bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of dietary fish oil (FO) supplementation on the skeletal muscle blood flow (BF) response to submaximal treadmill exercise
Authors: Hammel Lauren Elizabeth , Marshall Ben J , Hageman Karen S , Poole David C , Stebbins Charles L , Musch Timothy I
Published: June 2021

Dietary supplementation (SUPP) with FO containing docosahexaenoic (DHA) and eicosapentaenoic acid (EPA) has been demonstrated to produce advantageous effects on vascular function. Specifically, FO SUPP has resulted in enhanced brachial artery dilation during rhythmic handgrip exercise (EX). The effects of FO on BF during dynamic full body EX, however, remain unknown.

PURPOSE:

To test our hypothesis that 6 weeks of dietary SUPP with DHA and EPA enhances the regional BF response to submaximal treadmill EX in the rat hindlimb.

METHODS:

Following 6 weeks of dietary SUPP with safflower oil (SO) (control; n = 9) or FO (n = 8), heart rate (HR), mean arterial pressure (MAP), and BF to the hindlimb were measured at rest and during submaximal treadmill EX (20 m/min, 10%) via radiolabeled microspheres in male Sprague‐Dawley rats.

RESULTS:

HR and MAP were not different between SO and FO at rest or EX (P<0.05). BF was not different between SO and FO at rest. During EX, FO exhibited greater BF in 8 of the 28 muscle parts measured as well greater BF (158±9) to the total hindlimb musculature than SO (128±10ml/min/100g) (P<0.05).

CONCLUSION:

These results demonstrate that 6 weeks of dietary SUPP with FO results in enhanced BF to the hindlimb during submaximal EX. Thus, SUPP with FO may have therapeutic effects on oxygen delivery and vascular function in patients with impaired vascular function and EX tolerance.
Micronutrient Deficiencies in a Long Term Acute Care Hospital
Authors: Parkin Courtney , Rollins Carol
Published: June 2021

Background: The roles of micronutrients and the consequences of their deficiencies in human metabolism, immune function and wound healing are well documented. However, the prevalence of micronutrient deficiencies within the hospital setting is unknown.

Objectives: To identify the prevalences of zinc, selenium, vitamin A, vitamin D, and vitamin B12 deficiencies in a long term acute care hospital setting.

Results: The number and percent of serum levels indicative of micronutrient deficiency are shown in Table 1.

Conclusions: Micronutrient deficiencies within the long term acute care environment appear to be widespread. Patients with a history of poor dietary habits and chronic illness are at high risk of nutrient deficiencies, including micronutrient deficiencies.
Association between quantitative measures of skin color & plasma 25‐hydroxyvitamin D (25OHD)
Authors: Rockell Jennifer , Skeaff Murray , Green Tim
Published: June 2021

Most vitamin D is obtained through UV synthesis in the skin. Dark‐skin ethnic groups have lower 25OHD concentrations than light skin groups; due to more skin melanin, which acts as a UV filter. Ethnicity is a proxy measure of skin color & within an ethnic group skin color varies greatly. It would be expected that the color of unexposed skin would be negatively associated with 25OHD. Conversely, as skin color darkens with sun exposure, a measure of tanning would be positively associated with 25OHD. Skin color can be quantified using reflectance colorimetry with higher numbers indicating darker skin. We aimed to determine the association between constitutive (upper inner‐arm) & sun‐induced (forearm) skin color & 25OHD in a group of Pacific Islanders (n=82) & Europeans (n=239) living in NZ (47°S) in summer. Mean (SD) serum 25OHD was 88 (31) nmol/L for Europeans and 75 (34) nmol/L for Pacific Islanders. Based on constitutive skin color measurements 36% of participants were very light, 45% light, 15% intermediate, 4% tanned, & 1% dark. Tanning at the forearm but not constitutive skin color was a significant predictor of 25OH D. Each 10 unit higher value at the forearm (less tanning) was associated with a ~4.9 nmol/L lower 25OHD (P<0.001). Tanning was a more important predictor of 25OHD than constitutive skin color in this population. Further study is needed in a population with a wider range of constitutive skin color.
Spiroplasma experimentally induce astrogliosis and retinopathy in ruminants characteristic of transmissible spongiform encephalopathies
Authors: Bastian Frank O. , Todd William J. , French Hilari M. , Boudreaux Charles M. , Enright Fred M. , Elzer Philip H.
Published: June 2021

