, TC levels of WT mice were increased but no change in apoE KO mice compared to normal chow diet (Fig. [",
"1a](#Fig1){ref-type=\"fig\"}). ",
"The same trend was also found in HDL-C (Fig. [",
"1c](#Fig1){ref-type=\"fig\"}). ",
"In spite of this, there were no aortic lesions in WT mice on both a chow and Western diet but atherosclerosis was observed in apoE KO mice on both a chow and Western diet (Fig. [",
"1d](#Fig1){ref-type=\"fig\"}, [e](#Fig1){ref-type=\"fig\"} and [f](#Fig1){ref-type=\"fig\"}). ",
"Atherosclerotic lesions were significantly increased in Western diet-fed apoE KO mice compared with chow-fed apoE KO mice (Fig. [",
"1d](#Fig1){ref-type=\"fig\"}, [e](#Fig1){ref-type=\"fig\"} and [f](#Fig1){ref-type=\"fig\"}). ",
"Moreover, the macrophage and smooth muscle cells increased in high cholesterol feeding apoE KO mice compared to normal chow diet (Fig. [",
"1e](#Fig1){ref-type=\"fig\"}).Fig. ",
"1Western diet and apoE gene deletion elevate blood lipids and aggravate atherosclerosis. **",
"a** Total cholesterol (TC); (**b**) Triglycerides (TG); (**c**) High-density lipoprotein cholesterol (HDL-C) levels at 12 weeks in the plasma samples from four groups of mice; (**d**) Cross sections of aortic roots from four groups of mice were stained with Oil-Red-O; (**e**) Macrophage and smooth muscle cells staining of apoE KO mice; (**f**) En face Oil-Red-O staining of aortic specimens with the quantitive analysis on the right panel. ",
"Data are expressed as the mean ± SEM, *n* = 7\\~ 8 for each group. ",
"^∗^*P* \\< 005, ^∗∗∗^*P* \\< 0001 versus wild type (WT) mice\n\nThe gut microbiota alteration in apoE KO mice {#Sec13}\n---------------------------------------------\n\nTo shed light on the gut microbiota distribution in mice, we sequenced V4-V5 amplicons of 16S rRNA genes. ",
"After quality control, we obtained a total of 2,766,806 sequences (69,170 ± 26,490 per sample) with the average length of 327 bp (Additional file [1](#MOESM1){ref-type=\"media\"}: Figure S1). ",
"As expected, phylum-level composition was dominated by three phyla (*Bacteroidetes*, *Firmicutes*, and *Proteobacteria*), accounting for 84.7%\\~ 87.1% of the gut bacterial community (Additional file [2](#MOESM2){ref-type=\"media\"}: Figure S2). ",
"Of these phyla, the most dominant phylum was *Bacteroidetes* with a richness of 36.4%\\~ 50.9%, followed by *Firmicutes* with 17.6%\\~ 30.6%, and *Proteobacteria* with 15.6\\~ 20.1% (Additional file [2](#MOESM2){ref-type=\"media\"}: Figure S2). ",
"The other phyla such as *Actinobacteria*, *Deferribacteres* and *Verrucomicrobia* also were classified, but these bacteria were present at relatively very low abundance (less than 0.5%). ",
"Family-level composition was dominated by *Lachnospiraceae, Porphyromonadaceae*, *Prevotellaceae*, *Ruminococcaceae*, *Geobacteraceae*, *Desulfovibrionaceae* and *Bacteroidaceae* (Additional file [3](#MOESM3){ref-type=\"media\"}: Figure S3). ",
"Estimation of Shannon-Weiner index, a measurement of the α-diversity of the gut microbiota, demonstrated that apoE KO mice had a tendency for decreasing the microbiota diversity when challenged with a Western diet (Fig.",
" [2a](#Fig2){ref-type=\"fig\"}) whereas it was unchanged in WT mice fed on a Western diet. ",
"Principal coordinate analysis based on weighted-unifrac distance showed a high degree of divergence between apoE KO and WT mice on the first vector, which accounts for 58.6% of the gut microbiota profile (Fig. [",
"2b](#Fig2){ref-type=\"fig\"}) although the gut microbiota change induced by different diets was indistinguishable (Fig. [",
"2b](#Fig2){ref-type=\"fig\"}). ",
"On normal chow diet, apoE KO mice showed significant variance of two phyla (*Firmicutes* and *Bacteroidetes*) and six families (*Lachnospiraceae, Ruminococcaceae, Desulfovibrionaceae, Helicobacteraceae, Erysipelotrichaceae* and *Veillonellaceae*) (Fig.",
" [3](#Fig3){ref-type=\"fig\"}) compared to WT mice. ",
"Western diet feeding led to the increase of *Firmicutes* and *Erysipelotrichaceae* in WT mice but *Erysipelotrichaceae* was not changed in apoE KO mice (Fig. [",
"3](#Fig3){ref-type=\"fig\"}).Fig. ",
"2The gut microbiota diversity and PCoA analysis of four groups of mice. **",
"a** The comparison of Shannon-Weiner index, representing the microbiota diversity for four groups of mice; (**b**) Principal coordinate analysis (PCoA) for gut microbiota of four groups of miceFig. ",
"3Comparison of gut microbiota among four groups of mice. *",
"T*-test was applied to detect the statistical significance. ",
"Data are expressed as the mean ± SEM, *n* = 7\\~ 8 for each group. ",
"^∗^*P* \\< 005 versus wild type (WT) mice\n\nLEfSe and correlation analysis {#Sec14}\n------------------------------\n\nThe strict version of LEfSe (all against all) detected eight gut bacterial clades showing significant differences between aopE KO mice fed a Western diet and WT mice on a normal chow diet with LDA scores higher than two (Fig.",
" [4a](#Fig4){ref-type=\"fig\"}). ",
"The phylum of *Firmicutes* and *Lachnospiraceae* (from class to family) were found to be enriched in apoE KO mice on a Western diet, while *Bifidobacteriales* (from order to family), *Negativicutes*, *Selenomonadales* and *Veillonellaceae* were enriched in WT mice on a normal chow diet (Fig. [",
"4a](#Fig4){ref-type=\"fig\"}). ",
"In order to explore whether gut microbiota have any correlations with atherosclerosis in apoE KO mice, we performed a correlation analysis of family-level gut microbiota and aortic atherosclerosis lesion area. ",
"The relative abundances of *Lachnospiraceae* and *Ruminococcaceae* were found to be positively correlated with atherosclerosis lesion area (*P* \\< 0.05) (Fig. [",
"4b](#Fig4){ref-type=\"fig\"} and [c](#Fig4){ref-type=\"fig\"}).Fig. ",
"4Linear regression plots and LDA Effect Size (LEfSe) analysis. **",
"a** Cladogram generated with Linear Discriminant Analysis (LAD) Effect Size (LEfSe) analysis illustrating significant shifts in the gut microbiota in the four groups of mice (n = 7\\~ 8). ",
"Size of yellow circles is proportionated to each taxon's mean relative abundance; (**b**) Linear regression of *Lachnospiracea* correlated with relative atherosclerosis lesions; (**c**) Linear regression of *Ruminococcaceae* correlated with relative atherosclerosis lesions. *",
"P* \\< 0.05 and Pearson's correlation coefficient (r) \\> 0.6\n\nDiscussion {#Sec15}\n==========\n\nIn the current study, we analyzed the gut microbiota of apoE KO mice and compared with WT mice on either a normal chow or Western diet. ",
"It is well known that apoE KO mice can develop atherosclerosis even on a normal chow diet and moreover, a Western diet is very atherogenic and accelerates the development of atherosclerosis in apoE KO mice. ",
"In spite of this, it is not clear whether Western diet feeding influences the gut microbiota in apoE KO mice or whether there are any associations between the altered gut microbiota profiling with atherosclerosis. ",
"Our results showed that the gut microbiota diversity and composition were remarkably changed in apoE KO mice, especially on a Western diet. *",
"Firmicutes* and *Clostridia* (from class to family) were found to be enriched in apoE KO mice on a Western diet by LEfSe analysis. ",
"It has been reported that the relative abundance of *Firmicutes* could lead to the formation of increased amounts of metabolic endotoxins like lipopolysaccharides, which was able to enter the blood steam and cause the chronic inflammation \\[[@CR33]\\]. ",
"Three species, including *Clostridium asparagiforme*, *Clostridium hathewayi*, and *Clostridium sporogenes* in the family of *Clostridiales*, were also reported to be correlated with the accumulation of trimethylamine N-oxide (TMAO) \\[[@CR34], [@CR35]\\]. ",
"Atherosclerosis is considered as a chronic inflammatory process and previous studies have identified that the gut microbiota-dependent TMAO production directly contributed to the progression of atherosclerosis, indicating that gut microbiota participates in the development of atherosclerosis \\[[@CR24], [@CR35]--[@CR37]\\]. ",
"TMAO was able to accelerate atherosclerosis by disrupting the cholesterol transport and modifying bile acids composition \\[[@CR38]\\], enhancing thrombosis formation by activating platelets \\[[@CR39]\\], and increasing high blood pressure induced heart failure by adverse cardiac remodeling \\[[@CR40]\\]. ",
"In our study, *Lachnospiraceae* and *Ruminococcaceae*, the two most abundant families in the order of *Clostridiales*, were found to be correlated with the atherosclerosis size. ",
"Although it is not possible to ascertain whether such correlation is causal or casual, the further studies are required to investigate its significance in future. ",
"Nevertheless, it has also been reported that the members of *Lachnospiraceae* were able to cause diabetes mellitus in germ-free mice \\[[@CR41]\\]. ",
"Therefore, the gut microbiota transplantation would be performed using germ-free mice to determine the causal relationship in the future.",
"\n\nConclusions {#Sec16}\n===========\n\nCollectively, our study showed that there are marked changes in the gut microbiota of apoE KO mice, particularly challenged with a Western diet and these alterations may be possibly associated with atherosclerosis.",
"\n\nAdditional files\n================\n\n {#Sec17}\n\nAdditional file 1:**Figure S1.** ",
"Reads number distribution of each sample based on 16S rRNA gene sequencing. (",
"JPG 262 kb) Additional file 2:**Figure S2.** ",
"Dominant gut microbiota compositions of samples at phyla level. (",
"JPG 96 kb) Additional file 3:**Figure S3.** ",
"Dominant gut microbiota compositions of samples at family level. (",
"JPG 144 kb)\n\nBaoning Liu and Yali Zhang contributed equally to this work.",
"\n\n**Electronic supplementary material**\n\nThe online version of this article (10.1186/s12944-018-0811-8) contains supplementary material, which is available to authorized users.",
"\n\nFunding {#FPar1}\n=======\n\nThis work was supported in part by the National Natural Science Foundation of China (No. ",
"81270348, EL), the Natural Science Foundation of Shaanxi Province (2014FWPT017, EL), the China Postdoctoral Science Foundation (2014 M562425, RW).",
"\n\nAvailability of data and materials {#FPar2}\n==================================\n\nThe data supporting the findings of this study are available from the corresponding author upon request.",
"\n\nBL performed bioinformatics analysis and wrote the manuscript; YZ performed the experiments and wrote the manuscript. ",
"YA, WG, LB and YL performed the experiments; RW and SZ interpreted the data and revised the manuscript; EL and JF designed the experiments and read the manuscript. ",
"All authors have approved the final version to be published.",
"\n\nEthics approval {#FPar3}\n===============\n\nAnimal care and experimental procedures were approved by the Animal Care and Use Committee of Xi'an Jiaotong University.",
"\n\nConsent for publication {#FPar4}\n=======================\n\nNot applicable.",
"\n\nCompeting interests {#FPar5}\n===================\n\nThe authors declare that they have no competing interests.",
"\n\nPublisher's Note {#FPar6}\n================\n\nSpringer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.",
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"Bulldogs Continue Competition on Day Two at Penn Relays\n\nPHILADELPHA – The Bulldogs had their\nbusiest day of the Penn Relays on Friday, competing in four field\nevents and three relays. ",
"The Penn Relays were an opportunity for a\nselect group of Bulldogs to race at one of the most renowned track\nand field events in the U.S., and several of them took some of the\ntop spots in their divisions.",
"\n\nFreshman Mike Levine placed third in the\nChampionship division of the discus, coming through with a huge PR\nof 54.41m to pass his previous best by over nine feet. ",
"Senior\nDavid Smith also competed in the Eastern division\nof the shot put, placing 21st with a throw of 14.61m.\nSenior Eric DePalo made an impressive fifth-place\nfinish in the Eastern division of the pole vault with a mark of\n4.80m. In the long jump, senior Reynolds Holmes\nalso competed in the Eastern division and placed 20th\nwith a jump of 6.78m.\n\nThe relay teams also ran several top performances. ",
"In their\nfirst race of the outdoor season, the distance medley relay took\nfifth in the college men's division with seniors Chris\nLabosky and Remi Ray, sophomore\nConor Dooney and freshman Chris\nRamsey running for the Bulldogs. ",
"Their time of 9:50.44\nalso beat fellow Ivy League schools UPenn and Dartmouth. ",
"The 4x100\nmeter relay team of Holmes, junior Marty Evans,\njunior Chris Stanley and sophomore Matt\nBieszard placed sixth in their heat and 37th\noverall out of 87 teams in the college men's division (42.22), only\nfive spots away from qualifying for the College final on Saturday.",
"\nEvans, Stanley and Bieszard came back later in the day along with\nsenior David Soiles to place fourth in the\nHeptagonal section of the 4x400-meter relay, which is reserved for\nIvy League schools. ",
"The team ran 3:18.74 to beat Brown and\nDartmouth.",
"\n\nThe rest of the team will compete at home on Sunday at the\nYale Springtime Invitational before training for Heptagonals\nChampionships on May 8-9."
