bluuebunny/crossref_metadata_2025 huggingface dataset:
The Coronavirus Endoribonuclease Nsp15 Interacts with Retinoblastoma Tumor Suppressor Protein
Authors: Bhardwaj Kanchan , Liu Pinghua , Leibowitz Julian L. , Kao C. Cheng
Published: February 2012

Coronaviruses encode an endoribonuclease, Nsp15, which has a poorly defined role in infection. Sequence analysis revealed a retinoblastoma protein-binding motif (LXCXE/D) in the majority of the Nsp15 of the severe acute respiratory syndrome coronavirus (SARS-CoV) and its orthologs in the alpha and beta coronaviruses. The endoribonuclease activity of the SARS-CoV Nsp15 (sNsp15) was stimulated by retinoblastoma protein (pRb)
in vitro
, and the two proteins can be coimmunoprecipitated from cellular extracts. Mutations in the pRb-binding motif rendered sNsp15 to be differentially modified by ubiquitin in cells, and cytotoxicity was observed upon its expression. Expression of the sNsp15 in cells resulted in an increased abundance of pRb in the cytoplasm, decreased overall levels of pRb, an increased proportion of cells in the S phase of the cell cycle, and an enhanced expression from a promoter normally repressed by pRb. The endoribonuclease activity of the mouse hepatitis virus (MHV) A59 Nsp15 was also increased by pRb
in vitro
, and an MHV with mutations in the LXCXE/D-motif, named vLC, exhibited a smaller plaque diameter and reduced the virus titer by ∼1 log. Overexpression of pRb delayed the viral protein production by wild-type MHV but not by vLC. This study reveals that pRb and its interaction with Nsp15 can affect coronavirus infection and adds coronaviruses to a small but growing family of RNA viruses that encode a protein to interact with pRb.
Interleukin-27 Inhibits Vaccine-Enhanced Pulmonary Disease following Respiratory Syncytial Virus Infection by Regulating Cellular Memory Responses
Authors: Zeng Ruihong , Zhang Huixian , Hai Yan , Cui Yuxiu , Wei Lin , Li Na , Liu Jianxun , Li Caixia , Liu Ying
Published: February 2012

Respiratory syncytial virus (RSV) is the most important cause of lower respiratory tract disease in young children. In the 1960s, infants vaccinated with formalin-inactivated RSV developed a more severe disease characterized by excessive inflammatory immunopathology in lungs upon natural RSV infection. The fear of causing the vaccine-enhanced disease (VED) is an important obstacle for development of safe and effective RSV vaccines. The recombinant vaccine candidate G1F/M2 immunization also led to VED. It has been proved that cellular memory induced by RSV vaccines contributed to VED. Interleukin-27 (IL-27) and IL-23 regulate Th1, Th17, and/or Th2 cellular immune responses. In this study, mice coimmunized with pcDNA3–IL-27 and G1F/M2 were fully protected and, importantly, did not develop vaccine-enhanced inflammatory responses and immunopathology in lungs after RSV challenge, which was correlated with moderate Th1-, suppressed Th2-, and Th17-like memory responses activated by RSV. In contrast, G1F/M2- or pcDNA3–IL-23+G1F/M2-immunized mice, in which robust Th2- and Th17-like memory responses were induced, developed enhanced pulmonary inflammation and severe immunopathology. Mice coimmunized with G1F/M2 and the two cytokine plasmids exhibited mild inflammatory responses as well as remarkable Th1-, suppressed Th2-, and Th17-like memory responses. These results suggested that Th1-, Th2-, and Th17-like memory responses and, in particular, excessive Th2- and Th17-like memory responses were closely associated with VED; IL-27 may inhibit VED following respiratory syncytial virus infection by regulating cellular memory responses.
Enantioselective [2+2+2] cycloisomerisation of alkynes in the synthesis of helicenes: The search for effective chiral ligands
Authors: Stará Irena G. , Andronova Angelina , Kollárovič Adrian , Vyskočil Štěpán , Jugé Sylvain , Lloyd-Jones Guy C. , Guiry Patrick J. , Starý Ivo
Published: February 2012

The enantioselective [2+2+2] cycloisomerisation of the aromatic triynes under nickel(0) catalysis to afford nonracemic [6]- and [7]helicene derivatives has been systematically studied. A collection of mono- and bidentate phosphines, phosphites, phosphinites and phosphinous amides possessing stereogenic units such as chiral centre, axis or plane (or their combinations) has been tested and axially chiral binaphthyl-derived monodentate MOP-type phosphine ligands were the optimal class of ligands. Nickel complexes of these ligands afforded nonracemic tetrahydro[6]helicene in up to 64% ee in a model reaction.
Multimodal Stepped Care Approach with Acupuncture and Ppar-α Agonist Palmitoylethanolamide in the Treatment of a Patient with Multiple Sclerosis and Central Neuropathic Pain
Authors: Kopsky David J , Hesselink Jan M Keppel
Published: February 2012

Central neuropathic pain is a common debilitating symptom in patients with multiple sclerosis. Side effects of analgesics often limit reaching therapeutic dosages. In this case report, a 61-year-old woman with chronic central neuropathic pain due to multiple sclerosis is described. Acupuncture could only partly and temporarily reduce the pain. However, after adding the natural compound palmitoylethanolamide, a glial modulator and peroxisome proliferator-activated receptor-α agonist, pain reduction was more pronounced and the interval between acupuncture sessions could be increased. A multimodal stepped care approach is demonstrated, with acupuncture and palmitoylethanolamide both influencing non-neuronal cells, such as activated glial cells, which are key factors in the development and maintenance of neuropathic pain.
Does somatic illness explain the association between common mental disorder and elevated mortality? Findings from extended follow-up of study members in the UK Health and Lifestyle Survey: Table 1
Authors: Batty G David , Hamer Mark , Der Geoff
Published: February 2012

Background

Common mental disorder (psychological distress) is associated with an increased risk of disease-specific mortality. Given that physical illness is related to both exposure and outcome, it may explain this relation through confounding or mediation.