Spiroplasma mirum, tiny motile wall‐less bacterium, is a candidate causal agent of transmissible spongiform encephalopathy (TSE), which includes scrapie in sheep, chronic wasting disease in deer and Creutzfeldt‐Jakob disease (CJD) in humans. Spiroplasma spp. have been cultured from TSE‐affected brains via passage in embryonated eggs into cell‐free media. When inoculated into ruminants, spiroplasma induced spongiform encephalopathy with clinical signs of TSE. Here, brain tissues and eyes from the reported study (Bastian et al., JMM, 2007) were examined by immunohistochemistry for glial fibers and distribution of the organisms. Spiroplasma induced a striking astrogliosis with hypertrophic astrocytes and retinopathy characteristic of TSE. Spiroplasma were demonstrated in neurons throughout the brain tissues and retina. Cell‐free spiroplasma were seen within the vitreous and organisms immunostained within vacuolated corneal endothelia. The infected corneal endothelia may be the source for iatrogenic transmission of CJD via corneal transplants, and the presence cell‐free organisms within the vitreous may account for contamination of instruments secondary to ophthalmic surgical procedures involving the posterior eye. Supported in part by NIH grant R01‐NS044003, donations from CJD‐affected families and LSU Agricultural Center Research funds.
Predictors of 5 y change in cardio‐metabolic risk among Guatemalan adults
Authors: Gregory Cria O. , Ramirez‐Zea Manuel , Martorell Reynaldo , Stein Aryeh D.
Published: June 2021

We examined 376 Guatemalan adults in 1997â8 (age 24Â±3 y) and in 2002â4. Overweight (BMIâ¥25 kg/m2) and obesity (BMIâ¥30 kg/m2) prevalence increased (23% to 46% and 6% to 15%, respectively, both p<0.01), as did percent body fat (%BF; 23.3Â±9.8 to 27.0Â±10.2 %), triglycerides (129.2Â±64.5 to 161.0Â±80.2 mg/dL), the ratio of total to HDL cholesterol (TC:HDL‐c; 4.3Â±1.0 to 4.5Â±1.5 mg/dL), glucose (85.7Â±9.3 to 92.1Â±18.3 mg/dL), and diastolic blood pressure (DBP; 68.8Â±8.7 to 70.0Â±8.5 mmHg) (all P<0.05). 5y increase in %BF was associated with lower physical activity level at baseline and was higher among men; women had greater 5y increase in DBP. Age, rural/urban residence, schooling, socio‐economic status, smoking, and dietary energy and fat intakes were not associated with 5y changes in any risk factor examined. In adjusted models, 5y change in %BF was associated with changes in triglycerides (Î²= 4.6; 95% CI=2.3, 6.8 mg/dL per %), TC:HDL‐c (Î²= 0.09; 95% CI=0.05, 0.1 per %), and DBP (0.7; 95% CI=0.5, 1.0 mmHg per %). The 5y increases in cardio‐metabolic risk factors were primarily predicted by changes in %BF. Promotion of increased physical activity may help prevent adverse changes in body composition and consequent cardio‐metabolic risk.

Funding: NIH TW005598; AHA Pre‐doctoral fellowship (COG)
Bone mineralization and vitamin D/calcium intake of infants fed breast‐milk, milk‐based formula or soy‐based formula
Authors: ANDRES Aline , SMITH Stacee A. , BADGER Thomas M. , GILCHRIST Janet M.
Published: June 2021

A cross‐sectional analysis of bone mineralization during the first year of life of infants (N=107) exclusively fed breast‐milk (BF), milk‐based formula (MF) or soy‐based formula (SF) for at least the first 4 months of life was conducted. Participants were part of the longitudinal Beginnings study. Bone mineral content (BMC) was assessed using dual‐energy X‐ray absorptiometry at 3, 6, 9 and 12 months‐of‐age. Vitamin D and calcium intake were also assessed. At 3 and 6 months, BMC/mass ratio was significantly lower for SF compared to BF and MF groups. At 9 and 12 months, MF had higher BMC/mass ratio compared to BF and SF infants (p<0.05), but BF and SF were similar. BMC significantly increased in each feeding group over time but at a higher rate for SF (30% vs. 21% and 27% for BF and MF, respectively). These results suggest that although SF infants had a lower initial BMC, their rate of bone growth was faster than in BF infants. Vitamin D and calcium intake were significantly lower in BF compared with MF and SF at all time points. There was no difference in vitamin D and calcium intake between MF and SF infants except for calcium intake at 3 months. Further work is needed to delineate the characteristics of bone mineralization in each feeding group relative to their serum vitamin D concentration and calcium deposition. Funded by USDA‐ARS CRIS # 6251‐51000‐005‐03S.
Genetic Polymorphism Of Glutathione S‐Transferase (GST) And Susceptibility To And Severity Of Chronic Obstructive Pulmonary Disease (COPD)
Authors: Hemimi Neveen Salah El Din , Attar May Mahmoud Al , Elwahab Mahmoud M Adb
Published: June 2021

