EGFR Mutations in Latinos From the United States and Latin America.
Epidermal growth factor receptor (EGFR) mutations confer sensitivity to EGFR tyrosine kinase inhibitors in patients with advanced non-small-cell lung cancer (NSCLC). There are limited and conflicting reports on the frequency of EGFR mutations in Latinos. Samples from 642 patients with NSCLC from seven institutions in the United States and Latin America were assessed for EGFR mutations (exons 18 to 21) at Clinical Laboratory Improvement Amendments-certified central laboratories. EGFR mutation analysis was successfully performed in 480 (75%) of 642 patients; 90 (19%) were Latinos, 318 (66%) were non-Latino whites, 35 (7%) were non-Latino Asians, 30 (6%) were non-Latino blacks, and seven (2%) were of other races or ethnicities. EGFR mutations were found in 21 (23%) of 90 Latinos with varying frequencies according to the country of origin; Latinos from Peru (37%), followed by the United States (23%), Mexico (18%), Venezuela (10%), and Bolivia (8%). In never-smoker Latinos and Latinos with adenocarcinoma histology, EGFR mutation frequencies were 38% and 30%, respectively. There was a significant difference in the frequency of EGFR mutations among the different racial and ethnic subgroups analyzed (P < .001), with non-Latino Asians having the highest frequency (57%) followed by Latinos (23%), non-Latino whites (19%), and non-Latino blacks (10%). There was no difference between Latinos (23%) and non-Latinos (22%; P = .78) and Latinos and non-Latino whites (P = .37). Patients from Peru had an overall higher frequency of mutations (37%) than all other Latinos (17%), but this difference only exhibited a trend toward significance (P = .058). There was no significant difference between the frequency of EGFR mutations in NSCLC in Latinos and non-Latinos.


Paraffin spheres as porogen to fabricate poly(L-lactic acid) scaffolds with improved cytocompatibility for cartilage tissue engineering.
Three-dimensional poly(L-lactic acid) (PLLA) scaffolds with high porosity and pore size ranging from 150 to 700 microm were conveniently prepared with paraffin spheres used as porogen. PLLA/1,4-dioxane solution containing a given amount of paraffin spheres was frozen at -25 degrees C to obtain a solidified mixture, followed with freeze drying and subsequent leaching with hexane to remove the 1,4-dioxane and paraffin spheres, respectively. The fabricated PLLA scaffolds were highly porous with evenly distributed and interconnected pores. The microstructures of the PLLA scaffolds as a function of paraffin-sphere size, paraffin-sphere dosage, and PLLA concentration were characterized by confocal laser scanning microscopy (CLSM) and scanning-electronic microscopy (SEM). To improve the cytocompatibility of the bioinert material, a hybrid PLLA scaffold containing Type I collagen was prepared by pressing the collagen solution into the scaffold under reduced pressure. The amounts of the collagen introduced in the scaffolds were detected by ninhydrin method. The distribution of the collagen in the scaffolds was studied with CLSM. Finally, in vitro cell culture was performed by injecting a chondrocyte suspension into the scaffolds. The results showed that the chondrocytes were more evenly distributed and more spread out in the collagen-modified PLLA scaffolds than in the unmodified ones.


Aberrant high expression of immunoglobulin G in epithelial stem/progenitor-like cells contributes to tumor initiation and metastasis.
High expression of immunoglobulin G (IgG) in many non-B cell malignancies and its non-conventional roles in promoting proliferation and survival of cancer cells have been demonstrated. However, the precise function of non-B IgG remains incompletely understood. Here we define the antigen specificity of RP215, a monoclonal antibody that specifically recognizes the IgG in cancer cells. Using RP215, our study shows that IgG is overexpressed in cancer cells of epithelial lineage, especially cells with cancer stem/progenitor cell-like features. The RP215-recognized IgG is primarily localized on the cell surface, particularly lamellipodia-like structures. Cells with high IgG display higher migration, increased invasiveness and metastasis, and enhanced self-renewal and tumorgenecity ability in vitro and in vivo. Importantly, depletion of IgG in breast cancer leads to reduced adhesion, invasion and self-renewal and increased apoptosis of cancer cells. We conclude that high expression of IgG is a novel biomarker of tumor progression, metastasis and cancer stem cell maintenance and demonstrate the potential therapeutic benefits of RP215-recognized IgG targeted strategy.


Kluyveromyces marxianus supplementation ameliorates alcohol-induced liver injury associated with the modulation of gut microbiota in mice.
The aim of this study was to evaluate the intervention effect of the potential probiotic Kluyveromyces marxianus YG-4 isolated from Tibetan kefir grains on alcoholic liver disease (ALD). Eight-week-old male C57BL/6J mice were fed with a Lieber-DeCarli (LDC) diet containing ethanol with a progressively increasing concentration from 1% to 4% (vol/vol) to establish an ALD mouse model. Our results suggested that K. marxianus treatment improved ALD, as demonstrated by the reduction of serum ALT and AST levels and the suppression of TLR4/NF-κB-mediated inflammatory response in the liver. K. marxianus administration significantly elevated antioxidant activities of SOD, CAT and GSH-Px, and reduced the MDA level in mice. K. marxianus supplementation repaired the gut barrier by increasing tight junction proteins and the number of goblet cells in the colon of ALD mice. In addition, treatment with K. marxianus restored alcohol-induced gut dysbiosis. Specifically, K. marxianus administration depleted the abundance of Lactobacillus, Coriobacteriaceae_UCG-002 and Candida, while increased that of Allobaculum, Dubosiella and Epicoccum in mice. Our findings open new possibilities for K. marxianus application in ALD treatment.


Changes in Cardiovascular Disease Risk Factors With Immediate Versus Deferred Antiretroviral Therapy Initiation Among HIV-Positive Participants in the START (Strategic Timing of Antiretroviral Treatment) Trial.
HIV infection and certain antiretroviral therapy (ART) medications increase atherosclerotic cardiovascular disease risk, mediated, in part, through traditional cardiovascular disease risk factors. We studied cardiovascular disease risk factor changes in the START (Strategic Timing of Antiretroviral Treatment) trial, a randomized study of immediate versus deferred ART initiation among HIV-positive persons with CD4+ cell counts >500 cells/mm3. Mean change from baseline in risk factors and the incidence of comorbid conditions were compared between groups. The characteristics among 4685 HIV-positive START trial participants include a median age of 36 years, a CD4 cell count of 651 cells/mm3, an HIV viral load of 12 759 copies/mL, a current smoking status of 32%, a median systolic/diastolic blood pressure of 120/76 mm Hg, and median levels of total cholesterol of 168 mg/dL, low-density lipoprotein cholesterol of 102 mg/dL, and high-density lipoprotein cholesterol of 41 mg/dL. Mean follow-up was 3.0 years. The immediate and deferred ART groups spent 94% and 28% of follow-up time taking ART, respectively. Compared with patients in the deferral group, patients in the immediate ART group had increased total cholesterol and low-density lipoprotein cholesterol and higher use of lipid-lowering therapy (1.2%; 95% CI, 0.1-2.2). Concurrent increases in high-density lipoprotein cholesterol with immediate ART resulted in a 0.1 lower total cholesterol to high-density lipoprotein cholesterol ratio (95% CI, 0.1-0.2). Immediate ART resulted in 2.3% less BP-lowering therapy use (95% CI, 0.9-3.6), but there were no differences in new-onset hypertension or diabetes mellitus. Among HIV-positive persons with preserved immunity, immediate ART led to increases in total cholesterol and low-density lipoprotein cholesterol but also concurrent increases in high-density lipoprotein cholesterol and decreased use of blood pressure medications. These opposing effects suggest that, in the short term, the net effect of early ART on traditional cardiovascular disease risk factors may be clinically insignificant." URL: http://www.clinicaltrials.gov. Unique identifier: NCT00867048.


A new fossil dolphin Dilophodelphis fordycei provides insight into the evolution of supraorbital crests in Platanistoidea (Mammalia, Cetacea).
Many odontocete groups have developed enlarged facial crests, although these crests differ in topography, composition and function. The most elaborate crests occur in the South Asian river dolphin (Platanista gangetica), in which they rise dorsally as delicate, pneumatized wings anterior of the facial bones. Their position wrapping around the melon suggests their involvement in sound propagation for echolocation. To better understand the origin of crests in this lineage, we examined facial crests among fossil and living Platanistoidea, including a new taxon, Dilophodelphis fordycei, nov. gen. and sp., described herein, from the Early Miocene Astoria Formation of Oregon, USA. We measured the physical extent and thickness of platanistoid crests, categorized their relative position and used computed tomography scans to examine their internal morphology and relative bone density. Integrating these traits in a phylogenetic context, we determined that the onset of crest elaboration or enlargement and the evolution of crest pneumatization among the platanistoids were separate events, with crest enlargement beginning in the Oligocene. However, we find no evidence for pneumatization until possibly the Early Miocene, although certainly by the Middle Miocene. Such an evolutionary context, including data from the fossil record, should inform modelling efforts that seek to understand the diversity of sound generation morphology in Odontoceti.