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"/*\n * This file is part of the GROMACS molecular simulation package.",
"\n *\n * Copyright (c) 2010-2018, The GROMACS development team.",
"\n * Copyright (c) 2019, by the GROMACS development team, led by\n * Mark Abraham, David van der Spoel, Berk Hess, and Erik Lindahl,\n * and including many others, as listed in the AUTHORS file in the\n * top-level source directory and at http://www.gromacs.org.",
"\n *\n * GROMACS is free software; you can redistribute it and/or\n * modify it under the terms of the GNU Lesser General Public License\n * as published by the Free Software Foundation; either version 2.1\n * of the License, or (at your option) any later version.",
"\n *\n * GROMACS is distributed in the hope that it will be useful,\n * but WITHOUT ANY WARRANTY; without even the implied warranty of\n * MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. ",
" See the GNU\n * Lesser General Public License for more details.",
"\n *\n * You should have received a copy of the GNU Lesser General Public\n * License along with GROMACS; if not, see\n * http://www.gnu.org/licenses, or write to the Free Software Foundation,\n * Inc., 51 Franklin Street, Fifth Floor, Boston, MA 02110-1301 USA.",
"\n *\n * If you want to redistribute modifications to GROMACS, please\n * consider that scientific software is very special. ",
"Version\n * control is crucial - bugs must be traceable. ",
"We will be happy to\n * consider code for inclusion in the official distribution, but\n * derived work must not be called official GROMACS. ",
"Details are found\n * in the README & COPYING files - if they are missing, get the\n * official version at http://www.gromacs.org.",
"\n *\n * To help us fund GROMACS development, we humbly ask that you cite\n * the research papers on the package. ",
"Check out http://www.gromacs.org.",
"\n */\n/*! ",
"\\internal \\file\n * \\brief\n * Implements gmx::CommandLineHelpWriter.",
"\n *\n * \\author Teemu Murtola <teemu.murtola@gmail.com>\n * \\ingroup module_commandline\n */\n#include \"gmxpre.h\"\n\n#include \"cmdlinehelpwriter.h\"\n\n#include <cstring>\n\n#include <algorithm>\n#include <string>\n\n#include \"gromacs/commandline/cmdlinehelpcontext.h\"\n#include \"gromacs/onlinehelp/helpwritercontext.h\"\n#include \"gromacs/options/basicoptions.h\"\n#include \"gromacs/options/filenameoption.h\"\n#include \"gromacs/options/options.h\"\n#include \"gromacs/options/optionsvisitor.h\"\n#include \"gromacs/options/timeunitmanager.h\"\n#include \"gromacs/utility/arrayref.h\"\n#include \"gromacs/utility/exceptions.h\"\n#include \"gromacs/utility/stringutil.h\"\n#include \"gromacs/utility/textwriter.h\"\n\n#include \"shellcompletions.h\"\n\nnamespace gmx\n{\n\nnamespace\n{\n\n//! ",
"\\addtogroup module_commandline\n//! \\{",
"\n\n/********************************************************************\n * IOptionsFormatter\n */\n\n/*! ",
"\\brief\n * Interface for output format specific formatting of options.",
"\n *\n * \\see OptionsFilter\n */\nclass IOptionsFormatter\n{\npublic:\n virtual ~IOptionsFormatter() {}\n\n //! ",
"Formats a single option option.",
"\n virtual void formatOption(const OptionInfo& option) = 0;\n};\n\n/********************************************************************\n * OptionsFilter\n */\n\n/*! ",
"\\brief\n * Output format independent processing of options.",
"\n *\n * Together with code in CommandLineHelpWriter::writeHelp(), this class\n * implements the common logic for writing out the help.",
"\n * An object implementing the IOptionsFormatter must be provided to the\n * constructor, and does the actual formatting that is specific to the output\n * format.",
"\n */\nclass OptionsFilter : public OptionsVisitor\n{\npublic:\n //! ",
"Specifies the type of output that formatSelected() produces.",
"\n enum FilterType\n {\n eSelectInputFileOptions,\n eSelectInputOutputFileOptions,\n eSelectOutputFileOptions,\n eSelectOtherOptions\n };\n\n /*! ",
"\\brief\n * Creates a new filtering object.",
"\n *\n * Does not throw.",
"\n */\n OptionsFilter() : formatter_(nullptr), filterType_(eSelectOtherOptions), bShowHidden_(false) {}\n\n //! ",
"Sets whether hidden options will be shown.",
"\n void setShowHidden(bool bShowHidden) { bShowHidden_ = bShowHidden; }\n\n //! ",
"Formats selected options using the formatter.",
"\n void formatSelected(FilterType type, IOptionsFormatter* formatter, const Options& options);\n\n void visitSection(const OptionSectionInfo& section) override;\n void visitOption(const OptionInfo& option) override;\n\nprivate:\n IOptionsFormatter* formatter_;\n FilterType filterType_;\n bool bShowHidden_;\n\n GMX_DISALLOW_COPY_AND_ASSIGN(OptionsFilter);\n};\n\nvoid OptionsFilter::formatSelected(FilterType type, IOptionsFormatter* formatter, const Options& options)\n{\n formatter_ = formatter;\n filterType_ = type;\n visitSection(options.rootSection());\n}\n\nvoid OptionsFilter::visitSection(const OptionSectionInfo& section)\n{\n OptionsIterator iterator(section);\n iterator.acceptSections(this);\n iterator.acceptOptions(this);\n}\n\nvoid OptionsFilter::visitOption(const OptionInfo& option)\n{\n if (!",
"bShowHidden_ && option.isHidden())\n {\n return;\n }\n const FileNameOptionInfo* const fileOption = option.toType<FileNameOptionInfo>();\n if (fileOption !",
"= nullptr && fileOption->isInputFile())\n {\n if (filterType_ == eSelectInputFileOptions)\n {\n formatter_->formatOption(option);\n }\n return;\n }\n if (fileOption !",
"= nullptr && fileOption->isInputOutputFile())\n {\n if (filterType_ == eSelectInputOutputFileOptions)\n {\n formatter_->formatOption(option);\n }\n return;\n }\n if (fileOption !",
"= nullptr && fileOption->isOutputFile())\n {\n if (filterType_ == eSelectOutputFileOptions)\n {\n formatter_->formatOption(option);\n }\n return;\n }\n if (filterType_ == eSelectOtherOptions)\n {\n formatter_->formatOption(option);\n return;\n }\n}\n\n/********************************************************************\n * CommonFormatterData\n */\n\nclass CommonFormatterData\n{\npublic:\n explicit CommonFormatterData(const char* timeUnit) : timeUnit(timeUnit) {}\n\n const char* timeUnit;\n};\n\n/********************************************************************\n * Helper functions\n */\n\n//! ",
"Formats option name and value.",
"\nvoid formatOptionNameAndValue(const OptionInfo& option, std::string* name, std::string* value)\n{\n *name = option.name();\n *value = \"<\" + option.type() + \">\";\n if (option.isType<FileNameOptionInfo>())\n {\n // TODO: Make these work also for other option types.",
"\n if (option.maxValueCount() !",
"= 1)\n {\n *value += \" [...]\";\n }\n if (option.minValueCount() == 0)\n {\n *value = \"[\" + *value + \"]\";\n }\n }\n if (option.isType<BooleanOptionInfo>())\n {\n *name = \"[no]\" + *name;\n // Old command-line parser doesn't accept any values for these.",
"\n // value = \"[\" + value + \"]\";\n value->clear();\n }\n}\n\n//! ",
"Formats the default option value as a string.",
"\nstd::string defaultOptionValue(const OptionInfo& option)\n{\n return joinStrings(option.defaultValuesAsStrings(), \" \");\n}\n\n//! ",
"Formats the flags for a file option as a string.",
"\nstd::string fileOptionFlagsAsString(const FileNameOptionInfo& option, bool bAbbrev)\n{\n std::string type;\n if (!",
"option.isRequired())\n {\n type = bAbbrev ? \"",
"Opt.\" : \"",
"Optional\";\n }\n if (option.isLibraryFile())\n {\n if (!",
"type.empty())\n {\n type.append(\", \");\n }\n type.append(bAbbrev ? \"",
"Lib.\" : \"",
"Library\");\n }\n return type;\n}\n\n//! ",
"Formats the description for an option as a string.",
"\nstd::string descriptionWithOptionDetails(const CommonFormatterData& common, const OptionInfo& option)\n{\n std::string description(option.formatDescription());\n\n const FloatOptionInfo* floatOption = option.toType<FloatOptionInfo>();\n const DoubleOptionInfo* doubleOption = option.toType<DoubleOptionInfo>();\n if ((floatOption !",
"= nullptr && floatOption->isTime())\n || (doubleOption !",
"= nullptr && doubleOption->isTime()))\n {\n // TODO: It could be nicer to have this in basicoptions.cpp.",
"\n description = replaceAll(description, \"%t\", common.timeUnit);\n }\n\n return description;\n}\n\n/********************************************************************\n * OptionsSynopsisFormatter\n */\n\n/*! ",
"\\brief\n * Formatter implementation for synopsis.",
"\n */\nclass SynopsisFormatter : public IOptionsFormatter\n{\npublic:\n //! ",
"Creates a helper object for formatting the synopsis.",
"\n explicit SynopsisFormatter(const HelpWriterContext& context) :\n context_(context),\n bFormatted_(false),\n lineLength_(0),\n indent_(0),\n currentLength_(0)\n {\n }\n\n //! ",
"Starts formatting the synopsis.",
"\n void start(const char* name);\n //! ",
"Finishes formatting the synopsis.",
"\n void finish();\n\n void formatOption(const OptionInfo& option) override;\n\nprivate:\n const HelpWriterContext& context_;\n bool bFormatted_;\n int lineLength_;\n int indent_;\n int currentLength_;\n\n GMX_DISALLOW_COPY_AND_ASSIGN(SynopsisFormatter);\n};\n\nvoid SynopsisFormatter::start(const char* name)\n{\n currentLength_ = std::strlen(name) + 1;\n indent_ = std::min(currentLength_, 13);\n TextWriter& file = context_.outputFile();\n switch (context_.outputFormat())\n {\n case eHelpOutputFormat_Console:\n lineLength_ = 78;\n file.writeString(name);\n break;\n case eHelpOutputFormat_Rst:\n bFormatted_ = true;\n lineLength_ = 74;\n indent_ += 4;\n file.writeLine(\".. parsed-literal::\");\n file.writeLine();\n file.writeString(\" \");\n file.writeString(name);\n break;\n default:\n GMX_THROW(NotImplementedError(\n \"Synopsis formatting not implemented for this output format\"));\n }\n}\n\nvoid SynopsisFormatter::finish()\n{\n context_.outputFile().ensureLineBreak();\n}\n\nvoid SynopsisFormatter::formatOption(const OptionInfo& option)\n{\n std::string name, value;\n formatOptionNameAndValue(option, &name, &value);\n int totalLength = name.length() + 4;\n std::string fullOptionText = formatString(\" [%s-%s\", bFormatted_ ? \":",
"strong:`\" : \"\", name.c_str());\n if (!",
"value.empty())\n {\n fullOptionText.append(bFormatted_ ? \"",
"` :emphasis:`\" : \" \");\n fullOptionText.append(value);\n totalLength += value.length() + 1;\n }\n fullOptionText.append(bFormatted_ ? \"",
"`]\" : \"]\");\n\n TextWriter& file = context_.outputFile();\n currentLength_ += totalLength;\n if (currentLength_ >= lineLength_)\n {\n file.writeString(formatString(\"\\n%*c\", indent_ - 1, ' '));\n currentLength_ = indent_ - 1 + totalLength;\n }\n file.writeString(fullOptionText);\n}\n\n/********************************************************************\n * OptionsListFormatter\n */\n\n/*! ",
"\\brief\n * Formatter implementation for help export.",
"\n */\nclass OptionsListFormatter : public IOptionsFormatter\n{\npublic:\n //! ",
"Creates a helper object for formatting options.",
"\n OptionsListFormatter(const HelpWriterContext& context, const CommonFormatterData& common, const char* title);\n\n //! ",
"Initiates a new section in the options list.",
"\n void startSection(const char* header)\n {\n header_ = header;\n bDidOutput_ = false;\n }\n //! ",
"Finishes a section in the options list.",
"\n void finishSection()\n {\n if (bDidOutput_)\n {\n context_.writeOptionListEnd();\n }\n }\n\n void formatOption(const OptionInfo& option) override;\n\nprivate:\n void writeSectionStartIfNecessary()\n {\n if (title_ !",
"= nullptr)\n {\n context_.writeTitle(title_);\n title_ = nullptr;\n }\n if (!",
"bDidOutput_)\n {\n if (header_ !",
"= nullptr)\n {\n context_.paragraphBreak();\n context_.writeTextBlock(header_);\n context_.paragraphBreak();\n }\n context_.writeOptionListStart();\n }\n bDidOutput_ = true;\n }\n\n const HelpWriterContext& context_;\n const CommonFormatterData& common_;\n const char* title_;\n const char* header_;\n bool bDidOutput_;\n\n GMX_DISALLOW_COPY_AND_ASSIGN(OptionsListFormatter);\n};\n\nOptionsListFormatter::OptionsListFormatter(const HelpWriterContext& context,\n const CommonFormatterData& common,\n const char* title) :\n context_(context),\n common_(common),\n title_(title),\n header_(nullptr),\n bDidOutput_(false)\n{\n}\n\nvoid OptionsListFormatter::formatOption(const OptionInfo& option)\n{\n writeSectionStartIfNecessary();\n std::string name, value;\n formatOptionNameAndValue(option, &name, &value);\n std::string defaultValue(defaultOptionValue(option));\n std::string info;\n const FileNameOptionInfo* fileOption = option.toType<FileNameOptionInfo>();\n if (fileOption !",
"= nullptr)\n {\n const bool bAbbrev = (context_.outputFormat() == eHelpOutputFormat_Console);\n info = fileOptionFlagsAsString(*fileOption, bAbbrev);\n }\n std::string description(descriptionWithOptionDetails(common_, option));\n context_.writeOptionItem(\"-\" + name, value, defaultValue, info, description);\n}\n\n//! \\}",
"\n\n} // namespace\n\n/********************************************************************\n * CommandLineHelpWriter::Impl\n */\n\n/*! ",
"\\internal \\brief\n * Private implementation class for CommandLineHelpWriter.",
"\n *\n * \\ingroup module_commandline\n */\nclass CommandLineHelpWriter::Impl\n{\npublic:\n //! ",
"Sets the Options object to use for generating help.",
"\n explicit Impl(const Options& options) : options_(options) {}\n\n //! ",
"Format the list of known issues.",
"\n void formatBugs(const HelpWriterContext& context);\n\n //! ",
"Options object to use for generating help.",
"\n const Options& options_;\n //! ",
"Help text.",
"\n std::string helpText_;\n //! ",
"List of bugs/knows issues.",
"\n std::vector<std::string> bugs_;\n};\n\nvoid CommandLineHelpWriter::Impl::formatBugs(const HelpWriterContext& context)\n{\n if (bugs_.empty())\n {\n return;\n }\n context.writeTitle(\"Known Issues\");\n for (const auto& i : bugs_)\n {\n const char* const bug = i.c_str();\n context.writeTextBlock(formatString(\"* %s\", bug));\n }\n}\n\n\n/********************************************************************\n * CommandLineHelpWriter\n */\n\nCommandLineHelpWriter::CommandLineHelpWriter(const Options& options) : impl_(new Impl(options)) {}\n\nCommandLineHelpWriter::~CommandLineHelpWriter() {}\n\nCommandLineHelpWriter& CommandLineHelpWriter::setHelpText(const std::string& help)\n{\n impl_->helpText_ = help;\n return *this;\n}\n\nCommandLineHelpWriter& CommandLineHelpWriter::setHelpText(const ArrayRef<const char* const>& help)\n{\n impl_->helpText_ = joinStrings(help, \"\\n\");\n return *this;\n}\n\nCommandLineHelpWriter& CommandLineHelpWriter::setKnownIssues(ArrayRef<const std::string> bugs)\n{\n impl_->bugs_ = std::vector<std::string>(bugs.begin(), bugs.end());\n return *this;\n}\n\nCommandLineHelpWriter& CommandLineHelpWriter::setKnownIssues(const ArrayRef<const char* const>& bugs)\n{\n impl_->bugs_ = std::vector<std::string>(bugs.begin(), bugs.end());\n return *this;\n}\n\nvoid CommandLineHelpWriter::writeHelp(const CommandLineHelpContext& context)\n{\n if (context.isCompletionExport())\n {\n context.shellCompletionWriter().writeModuleCompletions(context.moduleDisplayName(), impl_->options_);\n return;\n }\n const HelpWriterContext& writerContext = context.writerContext();\n OptionsFilter filter;\n filter.setShowHidden(context.showHidden());\n\n {\n writerContext.writeTitle(\"Synopsis\");\n SynopsisFormatter synopsisFormatter(writerContext);\n synopsisFormatter.start(context.moduleDisplayName());\n filter.formatSelected(OptionsFilter::eSelectInputFileOptions, &synopsisFormatter, impl_->options_);\n filter.formatSelected(OptionsFilter::eSelectInputOutputFileOptions, &synopsisFormatter,\n impl_->options_);\n filter.formatSelected(OptionsFilter::eSelectOutputFileOptions, &synopsisFormatter, impl_->options_);\n filter.formatSelected(OptionsFilter::eSelectOtherOptions, &synopsisFormatter, impl_->options_);\n synopsisFormatter.finish();\n }\n\n if (!",
"impl_->helpText_.empty())\n {\n writerContext.writeTitle(\"Description\");\n writerContext.writeTextBlock(impl_->helpText_);\n }\n CommonFormatterData common(TimeUnitManager().timeUnitAsString());\n OptionsListFormatter formatter(writerContext, common, \"Options\");\n formatter.startSection(\"Options to specify input files:\");\n filter.formatSelected(OptionsFilter::eSelectInputFileOptions, &formatter, impl_->options_);\n formatter.finishSection();\n formatter.startSection(\"Options to specify input/output files:\");\n filter.formatSelected(OptionsFilter::eSelectInputOutputFileOptions, &formatter, impl_->options_);\n formatter.finishSection();\n formatter.startSection(\"Options to specify output files:\");\n filter.formatSelected(OptionsFilter::eSelectOutputFileOptions, &formatter, impl_->options_);\n formatter.finishSection();\n formatter.startSection(\"Other options:\");\n filter.formatSelected(OptionsFilter::eSelectOtherOptions, &formatter, impl_->options_);\n formatter.finishSection();\n\n impl_->formatBugs(writerContext);\n}\n\n} // namespace gmx\n"
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"Future of Patients in Healthcare Evaluation: The Patient-Informed Reference Case.",
"\nThe \"Reference Case\" was developed to facilitate comparability among published cost-effectiveness analyses intended to contribute to decisions about the broad allocation of healthcare resources. ",
"Although the societal perspective is recommended for Reference Case analyses, empirical estimations rarely adequately represent the patient perspective, and more often, healthcare system or payer perspectives are used. ",
"In this commentary, we discuss the evolution of the Reference Case over the past 20 years and how it now needs to further evolve. ",
"This should begin with a patient-informed societal perspective. ",
"A realignment of the societal perspective to better include patient perspectives in CEA creates a conduit for patient inclusion. ",
"Engaging patients to both derive patient-informed value elements and prioritize value elements using stated preference methods will lead to patient inclusion in the societal perspective and a patient-informed Reference Case analysis."