Methods

The authors used a 20-year follow-up of the UK Health and Lifestyle Survey (6127 men and women) in which common mental disorder was ascertained at baseline using the 30 item General Health Questionnaire and physical illness using a range of enquiries. Study members were an average of 45.2 years (SD 17.0) at study induction.

Results

In age-adjusted analyses, a 1 SD increase in common mental disorder score was associated with an elevated risk of mortality outcomes coronary heart disease (CHD) in men (HR 1.11, 95% CI 0.96 to 1.27), CHD in women (1.33, 1.16 to 1.51); plus, in men and women combined, stroke (1.13, 0.96 to 1.30), respiratory disease (1.31, 1.15 to 1.48), lung cancer (1.11, 0.92 to 1.33), ‘other’ cancer (1.14, 1.03 to 1.26) and all causes (1.18, 1.12 to 1.23). Controlling for prior physical illness effectively eliminated the common mental disorder–mortality relation in all analyses with the exception of CHD in women.

Conclusion

That physical illness largely explains the link between common mental disorder and mortality in the present cohort is compatible with either a confounding or mediation explanation.
Combined insulin treatment and intense exercise training improved basal cardiac function and Ca2+-cycling proteins expression in type 1 diabetic rats
Authors: Le Douairon Lahaye Solène , Gratas-Delamarche Arlette , Malardé Ludivine , Zguira Sami , Vincent Sophie , Lemoine Morel Sophie , Carré François , Rannou Bekono Françoise
Published: February 2012

This study investigated the effects of 8 weeks of intense exercise training combined with insulin treatment on the Ca2+-cycling protein complex expression and their functional consequences on cardiac function in type 1 diabetic rat hearts. Diabetic Wistar rats were randomly assigned into the following groups: received no treatment, insulin-treated diabetic, trained diabetic, and trained insulin-treated diabetic. A control group was also included. Insulin treatment and (or) treadmill intense exercise training were conducted over 8 weeks. Basal cardiac function was evaluated by Langendorff technique. Cardiac expression of the main Ca2+-cycling proteins (RyR2, FKBP 12.6, SERCA2, PLB, NCX1) was assessed by Western blot. Diabetes altered basal cardiac function (±dP/dt) and decrease the expression of the main Ca2+-cycling proteins expression: RyR2, SERCA2, and NCX1 (p < 0.05). Whereas combined treatment was not able to normalize –dP/dt, it succeeded to normalize +dP/dt of diabetic rats (p < 0.05). Moreover, both insulin and intense exercise training, applied solely, increased the expression of the Ca2+-cycling proteins: RyR2, SERCA2, PLB. and NCX1 (p < 0.05). But this effect was higher when the 2 treatments were combined. These data are the first to show that combined insulin treatment and intense exercise training during diabetes synergistically act on the expression of the main Ca2+-cycling proteins, providing insights into mechanisms by which the dual treatment during diabetes improves cardiac function.
A socioecological framework to understand weight-related issues in Aboriginal children in Canada
Authors: Willows Noreen D. , Hanley Anthony J.G. , Delormier Treena
Published: February 2012

Obesity prevention efforts in Aboriginal (First Nations, Métis, or Inuit) communities in Canada should focus predominantly on children given their demographic significance and the accelerated time course of occurrence of type 2 diabetes mellitus in the Aboriginal population. A socioecological model to address childhood obesity in Aboriginal populations would focus on the numerous environments at different times in childhood that influence weight status, including prenatal, sociocultural, family, and community environments. Importantly, for Aboriginal children, obesity interventions need to also be situated within the context of a history of colonization and inequities in the social determinants of health. This review therefore advocates for the inclusion of a historical perspective and a life-course approach to obesity prevention in Aboriginal children in addition to developing interventions around the socioecological framework. We emphasize that childhood obesity prevention efforts should focus on promoting maternal health behaviours before and during pregnancy, and on breastfeeding and good infant and child nutrition in the postpartum and early childhood development periods. Ameliorating food insecurity by focusing on improving the sociodemographic risk factors for it, such as increasing income and educational attainment, are essential. More research is required to understand and measure obesogenic Aboriginal environments, to examine how altering specific environments modifies the foods that children eat and the activities that they do, and to examine how restoring and rebuilding cultural continuity in Aboriginal communities modifies the many determinants of obesity. This research needs to be done with the full participation of Aboriginal communities as partners in the research.
Age-related changes in oxidative capacity differ between locomotory muscles and are associated with physical activity behavior
Authors: Larsen Ryan G. , Callahan Damien M. , Foulis Stephen A. , Kent-Braun Jane A.
Published: February 2012