The ulcerative colitis marker protein WAFL interacts with accessory proteins in endocytosis.
Ulcerative colitis (UC) is one of the major forms of inflammatory bowel disease with unknown cause. A molecular marker, WAFL, has recently been found to be up-regulated in the inflamed colonic mucosa of UC patients. Towards understanding biological function of WAFL, we analyzed proteins interacting with WAFL in HEK-293 cells by immunoprecipitation and mass spectrometry. Among four proteins found to specifically interact with WAFL, both KIAA0196 and KIAA1033 bind to alpha-appendage of the adaptor protein complex 2 (AP2), which acts as an interaction hub for accessory proteins in endocytosis mediated by clathrin-coated vesicle (CCV). The specific interaction between WAFL and KIAA0196 was also confirmed in human colorectal carcinoma HCT-116 cells by co-immunoprecipitation with specific antibodies. Meta-analyses of the databases of expressed genes suggest that the three genes are co-expressed in many tissues and cell types, and that their molecular function may be classified in the category of 'membrane traffic protein'. Therefore, these results suggest that WAFL may play an important role in endocytosis and subsequent membrane trafficking by interacting with AP2 through KIAA0196 and KIAA1033.


[Perfusion, Metabolism and Regional Myocardial Contractility in Patients With Valvular Heart Disease: a Long-Term Follow-Up After Valve Replacement].
To perform complex assessment of myocardial perfusion, metabolism, and regional contractility in patients with valvular heart disease (VHD) in remote period after surgical correction of heart defects. We examined 30 patients with VHD in 1-8 years after heart valve replacement because of mitral (n=13) or aortic (n=17) valve defects of the following etiology: rheumatic heart disease (n=10), degeneration of connective tissue (n=13), infective endocarditis (n=7). For assessment of myocardial perfusion, metabolism, and regional contractility, we used single photon emission computed tomography (SPECT). Myocardium of patients who had undergone surgical correction of heart valve defects was characterized by dominating diffuse sclerotic changes localized mostly in basal segments. Hibernating myocardium was a less frequent finding. There were segments of borderline state with normal parameters of metabolism but slight impairment of perfusion without pronounced disturbances of kinetics. Patients with infectious etiology of valvular defects had combined perfusion and metabolic changes in multiple myocardial segments. Extensive derangements of perfusion and metabolism were characteristic for patients with connective tissue degeneration. Signs of cardiac failure caused by disturbances of myocardial perfusion and metabolism in patients with valvular heart disease can persist even after successful correction of valvular defect with normalization of intracardiac hemodynamics.


[Adenocarcinoma following restorative proctocolectomy for ulcerative colitis: a case report and review of the literature].
Restorative proctocolectomy with ileal pouch-anal anastomosis has been accepted as the surgical treatment of choice for most patients with ulcerative colitis. The occurrence of adenocarcinoma arising near or into the ileal pouch is rare. Only 19 such cases have been reported so far. The authors report a case of a 67-year old male who developed an adenocarcinoma in the small rectal stump 12 years after a restorative proctocolectomy with double stapled ileal pouch-low rectal anastomosis for ulcerative colitis unresponsive to medical treatment. They, after a literature review, examine same steps of the procedure and emphasize the importance of regular and prolonged follow-up for all patients having restorative proctocolectomy for ulcerative colitis.


Morpho-Phonetic Effects in Speech Production: Modeling the Acoustic Duration of English Derived Words With Linear Discriminative Learning.
Recent evidence for the influence of morphological structure on the phonetic output goes unexplained by established models of speech production and by theories of the morphology-phonology interaction. Linear discriminative learning (LDL) is a recent computational approach in which such effects can be expected. We predict the acoustic duration of 4,530 English derivative tokens with the morphological functions DIS, NESS, LESS, ATION, and IZE in natural speech data by using predictors derived from a linear discriminative learning network. We find that the network is accurate in learning speech production and comprehension, and that the measures derived from it are successful in predicting duration. For example, words are lengthened when the semantic support of the word's predicted articulatory path is stronger. Importantly, differences between morphological categories emerge naturally from the network, even when no morphological information is provided. The results imply that morphological effects on duration can be explained without postulating theoretical units like the morpheme, and they provide further evidence that LDL is a promising alternative for modeling speech production.


The association between cholesterol and mortality in heart failure. Comparison between patients with and without coronary artery disease.
Hypercholesterolemia is a risk factor for development of coronary artery disease (CAD), however, several reports have suggested that low serum cholesterol is associated with a worse prognosis in patients with congestive heart failure (CHF). The objective of this study was to determine the prognostic value of cholesterol for CHF. The study subjects consisted of 133 consecutive patients hospitalized in our institution for progressive heart failure from April 2000 to March 2003. Thirty-two percent of the patients had CAD. After improvement of congestive heart failure and discharge from the hospital, lipid profiles, including serum total cholesterol (TC), triglycerides, and high and low density lipoprotein cholesterol (HDL, LDL, respectively), were obtained. During the follow-up period (2.3 +/- 0.9 years), 21 patients died. There was a significant difference between survivors and nonsurvivors in HDL (53 +/- 15, 43 +/- 15 mg/dL, P = 0.01), but no differences were observed in other variables. In patients with CAD, survivors had significantly lower TC concentrations (179 +/- 30 versus 246 +/- 55 mg/dL, P = 0.004), although in patients without CAD, survivors had significantly higher TC concentrations (203 +/- 37 versus 170 +/- 40 mg/dL, P = 0.02). Multivariate analysis showed high TC predicted a worse outcome in patients with CAD (odds ratio (OR) = 1.052, 95% confidence interval (CI) 1.002-1.104, P = 0.04), but a better outcome in patients without CAD (OR = 0.972, 95% CI 0.948-0.997, P = 0.03), independent of age, gender, medication, and complications. Thus, low serum cholesterol is associated with an improved outcome in patients with CAD, while it predicts a worse outcome in patients without CAD.


Molecular analysis of the massive GSH transport mechanism mediated by the human Multidrug Resistant Protein 1/ABCC1.
The transporter Multidrug Resistance Protein 1 (MRP1, ABCC1) is implicated in multidrug resistant (MDR) phenotype of cancer cells. Glutathione (GSH) plays a key role in MRP1 transport activities. In addition, a ligand-stimulated GSH transport which triggers the death of cells overexpressing MRP1, by collateral sensitivity (CS), has been described. This CS could be a way to overcome the poor prognosis for patients suffering from a chemoresistant cancer. The molecular mechanism of such massive GSH transport and its connection to the other transport activities of MRP1 are unknown. In this context, we generated MRP1/MRP2 chimeras covering different regions, MRP2 being a close homolog that does not trigger CS. The one encompassing helices 16 and 17 led to the loss of CS and MDR phenotype without altering basal GSH transport. Within this region, the sole restoration of the original G1228 (D1236 in MRP2) close to the extracellular loop between the two helices fully rescued the CS (massive GSH efflux and cell death) but not the MDR phenotype. The flexibility of that loop and the binding of a CS agent like verapamil could favor a particular conformation for the massive transport of GSH, not related to other transport activities of MRP1.


Spanish pacemaker registry. 20th official report of the Heart Rhythm Association of the Spanish Society of Cardiology (2022).
This article reports the cardiac pacing activity performed in 2022, including the total number of implants, adherence to remote monitoring, demographic and clinical factors, and the characteristics of the implanted devices. The information sources were the CardioDispositivos online platform, the European pacemaker patient identification card, and data provided by the manufacturers. The rates of conventional pacemakers and low-energy resynchronizers were 866 and 34 units per million population, respectively. A total of 815 leadless pacemakers were implanted. In all, 16426 procedures performed in 82 hospitals were reported (9407 through CardioDispositivos), representing 40% of the activity. The mean age was 78.6 years, with a predominance of men (60.3%). The most frequent disorder was atrioventricular block, and 14.5% of the patients had atrial fibrillation. There was a predominance of the DDD/R pacing mode (55.6%), and pacing mode was influenced by age, such that more than one-third of patients older than 80 years in sinus rhythm received single-chamber ventricular pacing. The remote monitoring program included 35% of conventional pacemakers and 55% of low-energy resynchronization pacemakers. The number of conventional pacemakers increased by 5.6%, low-energy resynchronizers by 16%, and leadless pacemakers by 25%. Adherence to remote monitoring was stable. The number of procedures included in CardioDispositivos increased by 11%, although the sample volume decreased. In the coming years, the widespread use of the platform will likely lead to a high-quality registry.


[Comparison of different methods for examining the feces of suckling piglets for Isospora suis].
A highly sensitive diagnostic method is of great importance for detection of I. suis (Isospora suis). In the present study KSFV (combined sedimentation-flotation method) using different flotation solutions (NaCl, ZnSO4, NaCl-sucrose) was investigated regarding sensitivity, costs and expenditure of human labour in comparison to FV (flotation method) using NaCl-sucrose solution and to AM (autofluorescence microscopy). Ninety faecal samples were examined. Using AM, oocysts were detected in 83.3% of the samples. In cases of KSFV and FV, respectively, 41.1% to 53.3% of samples were tested positive. The quantity of detected oocysts was markedly higher in AM than in KSFV or FV. Therefore AM is the most sensitive of the comprised methods. The expenditure of labour is lowest in AM, time exposure is lower in FV than in KSFV. For performance of AM a fluorescence microscope is required contrary to KSFV and FV.