Background: Chronic tobacoo smoking is a major risk factor in the development of COPD. However, it is estimated that only 10–20% of chronic heavy smokers will develop symptomatic COPD. This indicates the possible contribution of environmental or genetic cofactors to the development of COPD in smokers.

Aim: To study the relationship between GST polymorphism and susceptibility to and severity of COPD in smokers. Method: A case control study was done on 140 patients with COPD and 140 matched controls. All subjects were smokers or exsmokers. The GSTM1 and GSTT1 genotypes were identified by polymerase chain reaction in peripheral blood DNA samples. Analysis of data was done by IBM computer using SPSS program.

Results: The proportion of GSTM1‐null genotypes was significantly higher in patients with COPD than in control subjects (62.2% versus 32.2%). The odds ratio was 3.5 (95% confidence interval (CI) =2.1–5.7). Moreover the patients with GSTM1 null genotype were at high risk of developing the severe type of COPD. The odds ratio was 3.14 and (95% CI= 1.5–6.7). However the genotype frequencies of GSTT1‐null genotype did not show significant difference between groups.

Conclusion: Our data provide evidence that smokers with null genotype of GSTM1 were more susceptible to develop the severe type of COPD. Keyword: Genetic polymorphism, COPD, GSTT1, GSTM1. Source of Research support: Dubai Medical College for Girls.
Mechanoregulated expression of smooth muscle alpha‐actin in myofibroblasts is mediated by MRTF‐A localization and actin dynamics
Authors: Tomasek James J , Haaksma Carol J , McRae Joel K , Risinger George M , Howard Eric W
Published: June 2021

Expression of smooth muscle α‐actin (SMAA), which promotes myofibroblast contractile function and wound closure, is mechanoregulated. We hypothesize that mechanoregulated actin dynamics control the localization of myocardin related transcription factor‐A (MRTF‐A) thereby mechanoregulating SMAA expression. G‐actin has been shown to bind MRTF‐A, thus sequestering the MRTF‐A in the cytoplasm and blocking MRTF‐A activated SMAA expression. MRTF‐A is necessary for SMAA expression as demonstrated by shRNA knockdown. Decreasing myofibroblast tension with blebbistatin, or culturing myofibroblasts on a compliant collagen matrix, resulted in a loss of stress fibers, a higher G‐/F‐actin ratio, MRTF‐A localization to the cytoplasm, and decreased SMAA promoter activity. Treating cells on a compliant matrix with jasplakinolide or cytochalasin D resulted in increased F‐actin, MRTF‐A localization to the nucleus and increased SMAA promoter activity. Together, these results provide a model for mechanoregulated SMAA expression; in response to a stiff substratum G‐actin is assembled into F‐actin resulting in the release of MRTF‐A, expression of SMAA, and myofibroblast differentiation. Funded by NIH grant R01 GM60651
TGFb‐HIPK2 Signaling Pathway in the Survival of Dopamine Neurons During Toxin‐induced Degeneration
Authors: Zhang Jiasheng , Huang Eric J
Published: June 2021