There is discrepancy in the literature regarding the degree to which old age affects muscle bioenergetics. These discrepancies are likely influenced by several factors, including variations in physical activity (PA) and differences in the muscle group investigated. To test the hypothesis that age may affect muscles differently, we quantified oxidative capacity of tibialis anterior (TA) and vastus lateralis (VL) muscles in healthy, relatively sedentary younger (8 YW, 8 YM; 21–35 years) and older (8 OW, 8 OM; 65–80 years) adults. To investigate the effect of physical activity on muscle oxidative capacity in older adults, we compared older sedentary women to older women with mild-to-moderate mobility impairment and lower physical activity (OIW, n = 7), and older sedentary men with older active male runners (OAM, n = 6). Oxidative capacity was measured in vivo as the rate constant, kPCr, of postcontraction phosphocreatine recovery, obtained by31P magnetic resonance spectroscopy following maximal isometric contractions. While kPCrwas higher in TA of older than activity-matched younger adults (28%; p = 0.03), older adults had lower kPCrin VL (23%; p = 0.04). In OIW compared with OW, kPCrwas lower in VL (∼45%; p = 0.01), but not different in TA. In contrast, OAM had higher kPCrthan OM (p = 0.03) in both TA (41%) and VL (54%). In older adults, moderate-to-vigorous PA was positively associated with kPCrin VL (r = 0.65, p < 0.001) and TA (r = 0.41, p = 0.03). Collectively, these results indicate that age-related changes in oxidative capacity vary markedly between locomotory muscles, and that altered PA behavior may play a role in these changes.
Role of pyruvate dehydrogenase kinase 4 in regulating PDH activation during acute muscle contraction
Authors: Herbst Eric A.F. , Dunford Emily C.E. , Harris Robert A. , Vandenboom Rene , LeBlanc Paul J. , Roy Brian D. , Jeoung Nam Ho , Peters Sandra J.
Published: February 2012

The oxidation of carbohydrates in mammals is regulated by the pyruvate dehydrogenase (PDH) complex, which is covalently regulated by four PDH kinases (PDK1–4) and two PDH phosphatases (PDP1–2) unique to the PDH complex. To investigate the role that PDK4 plays in regulating PDH activation (PDHa) during muscle contraction, mouse extensor digitorum muscle was removed from wild type (WT) and PDK4-knockout (PDK4-KO) mice after a 24 h fast and stimulated for 3 min either at 10 Hz (low-intensity contraction), 40 Hz (moderate-intensity contraction), or allowed to rest. Force was recorded and muscle PDHa activity and metabolite concentrations were measured. PDHa activity was ∼2.5-fold higher at rest in PDK4-KO mice than WT mice (P = 0.009) and ∼2-fold higher in PDK4-KO mice at both 10 Hz (P < 0.001) and 40 Hz (P < 0.001). Force relative to muscle weight was similar at 10 Hz, but was 5.8 ± 0.7 mN·g–1 in PDK4-KO mice and 3.5 ± 0.7 mN·g–1 in WT mice at 40 Hz (P < 0.001), with a similar rate of fatigue in both genotypes. From these results it was concluded that PDK4 plays a role in reducing PDHa activity during low to moderate-intensity muscle stimulation, and that absence of PDK4 and the subsequent changes in carbohydrate utilization may alter force production.
An evaluation of the physical activity and health status of British Columbian Aboriginal populations
Authors: Foulds Heather J.A. , Bredin Shannon S.D. , Warburton Darren E.R.
Published: February 2012

Aboriginal people have experienced elevated rates of obesity, diabetes, cardiovascular disease, and other chronic conditions that are disproportionate to the general Canadian population. We sampled British Columbian Aboriginal populations to determine the current health status of this population. A total of 882 Aboriginal adults ≥16 years of age from 25 locations around the province were sampled from 2007–2011. Health measurements evaluated included body mass index, waist circumference, physical activity, smoking behaviours, and resting blood pressure as well as histories of diabetes, cardiovascular disease, and hypertension. Nonfasting measures of blood sugar, total cholesterol, and high-density lipoprotein cholesterol were also recorded. We used logistical regression to quantify variations in diabetes and cardiovascular disease risk factors with age, gender, geographic location, on- and off-reserve and urban–rural areas of residence, smoking, and physical activity behaviour. The prevalence of diabetes, hypertension, and high total cholesterol were found to be greater among males, while females experienced greater rates of abdominal obesity and physical inactivity. Rates of chronic conditions were similar across on- and off-reserve, urban–rural areas, and geographic region residences, though rural, on-reserve, and northern residents experienced greater risk of poor health status. Larger proportions of on-reserve, rural, and interior individuals were found to be more physically inactive. Aboriginal populations still experience substantially poorer health status in comparison with the general population. Initiatives to improve the health of this population are urgently needed addressing all sectors of the Aboriginal population, all geographic regions, and all areas of residence, with special emphasis on rural, on-reserve, and northern populations.
Glucose tolerance is affected by visceral adiposity and sex, but not birth weight, in Yucatan miniature pigs
Authors: McKnight Leslie L. , Myrie Semone B. , MacKay Dylan S. , Brunton Janet A. , Bertolo Robert F.
Published: February 2012

Epidemiological studies have linked small birth weight and lack of breastfeeding to type 2 diabetes mellitus. This study aimed to determine if (i) small birth weight promotes and (ii) suckling prevents the development of adiposity and diabetes biomarkers in a Yucatan miniature pig model. At 3 days of age, the intrauterine growth-restricted (IUGR) piglet (n = 6) was paired with a normal weight (NW), same-sex littermate (n = 6) and fed milk replacer for 4 weeks. A sow-fed normal weight littermate (n = 6) was also compared with the NW littermate to assess the effects of suckling. All pigs were fed a standard diet ad libitum for 5 h·day–1from week 4. At 9.5 months, pigs underwent intravenous glucose tolerance (IVGTT) and insulin sensitivity tests (IST). At 10 months, tissues were harvested for fat analysis and pancreas histology. IUGR pigs demonstrated compensatory growth before sexual maturity and had greater subcutaneous fat depth; birth weight also negatively correlated with visceral fat content. Visceral and subcutaneous adiposity were greater in females than males. IVGTT and IST outcomes were not different due to birth weight or suckling. However, visceral adiposity was associated with several glucose tolerance outcomes and females were more glucose intolerant due to their greater adiposity. Pancreas insulin content or histology outcomes were not different. This model did not develop markers of type 2 diabetes mellitus because of small birth weight or formula feeding. However, visceral adiposity and sex were associated with glucose intolerance, which is consistent with data in humans.
Acute l-arginine supplementation increases muscle blood volume but not strength performance
Authors: Álvares Thiago Silveira , Conte Carlos Adam , Paschoalin Vânia Margaret Flosi , Silva Joab Trajano , Meirelles Cláudia de Mello , Bhambhani Yagesh N. , Gomes Paulo Sergio Chagas
Published: February 2012