Density-of-states Monte Carlo simulation of a binary glass.
A newly proposed Monte Carlo formalism has been used to simulate a glass-forming liquid above and below the glass transition temperature. The heat capacity exhibits a sharp peak at a temperature lower than that reported from extensive molecular dynamics simulations. Its height is larger than that reported earlier. At temperatures below mode coupling, the average inherent-structure energy of the configurations generated in this work is significantly lower than that reported in the literature. The entropy of the supercooled liquid is calculated directly from our simulations and that of a disordered solid is calculated by a normal-mode analysis. We find that at low temperatures these two entropy curves become essentially parallel. They do not intersect each other, raising questions about the existence of a Kauzmann temperature in this glass-forming mixture.


Anti-idiotype antibody in the primate. 1. Characterization and specificity against cells primed for histocompatibility determinants.
An anti-idiotype serum was raised in a chimpanzee (A) by immunization with autologous lymphocytes primed in vitro against an unrelated chimp (B). This autoantiserum in the presence of complement was cytotoxic for 5 to 7% of the resting lymphocytes from chimp A and for 30 to 45% of the mixed lymphocyte culture (MLC) primed cells (A X Bx), but was not reactive against the lymphocytes of the priming chimp (B). Anti-idiotype antibody and complement treatment of autologous resting or primed cells blocked the ability of these cells to respond in MLC or primed lymphocyte test (PLT) to the stimulator cells from chimp B, but not to cells from a third chimp. When cells from the immunized animal (A) were incubated with the autologous antiserum in vitro, they were stimulated, thus producing cells which had the same activity in PLT as did cells primed against stimulator cells of chimp B. Thus, an autoanti-idiotype serum has been raised in a primate system which identifies the recognition structure on autologous T cells directed against antigeneic determinants on the stimulator cells of a histoincompatible donor.


[Vertical distribution characteristics of organochlorine pesticides and polychlorinated biphenyls in sediment core from Lake Nansihu].
The organochlorinne pesticides (OCPs) and the polychlorinated biphenyls (PCBs) in sediment core collected from Lake Nansihu were quantificationally analyzed by GC-ECD. The pollution history of OCPs and PCBs in this area was reconstructed by analysis of the concentrations of OCPs and PCBs connected with the deposition time by 210Pb isotope dating method. The results indicated that the concentration ranges of OCPs, HCHs, DDTs and PCBs were 1.64 - 17.9, 0.66 - 12.5, 0.24 - 2.99 and 7.84 - 42.8 ng g(-1) respectively. The sediment rate and the average mass sedimentation were calculated to be 0.330 cm a(-1) and 0.237 g (cm2 a)(-1), and the sediment time of this core sample ranged from 1899 to 2000. OCPs and PCBs had the peak value at 1960s and 1970s. The source analysis showed HCH came from HCH products and lindane, and HCH and DDT came from historic using. The source of PCBs had been deduced to be the blench process of paper making and incinerator emission. Except the concentration of beta-HCH, gamma-HCH and aldrin at 14 - 16 cm depth, the other concentrations of OCPs and PCBs were all less than the low ecologic risk value. The whole sediment core of Lake Nansihu belonged to the low ecologic risk level.


Delayed effect of QUIN-neurotoxicity. Ultrastructural study.
The ultrastructural studies of QUIN-injected rat hippocampus were performed in long-term experiment in order to determine delayed neurotoxic effect of this compound. QUIN, in a single dose, was administered stereotaxically into the dorsal hippocampus. The morphological changes were characterized by marked atrophy of neuropil accompanied by remarkable glial pathology. The neuronal changes consisted in loss of many nerve cells, abnormalities of nuclear chromatin in the persisting pyramidal neurons and degeneration of postsynaptic dendrites. The glial changes were represented by electron-lucency of astrocytic cytoplasm and their processes. The mechanism of this reaction is unclear and the suggestion, that electron-lucent astrocytes represent a form of transformed glia especially sensitive to amino acids, releasing from degenerating neurons is discussed.


Intermolecular binding of blueberry pectin-rich fractions and anthocyanin.
Pectin was extracted from blueberry powder into three fractions of water soluble (WSF), chelator soluble (CSF) and sodium carbonate soluble (NSF). The fractions were incubated with cyanidin-3-glucoside (C3G), a mixture of five anthocyanidins (cyanidin, pelargonidin, malvidin, petunidin and delphinidin) or blueberry juice at pH 2.0-4.5. Free anthocyanins and bound anthocyanin-pectin mixtures were separated by ultrafiltration. WSF bound the least amount of anthocyanin at all pH values. CSF had stronger anthocyanin binding ability at pH 2.0-3.6, while NSF had stronger anthocyanin binding ability at pH 3.6-4.5. The pectin and anthocyanin binding was lowest at pH 4.5 and higher at pH 2.0-3.6. Nearly doubling C3G pigment content increased bound anthocyanin percentage by 16-23% at pH 3.6, which favored anthocyanin aromatic stacking, compared to 3-9% increase at pH 2.0. Ionic interaction between anthocyanin flavylium cations and free pectic carboxyl groups, and anthocyanin stacking may be two major mechanisms for pectin and anthocyanin binding.


Association of Lead aVR T-wave Amplitude With Cardiovascular Events or Mortality Among Prevalent Dialysis Patients.
In dialysis patients, electrocardiogram (ECG) abnormalities are common. However, the associations between the T-wave of the lead aVR (aVRT) amplitude and cardiovascular (CV) events or total mortality are unknown. We performed a prospective, observational cohort study of prevalent hemodialysis patients (N = 474), followed for 4 years. Outcomes were composite CV events and all-cause mortality. Predictors were baseline aVRT and other ECG findings. ECG parameters were analyzed in three models: model 1, univariate; model 2, basic adjustments; and model 3, model 2 plus serum albumin, C-reactive protein level, and NT-proBNP. By Cox analysis, aVRT was best associated with both endpoints through model 1 to 3 compared to other ECG findings. Patients categorized according to aVRT amplitude showed a step-by-step increase in hazard ratios for both endpoints. The aVRT amplitude level was significantly associated with not only composite CV events but also with all-cause mortality in prevalent dialysis patients.


Effects of sucrose supplementation on dry matter intake, milk yield, and blood metabolites of periparturient Holstein dairy cows.
Thirty-four multiparous Holstein dairy cows (780 +/- 17.2 kg body weight; 3.4 +/- 0.08 body condition score) were used in a completely randomized design to evaluate the effects of fermentable carbohydrate source on dry matter (DM) intake, milk production, and blood metabolites of transition cows. Treatments were initiated 30 d before expected calving date. After calving, all cows went onto a similar lactating cow diet. Dry matter intake was measured daily, and milk production and composition were measured weekly for 56 d after calving The control diet consisted of 11.5% ground corn, whereas the treatment diet consisted of sucrose replacing 2.7% of ground corn on a DM basis. Prepartum plasma glucose concentrations were higher (66.3 vs. 69.3 +/- 1.13 mg/dl) for cows fed the diet containing sucrose. Glucose concentrations were not different postpartum. Prepartum and postpartum nonesterified fatty acids, blood urea N, and insulin did not differ between treatments. Substitution of sucrose as a readily available carbohydrate source for ground corn did not affect prepartum or postpartum DM intake. Milk yield, 3.5% fat-corrected milk yield, and milk components did not differ between treatments. Results from this research demonstrated that partially replacing ground corn with sucrose did not enhance prepartum or postpartum intake or performance.


[A new method for purification and identification of hepatocellular carcinoma stem cell of SMMC-7721].
To explore a new efficient purification method of hepatocellular carcinoma (HCC) stem cells and identify their features. Human hepatocellular carcinoma cell line SMMC-7721 was cultured in sphere-culture system of polyhedra-treated dish and tumor stem cell specific medium. Upon the formation of cellular sphere, the cells were inoculated subcutaneously into immunocompromised mice and received the interventions of different concentrations of cisplatin. Then the drug-resistant cells were purified and re-cultured in TSC medium. Finally the stem cell markers and tumor stem cell markers were determined through real-time polymerase chain reaction (PCR), immunofluorescence method and flow cytometry. Through the double filter of TSC medium and cisplatin-resistance, SMMC-7721 stem cells could be grown in a suspended form and formed spheres in TSC medium. The stem cell markers (NANOG, OCT-4, SOX-2 and Notch) and tumor stem cell markers (CD24, 90.0%; CD133, 6.1%; CD90, 4.8%) were all over-expressed in purified cancer stem cells as compared with ordinary cells. And the over-expression of CD24 was the most obvious. The combination of in vitro cell culture with TSC medium, in vivo proliferation and cisplatin resistance test is a new efficient method of purifying hepatocellular carcinoma stem cells. Tumor stem cell with stem cell characteristics and an over-expression of CD24 may be cloned from SMMC-7721.