The current study investigates TGFb‐HIPK2 signaling pathway in MPTP‐induced degeneration of DA neurons. Our results indicate that loss of HIPK2 leads to a near complete protection of DA neurons from MPTP‐induced cell death. Whereas MPTP treatment induces a robust increase in c‐jun phosphorylation after MPTP in wild type DA neurons, no phospho‐c‐jun is detected in the DA neurons of Hipk2 mutants. To determine if HIPK2 is sufficient for MPTP‐induced cell death, we used a genetic approach to selectively express additional copies of HIPK2 in DA neurons. Our results indicate that DA neurons in conditional mouse mutants expressing extra copies of HIPK2 show a dose‐dependent increase in MPTP‐induced cell loss. Taken together, these results support the notion that, HIPK2 is a key upstream regulator in JNK‐dependent activation of c‐jun phosphorylation and MPTP‐induced cell death in DA neurons. Our results further suggest that TGFb‐HIPK2 signaling pathway may serve as a therapeutic target to support the survival of DA neurons in MPTP‐induced degeneration.
Development of Methods for Exposure of Cultured Cells to Direct and Secondhand Cigarette Smoke
Authors: Mukherjee Shyamali , Das Salil K
Published: June 2021

Several epidemiological studies have linked cigarette smoking to breast cancer, particularly in young women. However, there is no scientific validity of this link between either direct or secondhand smoking and breast cancer. Because of difficulties in conducting such studies in humans, it is necessary to develop a smoke exposure model system for cells in culture. We have developed and standardized a smoke exposure model system for human breast cell lines. This is a modified version of CULTEX cell culture smoke exposure system using the principle of the air/liquid exposure technique. Cells were grown in culture media (DMEM/F‐12) and exposed to both direct and secondhand smoke generated from 1R3F cigarettes in a THRI MS‐SS smoke exposure system. A sham control group was also used where cells were exposed to the same environmental condition without smoke exposure. For the dose response study breast cells were exposed to 0.03, 0.06, 0.12 and 0.18 puff/ml and incubated for 24 h. For time response study, cells were exposed to 0.18 puff/ml and then incubated for 6, 12, 24 and 36 h. Our results indicated that both direct and secondhand smoke exposure caused an increase in cell proliferation and a morphological change at low dose and cellular apoptosis at higher dose. (Supported by 2006 FAMRI Clinical Innovator Award).
Forms of Vitamin E have Opposing Effects on Experimental Asthma
Authors: Cook‐Mills Joan Marie , Berdnikovs Sergejs , Abdala‐Valencia Hiam , McCary Christine , Somand Michelle , Bryce Paul
Published: June 2021

We previously reported that the endothelial cell adhesion molecule VCAM‐1 signals through reactive oxygen species generated by NOX2 and that this regulates leukocyte trafficking in vitro and in vivo. Therefore, we determined whether leukocyte migration is modulated by two major dietary forms of the antioxidant vitamin E, (d)‐α‐tocopherol (aT) and (d)‐γ‐tocopherol (gT). Mice were treated daily with physiological levels of aT or gT after sensitization and before challenge with the antigen ovalbumin, a model of experimental asthma. aT significantly inhibited inflammation in the lung and inhibited airway hyperresponsiveness (AHR). Surprisingly, gT exacerbated inflammation and AHR in the lung. Most interestingly, when gT and aT were co‐administered, gT ablated the benefit of aT. In addition, gT antagonized aT inhibition of leukocyte transendothelial migration in vitro. In summary, aT and gT have opposing effects on inflammation. These data have implications on the design and interpretation of clinical studies with mixed tocopherols in supplements and diets. Reported studies indicate that aT is beneficial for experimental asthma in mice and for asthma in Europeans who consume diets low gT but not for Americans who consume diets high in gT, suggesting that differences in the outcomes in these reports on vitamin E may, in part, reflect the opposing effects of α‐tocopoherol and γ‐tocopherol.

(NIH R01 HL069428 to JC‐M)
Human defensin alpha‐1 causes Trypanosoma cruzi membrane pore formation and induces DNA fragmentation, which leads to trypanosome destruction
Authors: Madison Marisa Nia , Kleshchenko YY , Nde PN , Simmons KJ , Lima MF , Villalta F
Published: June 2021