l-Arginine (L-arg) is an amino acid precursor to nitric oxide (NO). Dietary supplements containing L-arg have been marketed with the purpose of increasing vasodilation, thereby elevating blood flow to the exercising muscle and enhancing the metabolic response to exercise. Our goal was to identify the acute effect of L-arg supplementation on biceps strength performance, indicators of NO production (nitrite and nitrate – NOx), and muscle blood volume (Mbv) and oxygenation (Mox) during recovery from 3 sets of resistance exercise. Fifteen males participated in a randomized, double-blind, placebo-controlled study. After withdrawing resting blood samples, the subjects were supplemented with 6 g of L-arg (ARG) or placebo (PLA). Monitoring of Mbv and Mox with near-infrared spectroscopy began 30 min after supplementation and lasted for 60 min. The exercise protocol (3 sets of 10 maximal voluntary contractions of isokinetic concentric elbow extension at 60°·s–1, 2-min rest between sets) was initiated 80 min after supplementation. Blood samples were drawn at 30, 60, 90, and 120 min after supplementation. Repeated measures ANOVA showed that Mbv significantly (p ≤ 0.05) increased in ARG compared with the PLA during the recovery period of each set of resistance exercise. NOx, Mox, peak torque, total work, and set total work were not significantly different between groups. We found that acute L-arg supplementation increases Mbv during recovery from sets of resistance exercise with no increase in strength performance. It is still premature to recommend nutritional supplements containing L-arg as an ergogenic aid to increase muscle strength during resistance training in healthy subjects.
Benefits of pedometer-measured habitual physical activity in healthy women
Authors: Graff Scheila Karen , Alves Bruna Cherubini , Toscani Mariana Kirjner , Spritzer Poli Mara
Published: February 2012

This cross-sectional study aimed at (i) characterizing pedometer-determined physical activity and (ii) examining its associations with dietary intake and anthropometric and metabolic profile in healthy women. Anthropometric and metabolic profile was evaluated in 68 healthy women of reproductive age. Habitual physical activity was assessed using a pedometer for 6 consecutive days, including weekends. Participants were stratified into active and inactive according to the mean steps·day–1(≥6000 and <6000, respectively). Food consumption was evaluated by 24-h recall in a subsample of 35 participants. Thirty-eight women were defined as active and had significantly lower body mass index (BMI), fat percentage, waist circumference, sum of skinfold thickness, insulin, and HOMA than the sedentary group. Mean BMI was 27 kg·m–2(overweight) in active participants and 31 kg·m–2(class I obesity) in inactive participants. Active women consumed more carbohydrates (55.5% ± 9.4% vs. 46.3% ± 7.6%) and calories (2138 ± 679 vs. 1664 ± 558 kcal), and less protein (15.4% ± 4.2% vs. 19.9% ± 5.8%) and lipids (29.0% ± 7.2% vs. 33.8% ± 6.2%) than inactive individuals (p < 0.05). Fiber, cholesterol, and fatty acid intake was similar in both groups. The number of steps was lower on Sunday than on weekdays for the overall group. Using a pedometer for 3 days was sufficient to determine habitual physical activity (sensitivity: 94%; specificity 91% vs. 6 days of pedometer use). In the present study, nonstructured physical activity was associated with more adequate dietary consumption and contributed toward a healthier anthropometric and metabolic profile in young women, despite the high prevalence of overweight.
Validation of a new mixing chamber system for breath-by-breath indirect calorimetry
Authors: Kim Do-yeon , Robergs Robert Andrew
Published: February 2012

Limited validation research exists for applications of breath-by-breath systems of expired gas analysis indirect calorimetry (EGAIC) during exercise. We developed improved hardware and software for breath-by-breath indirect calorimetry (NEW) and validated this system as well as a commercial system (COM) against 2 methods: (i) mechanical ventilation with known calibration gas, and (ii) human subjects testing for 5 min each at rest and cycle ergometer exercise at 100 and 175 W. Mechanical calibration consisted of medical grade and certified calibration gas ((4.95% CO2, 12.01% O2, balance N2), room air (20.95% O2, 0.03% CO2, balance N2), and 100% nitrogen), and an air flow turbine calibrated with a 3-L calibration syringe. Ventilation was mimicked manually using complete 3-L calibration syringe manouvres at a rate of 10·min–1from a Douglas bag reservoir of calibration gas. The testing of human subjects was completed in a counterbalanced sequence based on 5 repeated tests of all conditions for a single subject. Rest periods of 5 and 10 min followed the 100 and 175 W conditions, respectively. COM and NEW had similar accuracy when tested with known ventilation and gas fractions. However, during human subjects testing COM significantly under-measured carbon dioxide gas fractions, over-measured oxygen gas fractions and minute ventilation, and resulted in errors to each of oxygen uptake, carbon dioxide output, and respiratory exchange ratio. These discrepant findings reveal that controlled ventilation and gas fractions are insufficient to validate breath-by-breath, and perhaps even time-averaged, systems of EGAIC. The errors of the COM system reveal the need for concern over the validity of commercial systems of EGAIC.
Impaired superficial femoral artery vasodilation and leg blood flow in young obese women following an oral glucose tolerance test
Authors: Olver T.D. , Hazell T.J. , Hamilton C.D. , Shoemaker J.K. , Lemon P.W.R.
Published: February 2012