Immunohistochemical markers for arthritis in psoriasis.
To examine the immunohistological features in the involved skin of patients with psoriatic arthritis (PA) (n = 15), compared with those in involved skin from patients with psoriasis but no arthritis (n = 5), and with a group with normal skin (n = 4), to identify markers for arthritis in psoriasis. Skin was obtained from patients by 6 mm punch biopsy and normal skin was provided by the department of plastic surgery. Samples were stained with monoclonal antibodies against T cells (CD3, CD8, CD4, CD45Ro), B cells (CD20), macrophages (mac387), vascular endothelium (FVIII-related antigen) and a Langerhan's cell marker (p155). The number of cells/vessels staining with each monoclonal antibody was calculated and serial sections of skin were examined to estimate the presence of DR+ keratinocytes. There were significantly more CD45Ro T-cells and blood vessels in patients with psoriatic arthritis compared with both psoriasis alone, and with normal controls (p < 0.02). While B-cells were not seen in psoriasis without arthritis or in normal skin, a small but significant number were observed in PA (p < 0.02). Furthermore, while DR+ keratinocytes were present in both psoriatic arthritis and psoriasis skin, there were significantly more DR+ cells in the psoriatic arthritis epidermis compared with psoriasis alone (p < 0.02). This study suggests that increased numbers of CD45Ro T-cells, greater vascularity, the presence of B-cells, and increased numbers of DR+ epidermal cells are markers for arthritis in patients with psoriasis.


Short- and long-term results of stapedectomy in children.
Studies have indicated that stapedectomy can be an effective procedure in children for correcting conductive hearing losses due to juvenile otosclerosis. However, because childhood otosclerosis is rare and children commonly choose to use hearing aids in lieu of undergoing surgery, little outcome data are available. The purpose of this retrospective study was to provide additional outcome data in both the short and the long term. Stapedectomies were performed on 47 children. Preoperative hearing results were compared with 6-month postoperative hearing results. Hearing results for the children who had long-term follow-up (5 years or more) were compared with the 6-month postoperative results. Stapedectomy was successful (postoperative air conduction pure-tone average [PTA] within 10 dB of the preoperative bone conduction PTA) in 91.7% of the cases. The mean overclosure of the preoperative bone conduction PTA by the postoperative air conduction PTA was 0.2 dB. The mean PTA hearing improvement was 32.8 dB. Results from the 21 children (28 ears) who had long-term follow-up indicated an average 0.7 dB/year PTA worsening from the 6-month postoperative PTA. Results from this study provide additional evidence that stapedectomy can be an effective procedure for correcting conductive hearing losses due to juvenile otosclerosis.


Murine leukemia inhibitory factor gene disruption attenuates the hypothalamo-pituitary-adrenal axis stress response.
Recently, we have shown that human fetal pituitary, mouse corticotroph AtT20 cells, and murine hypothalamus and pituitary express leukemia inhibitory factor (LIF). LIF knockout mice (LIFKO), heterozygous and wild type (wt), of B6D2F1 genetic background were used to examine whether LIF may play a role in the regulation of the hypothalamo-pituitary-adrenal axis in vivo. Resting levels of plasma ACTH and corticosterone were similar in all three genotypes. However, LIFKO mice did not respond to 30-min restraint and 45-min immobilization stress with increased plasma ACTH. Increased circulating ACTH was only observed in LIFKO mice after very short immobilization stress (15 min), but this ACTH level was lower than in wt animals (P < 0.05). Injection of mycobacterial adjuvant resulted in a 2-fold increase of corticosterone levels 7 days after treatment in wt, but not LIFKO, mice (P < 0.05). Pituitary POMC messenger RNA (mRNA) levels were very low in LIFKO animals. Although 15 and 45 min of immobilization stress resulted in enhanced POMC mRNA content in all three groups, this elevation was lowest in LIFKO mice. Injection of 12 microg murine LIF to LIFKO and normal C57BL/6 animals resulted in increased plasma ACTH and corticosterone levels and elevated pituitary POMC mRNA levels in both LIF-repleted and LIF-depleted mice. Thus, LIF appears to play an important role in activating the hypothalamo-pituitary-adrenal axis during stress and inflammation.


[The renin-angiotensin system in diabetic patients].
We review available data on the activity of the renin-angiotensin system (RAS), responsiveness to angiotensin II (ANG II), ANG II receptor number, and effects of inhibition of the RAS by angiotensin I converting enzyme (ACE) inhibitors in patients with diabetes mellitus. Most authors, including ourselves, observed a normal or enhanced activity of the RAS in metabolically stable diabetics. Increased but also reduced activity of the RAS was described in nephropathic diabetes. This is in contrast to the common suggestion that the RAS of diabetics is generally suppressed and functionally inactive. The last assumption was mainly based on the finding of reduced ANG II receptor numbers in anorectic, severely hyperglycemic rats. These findings could not be reproduced in man, and a higher ANG II receptor concentration on platelets of diabetics goes in parallel with the frequent finding of an enhanced pressor response to infused ANG II in diabetes. This increased responsiveness is most probably of functional importance since the RAS is not suppressed - as one would expect - in the face of a supranormal body sodium content. A number of data also indicate that renal resistance vessels display increased responsiveness to ANG II in diabetics. This may be a reason for hyperfiltration. This notion is further supported by the reduction of albuminuria which is usually observed following inhibition of the RAS with ACE inhibitors, and which may be an index of reduction of glomerular capillary pressure in human diabetes.


Role of tyrosine phosphorylation in the regulation of cleavage secretion of angiotensin-converting enzyme.
Both germinal (gACE) and somatic (sACE) isozymes of angiotensin-converting enzyme (ACE) are type I ectoproteins whose enzymatically active ectodomains are cleaved and shed by a membrane-bound protease. Here, we report a role of protein tyrosine phosphorylation in regulating this process. Strong enhancements of ACE cleavage secretion was observed upon enhancing protein Tyr phosphorylation by treating gACE- or sACE-expressing cells with pervanadate, an inhibitor of protein Tyr phosphatases. Secreted gACE, cell-bound mature gACE and its precursors were all Tyr-phosphorylated, as was the endoplasmic reticulum protein, immunoglobulin heavy chain-binding protein, that co-immunoprecipitated with ACE. The enhancement of cleavage secretion by pervanadate did not require the presence of the cytoplasmic domain of ACE, and it was not accomplished by enhancing the rate of intracellular processing of the protein. The observed enhancement of cleavage secretion of ACE in pervanadate-treated cells was specifically blocked by an inhibitor of the p38 mitogen-activated protein (MAP) kinase but not by inhibitors of many other Ser/Thr and Tyr protein kinases, including a specific inhibitor of protein kinase C that, however, could block the enhancement of cleavage secretion elicited by phorbol ester. These results indicate that ACE Tyr phosphorylation, probably in the endoplasmic reticulum, enhances the rate of its cleavage secretion at the plasma membrane using a regulatory pathway that may involve p38 MAP kinase.


Host-parasite coevolution in populations of constant and variable size.
The matching-allele and gene-for-gene models are widely used in mathematical approaches that study the dynamics of host-parasite interactions. Agrawal and Lively (Evolutionary Ecology Research 4:79-90, 2002) captured these two models in a single framework and numerically explored the associated time discrete dynamics of allele frequencies. Here, we present a detailed analytical investigation of this unifying framework in continuous time and provide a generalization. We extend the model to take into account changing population sizes, which result from the antagonistic nature of the interaction and follow the Lotka-Volterra equations. Under this extension, the population dynamics become most complex as the model moves away from pure matching-allele and becomes more gene-for-gene-like. While the population densities oscillate with a single oscillation frequency in the pure matching-allele model, a second oscillation frequency arises under gene-for-gene-like conditions. These observations hold for general interaction parameters and allow to infer generic patterns of the dynamics. Our results suggest that experimentally inferred dynamical patterns of host-parasite coevolution should typically be much more complex than the popular illustrations of Red Queen dynamics. A single parasite that infects more than one host can substantially alter the cyclic dynamics.


A landscape ecology assessment of land-use change on the Great Plains-Denver (CO, USA) metropolitan edge.
For better or worse, in those parts of the world with a widespread farming, livestock rising, and urban expansion, the maintenance of species richness and ecosystem services cannot depend only upon protected natural sites. Can they rely on a network of cultural landscapes endowed with their own associated biodiversity? We analyze the effects of land-cover change on landscape ecological patterns and processes that sustain bird species richness associated to cropland-grassland landscapes in the Great Plains-Denver metropolitan edge. Our purpose is to assess the potential contribution to bird biodiversity maintenance of Great Plain's cropland-grassland mosaics kept as farmland green belts in the edge of metropolitan areas. We present a quantitative landscape ecology assessment of land-cover changes (1930-2010) experienced in five Great Plains counties in Colorado. Several landscape metrics assess the diversity of land-cover patterns and their impact on ecological connectivity indices. These metrics are applied to historical land-cover maps and datasets drawn from aerial photos and satellite imagery. The results show that the cropland-grassland mosaics that link the metropolitan edge with the surrounding habitats sheltered in less human-disturbed areas provide a heterogeneous land matrix were a high bird species richness exists. They also suggest that keeping multifunctional farmland-grassland green belts near the edge of metropolitan areas may provide important ecosystem services, supplementing traditional conservation policies. Our maps and indicators can be used for selecting certain types of landscape patterns and priority areas on which biodiversity conservation efforts and land-use planning can concentrate.