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"`Phone Booth' is No. ",
"1 movie\n\nCBS.MarketWatch.com\n\nNEW YORK (CBS.MW) --Twentieth Century Fox’s thriller 'Phone Booth' led in the box office results this weekend, according to an early tally by Reuters. ",
"See full story.",
"\n\nChuck Jaffe: If you didn't make that money, who did?",
"\n\nBOSTON (CBS.MW) -- Chances are you didn't get an 8 percent pay raise last year, in the middle of the bear market. ",
"You probably didn't even get an 8 percent return on even the safest of your investments. ",
"But the guys who represent you as overseers on your mutual funds did. ",
"See full story.",
"\n\nStocks to Watch: Stocks to watch Monday\n\nEarnings season revs up next week, though schedules are light Monday and Tuesday. ",
"See full story.",
"\n\nU.S. says 3,000 Iraqis soldiers killed this weekend\n\nNEW YORK (CBS.MW) -- The U.S. Central Command said Sunday that 2,000 to 3,000 Iraqi soldiers were killed in “show-of-force” attacks in northern Iraq by American armored vehicles, according to the Associated Press. ",
"See full story.",
"\n\nSpotting an investment seminar scam\n\nLOS ANGELES (CBS.MW) -- Like many of their friends, Jerry and Linda Chessler found themselves at a crossroads in life, sandwiched between helping their kids financially and caring for elderly parents. ",
"See full story.",
"\n\nWeekend Investor: Spotting an investment seminar scam\n\nLOS ANGELES (CBS.MW) -- Like many of their friends, Jerry and Linda Chessler found themselves at a crossroads in life, sandwiched between helping their kids financially and caring for elderly parents. ",
"See full story.",
"\n\nIntraday Data provided by SIX Financial Information and subject to terms of use.",
"\nHistorical and current end-of-day data provided by SIX Financial Information.",
"\nAll quotes are in local exchange time. ",
"Real-time last sale data for U.S. stock quotes reflect trades reported through Nasdaq only.",
"\nIntraday data delayed at least 15 minutes or per exchange requirements."
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"\n\nTrack listing\n\nPersonnel\nTheo Loomans - vocals, bass guitar, guitars\nRonny Van Der Wey - guitar\nBob Bagchus - drums\n\nReferences\n\nCategory:Asphyx albums\nCategory:1996 albums\nCategory:Century Media Records albums"
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"Background\n==========\n\nPsychological disorders account for over 15% of the total burden of disease within established economies, a significant proportion of which manifests in depressive and anxiety-related disorders \\[[@B1]\\]. ",
"For these disorders, effective treatment options often include non-pharmacological as well as pharmacological interventions. ",
"Consensus guidelines recommend the use of cognitive-behavioral and interpersonal therapies for major depression \\[[@B2]-[@B4]\\] and cognitive behavioral therapy for panic disorder \\[[@B5]\\]. ",
"Research has also highlighted the potential value of these approaches in the treatment of dysthymia, phobia and generalized anxiety disorder \\[[@B6]\\].",
"\n\nDespite evidence of its efficacy however, most adults diagnosed with depressive or anxiety-related disorders do not receive psychotherapeutic care \\[[@B6],[@B7]\\]. ",
"Insufficient numbers of mental health professionals impede access to effective interventions, with people living in remote or under-served areas often having to travel long distances to obtain face-to-face services \\[[@B8]\\]. ",
"Other recognized barriers include time and economic constraints, caring responsibilities, psychological or physical impairment and concerns regarding the potential stigma of attending outpatient appointments \\[[@B9]-[@B11]\\].",
"\n\nRemote communication technologies such as the telephone, internet or videophone have the potential to mitigate many of these inequalities \\[[@B12],[@B13]\\]. ",
"The telephone in particular is a widely available telecommunication technology \\[[@B14]\\] that is being used increasingly as a mechanism for support and treatment delivery \\[[@B15],[@B16]\\]. ",
"The establishment of 1--900 counseling services alone illustrates a long but ongoing commitment to its use within psychotherapy service provision. ",
"More recently, the emergence of computer-aided technology alongside growth in the popularity of the Internet have increased multifold the opportunities for real time, long-distance consultations \\[[@B17]\\].",
"\n\nConventional wisdom still insists however, that for most purposes, psychological therapies should be delivered face-to-face. ",
"The central premise to this argument is that the effectiveness of such interventions depends upon on the development of a high quality therapeutic alliance between therapist and client \\[[@B18]\\]. ",
"Within the context of this relationship, visual as well as auditory information is reflected in behaviours such as eye contact, physical expression, posture and voice \\[[@B19]\\]. ",
"The use of the telephone or internet will invariably eliminate many of these cues whilst the use of teleconferencing will limit physical presence and touch. ",
"Whether or not an effective alliance can be delivered in the absence of interpersonal contact thus remains a topic of some debate \\[[@B20]-[@B22]\\].",
"\n\nEvidence to support the efficacy of non face-to-face psychotherapy service models is accumulating. ",
"Recent studies of collaborative care provision, in which telephone support is often provided as an adjunct to other interventions, have documented positive outcomes \\[[@B23]-[@B27]\\]. ",
"Reviews and meta-analyses of computer-delivered CBT packages \\[[@B28],[@B29]\\] and self-help treatments \\[[@B30]-[@B33]\\] also provide promising results. ",
"These interventions however are often considered as stand-alone technologies requiring little or no contact between therapist and client. ",
"One recent meta-analysis of internet-based cognitive behavioral therapy for depression and anxiety \\[[@B34]\\] has suggested that the efficacy of these psychological interventions may ultimately be higher where therapist support is available.",
"\n\nThe ability to deliver more intensive psychological therapy via remote communication media has the potential to confer multiple benefits for patients by combining real time, scheduled contact with increased accessibility. ",
"As yet however, the efficacy of delivering psychotherapy via such means has received comparatively little attention \\[[@B19]\\]. ",
"This article reports the findings of a systematic review conducted to determine the clinical effectiveness of remote psychotherapy.",
"\n\nMethods\n=======\n\nDesign\n------\n\nThis research was conducted using systematic review techniques \\[[@B35]\\] and meta-analysis to assess the clinical effectiveness of psychotherapy delivered via remote communication.",
"\n\nInclusion criteria\n------------------\n\n### Study design\n\nStudies eligible for inclusion were published randomised controlled trials (RCTs). ",
"Due to a lack of financial resources for translation facilities, the review was restricted to English language publications. ",
"There were no specific methodological quality criteria for inclusion in the review. ",
"Instead data were extracted from all studies on key methodological issues (see \\'Methods of the review\\' below).",
"\n\n### Interventions\n\nInterventions eligible for inclusion included any treatment incorporating a psychological intervention mediated by remote communication. ",
"Psychological interventions were defined as treatments with an explicit psychological orientation \\[[@B36]\\]. ",
"\\'Mediated by remote communication\\' was defined as a treatment where all or the majority (i.e. with the exception of an initial or final contact meeting) of the psychological intervention was delivered by a therapist to a patient on a scheduled and repeated one-to-one basis through an appropriate technology (rather than face-to-face). ",
"Group and marital therapies or any other intervention involving interaction between a therapist and more than one client simultaneously were excluded from the review. ",
"Trials examining emergency crisis interventions (e.g. telephone helplines) or the efficacy of medication management programs in the absence of psychotherapy were excluded. ",
"Minimal interventions in which the majority of therapy was delivered i) outside of client-therapist interaction time (e.g. self-help therapy) or ii) via technology with little or no therapist contact (e.g. computerized CBT) were also excluded. ",
"Studies measuring changes in mental health symptoms as a by-product of counseling for physical illness were excluded.",
"\n\n### Populations\n\nPopulations eligible for inclusion included any group seeking treatment for a mood disorder or functional (non-organic) mental health problem recognized by ICD-10 \\[[@B37]\\] or DSM criteria \\[[@B38]\\]. ",
"Studies involving therapy for substance misuse or addictions were excluded. ",
"Trials examining the efficacy of remote therapy for healthy populations *at risk*of mental health difficulties were also excluded.",
"\n\nSearch strategy\n---------------\n\nElectronic literature searches were performed using the Ovid electronic database on the MEDLINE (1966--2006), PsycInfo (1967--2006), EMBASE (1980--2006) and CINAHL databases (1982--2006). ",
"The Cochrane Central Register of Controlled Trials (CENTRAL) and the Cochrane Collaboration Depression, Anxiety and Neurosis Controlled Trials Register (CCDAN-CTR) were also searched. ",
"All searches were conducted to July 2006. ",
"Subject headings were used to identify all papers indexed as containing material relevant to mental/psychiatric health, psychological therapies/interventions and communication technologies. ",
"These headings were augmented by text words that included the full and abbreviated names of specific mental disorders, types of psychological therapy and modes of communication. ",
"Full details of the search terms used are appended.",
"\n\nAuthors of published and ongoing studies were contacted for further studies and information on the progress of ongoing work: 50% replied with information. ",
"Reference lists from identified papers and from reviews in the area were searched by hand.",
"\n\nMethods of the review\n---------------------\n\nEligibility judgements and data extraction were done independently by two reviewers. ",
"No formal measure of the reliability of data extraction was calculated but disagreements were resolved by discussion with other members of the project team.",
"\n\nA standardised data extraction recorded information on study context, population, interventions, outcomes and methodological quality. ",
"Methodological quality was assessed using standard criteria originally developed by the Cochrane Collaboration for Depression, Anxiety and Neurosis (CCDAN). ",
"The CCDAN quality rating scale \\[[@B39]\\] scores each study according to 23 elements of design and conduct including randomisation methods, sample size, follow-up period, power and appropriateness of analysis. ",
"Each criterion is scored from 0--2, giving a maximum score of 46.",
"\n\nThe CCDAN criteria, like other validated scales of RCT quality \\[[@B40]\\], include a score pertaining to the adequacy of blinding procedures. ",
"Since it is not feasible to blind patients to an active intervention, this criterion was not relevant to the present review. ",
"A second criterion pertaining to the recording of pharmacological side-effects was similarly omitted from the review. ",
"Thus overall quality scores for the CCDAN criteria were in this study only able to reach a maximum of 42.",
"\n\nMethods of analysis\n-------------------\n\nAll analyses were conducted using Comprehensive Meta Analysis \\[[@B41]\\]. ",
"Continuous measures were translated into a standardised effect size, Cohen\\'s *d*\\[[@B42]\\]. ",
"Dichotomous variables were translated into standardised effect sizes by computing the log odds ratio, and then conversion to *d*using procedures in the meta analytic package. ",
"Initial meta-analyses used a fixed effect model \\[[@B43]\\] to provide an overall pooled measure of effect. ",
"Between-study heterogeneity was assessed using the I^2^statistic \\[[@B44]\\], which describes the percentage of total variation across studies that is due to heterogeneity rather than chance. ",
"The I^2^statistic has several advantages over other measures of heterogeneity, including greater statistical power to detect clinical heterogeneity when fewer studies are available. ",
"As a guide, I^2^values of 25% may be considered \\'low\\', 50% \\'moderate\\' and 75%, \\'high\\'. ",
"Where heterogeneity was \\'high\\' (i.e. I^2^values of 75% or above), a random effects model was employed.",
"\n\nFor the purposes of analysis, three main comparison groups were created: i) remote communication therapy versus a control group (e.g. usual care, waiting list); ii) remote communication therapy versus conventional face-to-face therapy, and iii) different types of remotely communicated therapy. ",
"Where studies compared two different types of remote therapy with a control, both comparisons were entered into the meta-analysis separately, but control group sample sizes were halved to ensure that they were not double counted \\[[@B45]\\].",
"\n\nStudies examining depression underwent a separate analysis to those focussing on anxiety or anxiety-related disorders (e.g. anxiety, agoraphobia with panic, OCD). ",
"Studies assessed outcome over a range of time points. ",
"For the purposes of analysis, outcome assessments were categorised into two time periods, representing outcomes in the short (0--6 months), and longer term (7 months and over). ",
"Where studies reported assessments at multiple time points, data with the longest duration were used. ",
"Effect sizes were only calculated for the primary outcome measure (where specified), or the measure deemed most relevant to the mental health disorder under study (judged by KL and LG). ",
"When studies reported more than one relevant measure (e.g. BDI and HRSD), these were combined by taking an arithmetic mean \\[[@B46]\\].",
"\n\nResults\n=======\n\nThirteen studies were included in the review. ",
"Of 40 studies identified but subsequently excluded, the most common reasons for rejection were i) a lack of a mental health primary outcome measure (n = 11); ii) the use of a minimal intervention (e.g. guided self help) or an intervention with no clear psychological orientation (n = 10); iii) the use of a technology-based therapy requiring no direct therapist involvement (n = 18) or iv) group therapy (n = 1). ",
"One further study was identified as ongoing and this may be of relevance for later versions of the review. ",
"Summaries of the included studies are given in table [1](#T1){ref-type=\"table\"}, with corresponding methodological details in table [2](#T2){ref-type=\"table\"}. ",
"A list of excluded studies is available from the authors.",
"\n\n###### \n\nInterventions in the review\n\n **Study** **Target population** **Study groups** **Description of intervention in each group**\n ----------------- ---------------------------------------------------------------------------- -------------------------------------------------------- ----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------\n Hunkeler (2000) Depressed primary care patients Usual care plus telephone support & peer care \\'Good care\\' incorporating regular GP visits, continued antidepressant prescribing and any other referral thought usual by GP. ",
"Augmented by telephone-delivered medication adherence support, side-effect discussions and behavioural activation plans (mean of 10.1 × 5.6 min sessions over 16 wks) plus one or more telephone or face-to-face (6/62 participants) peer support contacts.",
"\n Usual care plus telephone support As above, minus peer support\n Usual care As above minus telephone & peer support.",
"\n \n Lange (2001) Psychology students with trauma experience Internet-mediated writing therapy 30 web-pages of psychoeducation followed by 10 × 45-min writing sessions over 5 wks (2/wk), therapist feedback (appro× 450 words) provided on 7 occasions across 3 treatment phases (self-confrontation, cognitive re-appraisal, sharing & farewell ritual).",
"\n Waiting list 30 web pages of psychoeducation only\n \n Lange (2003) Individuals with mild-relatively severe trauma symptoms Internet-mediated writing therapy 30 web-pages of psychoeducation followed by 10 × 45-min writing sessions over 5 wks (2/wk), therapist feedback (approx 450 words) provided on 7 occasions across 3 treatment phases (self-confrontation, cognitive re-appraisal, sharing & farewell ritual).",
"\n Delayed treatment As above, but only received once the intervention group had completed treatment.",
"\n \n Lovell (2006) Secondary care outpatients with OCD Face-to-face CBT 10 × 1-hr sessions using exposure & response prevention. ",
"Sessions incorporated the establishment of fear hierarchies, use of family co-therapist, weekly exposure targets (to be practised between sessions for at least 1-hr/dy), homework reviews and collaborative problem solving.",
"\n Telephone CBT 8 weekly telephone calls of up to 30-mins in length with treatment content identical to above. ",
"Homework sheets posted to participants. ",
"Initial 1-hr face-to-face session covering the same material as the face-to-face arm plus 1 × 1-hr final session face-to-face\n \n Lynch (1997) Primary care patients with minor depression Telephone counselling 6 × 20-min sessions based on problem-solving for depression; homework comprising of 5 steps of treatment including a demonstration of the connection between depressed mood and problems, expressing problems in a form that facilitates solutions, evaluating and modifying these solutions.",
"\n Comparison group No further details provided\n \n Lynch (2004) Primary care patients with minor depression Telephone problem solving Nezu\\'s problem solving therapy adapted for telephone use and administered over a 6-wk period\n Telephone stress management Treatment designed to serve as an attention control with topics including the identification of sources of stress, the importance of diet & exercise, ways of coping with stress\n Usual care Usual treatment deemed appropriate by primary care physician.",
"\n \n McName e (1989) Housebound agoraphobics with panic disorder Telephone self exposure Exposure goals set via 10 × 12-min telephone contacts with therapists. ",
"Subjects posted a self-help manual that encouraged use of coping strategies and family co-therapists.",
"\n Telephone relaxation therapy Subjects posted standard taped instructions of Jacobsen\\'s relaxation and instructed to listen for at least 1-hr/dy. ",