This study was designed to test the hypothesis that glucose ingestion following an overnight fast increases leg vascular conductance (LVCd) and superficial femoral artery (SFA) vasodilation in lean but not obese young women. Obese (23.5 ± 4.0 years, 84.7 ± 14.7 kg, 37.2% ± 6.4% fat; mean ± SD, n = 8) and lean (23.8 ± 2.4 years, 60.6 ± 4.0 kg, 22.3% ± 2.8% fat; n = 8) women arrived in the laboratory at 0830 h after a 12-h overnight fast for body composition (densitometry) assessment. Then, capillary blood glucose (BGlu), plasma insulin, heart rate, cardiac output, mean arterial pressure, leg blood flow (Doppler ultrasound), and LVCd were measured (after 15 min in the supine position), and at 30-min intervals for 2 h following glucose ingestion (75 g glucose load, 12.5% solution). Fasting BGlu concentration was not different between groups (obese = 5.1 ± 0.47 vs. lean = 4.9 ± 0.37 mmol·L–1, p = 0.71) but 60, 90, and 120 min postingestion BGlu was elevated (p ≤ 0.03) in the obese women. Insulin differences were not significant. Fasting LVCd was not different between groups (lean = 0.72 ± 0.49 vs. obese = 0.70 ± 0.19 mL·min–1·mm Hg–1; p = 0.48); however, LVCd, as well as Δ in SFA diameter were significantly elevated (p ≤ 0.04) in the lean compared with the obese group at 60, 90, and 120 min postglucose ingestion (LVCd, peak lean = 1.4 ± 0.5 vs. peak obese = 0.8 ± 0.1 mL·min–1·mm Hg–1; Δ in SFA, peak lean = 0.51 ± 0.30 vs. peak obese = 0.09 ± 0.45 mm). The reduced LVCd following glucose ingestion could contribute to impaired glucose tolerance. Further, the lack of SFA dilation may be evidence of impaired vascular insulin responsiveness in these obese young women.
Molecular assembly and dynamics of fluorescent protein-tagged single KCa1.1 channel in expression system and vascular smooth muscle cells
Authors: Yamamura Hisao , Ikeda Chikako , Suzuki Yoshiaki , Ohya Susumu , Imaizumi Yuji
Published: February 2012

The large-conductance Ca2+-activated K+ (KCa1.1, BK) channel has pivotal roles in the regulation of vascular tone. To clarify the molecular dynamics of BK channels and their functionally coupled protein on the membrane surface, we examined single-molecule imaging of fluorescent-labeled BK subunits in the plasma membrane using total internal reflection fluorescence (TIRF) microscopy. The dynamic mobility of yellow fluorescent protein (YFP)-tagged BKα subunit (BKα-YFP) expressed in human embryo kidney 293 (HEK) cells was detected in TIRF regions at the level of individual channels and their clusters on the plasma membrane with a diffusion coefficient of 6.7 × 103 nm2/s. When BKα-YFP was coexpressed with cyan fluorescent protein (CFP)-tagged BKβ1 subunit (BKβ1-CFP) in HEK cells, the mobility was reduced by ∼50%. Fluorescent image analyses suggest that green fluorescent protein (GFP)-tagged BKα subunit (BKα-GFP) expressed in vascular smooth muscle cells (VSMCs), at low density, preferentially formed a heterotetrameric molecular assembly with native BKα subunits, rather than homotetrameric BKα-GFP. Movement of BKα-YFP in VSMCs (0.29 × 103 nm2/s) was far more restricted than BKα-YFP/BKβ1-CFP in HEK cells (2.5 × 103 nm2/s). Actin disruption by pretreatment with cytochalasin D in VSMCs appeared to increase the mobile behavior of BKα-YFP, which was then significantly reduced by addition of jasplakinolide. Most BKα-YFP colocalized with caveolin 1 (Cav1)-CFP in VSMCs, but unexpectedly not frequently in HEK cells. Fluorescence resonance energy transfer analyses showed the direct interaction between BKα-YFP and Cav1-CFP, particularly in VSMCs. These results, obtained by single molecule imaging in living cells, indicate that the dynamics of BKα molecules on the membrane surface are strongly restricted or regulated by its auxiliary β-subunit, cytoskeleton, and direct interaction with Cav1 in VSMCs.
Abl regulates smooth muscle cell proliferation by modulating actin dynamics and ERK1/2 activation
Authors: Jia Li , Wang Ruping , Tang Dale D.
Published: February 2012

Abl is a nonreceptor tyrosine kinase that has a role in regulating migration and adhesion of nonmuscle cells as well as smooth muscle contraction. The role of Abl in smooth muscle cell proliferation has not been investigated. In this study, treatment with endothelin-1 (ET-1) and platelet-derived growth factor (PDGF) increased Abl phosphorylation at Tyr412 (an indication of Abl activation) in vascular smooth muscle cells. To assess the role of Abl in smooth muscle cell proliferation, we generated stable Abl knockdown cells by using lentivirus-mediated RNA interference. ET-1- and PDGF-induced cell proliferation was attenuated in Abl knockdown cells compared with cells expressing control shRNA and uninfected cells. Abl silencing also arrested cell cycle progression from G0/G1 to S phase. Furthermore, activation of smooth muscle cells with ET-1 and PDGF induced phosphorylation of ERK1/2 and Akt. Abl knockdown attenuated ERK1/2 phosphorylation in smooth muscle cells stimulated with ET-1 and PDGF. However, Akt phosphorylation upon stimulation with ET-1 and PDGF was not reduced. Because Abl is known to regulate actin polymerization in smooth muscle, we also evaluated the effects of inhibition of actin polymerization on phosphorylation of ERK1/2. Pretreatment with the actin polymerization inhibitor latrunculin-A also blocked ERK1/2 phosphorylation during activation with ET-1 and PDGF. The results suggest that Abl may regulate smooth muscle cell proliferation by modulating actin dynamics and ERK1/2 phosphorylation during mitogenic activation.
Detailed examination of Mg2+ and pH sensitivity of human TRPM7 channels
Authors: Chokshi Rikki , Matsushita Masayuki , Kozak J. Ashot
Published: February 2012