Green synthesis of silver nanoparticles using bovine skin gelatin and its antibacterial effect on clinical bacterial isolates.
Nanoparticles are being increasingly used in day-to-day life. Therefore, concerns have been raised regarding their interactions with the surrounding environment. This study focused on a simple green method for synthesizing silver nanoparticles (Ag-NPs) in an autoclave at 15 psi (103 kPa) and 121°C. An aqueous solution of AgNO3 as a precursor of Ag-NPs and gelatin (type B) reducing and/or stabilizing (capping) agent were used. The effect of various AgNO3 concentrations of certain gelatin concentration and various gelatin concentrations at constant AgNO3 concentration, and autoclaving time, was studied. UV-Vis spectra ascribed that the presence of localized surface plasmon resonance (SPR) of the synthesized Ag-NPs. TEM images and the selected area of electron diffraction confirmed, the formation of Ag-NPs with a diameter of approximately 5 ±0.35 nm. Furthermore, FT-IR revealed that a gelatin polymer matrix stabilized the synthesized Ag-NPs. The Well diffusion assay was used to test the effect of Ag-NPs on six clinical bacterial isolates, where Gram positive bacteria were more susceptible to Ag-NPs than Gram negative bacteria. Therefore, Ag-NPs capped by gelatin have remarkable potential effect as an antibacterial agent, and they not only have various medical applications but can also be used in biological, pharmaceutical and industrial fields.


Activation of AP-1 in primary B lymphocytes by surface immunoglobulin requires de novo Jun-B synthesis.
We demonstrate herein that resting primary B lymphocytes do not contain detectable levels of AP-1 (TRE)-binding activity. Upon cross-linking of surface immunoglobulin (sIg) receptors, TRE-binding activity is induced within 2 hr and its appearance requires de novo protein synthesis. Antisera to Jun-B inhibits the vast majority of TRE-binding activity, indicating that Jun-B is a primary component of B cell TRE-binding complexes. In keeping with this, Jun-B protein is not detectable in cytosol or nuclear extracts from resting B lymphocytes, as determined by immunoblotting with Jun-B antisera. However, the nuclear expression of Jun-B is induced within 2 hr following sIg cross-linking and is completely blocked by cycloheximide. 35S-labeling studies suggest that the increase in Jun-B expression results from de novo protein synthesis. Moreover, Jun-B migrates in SDS-polyacrylamide gels as two distinct electrophoretic proteins that correspond to a 41-kDa species and a phosphorylated 47-kDa form. These results suggest that the induction of AP-1-binding activity in primary B lymphocytes following sIg cross-linking does not result from post-translational phosphorylation of a preexisting cellular pool of Jun-B protein, but rather is coupled to the stimulation of de novo Jun-B synthesis. Thus Jun-B synthesis represents an integral event in the production of receptor-mediated AP-1 in B cells. The significance of these results with respect to the role of Jun-B in controlling gene expression during the activation of primary B cells is discussed.


Overexpression, purification, crystallization and preliminary X-ray analysis of putative molybdenum cofactor biosynthesis protein C (MoaC2) from Mycobacterium tuberculosis H37Rv.
Rv0864 (MoaC2) from Mycobacterium tuberculosis is one of the enzymes in the molybdenum cofactor (Moco) biosynthesis pathway. Together with MoaA, MoaC is involved in the conversion of guanosine triphosphate (GTP) to precursor Z, the first step in Moco synthesis. Full-length MoaC2 (17.5 kDa, 167 residues) was cloned in Escherichia coli and purified to homogeneity. Crystals of recombinant M. tuberculosis MoaC2 were grown by vapour diffusion using a hanging-drop setup. Diffracting crystals grew in a condition in which 3 µl protein solution at 10.5 mg ml(-1) was mixed with 1.5 µl reservoir solution (0.025 M potassium sodium tartrate tetrahydrate pH 8.0) and equilibrated against 1000 µl reservoir solution. Diffraction data extending to 2.5 Å resolution were collected at 100 K. The crystal belonged to the cubic space group P2(1)3, with unit-cell parameter 94.5 Å. Matthews coefficient (V(M)) calculations suggested the presence of two molecules in the asymmetric unit, corresponding to a solvent content of about 39%. Molecular-replacement calculations using the E. coli homologue as the search model gave an unambiguous solution.


Test-retest reliability of the magnetic mismatch negativity response to sound duration and omission deviants.
Mismatch negativity (MMN) is a neurophysiological measure of auditory novelty detection that could serve as a translational biomarker of psychiatric disorders, such as schizophrenia. However, the replicability of its magnetoencephalographic (MEG) counterpart (MMNm) has been insufficiently addressed. In the current study, test-retest reliability of the MMNm response to both duration and omission deviants was evaluated over two MEG sessions in 16 healthy adults. MMNm amplitudes and latencies were obtained at both sensor- and source-level using a cortically-constrained minimum-norm approach. Intraclass correlations (ICC) were derived to assess stability of MEG responses over time. In addition, signal-to-noise ratios (SNR) and within-subject statistics were obtained in order to determine MMNm detectability in individual participants. ICC revealed robust values at both sensor- and source-level for both duration and omission MMNm amplitudes (ICC = 0.81-0.90), in particular in the right hemisphere, while moderate to strong values were obtained for duration MMNm and omission MMNm peak latencies (ICC = 0.74-0.88). Duration MMNm was robustly identified in individual participants with high SNR, whereas omission MMNm responses were only observed in half of the participants. Our data indicate that MMNm to unexpected duration changes and omitted sounds are highly reproducible, providing support for the use of MEG-parameters in basic and clinical research.


Therapeutic Multidose Preparation of a Ready-to-Use 177Lu-PSMA-617 Using Carrier Added Lutetium-177 in a Hospital Radiopharmacy and Its Clinical Efficacy.
Introduction: [177Lu]Lu-prostate-specific membrane antigen (PSMA)-617 has emerged as a promising radiopharmaceutical for targeting PSMA in metastatic castrate-resistant prostate carcinoma (mCRPC). We have optimized the radiolabeling protocol for a multidose formulation (27-28.8 GBq equivalent to 6-7 patient-doses) of [177Lu]Lu-PSMA-617 using [177Lu]Lu3+ produced via 176Lu(n,γ)177Lu route with moderate specific activity (0.66-0.81 GBq/μg). Methods: [177Lu]Lu-PSMA-617 was synthesized using moderate specific activity [177Lu]LuCl3 (0.74 GBq/μg) with PSMA-617 having metal-to-ligand molar ratio ∼1: 2.5 in CH3COONH4 buffer (0.1 M) containing gentisic acid at pH 4.0-4.5. Human prostate carcinoma cell line LNCaP cell (high PSMA expression) was used for in vitro cell-binding studies and generating tumor xenograft models in nude mice for tissue biodistribution studies. Several batches of the present formulation have been clinically administered in mCRPC patients (single patient dose: 4.44-5.55 GBq per cycle). Results: In this study we report a consistent and reproducible protocol for multidose formulations of [177Lu]Lu-PSMA-617 for adopting in a hospital radiopharmacy setting. Although the radiochemical yield of [177Lu]Lu-PSMA-617 was found to be 97.30% ± 1.03%, the radiochemical purity was 98.24% ± 0.50% (n = 19). In vitro and serum stability of [177Lu]Lu-PSMA-617 was retained up to 72 and 120 h after radiolabeling and upon storage at -20°C with a radioactive concentration between 0.37 and 0.74 GBq/mL upon using stabilizer concentration as low as 43-48 μg/mCi. Preclinical cell-binding studies of [177Lu]Lu-PSMA-617 revealed specific binding with LNCaP cells of 17.4% ± 2.4%. The uptake in LnCaP xenografted tumor (nude mice) was 7.5 ± 2.6% ID/g for ∼1.5-2.0 cm3 tumor volume at 24-h post-injection. Post-therapy (24 h) SPECT image of mCRPC patients with prior orchidectomy and various hormone therapy showed specific localization of [177Lu]Lu-PSMA-617 in the tumor region. Conclusions: Formulation of a ready-to-use multidose formulation of [177Lu]Lu-PSMA-617 was successfully achieved and the procedure was optimized for routine preparation at a hospital radiopharmacy set-up. High degree of localization of [177Lu]Lu-PSMA-617 in post-therapy SPECT scan and the post-therapeutic response confirms its therapeutic efficacy. Clinical Trials.gov ID: RPC/51/Minutes/Final dated 16th October, 2019.


Verrucoid Variant of Invasive Squamous Cell Carcinoma in Oral Submucous Fibrosis: A Clinicopathological Challenge.
Verrucous carcinoma (VC) is an exophytic, low-grade, well-differentiated variant of squamous cell carcinoma. It is described as a lesion appearing in the sixth or seventh decade of life that has minimal aggressive potential and, in long-standing cases, has been shown to transform into squamous cell carcinoma. Oral submucous fibrosis (OSMF) is a potentially malignant disorder, and about one-third of the affected population develop oral squamous cell carcinoma. The histopathological diagnosis of verrucous carcinoma is challenging, and the interpretation of early squamous cell carcinoma requires immense experience. Here we present a rare case of a 24-year-old male with OSMF transforming to verrucous carcinoma with invasive squamous cell carcinoma. Even though the case had a straightforward clinical diagnosis, the serial sectioning done for pathological diagnosis disclosed the squamous cell carcinoma.