"Therapy augmented by 10 × 12-min telephone consultations.",
"\n \n Miller (2002) Women with history of recurrent/chronic depression Telephone interpersonal psychotherapy (IPT-T) 12 × 1-hr scheduled weekly sessions.",
"\n Usual care No treatment beyond usual care\n \n Mohr (2000) Depressed MS patients Telephone CBT 8 × 50-min sessions plus a workbook with assignments. ",
"Treatment delivered alongside access to usual care.",
"\n Usual care Any treatment given in the course of usual clinician care.",
"\n \n Mohr (2005) Depressed primary care patients with MS Telephone CBT (T-CBT) Weekly 50-min sessions completed over 16 wks.",
"\n Telephone supportive emotion focussed therapy (T-SEFT) Weekly 50-min sessions completed over 16 wks\n \n Nelson (2003) Depressed children aged 8--14 yrs Videoconferenc e CBT 8 sessions (1 × 90-min plus 7 × 60-min).",
"\n Face-to-face CBT 8 sessions (1 × 90-min plus 7 × 60-min).",
"\n \n Simon (2004) Depressed primary care patients Telephone psychotherapy 8 × 30--40 min CBT plus 1 mail contact and 3 × 10--15 min telephone sessions focussed on medication management, caseload tracking and structured assessment.",
"\n Telephone care management As above minus telephone CBT. ",
"Patients given CBT self-management booklet but no further support provided.",
"\n Usual care No further details given\n \n Swinson (1995) Rural primary care patients suffering from panic disorder with agoraphobia Telephone behaviour therapy Mailed psychometric package and educational workbook serving as an introduction to behavior therapy concepts (e.g. hierarchy construction, exposure exercises, record keeping); 8 × 1-hr scheduled therapy sessions completed over approx. ",
"10 wks. ",
"Therapy included exposure principles & exercises, long term goals, hierarchy construction, coping strategies, diary keeping, homework planning & reviewing.",
"\n Waiting list Initial psychometric package followed 10 wks later by an additional psychometric package and a workbook serving as an introduction to behavior therapy concepts (e.g. hierarchy construction, exposure exercises, record keeping).",
"\n\n###### \n\nCharacteristics of the included studies\n\n ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------\n **Study**\\ **Country**\\ **Target population**\\ **Recruitment**\\ **Sample**\\ **Outcomes**\\ **Follow-up**\\ **Follow-up**\\ **CCDAN**\\\n **size** **rate** **score**\n ------------------ -------------- ---------------------------------------------------------------------------- -------------------------------------------------------------------- ------------- ------------------------------ ------------------------------------------------------ ------------------------- ------------\n Hunkele r (2000) US Depressed primary care patients GP referral 302 HAMD, BDI, SF-12 Baseline, 6 w, 6 m 90% at 6 w, 85% at 6 m 25\n\n \n\n Lange (2001) Netherlands Psychology students with trauma experience From student pool in return for course credits 30 IES, SCL-90, POMS Baseline, 5 w, 11 w 83% at 5 w, 27% at 11 w 18\n\n \n\n Lange (2003) Netherlands Individuals with mild-relatively severe trauma symptoms Website contact 184 IES, SCL-90 Baseline, 5 w, 11 w 79% at 5 w, 31% at 11 w 21\n\n \n\n Lovell (2006) UK Secondary care outpatients with OCD Screening clinics 72 YBOCs, BDI Pre-baseline, baseline, 1 m, 3 m, 6 m 90% at 6 m 36\n\n \n\n Lynch (1997) US Primary care patients with minor depression Screening 29 BDI, HAMD, DHP, PSI Baseline, 7 w 55% at 7 w 20\n\n \n\n Lynch (2004) US Primary care patients with minor depression Screening 54 BDI, HAMD, DHP Baseline, 6 w 57% at 6 w 17\n\n \n\n McNam ee (1989) UK Housebound agoraphobics with panic disorder Telephone screening 23 BDI, FQ, PT, GP, SA, GI Baseline, 2 w, 4 w, 6 w, 8 w, 10 w, 12 w, 20 w, 32 w 78% at 6 w, 61% at 32 w 22\n\n \n\n Miller (2002) US Women with history of recurrent/chronic depression Ongoing longitudinal study 30 HRSD, GAS SAS-SR Baseline, 12 w 80% at 12 w 22\n\n \n\n Mohr (2000) US Depressed MS patients Telephone screening 32 POMS Baseline, 8 w 72% at 8 w 22\n\n \n\n Mohr (2005) US Depressed primary care patients with MS MS case registers & MS society newsletters 127 BDI, HDRS, PANAS Baseline, 8 wk, 16 w, 3 m, 6 m, 9 m, 12 m 91% at 16 w 31\n\n \n\n Nelson (2003) US Depressed children aged 8--14 yrs Not clear 38 K-SADS-P, CDI Baseline, 8 w 74% at 8 w 13\n\n \n\n Simon (2004) US Depressed primary care patients Computer records of patients starting new antidepressant treatment 600 SCL, PHQ Baseline, 6 w, 3 m, 6 m 89% at 6 m 38\n\n \n\n Swinson (1995) Canada Rural primary care patients suffering from panic disorder with agoraphobia GP/family physician referral 46 FQ, STAI-T, BDI, ASI, SCL-90 Baseline, 10 w, 3 m 91% at 10 w, 76% at 3 m 20\n ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------\n\nKey: POMS-Profile of Mood States; BDI-Beck Depression Inventory; HAMD/HRSD-Hamilton Rating Scale for Depression; DHP-Duke Health Profile; PSI-Problem Solving Inventory; SCL-90-Symptom Checklist 90; STAI-T-State-Trait Anxiety Inventory (Trait version); ASI-Anxiety Sensitivity Index; PHQ-Patient Health Questionnaire; SCL-Hopkins Symptom Checklist Depression Scale; IES-Impact of Events Scale; GAS-Global Assessment Score; SAS-SR Social Adjustment Scale; K-SADS-P-Schedule for Affective Disorders & Schizophrenia for School Age Children-Present Episode; CDI-Children\\'s Depression Inventory; PT-Phobic Targets; GP-Global Phobia; SA Social Adjustment; GI-Global Impression; FQ-Fear Questionnaire; YBOCs-Yale Brown Obsessive Compulsive Disorder Scale; PANAS-Positive & Negative Affect Scale; SF-12-Mental & Physical Composite Scales\n\nScope of the included studies\n-----------------------------\n\nNine studies reported 11 comparisons of technology-mediated therapy versus a control, three studies focussing on anxiety-related disorders \\[[@B47]-[@B49]\\] and six on depression \\[[@B50]-[@B55]\\]. ",
"Four of these studies used usual care as the comparative arm and four a waiting list or no treatment control. ",
"The remaining study did not provide a clear description of its control group. ",
"Two studies reported comparisons of technology-mediated therapy with conventional face-to-face interventions, one in an anxiety-related disorders (OCD) \\[[@B56]\\] and one in depression \\[[@B57]\\]. ",
"Three studies reported comparisons between different types of therapy mediated by the same technology, one in anxiety \\[[@B58]\\] and three in depression \\[[@B54],[@B55],[@B59]\\].",
"\n\nTen studies assessed the efficacy of psychological therapy mediated by telephone, two by internet and one by videoconference (Table [1](#T1){ref-type=\"table\"}). ",
"Participants were recruited via primary care screening or GP referral (five studies), secondary care outpatient screening (one study), by public advertisement (two studies), through case registers of Multiple Sclerosis patients (two studies), via longitudinal research (one study) and by access to a pool of psychology students (one study). ",
"One study focusing on depressed children did not provide a clear description of its recruitment context. ",
"Patients included those with PTSD, OCD, agoraphobia with panic disorder and depression (Table [2](#T2){ref-type=\"table\"}). ",
"The number of participants randomized within each trial ranged from 23 to 600 (mean 121), with losses at follow-up of between 9% and 73%. ",
"Length of follow-up ranged from 6 weeks to 1 year. ",
"No data on costs were reported in any of the studies.",
"\n\nQuality of the included studies\n-------------------------------\n\nAll studies were RCTs, although in two studies methods of allocation were only quasi-randomised with the possibility of bias \\[[@B50],[@B58]\\]. ",
"Only two out the 13 studies reported details of allocation concealment \\[[@B55],[@B56]\\]. ",
"Less than half of the included studies (n = 5) defined a main outcome *a priori*\\[[@B50],[@B51],[@B54]-[@B56]\\] and only five reported conducting a power analysis \\[[@B49],[@B54]-[@B57]\\]. ",
"Although all studies conducted largely or wholly appropriate analyses of their data this failed to include an intention to treat analysis in eight instances \\[[@B47]-[@B51],[@B54],[@B57],[@B58]\\]. ",
"Seven studies controlled for between-group differences at baseline \\[[@B47],[@B48],[@B51],[@B53],[@B55],[@B56],[@B59]\\] and all but two studies presented sufficient data for re-analysis \\[[@B50],[@B57]\\]. ",
"Only two studies involved more than 100 participants per trial arm \\[[@B51],[@B55]\\], the vast majority (n = 9) recruiting less than 50.",
"\n\nThe mean methodological quality score for the included studies was 23 (SD = 7.33). ",
"The highest scores were 38 \\[[@B55]\\], 36 \\[[@B56]\\] and 31 \\[[@B59]\\].",
"\n\nQuantitative results\n--------------------\n\nPooled effect sizes for each of the main comparison groups are presented in table [3](#T3){ref-type=\"table\"}. ",
"In both meta analyses using multiple studies, the I^2^statistic was zero (indicating no heterogeneity beyond that expected by chance) and thus fixed effect models were used.",
"\n\n###### \n\nResults of meta-analyses\n\n ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------\n **Intervention**\\ **Comparison**\\ **Disorder**\\ **Follow**\\ **No.**\\ **Total**\\ **Pooled**\\ **95% CI**\\\n \\ \\ \\ **-up**\\ **Comparisons**\\ **Participants**\\ **Effect**\\ \\\n **period** **Size** \n -------------------------------- -------------------------------------- ------------------ ------------- ------------------ ------------------- ------------- ----------------\n Remote psychotherapy Control Depression 0--6 m 7 726 0.44 0.29 to 0.59\n\n Remote psychotherapy Control Anxiety-related 0--6 m 3 168 1.15 0.81 to 1.49\n\n Remote psychotherapy Face-to-face psychotherapy Depression 0--6 m 1 28 0.55 -0.20 to 1.31\n\n Remote psychotherapy Face-to-face psychotherapy Anxiety-related 0--6 m 1 63 -0.11 -0.60 to 0.38\n\n Remote psychotherapy\\ Remote psychotherapy\\ Depression\\ 0--6 m\\ 1\\ 18\\ 0.38\\ -0.56 to 1.32\\\n (problem solving therapy) (stress management) \n\n Remote psychotherapy\\ Remote psychotherapy\\ Depression\\ 0--6 m\\ 1\\ 122\\ 0.39\\ 0.04 to 0.74\\\n (cognitive behavioral therapy) (supportive emotion focused therapy) \n\n Remote psychotherapy\\ Remote psychotherapy\\ Depression\\ 7 m+\\ 1\\ 117\\ 0.25\\ -0.12 to 0.62\\\n (cognitive behavioral therapy) (supportive emotion focused therapy) \n\n Remote psychotherapy\\ Remote psychotherapy\\ Anxiety-related\\ 0--6 m\\ 1\\ 18\\ 1.10\\ 0.10 to 2.10\\\n (exposure therapy) (relaxation therapy) \n\n Remote psychotherapy\\ Remote psychotherapy\\ Anxiety-related\\ 7 m+\\ 1\\ 14\\ 1.22\\ 0.06 to 2.38\\\n (exposure therapy) (relaxation therapy) \n ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------\n\nNote: Effect size has been coded so that a positive effect size indicates a study where patients in the intervention group have better outcomes than those in the comparison\n\nFor depressive disorders, the pooled effect size for remotely delivered therapy compared to control conditions (e.g. waiting list or usual care) was 0.44 (95% CI 0.29 to 0.59), demonstrating a statistically significant \\'medium\\' effect \\[[@B42]\\] (figure [1](#F1){ref-type=\"fig\"}).",
"\n\n{#F1}\n\nFor anxiety-related disorders, remotely delivered therapy demonstrated a \\'large\\' effect compared to control conditions (figure [2](#F2){ref-type=\"fig\"}, 3 comparisons, n = 168). ",
"The pooled effect size was 1.15 (95% CI 0.81 to 1.49) (figure [2](#F2){ref-type=\"fig\"}).",
"\n\n{#F2}\n\nComparisons between technology-mediated and face-to-face interventions and comparisons between different forms of technology-mediated therapy were limited by the small number of studies reporting relevant data (table [3](#T3){ref-type=\"table\"}).",
"\n\nDiscussion\n==========\n\nThe aim of this review was todetermine the clinical effectiven ess of psychotherapy delivered by remote communication technologies. ",
"Compared to control conditions, technology-mediated therapy demonstrated a \\'large\\' effect for anxiety-related disorders and a \\'medium\\' effect for depression. ",
"The majority of included studies suffered at least some methodological limitations and few comparisons of remote versus face-to-face psychotherapy were found.",
"\n\nOverall, the effect size calculations would suggest that remotely delivered psychotherapies do have the potential to be clinically effective, although the magnitude of this effect varies and may be more evident for anxiety than for depression. ",
"The comparison of technology-mediated psychotherapy against control conditions for depression produced a pooled effect size of 0.44 in favor of therapy. ",
"This finding remains in accordance with an effect size of 0.42 reported by an earlier meta-analysis \\[[@B60]\\] comparing conventional face-to-face therapy with no treatment controls in depression. ",
"A second more recent meta-analysis of face-to-face psychotherapy for depression reports standardized mean differences of 0.63 up to 3 months and 0.56 at 6--9 month follow-up in favor of face-to-face therapy over treatment as usual or waiting list controls \\[[@B36]\\].",
"\n\nFor comparative purposes, meta analyses of lower intensity psychological interventions (e.g. guided self help) for anxiety and depressive disorders report mean effect sizes of between 0.40 and 1.19 \\[[@B30],[@B31],[@B33],[@B34],[@B36],[@B61]\\], However, these reviews include studies of treatments that, by definition, use technology to reduce rather than mediate professional therapist input. ",
"The effects of such interventions have been shown to be greater for some disorders than others \\[[@B31],[@B33]\\], with certain problems (e.g. anxiety disorders) responding better to self-help packages supported by increased therapist contact \\[[@B30],[@B33]\\]. ",
"The characteristics of patients accessing such treatments are likely to differ substantially from those requiring more traditional and higher intensity therapies, not least in terms of their problem severity and/or motivation for treatment.",
"\n\nA definitive conclusion as to the relative efficacy of technology-mediated versus face-to-face administered psychotherapy can only be drawn from randomized controlled trials comparing the two interventions. ",
"A comparison of the efficacy of technology-mediated versus face-to-face psychotherapy was conducted within the present review, with a large but non-significant effect size of 0.55 for depression and a smaller difference of -0.11 for anxiety-related disorders being obtained. ",
"However, a shortage of literature limits the utility of this result. ",
"Only two of the 13 studies included in the review directly compared the efficacy of face-to-face psychotherapy with an equivalent intervention delivered via more remote means. ",
"It should be acknowledged that even in these studies, meaningful comparisons may be difficult. ",
"The ability for remote therapy to overcome multiple physical, psychological and geographical constraints means that the clinical populations who may wish to access such treatments may differ markedly from those in face-to-face services.",
"\n\nUltimately, the published evidence base for clinical effectiveness of psychotherapies delivered solely or largely via remote communication methods is limited, both in size and quality. ",
"Therefore, the effect size estimates reported in the current review can at best only be viewed as preliminary. ",
"The majority of identified studies were of relatively small sample size with common shortcomings noted in relation to allocation procedures, identification of primary outcome measures and statistical analyses. ",
"From a possible maximum score of 42, overall quality ratings for the included studies ranged from 13 to 38 with a median of 22 (SD = 7.33). ",
"Despite the above limitations, these scores remain slightly higher than those observed elsewhere, a recent meta-analysis of brief psychological treatments for depression reporting a mean score of 19 (SD 7.3) for 63 controlled trials assessed against the same quality criteria \\[[@B36]\\]. ",
"Given that the financial implications of remote therapy are likely to be a key factor of interest, it is disappointing that no economic data were available.",
"\n\nAttrition rates for the studies ranged from 9 to 73% depending upon the population studied, the nature of the intervention and the length of the follow-up period. ",
"This heterogeneity suggests that whilst technology-mediated psychotherapies have the potential to be more effective than delayed or usual care, different treatment modalities may differ in their perceived acceptability. ",
"Data from earlier systematic reviews suggest that attrition rates of alternative models of treatment delivery can also vary widely. ",
"Randomized trials of face-to-face therapy for depression report post-treatment drop-outs of between 0 and 60% \\[[@B7]\\]. ",
"Trials of written self help materials for anxiety and depressive disorders report comparable figures of between 0 and 61% \\[[@B62]\\], although attrition from open access online therapy is generally assumed to be higher and may be as much as 99% \\[[@B63]\\], Once again however, differences in study context, recruitment and sample characteristics can often prevent any direct comparison of attrition rates across different modes of treatment.",
"\n\nFour of the studies included in the review provided a quantitative measure of patient satisfaction, all indicating a preference for, or equivalent satisfaction with, technologically-mediated care as compared to face-to face delivery \\[[@B56],[@B57]\\] or usual care \\[[@B51],[@B55]\\]. ",
"Two other studies also provided data to suggest that levels of patient satisfaction with remote psychotherapy are high, although in these instances no comparable data from control conditions were available \\[[@B48],[@B53]\\]. ",
"Eighty-three per cent of participants in one trial expressed a favorable attitude towards the use of the telephone to deliver psychotherapy and 75% expressed a desire to continue treatment either now in the future through this medium \\[[@B53]\\]. ",
"However, whether or not these preferences remain consistent across different technologies and communication modalities is unclear. ",
"The vast majority of studies included in the present review focused on therapy delivered via the telephone, with very little research effort being directed towards newer and increasingly available resources such as the web. ",
"Only one study in the original review examined the efficacy of psychotherapy delivered via videoconferencing and only two studies used the internet, both of which were conducted by the same research team.",