TRPM7 channel kinase is a protein highly expressed in cells of hematopoietic lineage, such as lymphocytes. Studies performed in native and heterologous expression systems have shown that TRPM7 forms nonselective cation channels functional in the plasma membrane and activated on depletion of cellular Mg2+. In addition to internal Mg2+, cytosolic pH and the phospholipid phosphatidylinositol-( 4 , 5 )-bisphosphate [PI( 4 , 5 )P2] are potent physiological regulators of this channel: protons inhibit, while PI( 4 , 5 )P2 is required for TRPM7 channel activity. These channels are also inhibited from inside by other metal cations and polyamines. While the regulation of TRPM7 channels by internal metal ions, acidic pH, and PI( 4 , 5 )P2 is voltage independent, extracellular metal cations and polyamines block voltage dependently at micromolar concentrations and appear to occupy a distinct blocking site. In the present study we investigated intracellular Mg2+ and pH dependence of native TRPM7 currents using whole cell patch-clamp electrophysiology in human Jurkat T lymphocytes and HEK293 cells. Our main findings are 1) Mg2+ inhibition involves not one but two separate sites of high (∼10 μM) and low (∼165 μM) affinity; and 2) while sharing certain characteristics with Mg2+ inhibition, protons most likely inhibit through one inhibitory site, corresponding to the low-affinity Mg2+ site, with an estimated IC50 of pH 6.3. Additionally, we present data on amplitude distribution of preactivated TRPM7 currents in Jurkat T lymphocytes in the absence of prior Mg2+ or proton depletion.
Mouse SGLT3a generates proton-activated currents but does not transport sugar
Authors: Barcelona Stephanie , Menegaz Danusa , Díez-Sampedro Ana
Published: February 2012

Sodium-glucose cotransporters (SGLTs) are secondary active transporters belonging to the SLC5 gene family. SGLT1, a well-characterized member of this family, electrogenically transports glucose and galactose. Human SGLT3 (hSGLT3), despite sharing a high amino acid identity with human SGLT1 (hSGLT1), does not transport sugar, although functions as a sugar sensor. In contrast to humans, two different genes in mice and rats code for two different SGLT3 proteins, SGLT3a and SGLT3b. We previously cloned and characterized mouse SGLT3b (mSGLT3b) and showed that, while it does transport sugar like SGLT1, it likely functions as a physiological sugar sensor like hSGLT3. In this study, we cloned mouse SGLT3a (mSGLT3a) and characterized it by expressing it in Xenopus laevis oocytes and performing electrophysiology and sugar transport assays. mSGLT3a did not transport sugar, and sugars did not induce currents at pH 7.4, though acidic pH induced inward currents that increased in the presence of sugar. Moreover, mutation of residue 457 from glutamate to glutamine resulted in a Na+-dependent transport of sugar that was inhibited by phlorizin. To corroborate our results in oocytes, we expressed and characterized mSGLT3a in mammalian cells and confirmed our findings. In addition, we cloned, expressed, and characterized rat SGLT3a in oocytes and found characteristics similar to mSGLT3a. In summary, acidic pH induces currents in mSGLT3a, and sugar-induced currents are increased at acidic pH, but wild-type SGLT3a does not transport sugar.
Suppression of the cough reflex by inhibition of ERK1/2 activation in the caudal nucleus tractus solitarii of the rabbit
Authors: Mutolo Donatella , Bongianni Fulvia , Cinelli Elenia , Giovannini Maria Grazia , Pantaleo Tito
Published: February 2012

The caudal nucleus tractus solitarii (cNTS), the predominant site of termination of cough-related afferents, has been shown to be a site of action of some centrally acting antitussive agents. A role of ERK1/2 has been suggested in acute central processing of nociceptive inputs. Because pain and cough share similar features, we investigated whether ERK1/2 activation could also be involved in the central transduction of tussive inputs. For this purpose, we undertook the present research on pentobarbital sodium-anesthetized, spontaneously breathing rabbits by using microinjections (30–50 nl) of an inhibitor of ERK1/2 activation (U0126) into the cNTS. Bilateral microinjections of 25 mM U0126 caused rapid and reversible reductions in the cough responses induced by both mechanical and chemical (citric acid) stimulation of the tracheobronchial tree. In particular, the cough number and peak abdominal activity decreased. Bilateral microinjections of 50 mM U0126 completely suppressed the cough reflex without affecting the Breuer-Hering inflation reflex, the pulmonary chemoreflex, and the sneeze reflex. These U0126-induced effects were, to a large extent, reversible. Bilateral microinjections of 50 mM U0124, the inactive analog of U0126, at the same cNTS sites had no effect. This is the first study that provides evidence that ERK1/2 activation within the cNTS is required for the mediation of cough reflex responses in the anesthetized rabbit. These results suggest a role for ERK1/2 in the observed effects via nontranscriptional mechanisms, given the short time involved. They also may provide hints for the development of novel antitussive strategies.
Computer simulations of osmotic ultrafiltration and small-solute transport in peritoneal dialysis: a spatially distributed approach
Authors: Stachowska-Pietka Joanna , Waniewski Jacek , Flessner Michael F. , Lindholm Bengt
Published: February 2012