Nitric oxide synthesis modulation - a possible diagnostic and therapeutic target in colorectal cancer.
Considering the contradictory literature data about the role of nitric oxide (NO) in colon carcinogenesis, the purpose of this study was to examine the changes of L-arginine metabolites in colon cancer and surrounding tissue as possible molecular markers of tumor behavior after surgery and the possibility of NO synthesis modulation in new individualized therapeutic strategies. The study encompassed 50 patients who underwent surgery for colorectal cancer (CRC). The three tissue specimens were taken by surgery (tumor, adjacent and healthy tissue) and the concentrations of NO2+NO3, asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) were determined in the tissue specimens. The results proved higher NO2+NO3 concentrations in adjacent tissue compared to the tumor, implicating high angiogenic potential of the tumor-surrounding tissue, which could have clinical importance in the assessment of the probability of tumor local recurrence and metastasis. Increased ADMA concentrations in tumor tissue associated with low NO levels, could lead to new therapeutic strategies directed to the use of inhibitors of NO synthesis as ideal candidates for molecular therapy of CRC. ADMA concentration in adjacent tissue was an independent predictor of distant metastasis. The obtained results suggest that determination of the examined biomarkers in CRC and adjacent tissue samples could give useful information about tumor proliferative and angiogenic potential, which in turn could enable individualization of therapy and the choice of proper adjuvant therapy in patients with CRC.


Clinical Features and Genetic Analysis of 48 Patients with Chronic Granulomatous Disease in a Single Center Study from Shanghai, China (2005-2015): New Studies and a Literature Review.
Chronic Granulomatous Disease (CGD) is a rare inherited primary immunodeficiency, which is characterized by recurrent infections due to defective phagocyte NADPH oxidase enzyme. Nowadays, little is known about Chinese CGD patients. Here we report 48 CGD patients in our single center study, which is the largest cohort study from Mainland China. The ratio of male to female was 11 : 1. The mean onset age was 0.29 years old, and 52% patients had an onset within the 1st month of life. The mean diagnosis age was 2.24 years old. 11 patients (23%) had died with an average age of 2.91 years old. 13 patients (28%) had positive family histories. The most prevalent infectious sites were the lungs (77%), followed by gastrointestinal tract (54%), lymph nodes (50%), and skin (46%). In addition, septicopyemia, thrush, and hepatosplenomegaly were also commonly observed, accounting for 23%, 23%, and 40% of the cases. Lesions due to BCG vaccination occurred in more than half of the patients. X-linked CGD due to CYBB gene mutations accounted for 75% of the cases, and 11 of them were novel mutations. Autosomal recessive inheritance accounted for 6% patients, including 1 patient with CYBA, 1 with NCF1, and 1 with NCF2 gene mutations.


Turner's syndrome and inflammatory bowel disease: a case report with immunologic studies.
A patient with Turner's syndrome who developed ulcerative colitis (UC) is reported, and reports from the literature of 16 cases of Turner's syndrome with inflammatory bowel disease (IBD) are reviewed. Most of the patients previously described had severe disease. Half of the patients had ulcerative colitis and half had Crohn's disease (CD). Of those with Crohn's, most had colonic involvement. Of note, also, half of the patients had an isochromosome for the long arm of X which usually is found in only 17% of all patients with Turner's syndrome. A high incidence of autoimmune diseases is associated with this syndrome. We demonstrate the presence of specific anticolonic antibodies in this patient and propose some possible explanations for the association of this particular chromosomal disorder with autoimmune diseases.


Insulin-like growth factor-1 (IGF-1) enhances the osteogenic activity of bone morphogenetic protein-6 (BMP-6) in vitro and in vivo, and together have a stronger osteogenic effect than when IGF-1 is combined with BMP-2.
Bone morphogenetic protein-2 (BMP-2) is widely used in orthopedic surgery and bone tissue engineering because of its strong osteogenic activity. However, BMP-2 treatments have several drawbacks and many groups are actively exploring alternatives. Since BMP-6 has been demonstrated to be more osteoinductive, its use, either alone or together with other growth factors, might be an interesting option. In this work, we have compared the effect of BMP-2, BMP-6, or insulin-like growth factor-1 (IGF-1), either alone or in combination. Murine preosteoblasts were treated with 15 nM IGF-1 and/or 6 nM BMP-2 or -6 and the expression of osteogenic marker genes, proliferation, and alkaline phosphatase (ALP) activity in vitro were analyzed. The results showed that IGF-1 greatly enhanced the BMP-induced osteogenic differentiation of these cells in general and that the ALP activity in the cultures was higher when the combination was made with BMP-6 than with BMP-2. Furthermore, we tested the osteogenic potential of these treatments in vivo by loading 25 pmoles of IGF-1 and/or 10 pmoles of BMP-2 or -6 onto absorbable collagen sponges and implanting them into an ectopic bone formation model in rats. This study revealed that only BMP-6 was able to induce bone formation at the used dose and that the addition of IGF-1 contributed to an increase of the mineralization in the implants. Hence, the combination of BMP-6 with IGF-1 might be a better alternative than BMP-2 for orthopedic surgery or bone tissue engineering approaches. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1867-1875, 2017.


Subjective Nearness-To-Death and Retirement Anxiety Among Older Workers: A Three-Way Interaction With Work Group Identification.
Retirement anxiety represents a major challenge for older workers who hold negative expectations and concerns regarding the consequences of their future retirement. Although prior studies suggest that retirement is an age-related transition that may serve as a reminder that life is nearing its end, little is known about how subjective nearness-to-death is related to retirement anxiety, and the role of work group identification as a boundary condition. The current study draws on terror management and social identity theories to hypothesize and investigate these relationships. A three-way interaction model was tested on a sample of 327 Israeli older workers. We found a positive association between subjective nearness-to-death and retirement anxiety. Further, we demonstrated that the positive relationship between subjective nearness-to-death and retirement anxiety was more prominent for older workers whose work group self-definition and self-investment were both high or were both low. The theoretical and practical implications of these findings are discussed.


Molecular analysis of the human interferon-alpha gene family.
Fifteen DNA clones containing sequences related to human interferon-alpha cDNA were isolated from a human chromosomal gene bank (Nagata et al., Nature 287 (1980) 401-408) and characterized by restriction mapping, R-loop and heteroduplex analysis. Nine distinct DNA segments hybridized strongly with interferon-alpha 1 cDNA and formed R-loops with poly(A) RNA from interferon-producing human leukocytes; most if not all of these segments represent functional interferon genes. Five segments hybridized weakly with the probe and did not form R-loops with the poly(A) RNA; one of these was characterized as an interferon-alpha pseudogene. Several DNA segments overlap and define a region of 36 kilobase pairs (kb) that contains three strongly and three weakly hybridizing sequences. From our data and those of Goeddel et al. (Nature 290 (1981) 20-25) we conclude that there exist at least 11 distinct genes of gene-like sequences of the interferon-alpha type in the human genome, of which most likely represents an allelic variant, and at least five pseudogenes distantly related to the interferon-alpha genes.


Associative effects of wet distiller's grains plus solubles and tannin-rich peanut skin supplementation on in vitro rumen fermentation, greenhouse gas emissions, and microbial changes1.
Two sets of in vitro rumen fermentation experiments were conducted to determine effects of diets that included wet distiller's grains plus solubles (WDGS) and tannin-rich peanut skin (PS) on the in vitro digestibility, greenhouse gas (GHG) and other gas emissions, fermentation rate, and microbial changes. The objectives were to assess associative effects of various levels of PS or WDGS on the in vitro digestibility, GHG and other gas emissions, fermentation rate, and microbial changes in the rumen. All gases were collected using an ANKOM Gas Production system for methane (CH4), carbon dioxide (CO2), nitrous oxide (N2O), and hydrogen sulfide (H2S) analyses. Cumulative ruminal gas production was determined using 250 mL ANKOM sampling bottles containing 50 mL of ruminal fluid (pH 5.8), 40 mL of artificial saliva (pH 6.8), and 6 g of mixed diets after a maximum of 24 h of incubation. Fermenters were flushed with CO2 gas and held at 39 °C in a shaking incubator for 24 h. Triplicate quantitative real-time polymerase chain reaction (qPCR) analyses were conducted to determine microbial diversity. When WDGS was supplied in the diet, in the absence of PS, cumulative CH4 production increased (P < 0.05) with 40% WDGS. In the presence of PS, production of CH4 was reduced but the reduction was less at 40% WDGS. In the presence of PS, ruminal lactate, succinate, and acetate/propionate (A/P) ratio tended to be less with a WDGS interaction (P < 0.01). In the presence of PS and with 40% WDGS, average populations of Bacteroidetes, total methanogens, Methanobrevibacter sp. AbM4, and total protozoa were less. The population of total methanogens (R2 = 0.57; P < 0.01), Firmicutes (R2 = 0.46: P < 0.05), and Firmicutes/Bacteroidetes (F/B) ratio (R2 = 0.46; P < 0.03) were strongly correlated with ruminal CH4 production. Therefore, there was an associative effect of tannin-rich PS and WDGS, which suppressed methanogenesis both directly and indirectly by modifying populations of ruminal methanogens.