"\n\nA scoping search of literature published since the original searches were undertaken has returned only a small number of additional studies \\[[@B64]-[@B66]\\]. ",
"One presents data already included in the present review \\[[@B64]\\]. ",
"Another, examining telephone psychotherapy for depression and conducted as a follow-up to one of trials in the present meta-analysis reports maintenance of clinical benefits when compared to usual care \\[[@B65]\\]. ",
"A third trial that has recently compared face-to-face therapy with a comparable intervention delivered via telepsychiatry reports no significant differences in effect \\[[@B66]\\].",
"\n\nWhilst the telephone undoubtedly confers specific advantages in terms of its widespread availability and ease of operation, research suggests that videoconferencing may also be particularly suited to psychotherapeutic use. ",
"A recent systematic review has shown real time telepsychiatry to represent a highly feasible method of conducting mental health treatments and assessments, often with clinical outcomes and rates of attendance equivalent to those obtained face to face \\[[@B67]\\]. ",
"However, criteria for inclusion and exclusion in this review were not explicitly stated and therefore it is unclear whether or not these observations are based on high quality evidence from randomized trials.",
"\n\nThe present review has its limitations. ",
"It includes studies examining the effectiveness of psychotherapy delivered via remote communication technology on a scheduled and repeated one-to-one basis and as such, excludes trials examining the effectiveness of more minimal, technology-based interventions. ",
"A number of reviews have already been published examining the clinical effectiveness of self-help treatments delivered via technology, and thus the decision was made to conduct a more focused review of optimal relevance to more traditional therapy formats. ",
"Nevertheless, publication bias is a problem for any review of controlled trials and given the high dependence on electronic database searching it cannot be certain that unpublished studies do not exist. ",
"Due to the small number of studies eligible for the review, funnel plots to assess publication bias were not created in line with recommendations \\[[@B68]\\]. ",
"The analysis treated \\'usual care\\' and waiting list\\' controls as equivalent to \\'no treatment\\', although it should be noted that previous studies have suggested that effects in usual care may be significantly lower \\[[@B69]\\]. ",
"Translating dichotomous outcomes into standardised effect sizes for the purposes of the meta analysis does allow the most comprehensive summary of the outcome data, but these transformations are based on assumptions which may not always be warranted.",
"\n\nConclusion\n==========\n\nTechnology-mediated psychotherapy provision has the potential to overcome many of the barriers to care associated with more traditional face-to-face interventions. ",
"Data suggest that good treatment effects and by implication therapeutic alliance may not be dependent on a patient and therapist being co-located. ",
"However, the limited amount of evidence that is available and the restricted number of studies in any one subgroup as yet prevents any definitive conclusions from being drawn. ",
"Future research priorities should include overcoming the methodological shortcomings of published work by conducting large-scale trials that incorporate both clinical outcome and more process-orientated measures. ",
"In particular, these studies should seek to compare remote psychotherapy with more conventional face-to-face methods by quantifying levels of patient and therapist satisfaction and exploring the potential impact of different modes of communication on therapeutic outcome and examining the quality of the therapeutic alliance that is established,\n\nCompeting interests\n===================\n\nThe authors declare that they have no competing interests.",
"\n\nAuthors\\' contributions\n=======================\n\nPEB participated in the study design, search strategy development, retrieval of articles, screening of articles, quality assessment, data extraction, data synthesis, and writing of manuscript. ",
"PB participated in the study design, quality assessment, data extraction, meta-analysis, data synthesis, and writing of manuscript. ",
"KL, SG, DR and LG participated in the study design, quality assessment, data extraction, and writing of manuscript. ",
"PBarnes participated in the retrieval of articles, screening of articles, quality assessment, data extraction, and writing of manuscript. ",
"All authors read and approved the final manuscript.",
"\n\nAppendix\n========\n\nElectronic search strategy\n\n1\\. online therapy/or telemedicine/or Diagnosis, Computer-Assisted/or Therapy, Computer-Assisted/\n\n2\\. telecommunications/or telephone/or videoconferencing/or teleconferencing/\n\n3\\. internet/or computer assisted instruction/or virtual reality/or Electronic Mail/\n\n4\\. phone\\$.mp. ",
"or telephone\\$.mp. ",
"or telecommnicat\\$.mp\n\n5\\. (text and messag\\$).mp. ",
"or SMS.mp. ",
"or (short and messag\\$).mp.",
"\n\n6\\. videophone\\$.mp. ",
"or videoconferenc\\$.mp. ",
"or teleconferenc\\$.mp\n\n7\\. email.mp. ",
"or (electronic and mail).mp. ",
"or (electronic and communication).mp.",
"\n\n8.(virtual and reality).mp. ",
"or VR.mp.",
"\n\n9.(bulletin and board\\$).mp. ",
"or (discussion and board\\$).mp. ",
"or (discussion and list\\$).mp.",
"\n\n10\\. internet.mp. ",
"or online.mp. ",
"or worldwide web.mp. ",
"or web based.mp.",
"\n\n11\\. Emedicine.mp. ",
"or Etherapy.mp. ",
"or Ehealth.mp.",
"\n\n12\\. telemedicin\\$.mp. ",
"or teleconsult\\$.mp. ",
"or telepsychiatry.mp. ",
"or telehealth.mp. ",
"or teletherap\\$.mp.",
"\n\n13\\. (computer and based).mp.or (computer and mediated).mp. ",
"or (computer and assisted).mp.",
"\n\n14\\. (remote and consultation).mp.",
"\n\n15\\. 1 or 2 or 3 or 4 or 5 or 6 or 7 or 8 or 9 or 10 or 11 or 12 or 13 or 14\n\n16\\. Psychotherapy/or Brief psychotherapy or Individual Psychotherapy/or Psychotherapeutic Techniques/or Supportive Psychotherapy or or Group Psychotherapy/or Psychotherapeutic Processes/\n\n17\\. psychotherap\\$.mp. ",
"or psychodynamic therap\\$.mp. ",
"or PDT.mp. ",
"or psychoanalytic therap\\$.mp.",
"\n\n18\\. Behavior Therapy/or Cognitive Therapy/or Cognitive Behavior Therapy/or Rational Emotive Behavior Therapy/or Behavior Modification/\n\n19\\. cognitive behavio?r therap\\$.mp. ",
"or cognitive therap\\$.mp. ",
"or behavio?r therap\\$.mp. ",
"or CBT.mp. ",
"or cognitive analytic therap\\$.mp. ",
"or behavio?r modification.mp.",
"\n\n20\\. Relaxation Therapy/or relaxation therap\\$.mp.",
"\n\n21\\. Family Therapy/or Interpersonal Psychotherapy/or Psychodynamic Psychotherapy/or Marital Therapy or Group therapy/\n\n22\\. Interpersonal therap\\$.mp or IPT.mp.",
"\n\n23\\. family therap\\$.mp. ",
"or marital therap\\$.mp. ",
"or group therap\\$.mp. ",
"or support group\\$.mp.",
"\n\n24\\. GROUP COUNSELING/or PSYCHOTHERAPEUTIC COUNSELING/or EDUCATIONAL COUNSELING/or COUNSELING/or MARRIAGE COUNSELING/or COUNSELING PSYCHOLOGY/or PARENT COUNSELING/or FAMILY COUNSELING/or PATIENT COUNSELING/\n\n25\\. counsel?ing.mp.",
"\n\n26\\. Self Help/or Self Management/or exp Self Help Techniques/\n\n27\\. self treatment.mp. ",
"or self help.mp. ",
"or self directed.mp. ",
"or self management.mp.",
"\n\n28\\. Behavior Modification/or Relaxation Techniques/or Relaxation Training/\n\n29\\. relaxation training.mp. ",
"or relaxation techniques.mp.",
"\n\n30\\. psychosocial support.mp. ",
"or psychological therap\\$.mp. ",
"or psychological treatment\\$.mp.",
"\n\n31\\. exposure therapy.mp.",
"\n\n32\\. 16 or 17 or 18 or 19 or 20 or 21 or 22 or 23 or 24 or 25 or 26 or 27 or 28 or 29 or 30 or 31\n\n33\\. 15 and 32\n\n34\\. limit 33 to English\n\n35\\. remove duplicates from 34\n\nPre-publication history\n=======================\n\nThe pre-publication history for this paper can be accessed here:\n\n<http://www.biomedcentral.com/1471-244X/8/60/prepub>\n"
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"SXSW 2016 Report Card\n\nSXSW is a brand name when it comes to film festivals, but defining its identity is still problematic. ",
"It’s probably still second to Sundance as far as prestige for indie films looking to break out, and it comes too early in the year to attract international premieres or blockbuster headliners.",
"\n\nAs a regional festival, it continues to attract Texas-themed films, and its location in Austin makes it a natural for home-grown talent such as Jeff Nichols and Richard Linklater. ",
"It it’s line up isn’t as deep at TIFF or Telluride, it still attracts talent and never lacks for star power.",
"\n\nThis year’s festival lineup was a little light at the top, and documentaries tended to outshine narratives. ",
"That said, among the twenty films I screened, there was only one out and out dud…which of course ended up winning a jury prize.",
"\n\nHere’s my comprehensive report card from from the film side of America’s music festival\n\nNewtown — Kim Snyder’s unflinching look at the aftermath of mass murder at Sandy Hook Elementary is understandably hard to watch at times. ",
"What it gets right is not just the anger evoked by life’s sometimes brutal cruelty but the human sacred solemnity felt in the presence of survivors. (",
"A)\n\nAccidental Courtesy: Daryl Davis, Race & America — An impressive debut feature from Matt Ornstein profiles the outspoken musician who befriends Ku Klux Klan members. ",
"What stands out even more than Davis’s intelligence is his emotional differentiation. ",
"Only once does a conflict seem to rile him, and that is when he is confronted by other African-Americans who question the value of his methodology. (",
"A)\n\nFantastic Lies — This ESPN sponsored summary of the way Duke University, Mike Nifong, the media, and the rest of us rushed to judgment upon hearing accusations of sexual assault at a party for the high-profile lacrosse team suffers (but only slightly) from relying a little too heavily on parents of the falsely accused. ",
"Not that there are any remaining doubts about the players’ innocence, but the film might have benefited from a bit more introspection instead of summary. ",
"That this happened is shocking and depressing. ",
"Thinking a little more deeply about why it happened might have been a worthwhile enterprise. (",
"A-)\n\nClaire in Motion — Less flashy but more intelligent that the festivals more highly touted spousal grief film, Demolition, Claire in Motion benefits from a wonderfully restrained performance from Betsy Brandt. ",
"Claire needs to keep herself together for the benefit of her son, and while the circumstances that keep throwing her into the path of an art student with whom her husband might have had an emotional (but not sexual) affair are a bit contrived, Claire’s responses to the other woman always ring true. (",
"A-)\n\nIn a Valley of Violence — Before Ti West’s western revenge saga dissolves into Tarantino parroting in its last act, it a surprisingly effective genre piece. ",
"Ethan Hawke expands his repertoire and Taissa Farmiga practically steals the show. ",
"This is a familiar story, but West’s framing and composition are effective, and he lets his stars do the heavy lifting. (",
"B+)\n\nIn Pursuit of Silence — A documentary about the spiritual and psychological benefits from silence is a tricky thing. ",
"On the one hand, there are ample testimonials about the detriments of having fewer and fewer spaces devoid of near constant noise. ",
"On the other hand, silence may be one of those subjects that demands a more immersive treatment. ",
"The film attempts to both explain and embody silence. ",
"A better film might have done one or the other. (",
"B)\n\nFree in Deed — Jake Mahaffey’s depiction of an African-American man (David Harewood) seeking redemption for we know not what by trying to miraculously heal a boy who is most probably autistic was the most visually accomplished and striking film of the festival. ",
"Harewood is riveting as Abe, a man who Mahaffey said (in an interview) has a “talent” for healing but comes across as neither comfortable in his own skin or confident of his place in God’s kingdom. ",
"The film doesn’t rank higher only because it comes across as a fine act of imitation. ",
"It gets the surface textures of the charismatic religion right, but it holds us at arms length. ",
"Mahaffey appears to want the audience to extract its own meaning from the (inspired by a true-) story. ",
"That’s normally fine, but there is a difference between being circumspect and being opaque. (",
"B)\n\nMiss Stevens — Without fine performances from Lily Rabe as Miss Stevens and Rob Huebel as a drama student who may be cycling off his medication, Julia Hart’s feel-good melodrama could easily have come across as trite or even icky. ",
"Student and teacher have an emotional bond, and Miss Stevens lets it go farther than it should for reasons that aren’t fully explained until the end. ",
"A gay male student and a less talented female student both come along for the ride. ",
"The film only really soars when Huebel’s character delivers a monologue from Death of a Salesman, but Rabe is always interesting to watch. ",
"Also Hart’s script is good at showing how teachers, like politicians, are forced to make dozens of snap decisions every day, never really knowing which they will asked to account for. (",
"B-)\n\nThe Liberators — Cassie Bryant’s documentary about priceless artifacts stolen from Germany at the end of World War II takes too long to get to its most interesting questions, insisting on trying to make itself into a detective story even though the outcome is never really in doubt. ",
"High marks, however, for some very articulate talking heads. (",
"B-)\n\nEverybody Wants Some!! — ",
"I’ve warmed a tad, but only a tad, to Richard Linklater’s opening-night film. ",
"The mostly plotless depiction of three days at a Texas university before the start of classes is mostly an excuse for a very of well executed vignettes that feel like less than the some of its parts. ",
"That said, Me and Orson Welles is the only Linklater film outside the Before trilogy that I’ve truly liked, so your mileage may vary. (",
"C+)\n\nCameraperson — Is it fair to call a person’s memoir thirty minutes too long? ",
"Maybe not, but while Kirsten Johnson’s highlight reel does evidence a keen eye for framing, it usually feels more like a scrapbook than a memoir. ",
"I actually found myself wanting a little bit of Johnson talking about her work. (",
"C)\n\nMidnight Special — I am convinced that if Jeff Nichols’s sci-fi mash up is screened outside of Austin that maybe more people will eventually agree with me that it is underwritten. ",
"The actors are all great–Nichols rightly called it a dream cast in introducing the film–but nothing much happens until the end. ",
"By that time, the fate of these characters matters less to us than it should. (",
"C)\n\nLittle Sister — This film felt more like Tribeca than SXSW. ",
"It was the story of a dysfunctional family as seen through the eyes of one semi-normal member. ",
"Colleen is a nun-in-training who takes a leave to visit her family when her brother returns (wounded) from a war. ",
"The family dynamics are a bit too sitcommish, but Addison Timlin holds it together in the lead, and its always nice to see a film have its religious characters treat their religion seriously. (",
"C)\n\nI Am Not a Serial Killer — Novelist Dan Wells confirmed in an interview that he was very pleased with Billy O’Brien’s adaptation of his genre-bending series of John Wayne Cleaver books. ",
"My less sanguine response had much to do with the fact that the film greatly de-emphasizes the teen’s internal struggle to keep his own demons at bay while fighting a monster he thinks he is uniquely qualified to understand. ",
"Wells countered that Max Records does a fine job of communicating the internal struggle without the film resorting to voice-over, and he’s right about that. ",
"But he also insisted that the book is about someone trying to be good, and all I saw in the film was someone trying to destroy another evil. (",
"C)\n\nThe Master Cleanse — Almost. ",
"This deadpan comedy horror vehicle simultaneously skewers health nuts, New Age religion, and self-help seminars. ",
"It’s central conceit–making the purging of impurities literal rather than figurative–sort of works, but it takes too long to get to it. ",
"The tone is stuck somewhere between ironic detachment and enthusiastic embrace of farce. ",
"Another draft might have helped a great deal. (",
"C-)\n\nYarn — A documentary about how yarn is made and used and thought about ought to at least be charming. ",
"The film, unfortunately, can’t decide if it wants to focus more on the art projects–some of which are impressive–or the eccentric artists that create them. ",
"The latter aren’t always as articulate as you would like, and a few are actually cruder than you would expect in a film that might otherwise attract a wider audience. ",
"It’s almost as though Yarn doesn’t really believe anyone might actually be interested in its subject and so pushes away those who are while trying to attract…who exactly? (",
"D+)\n\nBodkin Ras — Kaweh Madiri told audiences at SXSW that he was fascinated by the characters in this Scottish town but mostly interested in Forres as a swamp from which nobody could escape. ",
"The mix of fictional character and documentary travelogue never quite gels, and even at 79 minutes, the film feels stretched. ",
"Before watching the film, I’ve always thought Scotland might be a cool place to visit. ",
"Afterwards, I wondered if everybody just hates living in small towns wherever they are. ",
"Surely that was not the intent? (",
"D)\n\nThe Arbalest — For the second time in three years, my least favorite movie at SXSW won the Grand Jury Prize. ",
"Not sure what that means. ",
"The Arbalest is a satire that isn’t sure what it is satrizing, a comedy that isn’t funny, and a film that uses violence in the laziest ways possible while expecting us be alternately horrified and amused by it. ",
"My only consolation is that Fort Tilden also received a Grand Jury prize and was rightfully never heard from again.",
"\n\nIt is the policy of this blog that if the editor or reviewer has received from the producers or marketers of a film a complementary screener, free admission to a public (or private) screening, or any form of direct or indirect compensation for expenses incurred (such as for travel) in the process of reviewing a film, it will be noted in the tags for that film's coverage."