The aim of this study was to simulate clinically observed intraperitoneal kinetics of dialysis fluid volume and solute concentrations during peritoneal dialysis. We were also interested in analyzing relationships between processes in the peritoneal cavity and processes occurring in the peritoneal tissue and microcirculation. A spatially distributed model was formulated for the combined description of volume and solute mass balances in the peritoneal cavity and flows across the interstitium and the capillary wall. Tissue local parameters were assumed dependent on the interstitial hydration and vasodilatation induced by glucose. The model was fitted to the average volume and solute concentration profiles from dwell studies in 40 clinically stable patients on chronic ambulatory peritoneal dialysis using a 3.86% glucose dialysis solution. The model was able to describe the clinical data with high accuracy. An increase in the local interstitial pressure and tissue hydration within the distance of 2.5 mm from the peritoneal surface of the tissue was observed. The penetration of glucose into the tissue and removal of urea, creatinine, and sodium from the tissue were restricted to a layer located within 2 mm from the peritoneal surface. The initial decline of sodium concentration (sodium dip) was observed not only in intraperitoneal fluid but also in the tissue. The distributed model can provide a precise description of the relationship between changes in the peritoneal tissue and intraperitoneal dialysate volume and solute concentration kinetics. Computer simulations suggest that only a thin layer of the tissue within 2–3 mm from the peritoneal surface participates in the exchange of fluid and small solutes between the intraperitoneal dialysate and blood.
Inducible kidney-specific Sgk1 knockout mice show a salt-losing phenotype
Authors: Faresse Nourdine , Lagnaz Dagmara , Debonneville Anne , Ismailji Adil , Maillard Marc , Fejes-Toth Geza , Náray-Fejes-Tóth Aniko , Staub Olivier
Published: February 2012

The expression of the serum- and glucocorticoid-regulated kinase 1 (Sgk1) is induced by mineralocorticoids and, in turn, upregulates the renal epithelial Na+ channel (ENaC). Total inactivation of Sgk1 has been associated with transient urinary Na+ wasting with a low-Na+ diet, while the aldosterone-mediated ENaC channel activation was unchanged in the collecting duct. Since Sgk1 is ubiquitously expressed, we aimed to study the role of renal Sgk1 and generated an inducible kidney-specific knockout (KO) mouse. We took advantage of the previously described TetOn/CreLoxP system, in which rtTA is under the control of the Pax8 promotor, allowing inducible inactivation of the floxed Sgk1 allele in the renal tubules (Sgk1fl/fl/Pax8/LC1 mice). We found that under a standard Na+ diet, renal water and Na+/K+ excretion had a tendency to be higher in doxycycline-treated Sgk1 KO mice compared with control mice. The impaired ability of Sgk1 KO mice to retain Na+ increased significantly with a low-salt diet despite higher plasma aldosterone levels. On a low-Na+ diet, the Sgk1 KO mice were also hyperkaliuric and lost body weight. This phenotype was accompanied by a decrease in systolic and diastolic blood pressure. At the protein level, we observed a reduction in phosphorylation of the ubiquitin protein-ligase Nedd4-2 and a decrease in the expression of the Na+-Cl−-cotransporter (NCC) and to a lesser extent of ENaC.
Combined inhibition of aromatase activity and dihydrotestosterone supplementation attenuates renal injury in male streptozotocin (STZ)-induced diabetic rats
Authors: Manigrasso Michaele B. , Sawyer R. Taylor , Hutchens Zachary M. , Flynn Elizabeth R. , Maric-Bilkan Christine
Published: February 2012

Our previous studies showed that streptozotocin (STZ)-induced diabetic male rats have increased estradiol and decreased testosterone levels that correlate with renal injury (Xu Q, Wells CC, Garman GH, Asico L, Escano CS, Maric C. Hypertension 51: 1218–1224, 2008). We further showed that either supplementing dihydrotestosterone (DHT) or inhibiting estradiol biosynthesis in these diabetic rats was only partially renoprotective (Manigrasso MB, Sawyer RT, Marbury DC, Flynn ER, Maric C. Am J Physiol Renal Physiol 301: F634–F640, 2011; Xu Q, Prabhu A, Xu S, Manigrassso MB, Maric C. Am J Physiol 297: F307–F315, 2009). The aim of this study was to test the hypothesis that the combined therapy of DHT supplementation and inhibition of estradiol synthesis would afford better renoprotection than either treatment alone. The study was performed in 12-wk-old male nondiabetic (ND), STZ-induced diabetic (D), and STZ-induced diabetic rats that received the combined therapy of 0.75 mg/day of DHT along with 0.15 mg·kg−1·day−1 of an aromatase inhibitor, anastrozole (Dta), for 12 wk. Treatment with the combined therapy resulted in attenuation of albuminuria by 84%, glomerulosclerosis by 55%, and tubulointerstitial fibrosis by 62%. In addition, the combined treatment decreased the density of renal cortical CD68-positive cells by 70% and decreased protein expression of transforming growth factor-β protein expression by 60%, collagen type IV by 65%, TNF-α by 55%, and IL-6 by 60%. We conclude that the combined treatment of DHT and blocking aromatase activity in diabetic male STZ-induced diabetic rats provides superior treatment than either treatment alone in the prevention of diabetic renal disease.
Sensitivity to temporal modulation rate and spectral bandwidth in the human auditory system: fMRI evidence
Authors: Overath Tobias , Zhang Yue , Sanes Dan H. , Poeppel David
Published: February 2012