Bone densitometry as an adjunct to GnRH agonist therapy.
To outline the optimal use of bone densitometry in women receiving gonadotropin releasing hormone (GnRH) agonist therapy. The use of bone densitometry during clinical trials of the use of GnRH agonists in endometriosis has resulted in a number of publications with varying conclusions about the effects of these agents on the skeleton. A review of this literature, taking into account known concepts of bone physiology and the varying quantities measured by different bone densitometry devices, was done to place these results into a common framework. Within this context, a proposal is made for when and how to use various bone densitometry methods in individual patients and how to interpret the results properly. Bone density in young women being treated for endometriosis is usually within the normal range, with even those 2 SD below normal not at significant risk of osteoporotic fracture. However, the clinical results show that most, if not all, women treated with GnRH agonists experience an increase in bone turnover due to estrogen deficiency during treatment, and this can result in a transient decrease in bone density. Even though the bone is regained after treatment is stopped, it may take years to recover fully. For those women who had low bone density to start, they may lose sufficient bone to increase their risk of osteoporotic fracture, and it is important to identify those individuals before treatment and to monitor them carefully. If the result of a peripheral bone density measurement (hand, forearm, heel) or central measurement (spine or hip) before treatment is higher than about 1 SD below a young normal mean, then even if a woman loses some bone, she will not have a significant increase in the risk of fracture. However, if a measurement is low (> 1 SD below the mean, putting her in the lower 15%), then it is advisable to have a spinal measurement by quantitative computed tomography or dual energy x-ray absorptiometry before and after therapy to determine what the bone changes are and to follow-up after the end of treatment. This is especially true if she decides to become pregnant, needs retreatment with a GnRH agonist or will be going through natural menopause in the near future; all can cause further bone loss. Different bone density methods give different information and must be interpreted properly when used in evaluating patients on GnRH agonist therapy. Any bone measurement may be used to assess skeletal status before treatment, but if a peripheral bone measurement is low, it should be followed with a spinal measurement by quantitative computed tomography or dual energy x-ray absorptiometry so that the effect of the treatment can be monitored. Bone loss is transient with GnRH treatment, but only spinal measurements have sufficient sensitivity to follow patients and evaluate the skeletal changes in a meaningful way.


Activatable Fluorescence Imaging and Targeted Drug Delivery via Extracellular Vesicle-Like Porous Coordination Polymer Nanoparticles.
Cancer imaging with minimal background signal and targeted intracellular drug delivery are of vital importance in clinical cancer diagnosis and therapy. Herein, we developed a biomimetic nanoprobe for activated fluorescence imaging and targeted drug delivery. pH-responsive porous coordination polymer nanoparticles (PCP NPs) were first synthesized by a codeposition method, anticancer drug doxorubicin (DOX) was then loaded into PCP NPs through physical and electrostatic adsorption (PCP-DOX), and finally the cell membranes extracted from Bel-7402 cancer cells were coated on the DOX-loaded PCP NPs (PCP-DOX-CM). The fluorescence of DOX was quenched due to the fluorescence resonance energy transfer between DOX and PCP NPs. Under acidic environment inside cancer cells, PCP NPs degraded, DOX was released from PCP-DOX-CM, and the fluorescence of DOX was activated, which was very specific for cancers with a high signal-to-noise ratio. Benefited from immune escaping and homologous targeting ability from cancer cell membranes, compared with PCP-CM and PCP-DOX, PCP-DOX-CM significantly enhanced the cellular endocytosis of DOX in Bel-7402 cancer cells and exhibited excellent cancer therapy effect in vitro. Together, our work provides a useful platform for an activated cancer imaging system and personalized cancer treatment.


Description and phylogenetic relationships of a new species of treefrog of the Dendropsophus leucophyllatus group (Anura: Hylidae) from the Amazon basin of Colombia and with an exceptional color pattern.
We describe Dendropsophus manonegra sp. nov. from the upper Amazon basin in the eastern foothills of the Andes in Colombia (1 degree 47' 42.2" N, 75 degrees 38' 48.7" W; 1040 m a.s.l.). Phylogenetic analysis of DNA sequences of 2582 aligned base pairs of the 12S and 16S rRNA genes recovered the new species as a member of the D. leucophyllatus species group and sister to D. bifurcus. Morphological traits, such as the presence of pectoral glands in males and females, support this hypothesis. The new species is readily distinguished from all other members of the species group by having bluish-black coloration on fingers, toes, webbing, axillary membranes, groin and hidden surfaces of arms and legs. Some aspects on the composition and systematics of the D. leucophyllatus group are discussed.


Sperm morphology and evidence for sperm competition among parrots.
Sperm competition is an important component of post-copulatory sexual selection that has shaped the evolution of sperm morphology. Previous studies have reported that sperm competition has a concurrently directional and stabilizing effect on sperm size. For example, bird species that show higher levels of extrapair paternity and larger testes (proxies for the intensity of sperm competition) have longer sperm and lower coefficients of variation in sperm length, both within and between males. For this reason, these sperm traits have been proposed as indexes to estimate the level of sperm competition in species for which other measures are not available. The relationship between sperm competition and sperm morphology has been explored mostly for bird species that breed in temperate zones, with the main focus on passerine birds. We measured sperm morphology in 62 parrot species that breed mainly in the tropics and related variation in sperm length to life-history traits potentially indicative of the level of sperm competition. We showed that sperm length negatively correlated with the within-male coefficient of variation in sperm length and positively with testes mass. We also showed that sperm is longer in sexually dichromatic and in gregarious species. Our results support the general validity of the hypothesis that sperm competition drives variation in sperm morphology. Our analyses suggest that post-copulatory sexual selection is also important in tropical species, with more intense sperm competition among sexually dichromatic species and among species that breed at higher densities.


Stability of vitamin B1 in Ultra Rice in the presence of encapsulated ferrous fumarate.
Food fortification with iron is effective in combating iron-deficiency anemia. As iron is reactive, it can destroy micronutrients, contribute to poor taste, and discolor the food. Encapsulation could be used to prevent the reaction of iron with food components. The stability of thiamin (vitamin B1) and its effects on organoleptic properties in Ultra Rice were investigated in the presence of encapsulated ferrous fumarate. The preparation of simulated rice grains did not affect the solubility of encapsulated ferrous fumarate in simulated gastric juice. After 20 weeks at 40 degrees C/ approximately 100% relative humidity, essentially all of the vitamin B1 and ferrous iron were retained. Antioxidants were effective in preventing the loss of thiamin and retarding oxidative rancidity catalyzed by ferrous iron. This study demonstrated the feasibility of incorporating both iron and vitamin B1 in a stable Ultra Rice formulation. Inclusion of other stable B vitamins in the formulation should be also feasible.


Influence of Abutment Design on Stress Distribution in Narrow Implants with Marginal Bone Loss: A Finite Element Analysis.
The aim of this study was to determine the effect of solid (one-piece) and two-piece abutments on the stress profile of narrow implants with marginal bone loss. Solid and two-piece abutments were connected to a conical internal octagon-connection implant (3.3 mm in diameter, 10 mm in length) and restored with a single crown. Three-dimensional finite element analysis was used to simulate the stress distribution in implant models with different levels of marginal bone resorption (0, 1, 2, and 3 mm). The effect of the design variables under increased bone resorption scenarios, including abutment screw length and diameter, was assessed. Static loading was applied to determine the mechanical response of the implant and cortical and trabecular bone. Marginal bone resorption levels dominated the mechanical response under static loading conditions. A marginal bone loss of 3 mm significantly increased stress values in the implant vicinity and abutment screw. Both abutment designs displayed similar stress distribution in the surrounding bone, but lower stress values were observed in the implant body with two-piece abutments. The abutment screw length was more effective in the resultant stress, as the longer screws reduced the stress in the implants. Marginal bone resorption magnitude is the crucial parameter in biomechanics to determine the mechanical behavior. As bone loss increases, resultant stress around implants under mastication forces may lead to implant failure, regardless of abutment type.


The epigenome as a molecular marker and target.
Tumor cell proliferation, de-differentiation, and progression depend on a complex combination of altered cell cycle regulation, excessive growth factor pathway activation, and decreased apoptosis. The understanding of these complex mechanisms should lead to the identification of potential molecular markers, targets, and molecular profiles that should eventually expand and improve therapeutic intervention. It now appears clear that methylation plays a central role in transformation, both in vitro and in vivo. However, the exact targets and mechanism(s) are not yet fully understood. This is partly due to the significant number of genes altered by changes in intracellular methyltransferase activity and the chemical agents used to modulate gene expression. The complex nature of methylation's role in regulating gene expression suggests that in addition to investigating individual genes, researchers should develop more comprehensive methods to examine gene expression patterns and their predictive value as this will likely be necessary in the future. If methylation plays a role in transformation, then it seems logical that genes regulating intracellular methylation status may be used as molecular markers to profile tumors by any new methods currently being developed. Perhaps more noteworthy is that DNMT genes may be found to be novel molecular targets for new factor-specific anticancer agents. This idea will be addressed.