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"That's right. ",
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"On 2: is it not true that women can also oppress women? ",
"See anti-abortion politicians, etc.",
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"There are no exceptions. ",
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"Plasticity of gastrointestinal vagal afferent satiety signals.",
"\nThe vagal link between the gastrointestinal tract and the central nervous system (CNS) has numerous vital functions for maintaining homeostasis. ",
"The regulation of energy balance is one which is attracting more and more attention due to the potential for exploiting peripheral hormonal targets as treatments for conditions such as obesity. ",
"While physiologically, this system is well tuned and demonstrated to be effective in the regulation of both local function and promoting/terminating food intake the neural connection represents a susceptible pathway for disruption in various disease states. ",
"Numerous studies have revealed that obesity in particularly is associated with an array of modifications in vagal afferent function from changes in expression of signaling molecules to altered activation mechanics. ",
"In general, these changes in vagal afferent function in obesity further promote food intake instead of the more desirable reduction in food intake. ",
"It is essential to gain a comprehensive understanding of the mechanisms responsible for these detrimental effects before we can establish more effective pharmacotherapies or lifestyle strategies for the treatment of obesity and the maintenance of weight loss."
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"\nPublication of the Quality in Australian Health Care Study in 1995 represented a defining moment for Australian health care providers. ",
"The high incidence and cost of preventable adverse events underscored a need for defined process, error recognition and audit cycle. ",
"Despite this, surgical audit has continued to emphasize clinical indicators relevant to technical performance. ",
"The greatest burden of preventable error can be traced to deficiencies in the process by which management expectations are supported. ",
"Recognizing this, the focus of clinical audit must be expanded. ",
"In particular, outcome assessment should be routine rather than sporadic, and should broadly encompass safety, effectiveness and efficiency. ",
"Devolving this responsibility to paraclinical groups is in itself insufficient. ",
"Quality and safety cannot be adequately addressed unless surgeons actively participate in audit cycle. ",
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"Fractures are often treated with screws or other fixation devices inserted into or through a bone to stabilize fractured portions thereof once they have been brought into corrective alignment. ",
"Trochanteric bone fixation treatments often comprise the insertion of an intramedullary nail into a medullary cavity of a bone and a subsequent insertion of a bone fixation nail into a condylar portion of the bone at an angle relative to the intramedullary nail. ",
"Once implanted conventional trochanteric bone fixation devices permit medial and lateral migration of the bone fixation nail within and sometimes out of an outer periphery of the bone. ",
"Furthermore, conventional bone fixation devices comprise multiple elements that add to the complexity of bone fixation procedures while minimizing the degree of adjustability of the components relative to one another. ",
"Accordingly, this prevents the tailoring of these bone fixation devices to individual requirements of various patients. ",
"Such systems therefore reduce the anchoring strength of the bone fixation devices increasing the likelihood of further fractures or other complications."
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"\nInert gas elimination studies in interstitial fibrosis ascribe all of the resting and most (58-83%) of the exercise (A-a)PO2 difference to ventilation-perfusion inequality. ",
"The previous paper (Hughes, J.M.B., D.N.A. Lockwood, H.A. Jones and R.J. Clark, (1991) Respir. ",
"Physiol. ",
"83:155-166) [corrected] suggests from estimates of global DLO2/Q beta ratios a larger role for diffusion limitation on exercise. ",
"Gas exchange data from that paper was analyzed at rest and on exercise for five patients with interstitial fibrosis. ",
"Hypoxemia at rest was attributed to VA/Q inequality which was quantified using a log-normal lung model. ",
"DLO2 was calculated by Bohr integration. ",
"The base 10 LogSDQ at rest averaged 0.5 +/- 0.1 (SEM). ",
"On the assumption that VA/Q inequality remained unchanged on exercise, DLO2 (exercise) was estimated to be 14.3 +/- 1.9 ml.min-1.Torr-1. ",
"At that level of DLO2, diffusion limitation accounted for 36% +/- 8(SEM)% of the total (A-a)PO2 difference using the log-normal VA/Q model. ",
"But estimates of DLO2/Q beta assuming a homogenous lung, ascribed 96% of the (A-a)PO2 gradient on exercise to diffusion limitation. ",
"This discrepancy was shown to be related to the shape of the oxygen equilibrium curve and high alveolar PO2 values. ",
"On the other hand, analysis in terms of oxygen contents showed that 68 +/- 5% of the (A-a) content difference was accounted for by diffusion limitation. ",
"This differs substantially from estimates based on partial pressure alone."
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"USS Unit (1862)\n\nUSS Unit (1862) was a steamer acquired by the Union Navy during the American Civil War.",
"\n\nShe was used as a tugboat by the Navy and she provided her services to ships in the blockade squadrons. ",
"She also served as a repair tender for ships needing her services.",
"\n\nService during the American Civil War \n\nUnit—steamer Union built at Philadelphia in 1862—was purchased at Boston on 2 June 1864. ",
"Unit was assigned to the North Atlantic Blockading Squadron and served as a tug and repair vessel in Hampton Roads, Virginia, for the duration of the Civil War. ",
"In June 1865, the tug was sent to New York City.",
"\n\nEnd-of-war decommission and continued maritime career \n\nUnit was sold at auction there on 12 July 1865 to C. and E. T. Peters. ",
"Redocumented as a merchant steamer on 6 September 1865, Unit remained in mercantile service until 1902.",
"\n\nSee also\n\nUnited States Navy\nList of United States Navy ships\n\nReferences \n\nCategory:Ships of the Union Navy\nCategory:Ships built in Philadelphia\nCategory:Steamships of the United States Navy\nCategory:Tugs of the United States Navy\nCategory:Tenders of the United States Navy\nCategory:American Civil War auxiliary ships of the United States\nCategory:1862 ships"
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") - (sqrt(200) + sqrt(200)*-1)) + sqrt(8))*4)*-5.",
"\n375*sqrt(7)/2\nSimplify (-2 + (sqrt(396) + 5*sqrt(396) + -5 + -3)*3)**2*-6.",
"\n-773880 + 33696*sqrt(11)\nSimplify (3 + ((sqrt(176) + sqrt(176) + -3)*1 - (sqrt(176) + (sqrt(176)*-2)**2 + sqrt(176) + sqrt(176) + -3)))*1.",
"\n-701 - 4*sqrt(11)\nSimplify (2*sqrt(70)*5 - sqrt(70))/(sqrt(250) - sqrt(250)*2) + (-4*3*-2*sqrt(21)/sqrt(3))**2.",
"\n-9*sqrt(7)/5 + 4032\nSimplify 1 + (3*sqrt(320))**2 - 1*(-2 + sqrt(320)) - ((-2 + (sqrt(320) - (3 + sqrt(320) + sqrt(320))))**2 + -3)*-1.",
"\n72*sqrt(5) + 3225\nSimplify ((sqrt(275) + -1)*2 + -3 + (sqrt(275) - (sqrt(275) - (sqrt(275) + -1)**2 - sqrt(275))))*-2 - (sqrt(1100) + 0 - (sqrt(77)/sqrt(7) + 4))*6.",
"\n-518 - 64*sqrt(11)\nSimplify ((sqrt(96)*-1 - (sqrt(24) - 2*sqrt(3456)))*3)/(6*-1*sqrt(648)*1).",
"\n-7*sqrt(3)/6\nSimplify ((-6*(sqrt(175) - (1 + sqrt(175) + sqrt(175) + 3)) + 2*(sqrt(175) + -1 + sqrt(175)) + sqrt(175) + sqrt(175) + 1 + -3)*3 + -3)**2.",
"\n20520*sqrt(7) + 230049\nSimplify ((1*(sqrt(84) + sqrt(84)*2*2)*5)/((sqrt(768) - (sqrt(768) - 2*sqrt(768))*6 - sqrt(768))*4))**2.",
"\n4375/36864\nSimplify (1 + 3 + (sqrt(1331) - (-1 + 5*sqrt(1331) + sqrt(1331)) - (sqrt(1331)*3 + 3)))*-5.",
"\n-10 + 440*sqrt(11)\nSimplify ((sqrt(72) - 6*(sqrt(72)*1 + sqrt(72)))*5 + -2)**2 - ((sqrt(72) + -1)*-1 - (sqrt(72) + -2*sqrt(72) + 3))**2*6.",
"\n1320*sqrt(2) + 217780\nSimplify (((sqrt(85)*1*3 - sqrt(85)) + sqrt(85))/sqrt(5)*5 - (sqrt(1377)*-5 + 4)*-4)**2*6.",
"\n-31680*sqrt(17) + 2778486\nSimplify ((sqrt(66) - (sqrt(66) - -4*(sqrt(66) + sqrt(66)*3))) + 1*sqrt(66)*-6)/(-4*(sqrt(150) + sqrt(6)) - (sqrt(588) + sqrt(12) - sqrt(12))/sqrt(2)).",
"\n22*sqrt(11)/31\nSimplify -5*(3 + sqrt(252)*-2*-3 + -2*sqrt(252) + -5 + -1 + -3)**2.",
"\n-20340 + 1440*sqrt(7)\nSimplify ((sqrt(1134) - (sqrt(1134) + sqrt(1134) + -1*sqrt(1134) + sqrt(1134))*-6 - sqrt(1134) - 2*sqrt(1134)*-6)/((sqrt(378)*1)/(sqrt(864)/sqrt(9))))**2.",
"\n165888\nSimplify (((sqrt(135)*-1 - sqrt(135)) + sqrt(135) + sqrt(135))*4 - sqrt(135)*1*-3)/((sqrt(2500) - sqrt(2500)*-2 - sqrt(25))/(sqrt(405)*2)).",
"\n162*sqrt(3)/29\nSimplify 1*-5*(((-2*(sqrt(693) + 1*sqrt(693) + sqrt(693)) - sqrt(693)) + sqrt(693))/(2*sqrt(275)))/(-6*(sqrt(36)/(sqrt(4) + sqrt(36)) - sqrt(54)/(1*sqrt(6)))).",
"\n2*sqrt(7)/3\nSimplify (-2 + 4 + -4*(-2 + (sqrt(931) + 3 - sqrt(931)))*4)*-1.",
"\n14\nSimplify (((sqrt(3328) - 3*sqrt(3328) - sqrt(3328)) + sqrt(3328) - 3*sqrt(416)/sqrt(8))/(sqrt(288)/sqrt(128)*-1))**2.",
"\n75088/9\nSimplify -1 + (5 + (4*-3*sqrt(320)*-2*3)**2 + -3)*6.",
"\n9953291\nSimplify (-3*(sqrt(704) + sqrt(704) + (2 + sqrt(704) - sqrt(704)) + sqrt(44) + -4 + sqrt(11)) - (-4 + sqrt(99)*2 + 0))**2.",
"\n-1260*sqrt(11) + 43759\nSimplify 0 + (2*sqrt(136)*5 - (sqrt(136) + -4*sqrt(136)*4 + sqrt(136) + sqrt(136) - sqrt(136)))/((sqrt(72)/sqrt(3)*5)/(1*sqrt(48))).",
"\n96*sqrt(17)/5\nSimplify ((1 + sqrt(68)/sqrt(4) - (2 + -3*sqrt(17))) + (-1*(sqrt(170) + 2*sqrt(170)))/(sqrt(70)/sqrt(112)))**2 + 3.",
"\n16*sqrt(17) + 1092\nSimplify 4*(-1 + (1*sqrt(171)*1 - (sqrt(171) + sqrt(171) + 2*sqrt(171))*-6)/(4*(sqrt(441)*-1 + sqrt(9)))).",
"\n-25*sqrt(19)/6 - 4\nSimplify (3*(sqrt(153) + sqrt(153)*-1*2 - 4*sqrt(153)*2)*6)/(2*-1*sqrt(9)*6 + sqrt(108)/sqrt(192)*-1).",
"\n648*sqrt(17)/49\nSimplify (((1*sqrt(396)*6)/sqrt(12)*-2)/(sqrt(1100) - -6*(sqrt(1100) + -1*sqrt(1100)) - 4*((sqrt(1100)*-2 - sqrt(1100)) + sqrt(1100)))*-4)**2.",
"\n64/75\nSimplify 4*((sqrt(288)*2)**2 - sqrt(2)) + -4 - ((-2 + -1 + sqrt(200))**2 - (5 + -1 + sqrt(200) + sqrt(200))**2).",
"\n216*sqrt(2) + 5211\nSimplify (sqrt(1188) - (-2*sqrt(1188) + sqrt(1188)))/(2*sqrt(432)) + -3*-5*(1 + sqrt(396))*3.",
"\n45 + 541*sqrt(11)/2\nSimplify (sqrt(228) - (-2*sqrt(228)*5 - sqrt(228)))/(sqrt(12) - (sqrt(12) + 4*sqrt(48) - sqrt(12))) + -2 + (-6*sqrt(228))/(2*sqrt(75)).",
"\n-102*sqrt(19)/35 - 2\nSimplify ((2 + -1*-1*sqrt(180)*-2 + (sqrt(180) + -5*-3*sqrt(180) - (sqrt(180)*2 + 3)))*-3)**2.",
"\n-1296*sqrt(5) + 233289\nSimplify 4*(-5 + ((sqrt(1053) + 3)*6 - (sqrt(1053) + sqrt(1053)*4)*6))**2 + -3.",
"\n-22464*sqrt(13) + 2426785\nSimplify (3 + -1 + 5 + 2*2*sqrt(68) + 5 + -1)**2.",
"\n176*sqrt(17) + 1209\nSimplify (2*(sqrt(891) + -3) + (1*(sqrt(891) + (sqrt(891) - sqrt(891)*3) - sqrt(891)) - sqrt(891)) - -3*-3*sqrt(891)*2*-5)**2.",
"\n-9612*sqrt(11) + 7057647\nSimplify ((sqrt(3200)*-2 - sqrt(3200))*5 - sqrt(3200) - (sqrt(3200) - -4*sqrt(3200)*-5) - 1*(sqrt(3200) - (4 + 2*sqrt(3200) + sqrt(3200)))*-5)**2.",
"\n11200*sqrt(2) + 157200\nSimplify (3 + sqrt(832) + -5 - (-2*(sqrt(832) + -2) + sqrt(832)))*-3 + 1 + 0.",
"\n-48*sqrt(13) + 19\nSimplify (-3*6*(sqrt(91) - (sqrt(91) - sqrt(91)*-2*-6)))/((-2*sqrt(350))/(sqrt(288) - (sqrt(288) + sqrt(288)*-2))).",
"\n2592*sqrt(13)/5\nSimplify -5*((2 + sqrt(2816) + 5)**2 - (sqrt(88)/sqrt(2) + -4 + sqrt(176) + -4)**2).",
"\n-12025 - 1600*sqrt(11)\nSimplify (-3 + ((sqrt(1216) - (sqrt(1216) + 1 - sqrt(1216))*6 - (4 + sqrt(1216)*1)) + (sqrt(1216)*1 + 2)*5 + 1 + 3)**2)*1.",
"\n320*sqrt(19) + 30413\nSimplify (-5*(3 + -4*(1*(sqrt(2057) + 1) - sqrt(2057)) + -6*(sqrt(68)*5 + sqrt(85)/sqrt(5) + -3)))**2.",
"\n-56100*sqrt(17) + 1858525\nSimplify (-5 + (5*(sqrt(1331) + -1) + 5 + 1 + 0)*-4 + 5)**2.",
"\n1760*sqrt(11) + 532416\nSimplify (0 + -3 + 2 + sqrt(320) + 0 - (1 + sqrt(320)*1*-5 + sqrt(320) + 5*(sqrt(320) + -2)))**2.",
"\n64\nSimplify (sqrt(17) + (sqrt(17) + 0)*4)*-4 + 3 + (sqrt(17)*-1*2 + (sqrt(17) - (sqrt(272) + 0)))*2.",