Hierarchical models of auditory processing often posit that optimal stimuli, i.e., those eliciting a maximal neural response, will increase in bandwidth and decrease in modulation rate as one ascends the auditory neuraxis. Here, we tested how bandwidth and modulation rate interact at several loci along the human central auditory pathway using functional MRI in a cardiac-gated, sparse acquisition design. Participants listened passively to both narrowband (NB) and broadband (BB) carriers (1/4- or 4-octave pink noise), which were jittered about a mean sinusoidal amplitude modulation rate of 0, 3, 29, or 57 Hz. The jittering was introduced to minimize stimulus-specific adaptation. The results revealed a clear difference between spectral bandwidth and temporal modulation rate: sensitivity to bandwidth (BB > NB) decreased from subcortical structures to nonprimary auditory cortex, whereas sensitivity to slow modulation rates was largest in nonprimary auditory cortex and largely absent in subcortical structures. Furthermore, there was no parametric interaction between bandwidth and modulation rate. These results challenge simple hierarchical models, in that BB stimuli evoked stronger responses in primary auditory cortex (and subcortical structures) rather than nonprimary cortex. Furthermore, the strong preference for slow modulation rates in nonprimary cortex demonstrates the compelling global sensitivity of auditory cortex to modulation rates that are dominant in the principal signals that we process, e.g., speech.
Effect of gravity-like torque on goal-directed arm movements in microgravity
Authors: Bringoux L. , Blouin J. , Coyle T. , Ruget H. , Mouchnino L.
Published: February 2012

Gravitational force level is well-known to influence arm motor control. Specifically, hyper- or microgravity environments drastically change pointing accuracy and kinematics, particularly during initial exposure. These modifications are thought to partly reflect impairment in arm position sense. Here we investigated whether applying normogravitational constraints at joint level during microgravity episodes of parabolic flights could restore movement accuracy equivalent to that observed on Earth. Subjects with eyes closed performed arm reaching movements toward predefined sagittal angular positions in four environment conditions: normogravity, hypergravity, microgravity, and microgravity with elastic bands attached to the arm to mimic gravity-like torque at the shoulder joint. We found that subjects overshot and undershot the target orientations in hypergravity and microgravity, respectively, relative to a normogravity baseline. Strikingly, adding gravity-like torque prior to and during movements performed in microgravity allowed subjects to be as accurate as in normogravity. In the former condition, arm movement kinematics, as notably illustrated by the relative time to peak velocity, were also unchanged relative to normogravity, whereas significant modifications were found in hyper- and microgravity. Overall, these results suggest that arm motor planning and control are tuned with respect to gravitational information issued from joint torque, which presumably enhances arm position sense and activates internal models optimally adapted to the gravitoinertial environment.
Mechanical signals at the base of a rat vibrissa: the effect of intrinsic vibrissa curvature and implications for tactile exploration
Authors: Quist Brian W. , Hartmann Mitra J. Z.
Published: February 2012

Rats actively tap and sweep their large mystacial vibrissae (whiskers) against objects to tactually explore their surroundings. When a vibrissa makes contact with an object, it bends, and this bending generates forces and bending moments at the vibrissa base. Researchers have only recently begun to quantify these mechanical variables. The present study quantifies the forces and bending moments at the vibrissa base with a quasi-static model of vibrissa deflection. The model was validated with experiments on real vibrissae. Initial simulations demonstrated that almost all vibrissa-object collisions during natural behavior will occur with the concave side of the vibrissa facing the object, and we therefore paid particular attention to the role of the vibrissa's intrinsic curvature in shaping the forces at the base. Both simulations and experiments showed that vibrissae with larger intrinsic curvatures will generate larger axial forces. Simulations also demonstrated that the range of forces and moments at the vibrissal base vary over approximately three orders of magnitude, depending on the location along the vibrissa at which object contact is made. Both simulations and experiments demonstrated that collisions in which the concave side of the vibrissa faces the object generate longer-duration contacts and larger net forces than collisions with the convex side. These results suggest that the orientation of the vibrissa's intrinsic curvature on the mystacial pad may increase forces during object contact and provide increased sensitivity to detailed surface features.
Time-frequency analysis of short-lasting modulation of EEG induced by intracortical and transcallosal paired TMS over motor areas
Authors: Manganotti Paolo , Formaggio Emanuela , Storti Silvia Francesca , De Massari Daniele , Zamboni Alessandro , Bertoldo Alessandra , Fiaschi Antonio , Toffolo Gianna Maria
Published: February 2012

Dynamic changes in spontaneous electroencephalogram (EEG) rhythms can be seen to occur with a high rate of variability. An innovative method to study brain function is by triggering oscillatory brain activity with transcranial magnetic stimulation (TMS). EEG-TMS coregistration was performed on five healthy subjects during a 1-day experimental session that involved four steps: baseline acquisition, unconditioned single-pulse TMS, intracortical inhibition (ICI, 3 ms) paired-pulse TMS, and transcallosal stimulation over left and right primary motor cortex (M1). A time-frequency analysis based on the wavelet method was used to characterize rapid modifications of oscillatory EEG rhythms induced by TMS. Single, paired, and transcallosal TMS applied on the sensorimotor areas induced rapid desynchronization over the frontal and central-parietal electrodes mainly in the alpha and beta bands, followed by a rebound of synchronization, and rapid synchronization of delta and theta activity. Wavelet analysis after a perturbation approach is a novel way to investigate modulation of oscillatory brain activity. The main findings are consistent with the concept that the human motor system may be based on networklike oscillatory cortical activity and might be modulated by single, paired, and transcallosal magnetic pulses applied to M1, suggesting a phenomenon of fast brain activity resetting and triggering of slow activity.