Conserved determinants for membrane association of nonstructural protein 5A from hepatitis C virus and related viruses.
Nonstructural protein 5A (NS5A) is a membrane-associated essential component of the hepatitis C virus (HCV) replication complex. An N-terminal amphipathic alpha helix mediates in-plane membrane association of HCV NS5A and at the same time is likely involved in specific protein-protein interactions required for the assembly of a functional replication complex. The aim of this study was to identify the determinants for membrane association of NS5A from the related GB viruses and pestiviruses. Although primary amino acid sequences differed considerably, putative membrane anchor domains with amphipathic features were predicted in the N-terminal domains of NS5A proteins from these viruses. Confocal laser scanning microscopy, as well as membrane flotation analyses, demonstrated that NS5As from GB virus B (GBV-B), GBV-C, and bovine viral diarrhea virus, the prototype pestivirus, display membrane association characteristics very similar to those of HCV NS5A. The N-terminal 27 to 33 amino acid residues of these NS5A proteins were sufficient for membrane association. Circular dichroism analyses confirmed the capacity of these segments to fold into alpha helices upon association with lipid-like molecules. Despite structural conservation, only very limited exchanges with sequences from related viruses were tolerated in the context of functional HCV RNA replication, suggesting virus-specific interactions of these segments. In conclusion, membrane association of NS5A by an N-terminal amphipathic alpha helix is a feature shared by HCV and related members of the family Flaviviridae. This observation points to conserved roles of the N-terminal amphipathic alpha helices of NS5A in replication complex formation.


Sonographic characteristics of presumptively normal canine medial iliac and superficial inguinal lymph nodes.
The medial iliac and superficial inguinal lymph nodes are not routinely palpable in the dog, and ultrasound imaging provides an alternate noninvasive technique to assess these lymph nodes, as well as to guide needle aspiration. Herein we describe the ultrasound characteristics of the medial iliac and superficial inguinal lymph nodes in 50 healthy dogs, as well as frequency and ease of node detection. The relationship between the size of the lymph nodes and the following variables was assessed: age, gender, body weight, body condition score, body length, and thoracic height and width. Right and left medial iliac lymph nodes were detected in 50 (100%) dogs, right superficial inguinal lymph node(s) in 49 (98%) dogs, and left superficial inguinal lymph node(s) in 47 (96%) dogs. In > 90% of both sets of lymph nodes, the echogenicity was hypoechoic or isoechoic to surrounding tissues, with a corticomedullary or homogenous echotexture, smooth, clearly defined margins, and a fusiform shape. Increasing weight, distance from the sternal manubrium to the ischium, and thoracic height and width were associated with increased lymph node size (P-values < 0.05). Average lymph node sizes and range of sizes provide preliminary reference values for the medial iliac and superficial inguinal lymph nodes in normal dogs.


Soilborne Phytophthora and Pythium Diversity From Rhododendron in Propagation, Container, and Field Production Systems of the Pacific Northwest.
Rhododendron root rot is a severe disease that causes significant mortality in rhododendrons. Information is needed about the incidence and identity of soilborne Phytophthora and Pythium species causing root rot in Pacific Northwest nurseries in order to better understand the disease etiology and to optimize disease control strategies. The last survey focusing solely on soilborne oomycete pathogens in rhododendron production was conducted in 1974. Since then, advances in pathogen identification have occurred, new species may have been introduced, pathogen communities may have shifted, and little is known about Pythium species affecting this crop. Therefore, a survey of root-infecting Phytophthora and Pythium species was conducted at seven nurseries from 2013 to 2017 to (i) document the incidence of root rot damage at each nursery and stage of production, (ii) identify soilborne oomycetes infecting rhododendron, and (iii) determine whether there are differences in pathogen diversity among nurseries and production systems. Rhododendrons from propagation, container, and field systems were sampled and Phytophthora and Pythium species were isolated from the roots and collar region. Root rot was rarely evident in propagation systems, which were dominated by Pythium species. However, severe root rot was much more common in container and field systems where the genus Phytophthora was also more prevalent, suggesting that Phytophthora species are the primary cause of severe root rot and that most contamination by these pathogens comes in after the propagation stage. In total, 20 Pythium species and 11 Phytophthora species were identified. Pythium cryptoirregulare, Pythium aff. macrosporum, Phytophthora plurivora, and Phytophthora cinnamomi were the most frequently isolated species and the results showed that Phytophthora plurivora has become much more common than in the past. Phytophthora diversity was also greater in field systems than in propagation or container systems. Risks for Phytophthora contamination were commonly observed during the survey and included placement of potting media in direct contact with field soil, the presence of dead plants that could serve as continuous sources of inoculum, and the presence of excess water as a result of poor drainage, overirrigation, or malfunctioning irrigation equipment. In the past, research on disease development and root rot disease control in rhododendron focused almost exclusively on Phytophthora cinnamomi. More research is needed on both of these topics for the other root-infecting species identified in this survey.


Impaired memory is more closely associated with brain beta-amyloid than leukoaraiosis in hypertensive patients with cognitive symptoms.
Hypertension is the strongest modifiable risk factor for subcortical ischemic changes and is also a risk factor for Alzheimer's dementia. We used neuroimaging to investigate the pathological basis of early cognitive symptoms in patients with hypertension. In this cross-sectional cohort study 67 patients age >60 years with hypertension and Clinical Dementia Rating scale score of 0.5 without dementia, and without history of symptomatic stroke, underwent MRI for measurement of subcortical vascular changes and positron emission tomography (PET) scan with Pittsburgh Compound B (PiB-PET) to detect beta-amyloid deposition. These imaging measures were related to neuropsychological tests of memory, executive function and processing speed. Mean age was 75.0 (standard deviation, SD, 7.3). Mean neuropsychological Z scores were: episodic memory -0.63 (SD 1.23), executive function -0.40 (SD 1.10), processing speed -0.24 (SD 0.88); 22 of the 67 subjects met criteria for mild cognitive impairment (MCI) and the remaining 45 subjects had subjective cognitive concerns only. In multivariable models adjusting for age and years of education, each 0.1 unit increase in mean cortical PiB-PET binding was associated with 0.14 lower mean Z score for episodic memory (95% CI -0.28 to -0.01). This means that for every 0.1 unit increase in mean cortical PiB-PET, episodic memory was 0.14 standard deviations lower. White matter hyperintensity volume, silent brain infarcts and microbleeds were not associated with neuropsychological test scores. Episodic memory was prominently affected in hypertensive participants with MCI or subjective cognitive concerns, and was associated with PiB-PET binding. This suggests a prominent role for Alzheimer pathology in cognitive impairment even in hypertensive participants at elevated risk for vascular cognitive impairment.


The effect of transformation on the virulence of Streptococcus pneumoniae.
Although pneumococcus is one of the most frequently encountered opportunistic pathogen in the world, the mechanisms responsible for its infectiveness have not yet been fully understood. In this paper, we have attempted to characterize the effects of pneumococcal transformation on the pathogenesis of the organism. We constructed three transformation-deficient pneumococcal strains, which were designated as Nos. 1d, 2d, and 22d. The construction of these altered strains was achieved via the insertion of the inactivated gene, comE, to strains 1, 2 and 22. We then conducted a comparison between the virulence of the transformation-deficient strains and that of the wild-type strains, via an evaluation of the ability of each strain to adhere to endothelial cells, and also assessed psaA mRNA expression, and the survival of hosts after bacterial challenge. Compared to what was observed with the wild-type strains, our results indicated that the ability of all of the transformation-deficient strains to adhere to the ECV304 cells had been significantly reduced (p < 0.05), the expression of psaA mRNA was reduced significantly (p < 0.05) in strains 2d and 22d, and the median survival time of mice infected with strains 1d and 2d was increased significantly after intraperitoneal bacterial challenge (p < 0.05). The results of our study also clearly indicated that transformation exerts significant effects on the virulence characteristics of S. pneumoniae, although the degree to which this effect is noted appears to depend primarily on the genetic background of the bacteria.


The distribution of lymphoid and macrophage like cell subsets of sarcoid and Kveim granulomata: possible mechanism of negative PPD reaction in sarcoidosis.
Immunohistological observations of lymphoid and non-lymphoid cell subsets in biopsies of sarcoid skin granulomas have been compared with positive Kveim tests and the sites of PPD injection in sarcoid patients. Monoclonal antibodies have been used in indirect immunofluorescence often in combination with histochemical methods for the detailed characterization of the cells involved. The antibodies used included two new reagents, RFD-1 and RFD-2, which react with interdigitating cells and acid phosphatase positive macrophages, respectively. Sarcoid granulomas had a distinctive pattern of organization though there was a heterogeneity of macrophage like and T lymphoid cells. In the centre, predominantly HLA-DR+, acid phosphatase positive macrophages (RFD-2+) were seen and the lymphoid cells were almost exclusively T4+. At the periphery of the granulomas the HLA-DR+ dendritic cells were ACP negative and RFD-1+. Here T8+ cells were admixed with the T4+ population. The Kveim granuloma had fewer RFD-2+, macrophages and therefore the RFD-1+ cells were more evenly distributed, but the other cells showed a similar distribution to the established lesions. The PPD injection sites contained fewer T cells than the normal control infiltrates in PPD positive healthy individuals. The T4+/T8+ ratios were about 3:2. The most likely explanation for the PPD anergy in sarcoidosis is the sluggish traffic of T4+ cells which could be due to the sequestration of T4+ cells in sites of ongoing inflammation.