"\n-30*sqrt(17) + 3\nSimplify (((sqrt(484)*1 - sqrt(484) - sqrt(484))*5)/(-2*sqrt(396)))/((sqrt(360)*6 - sqrt(40))/(sqrt(90)/(sqrt(9)*-2))).",
"\n-5*sqrt(11)/408\nSimplify -2*(-3 + (4*sqrt(891)*-3 + sqrt(891) + -3)*-3) + -5.",
"\n-594*sqrt(11) - 17\nSimplify 5 + -2*sqrt(1728)*-4 - ((sqrt(1728) - -5*(sqrt(1728) + (sqrt(1728) - sqrt(1728)*-1))) + -6*(-1 + sqrt(1728)))**2.",
"\n-172831 - 2688*sqrt(3)\nSimplify ((sqrt(200) + -3*(sqrt(200) - (sqrt(200) + (sqrt(200) - sqrt(200)*-2) + sqrt(200))))/(-2*sqrt(80)))/(-4*-1*sqrt(648)*-1) + 1.",
"\n13*sqrt(5)/288 + 1\nSimplify -3 + ((0 + sqrt(243))*-4 + -4)**2 + (-1*sqrt(243)*3 - sqrt(243) - (sqrt(243) - (1*sqrt(243) + -1)))**2 + 5.",
"\n360*sqrt(3) + 7795\nSimplify ((sqrt(2688) + sqrt(2688) + sqrt(2688)*1)*-6*5)/((sqrt(7776)*3 + sqrt(54))/(sqrt(72)/(sqrt(40)/sqrt(5)))).",
"\n-720*sqrt(7)/37\nSimplify ((1*sqrt(228))/(-2*sqrt(6)/sqrt(2))*-4)**2 - (4 + 2*sqrt(1216) + 5*(sqrt(38) - sqrt(342) - sqrt(38))/sqrt(2)).",
"\n-sqrt(19) + 300\nSimplify (((sqrt(60)/(-2*sqrt(5)))/(sqrt(128) - sqrt(128)*-1))/(sqrt(300) - (sqrt(300) - sqrt(300)*-1) - sqrt(27)*2))**2.",
"\n1/131072\nSimplify 6*(sqrt(234)/(1*sqrt(600)))/(sqrt(363)*1*1) + -5.",
"\n-5 + 3*sqrt(13)/55\nSimplify -5*(sqrt(425) + sqrt(425) + 2*sqrt(425) + sqrt(425) + -1 + sqrt(425))*2 - -3*(1 + sqrt(612) + sqrt(612) - (sqrt(272)*-6 - sqrt(17))).",
"\n-189*sqrt(17) + 13\nSimplify (((sqrt(510) + sqrt(510)*1*5)/(sqrt(160) - 2*sqrt(160)))/(-2*2*sqrt(3) + -6*sqrt(12)))**2.",
"\n153/1024\nSimplify -2*(3 + -5 + (6*2*sqrt(3564) + sqrt(3564) + 2 + 2)*4)**2.",
"\n-19274504 - 52416*sqrt(11)\nSimplify (-4 + -5 + -1 + sqrt(272))*-6 - (-2 + (sqrt(816)*-3)/(sqrt(18)/sqrt(96))).",
"\n62 + 24*sqrt(17)\nSimplify 2*(0 + -4*(4 + ((sqrt(65) - (sqrt(65) - (sqrt(65) - (sqrt(65) - (1*sqrt(65) - sqrt(65)) - sqrt(65) - sqrt(65))*-4)))/sqrt(5)*-1)**2)).",
"\n-968\nSimplify ((1 + sqrt(76) + 1 + sqrt(76) - ((sqrt(76) - (sqrt(76) + -2*-1*sqrt(76) + sqrt(76))) + sqrt(76) + sqrt(76))) + -3 + sqrt(133)/sqrt(7) + 5 + 3)**2 + -1.",
"\n98*sqrt(19) + 979\nSimplify -2*(2*1*1*sqrt(78)*-6)/(((sqrt(726) + sqrt(726)*-2 - sqrt(726))*1 + sqrt(726))*-5).",
"\n24*sqrt(13)/55\nSimplify ((sqrt(4)/sqrt(2) + -5 + 2 + sqrt(2)*2)*3 + 5 + 1*(0 + sqrt(98))*3)**2.",
"\n-240*sqrt(2) + 1816\nSimplify 4*((sqrt(5) + -4*sqrt(720))**2 + sqrt(5) + 0 + sqrt(45)) - 1*(-2*sqrt(125) + 1)*-1.",
"\n6*sqrt(5) + 44181\nSimplify (4*(sqrt(539) + 2) + sqrt(539)*1*-3)**2 + (5 + (sqrt(704) + (3 + sqrt(704) + sqrt(704) + sqrt(704) - sqrt(704) - sqrt(704)))**2 - sqrt(704))*-3.",
"\n-7887 - 152*sqrt(11)\nSimplify (-5 + 1*(-1 + sqrt(539)) - (-4*(1*sqrt(99) - sqrt(99)) - sqrt(99))/sqrt(9)*1)**2*1.",
"\n-96*sqrt(11) + 740\nSimplify sqrt(72) + 1 + -5 + 3 + -5 - ((sqrt(12)/(sqrt(18)/sqrt(3)) - sqrt(2))*-1 - -2*sqrt(72)*-1).",
"\n-6 + 18*sqrt(2)\nSimplify (sqrt(99)*-1*-6)/(sqrt(36)/(sqrt(16)/sqrt(4))) + 1*(sqrt(1331) + 2 + sqrt(1331) + 3).",
"\n5 + 28*sqrt(11)\nSimplify (6*(sqrt(70) + (sqrt(70) - -2*sqrt(70))) - sqrt(70))/(sqrt(10)*2*2)*4 - (sqrt(49) - sqrt(784)*-5)/(sqrt(7) - (sqrt(7) + -4*sqrt(49)/sqrt(7))).",
"\n71*sqrt(7)/4\nSimplify 0 + -2 + sqrt(2) + (-3*sqrt(24) + sqrt(24))/sqrt(12) + (sqrt(98) + -3*sqrt(98) "
] | {
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[
"Pleigne\n\nPleigne is a municipality in the district of Delémont in the canton of Jura in Switzerland.",
"\n\nHistory\nPleigne is first mentioned in 1179 as Plenna. ",
" The municipality was formerly known by its German name Pleen, however, that name is no longer used.",
"\n\nGeography\n\nPleigne has an area of . ",
" Of this area, or 47.2% is used for agricultural purposes, while or 49.2% is forested. ",
" Of the rest of the land, or 3.0% is settled (buildings or roads), or 0.4% is either rivers or lakes and or 0.1% is unproductive land.",
"\n\nOf the built up area, housing and buildings made up 1.6% and transportation infrastructure made up 1.2%. ",
" Out of the forested land, 46.1% of the total land area is heavily forested and 3.1% is covered with orchards or small clusters of trees. ",
" Of the agricultural land, 17.4% is used for growing crops and 11.0% is pastures and 18.5% is used for alpine pastures. ",
" All the water in the municipality is in lakes.",
"\n\nThe municipality is located in the Delemont district, on a terrace at an elevation of . ",
" The village is north-west of Delemont. ",
" It consists of the village of Pleigne, numerous former farms of Lucelle/Lützel Monastery, the Gasthaus Moulin-Neuf, the 18th century mill at Bavelier and the archeological Löwenburg site.",
"\n\nThe municipalities of Bourrignon, Châtillon, Courrendlin, Courtételle, Delémont, Develier, Ederswiler, Mettembert, Movelier, Pleigne, Rebeuvelier, Rossemaison and Vellerat are considering a merger on at a date in the future into the new municipality with an, , undetermined name.",
"\n\nCoat of arms\nThe blazon of the municipal coat of arms is Gules, between three Mullets of Five Or a Sword and a Staff Argent in Saltire and in chief on an Escutcheon of the last a Jay statant lined Sable.",
"\n\nDemographics\nPleigne has a population () of . ",
" , 2.6% of the population are resident foreign nationals. ",
" Over the last 10 years (2000–2010) the population has changed at a rate of 0%. ",
" Migration accounted for -3.6%, while births and deaths accounted for 2.8%.",
"\n\nMost of the population () speaks French (347 or 85.0%) as their first language, German is the second most common (58 or 14.2%) and Dutch is the third (2 or 0.5%).",
"\n\n, the population was 48.3% male and 51.7% female. ",
" The population was made up of 185 Swiss men (47.3% of the population) and 4 (1.0%) non-Swiss men. ",
" There were 194 Swiss women (49.6%) and 8 (2.0%) non-Swiss women. ",
" Of the population in the municipality, 200 or about 49.0% were born in Pleigne and lived there in 2000. ",
" There were 104 or 25.5% who were born in the same canton, while 69 or 16.9% were born somewhere else in Switzerland, and 15 or 3.7% were born outside of Switzerland.",
"\n\n, children and teenagers (0–19 years old) make up 29.2% of the population, while adults (20–64 years old) make up 59.6% and seniors (over 64 years old) make up 11.3%.",
"\n\n, there were 187 people who were single and never married in the municipality. ",
" There were 188 married individuals, 21 widows or widowers and 12 individuals who are divorced.",
"\n\n, there were 141 private households in the municipality, and an average of 2.8 persons per household. ",
" There were 30 households that consist of only one person and 25 households with five or more people. ",
" , a total of 137 apartments (78.7% of the total) were permanently occupied, while 29 apartments (16.7%) were seasonally occupied and 8 apartments (4.6%) were empty. ",
" , the construction rate of new housing units was 2.6 new units per 1000 residents. ",
" The vacancy rate for the municipality, , was 1.65%.",
"\n\nThe historical population is given in the following chart:\n\nHeritage sites of national significance\n\nThe former Priory of Löwenburg and the paleolithic settlement and neolithic flint mine at Löwenburg are listed as Swiss heritage site of national significance. ",
" The entire Löwenburg area is part of the Inventory of Swiss Heritage Sites.",
"\n\nPolitics\nIn the 2007 federal election the most popular party was the CVP which received 32.31% of the vote. ",
" The next three most popular parties were the SPS (23.46%), the FDP (21.15%) and the SVP (20.38%). ",
" In the federal election, a total of 133 votes were cast, and the voter turnout was 42.9%.",
"\n\nEconomy\n, Pleigne had an unemployment rate of 3.6%. ",
" , there were 51 people employed in the primary economic sector and about 16 businesses involved in this sector. ",
" 33 people were employed in the secondary sector and there were 3 businesses in this sector. ",
" 19 people were employed in the tertiary sector, with 9 businesses in this sector. ",
" There were 217 residents of the municipality who were employed in some capacity, of which females made up 40.1% of the workforce.",
"\n\n the total number of full-time equivalent jobs was 80. ",
" The number of jobs in the primary sector was 35, all of which were in agriculture. ",
" The number of jobs in the secondary sector was 32 of which 26 or (81.3%) were in manufacturing and 6 (18.8%) were in construction. ",
" The number of jobs in the tertiary sector was 13. ",
" In the tertiary sector; 3 or 23.1% were in wholesale or retail sales or the repair of motor vehicles, 5 or 38.5% were in the movement and storage of goods, 5 or 38.5% were in a hotel or restaurant, and .",
"\n\n, there were 38 workers who commuted into the municipality and 138 workers who commuted away. ",
" The municipality is a net exporter of workers, with about 3.6 workers leaving the municipality for every one entering. ",
" About 21.1% of the workforce coming into Pleigne are coming from outside Switzerland. ",
" Of the working population, 9.7% used public transportation to get to work, and 55.8% used a private car.",
"\n\nReligion\nFrom the , 305 or 74.8% were Roman Catholic, while 35 or 8.6% belonged to the Swiss Reformed Church. ",
" Of the rest of the population, there were 34 individuals (or about 8.33% of the population) who belonged to another Christian church. ",
" There were 3 (or about 0.74% of the population) who were Islamic. ",
" 26 (or about 6.37% of the population) belonged to no church, are agnostic or atheist, and 22 individuals (or about 5.39% of the population) did not answer the question.",
"\n\nEducation\nIn Pleigne about 139 or (34.1%) of the population have completed non-mandatory upper secondary education, and 33 or (8.1%) have completed additional higher education (either university or a Fachhochschule). ",
" Of the 33 who completed tertiary schooling, 60.6% were Swiss men, 27.3% were Swiss women.",
"\n\nThe Canton of Jura school system provides two year of non-obligatory Kindergarten, followed by six years of Primary school. ",
" This is followed by three years of obligatory lower Secondary school where the students are separated according to ability and aptitude. ",
" Following the lower Secondary students may attend a three or four year optional upper Secondary school followed by some form of Tertiary school or they may enter an apprenticeship.",
"\n\nDuring the 2009-10 school year, there were no students attending school in Pleigne.",
"\n\n, there were 9 students in Pleigne who came from another municipality, while 41 residents attended schools outside the municipality.",
"\n\nReferences\n\nCategory:Municipalities of the canton of Jura\nCategory:Cultural property of national significance in the canton of Jura"
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"Association of Fluid Overload with Mortality in Critically-ill Mechanically Ventilated Children.",
"\nTo study the association of fluid overload with mortality and morbidity in critically-ill mechanically ventilated children. ",
"Prospective observational study. ",
"Pediatric Intensive Care Unit (PICU) of a tertiary care hospital, New Delhi, India. ",
"118 children (age 1 mo - 15 y) requiring mechanical ventilation. ",
"Primary: Association of fluid overload with mortality. ",
"Secondary: Association of fluid overload with oxygenation, organ dysfunction, duration of mechanical ventilation and PICU stay. ",
"Cumulative fluid overload of ≥15% was observed in 74 (62.7%) children. ",
"About 50% of these children reached cumulative fluid overload of ≥15% within the first 5 days of PICU stay. ",
"The mortality was 40.5% in those with ≥15% cumulative fluid compared to 34% in the rest [OR (95% CI): 1.02 (0.97, 1.07)]. ",
"On multivariate analysis, after adjusting for confounders, cumulative fluid overload ≥15% was associated with higher maximum PELOD (pediatric logistic organ dysfunction) score (Median: 21 vs. 12; P = 0.03), longer median duration of mechanical ventilation (10 vs. 4 d; P <0.0001) and PICU stay (13.5 vs. 6 d; P <0.0001). ",
"There was no significant association of fluid overload with oxygenation index (P=0.32). ",
"There is no association of fluid overload with mortality. ",
"However, it is associated with poor organ function, longer duration of mechanical ventilation and PICU stay in critically-ill, mechanically ventilated children."
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"Laboratory-related outcomes in healthcare.",
"\nOutcomes studies, long common on the therapeutic side of medicine, are appearing in the diagnostic arena. ",
"Outcomes can be defined as results of medical interventions (therapies or tests) in terms of health or cost. ",
"The studies of outcomes are important because funding for medical interventions increasingly depends on them; a major accrediting agency even defines \"quality\" entirely in terms of outcomes. ",
"The study of laboratory-related outcomes is complex. ",
"Multiple steps occur between testing and outcomes, physicians act unpredictably on test results, and outcomes studies have high costs relative to potential profit from the test. ",
"Study design often must specify the action that is to follow a test result. ",
"The model outcomes study is a randomized controlled trial (RCT). ",
"The CONSORT statement, which is used as a guideline for RCTs of therapies, is largely applicable to studies of diagnostic interventions. ",
"Recent laboratory-related RCTs have addressed questions such as: \"Does routine testing before cataract surgery decrease morbidity or mortality?\" ",
"and \"Does fecal occult bleed testing decrease the incidence of colorectal cancer?\" ",
"RCTs of tests are sometimes impractical. ",
"Other approaches include simulation modeling and the use of intervention and control periods of testing. ",
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"This FEC audit may hold more weight The system of regulating campaign money is full of holes and needs to be replaced.",
"\n\nDecember 04, 1998|By Jack W. Germond and Jules Witcover\n\nWASHINGTON -- Moving at its usual snail's pace, the Federal Election Commission has suddenly interjected itself into the already complex issue of whether President Clinton violated the campaign finance laws in 1996.",
"\n\nThe short answer is that, according to FEC auditors, the Clinton campaign did indeed use \"soft money\" for purposes for which it was not supposed to be used. ",
"But so did the campaign of Bob Dole, Mr. Clinton's Republican foe.",
"\n\nThe FEC audit found that both sides paid for television commercials seeking support for the candidate specifically although the money they used was only supposed to be spent on generic party-building activities. ",
"These might include commercials supporting the entire party ticket, touting each candidate's political party in general terms or encouraging voter registration and turnout.",
"\n\nAnd because both sides broke the rules while accepting federal matching funds, the auditors decided the two campaigns should reimburse the Treasury the amounts to which they would not have been entitled if they had followed the law. ",
"If the FEC itself approves the recommendations -- which is far from certain with a commission evenly divided on party lines -- the Clinton campaign will have to pay $7 million, the Dole campaign more than $17 million.",
"\n\nWhat no one explained, of course, is why the FEC auditors required two full years to determine that this kind of ducking around the statute was being done. ",
"Everyone with any understanding of how presidential campaigns operate these days knew all along that both the soft and hard money were fully controlled by the presidential candidate's staff. ",
"And no one had any reason to be surprised when the spots specifically urging votes for Mr. Clinton or Mr. Dole looked so much like other spots that didn't make that request in specific language.",
"\n\nPolitical ties\n\nBut the fact is that the FEC has never been given the kind of staff that would allow its findings to be more timely. ",
"When, for example, candidates exceed the spending limits in a primary campaign, the FEC never nails them until long after the election and thus long after the time when they might pay a political price as well as a fine.",
"\n\nIn this case, however, the findings of the auditors may carry far more weight than usual because of the context in which they have been put forward.",
"\n\nAt the very least, the reports make things awkward for the Republicans on the House Judiciary Committee who have just voted to extend their inquiry into whether Mr. Clinton violated the campaign finance laws in 1996. ",
"The answer seems to be that maybe he did, but so did the Republicans. ",
"Is the committee going to examine that?",
"\n\nForeign money\n\nStrictly speaking, the report of the FEC auditors doesn't deal with all the ways the law might have been violated by either Mr. Clinton or Mr. Dole. ",
"The Republicans on the committee suspect, for example, that there may be evidence of illegal foreign money being channeled into the Clinton campaign. ",
"But two other congressional inquiries have failed to establish such a pattern, so you have to wonder how Judiciary Committee Chairman Henry Hyde expects to do so in a week or 10 days.",
"\n\nIn fact, the issues of ostensible concern to the committee already are being examined by the Justice Department as part of the criminal inquiry ordered by Attorney General Janet Reno on the basis of a preliminary FEC audit. ",
"The operative question here now is whether Ms. Reno should have turned the whole thing over to still another special prosecutor, as the Republicans have been urging.",
"\n\nAll of this is, of course, the kind of political game-playing that defies understanding. ",
"But there are some obvious conclusions that can be drawn.",
"\n\nTroubled system\n\nFirst, the system of regulating campaign money is full of holes and needs to be replaced. ",
"At a minimum, the FEC needs to be given more enforcement authority and the kind of appropriations to use that authority effectively.",
"\n\nSecond, the Republicans are in a particularly vulnerable position because of their demonstrated refusal to support any serious attempt at campaign-finance reform. ",
"That could change with the departure of House Speaker Newt Gingrich, but don't bank on it.",
"\n\nMeanwhile, both parties are free to play the cheapest kind of politics with the issue. ",
"That is what the Republicans on Judiciary are doing now with their 11th-hour attempt to broaden their inquiry -- an attempt undermined by the finding of those FEC auditors that breaking the rules in 1996 was a bipartisan exercise."
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"Activity of dendrimer-methotrexate conjugates on methotrexate-sensitive and -resistant cell lines.",
"\nDendritic nanostructures can play a key role in drug delivery, due to the high density and variety of surface functional groups that can facilitate and modulate the delivery process. ",
"We have investigated the effect of dendrimer end-functionality on the activity of polyamido amine (PAMAM) dendrimer-methotrexate (MTX) conjugates in MTX-sensitive and MTX-resistant human acute lymphoblastoid leukemia (CCRF-CEM) and Chinese hamster ovary (CHO) cell lines. ",
"Two amide-bonded PAMAM dendrimer-MTX conjugates were prepared using a dicyclohexylcarbodiimide (DCC) coupling reaction: one between a carboxylic acid-terminated G2.5 dendrimer and the amine groups of the MTX (conjugate A) and another between an amine-terminated G3 dendrimer and the carboxylic acid group of the MTX (conjugate B). ",
"Our studies suggest that conjugate A showed an increased drug activity compared to an equimolar amount of free MTX toward both sensitive and resistant cell lines, whereas conjugate B did not show significant activity on any of the cell lines. ",
"Despite substantially impaired MTX transport by MTX-resistant CEM/MTX and RII cells, conjugate A showed sensitivity increases of approximately 8- and 24-fold (based on IC50 values), respectively, compared to free MTX. ",
"Co-incubation of the cells with adenosine and thymidine along with either conjugate A or MTX resulted in almost complete protection, suggesting that the conjugate achieves its effect on dihyrofolate reductase (DHFR) enzyme through the same mechanism as that of MTX. ",
"The differences in cytotoxicity of these amide-bonded conjugates may be indicative of differences in the intracellular drug release from the cationic dendrimer (conjugate B) versus the anionic dendrimer (conjugate A), perhaps due to the differences in lysosomal residence times dictated by the surface functionality. ",
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"If the following claims are found valid, does that mean Bilski is null and void?",
"\n\n1. ",
"A computer system for the manipulation of the architecture and content of a document having a plurality of metacodes and content by producing a first map of metacodes and their addresses of use in association with mapped content; said system comprising:\n\nmetacode map distinct storage means;\n\nmeans for providing a menu of metacodes to said metacode storage means;\n\nand means for compiling said metacodes of the menu by locating, detecting and addressing the metacodes in the document to constitute the map and storing the map in the metacode storage means; and\n\nmeans for resolving the content and the metacode map into the document.",
"\n\n2. ",
"A system as claimed in claim 1 further comprising mapped content distinct storage means.",
"\n\n3. ",
"A system as claimed in claim 1 further comprising raw content distinct storage means.",
"\n\n4. ",
"A system as claimed in claim 1 further comprising means for reading a document having first mapped content and a first multiplicity of metacodes constituting in whole or in part, the menu and detecting, locating and storing the first multiplicity of metacodes, in whole or in part, to constitute the map.",
"\n\n5. ",
"A system as claimed in claim 1 further comprising means for comparing the multiplicity of metacodes in the map with a predetermined set of criteria.",
"\n\n6. ",
"A system as claimed in claim 1 further comprising means for amending the first map to produce a second map.",
"\n\n7. ",
"A system as claimed in claim 1 further comprising means for applying the first map to mapped content to provide first differentiated data.",
"\n\n8. ",
"A system as claimed in claim 7 wherein the mapped content is the first mapped content.",
"\n\n9. ",
"A system as claimed in claim 1 further comprising means for providing a multiplicity of maps in association with a mapped single content.",
"\n\n10. ",
"A system as claimed in claim 1 further comprising means for amending a plurality of maps in consequence of an amendment of mapped content.",
"\n\n11. ",
"A system as claimed in claim 1 further comprising means for amending a plurality of maps in consequence of an amendment of an associated metacode map.",
"\n\n12. ",
"A system as claimed in claim 1 comprising means for amending a first plurality of maps to produce a second plurality of amended maps in a plurality of distinct storage means.",
"\n\n13. ",
"A system as claimed in claim 1 wherein metacode is a description code.",
"\n\n14. ",
"A method for producing a first map of metacodes and their addresses of use in association with mapped content and stored in distinct map storage means, the method comprising:\n\nproviding the mapped content to mapped content storage means;\n\nproviding a menu of metacodes; and\n\ncompiling a map of the metacodes in the distinct storage means, by locating, detecting and addressing the metacodes; and\n\nproviding the document as the content of the document and the metacode map of the document.",
"\n\n15. ",
"A method as claimed in claim 14 further comprising:\n\ndetecting and locating a multiplicity of metacodes constituting the menu in a document;\n\nstoring the multiplicity of metacodes, in whole or in part, in the distinct storage means;\n\ndetecting and locating mapped content in the document;\n\nand storing the mapped content, in whole or in part, in the mapped content storage means.",
"\n\n16. ",
"A method as claimed in claim 15 further comprising amending the multiplicity of the metacodes to produce a second map.",
"\n\n17. ",
"A method as claimed in claim 16 wherein metacode is a description code.",
"\n\n18. ",
"A method as claimed in claim 14 further comprising comparing the multiplicity of metacodes in the map with a predetermined set of criteria.",
"\n\n19. ",
"A method as claimed in claim 18 further comprising applying the first map to the mapped content to provide a differentiated document.",
"\n\n20. ",
"A method for producing from a document made up of metacodes and content, a map of metacodes and their addresses of use in association with mapped content of the document and stored in distinct map storage means, the method comprising:\n\n(a) reading the content of the document until a metacode is found;\n\n(b) copying the content and storing the copied content in a mapped content storage;\n\n(c) noting in the map the found metacode and its position in the content;\n\n(d) repeating the processing of (a)-(c) until the entire document has been processed; and then\n\n(e) providing the document as the content of the document separately from the metacode map of the document"
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"Monday, July 20, 2009\n\nMonday, July 13, 2009\n\nAngry Woebots \"Wall of Fur & Fangs\" Stretched Canvas Print. ",
"After a huge demand for a larger print of this image, we decided to do a 20\" x 20\" stretched canvas print. ",
"It will only be an edition of 5, each individually hand accented by Angry Woebots himself. ",
"Each canvas print will be signed & numbered as well. ",
"These are ready to hang & will retail at $210 each. ",
"This will be the first public Angry Woebots stretched canvas print release. ",
"These will also release at the PFOM Pop-Up store & will be available online on our store after SDCC.",
"\n\nAngry Woebots SDCC Exclusive Print \"Cheeseburger\". ",
"Edition of Only 20 & will be released at the PFOM Pop-Up store. ",
"The Giclee' Print measures 11\"x14\" & retails for $70. ",
"Those who can not make it to SDCC this year & are a forum member, please PM us. ",
"Thank you & see you at SDCC!!! ",
"These will be available online on our store after SDCC.",
"\n\nWednesday, July 1, 2009\n\nLooks like we got an exclusive print from Angry Woebots. ",
"After seeing the original piece in person, we were blown away and had to make an exclusive print. ",
"The \"Wall of Fur & Fangs\" Series 1 Print will be a signed & numbered edition of 35, measuring 12\" x 12\" on archival museum paper. ",
"Retail will be $60 & will be released July 2nd at 1:00 Pacific Standard Time at Silent Stage Gallery."
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"There’s an interesting poem by Robert Fulghum from a book called All I Really Need to Know I Learned in Kindergarten. ",
"I first came across it while I was training to be a teacher. ",
"Working in a number of kindergarten classrooms made me realize that there was a lot of truth in the poem, but I didn’t fully appreciate its message until I had children of my own.",
"\n\nSince it’s Fall and the kids are back in school, I thought we might look at some of the major themes of the poem and see how they apply to personal finance. ",
"The bullet points under each theme are the actual kindergarten lessons mentioned in the poem. ",
"They are followed by a few important ways we might be able to use them as we navigate our financial life. ",
"Surprisingly, these Kindergarten lessons are still very applicable to our lives.",
"\n\n1. ",
"We’re All in This Together\n\nShare everything.",
"\n\nWhen you go out in the world, watch out for traffic, hold hands and stick together.",
"\n\nWe’ve evolved into an “every man for himself” kind of society over the past couple of decades. ",
"We tend to argue more than debate, and compete rather than collaborate. ",
"Given the financial challenges we face globally, I’d have to say that approach isn’t working very well for us right now.",
"\n\nOur greatest accomplishments are achieved when we work together. ",
"Sharing our money with those less fortunate than ourselves feels good, and it often comes back to us in ways that we can’t predict ahead of time. ",
"What goes around comes around.",
"\n\n2. ",
"Be a Good Citizen\n\nPlay fair.",
"\n\nDon’t hit people.",
"\n\nDon’t take things that aren’t yours.",
"\n\nSay you’re sorry when you hurt somebody.",
"\n\nAt first I thought the hitting advice was a little off-course for a personal finance post, but if you think about it, people have done a lot worse than hitting one another over money. ",
"Making money by taking it from others or otherwise “gaming” the system will not provide the satisfaction of earning money through consistent effort and prudent management. ",
"If bending the rules to get ahead at work, in business, or with your taxes moves you ahead, rest assured that success will be temporary. ",
"Again, what goes around comes around. ",
"If you do find yourself on the wrong side of accepted ethics, a sincere apology coupled with adequate reparations can go a long way toward making amends, and perhaps cementing a valuable relationship that might otherwise have been lost.",
"\n\n3. ",
"Organization & Maintenance Matter\n\nPut things back where you found them.",
"\n\nClean up your own mess.",
"\n\nWash your hands before you eat.",
"\n\nFlush.",
"\n\nI’m a huge fan of cleaning up your own mess – not just because it’s absolutely the right thing to do, but because it’s really empowering to dig yourself out of a hole of your own making. ",
"Still, an even better plan is to avoid that hole in the first place. ",
"Organizing your finances with the use of a proper filing, budgeting, and organization system can prevent “accidents” like overdraft charges, excessive debt, and overspending. ",
"Like washing your hands before eating, these tasks can be easy to forget and kind of tedious, but a healthier life and a stronger balance sheet are worth it.",
"\n\n4. ",
"Everything in Moderation\n\nWarm cookies and cold milk are good for you.",
"\n\nLive a balanced life – learn some and think some and draw and paint and sing and dance and play and work every day some.",
"\n\nTake a nap every afternoon.",
"\n\nBalance has become increasingly important to me over the years and the concept, like the others in this poem, applies to all areas of our lives – finance included. ",
"It’s great to be right on top of your budgeting, spending and investments. ",
"But let’s not forget why we’re going through all of this work in the first place. ",
"We want to enjoy the good things in life. ",
"Some of those things can be bought. ",
"Most cannot.",
"\n\nI don’t know of many working adults who can take a nap each afternoon or many who ever take the time to make a meal out of milk and cookies. ",
"But that doesn’t mean we can’t make the time to allocate money for the fun stuff every now and again. ",
"And yeah – sometimes a quick nap on a Saturday afternoon is just what you need to tackle those financial issues with renewed vigor and to simply be happy.",
"\n\n5. ",
"Pay Attention: Time Is of the Essence\n\nBe aware of wonder. ",
"Remember the little seed in the Styrofoam cup: the roots go down and the plant goes up and nobody really knows how or why, but we are all like that.",
"\n\nGoldfish and hamsters and white mice and even the little seed in the Styrofoam cup – they all die. ",
"So do we.",
"\n\nAnd then remember the Dick-and-Jane books and the first word you learned – the biggest word of all – LOOK.",
"\n\nWe can’t know what will happen in our lives, the economy, or the how the stock market will perform next year or next decade. ",
"In the end, all we can really do is our best. ",
"The main goals of our time in the kindergarten classroom, and of our limited time on this planet, are to learn as much as we can, have fun doing it, and leave it in better condition than we found it. ",
"If you can do all of those things, you’ll pass kindergarten – and life – with flying colors.",
"\n\nWhat are some of the basics you learned in kindergarten that have stayed with you?"
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