Human defensins play a fundamental role in the initiation of innate immune responses to some microbial pathogens. Here we show that mammalian cells rapidly up‐regulate the expression of human defensin alpha‐1 as an innate response to counter Trypanosoma cruzi infection. We discovered that defensin alpha‐1 displays a trypanocidal role against T. cruzi, the causative agent of Chagas' disease. The toxicity of human defensin alpha‐1 against T. cruzi is mediated by membrane pore formation and the induction of nuclear and mitochondrial DNA fragmentation, leading to trypanosome destruction. Exposure of trypomastigote and amastigote forms of T. cruzi to defensin alpha‐1 significantly reduced parasite viability in a peptide concentration‐dependent and saturable manner. Electron microscopic analysis of trypomastigotes exposed to defensin alpha‐1 revealed pore formation in the cellular and flagellar membranes, membrane disorganization, and blebbing as well as cytoplasmic vacuolization. Furthermore, human defensin alpha‐1 enters the trypanosome when membrane pores are present and is associated with later intracellular damage. Preincubation of trypomastigotes with defensin alpha‐1 followed by exposure to human epithelial cells significantly reduced T. cruzi infection in these cells. Thus, human defensin alpha‐1 is an innate immune molecule that causes severe toxicity to T. cruzi and plays an important role in reducing cellular infection. This is the first report showing that human defensin alpha‐1 causes membrane pore formation in a human parasite, leading to trypanosome destruction. This work was supported in part by NIH grants 1SC1 GM081168,GM 08037, GM 059994, AI 07281, AI 056667, HL 007737, MD 000104,RR 003032, and DK 20539.
Impact of diabetes peer counseling on glycosylated hemoglobin among Latinos enrolled in the DIALBEST Trial: Preliminary results
Authors: Perez‐Escamilla Rafael , Vega‐Lopez Sonia , Segura‐Perez Sofia , Damio Grace , Fernandez Maria Luz , Calle Mariana , Samuel Grace K , Chhabra Jyoti , D’Agostino Darrin
Published: June 2021

The ongoing Diabetes among Latinos Best Practices Trial (DIALBEST) has randomly assigned Latinos with type 2 diabetes to the standard of clinical care or the intervention group that receives home visits from peer counselors. Intervention group participants receive one weekly visit during the first month, biweekly during months 2 and 3, and monthly thereafter until month 12. Peer counselors teach culturally tailored nutrition, physical activity and adherence with glucose monitoring, medications and medical appointments. We report on 63 subjects with HbA1c data available at baseline and 3 months. The proportion of females in the intervention (n=30) and control groups (n=33) was 80.0% and 84.8%, respectively. The corresponding HbA1c values were 10.2 ± 1.8 % vs. 10.1 ± 1.8 %, and for glucose 11.3 ± 4.4 mmol/l vs. 11.6 ± 5.1 mmol/l. Baseline BMI was also equivalent in the intervention and control groups (34.7 ± 8.6 vs. 35.4 ± 8.6). As expected, there was a significant correlation between HbA1c and plasma glucose both at baseline (r=0.515, p<0.001) and 3 months (r=0.685, p<0.001). The intervention group had a greater improvement in HbA1c at 3 months compared to the control group (−1.2 ± 1.8 % vs. −0.5 ± 1.6 %, 1‐tailed p=0.035). Thus far the DIALBEST intervention is having a short term positive impact on glycemic control. Funded by The Connecticut Latino Health Disparities NIH EXPORT Center (NIH‐NCMHD grant # P20MD001765).
Consumption of three supplements with identical micronutrient content in pregnant Mexican women
Authors: Dominguez Clara P , Neufeld Lynnette M , Cabrera Amira Hernandez , Cordero Sonia Hernandez
Published: June 2021

The objective of this analysis was to compare consumption of three nutritional supplements with identical micronutrient content for pregnant women. Communities (18 per supplement) were randomly assigned to receive a fortified beverage (FB, 54 g of milk‐based powder dissolved in a glass of water), tablets (TB, 1 tablet) or Sprinkles (SK, 1 g dissolved in food). Pregnant women, beneficiaries of the Oportunidades program were recruited before 25 wk pregnancy. Supplements were delivered daily (6 mo) then weekly during pregnancy until 3 m postpartum (pp). Compliance and consumed amount (0, 1 for TB; 0, ¼, ½, ¾, 1 of portion served for FB and SK) was recorded. Appropriate statistical analyses for cluster randomized trials were used. 715 women were recruited (FB n=233; TB n=238; SK n=244). Supplement was consumed for 180±39 days and there was no difference in the mean proportion of days offered that any supplement was consumed between groups (91.0%). The mean proportion of days that the complete dose was consumed was significant lower in the FB group (86.4%) than the TB (91.1%) or SK (90.8%) groups. The proportion of days offered that any supplement was consumed was significantly higher pp than during pregnancy in the FB (+3.6%) and TB (+2.9%) groups, but there was no difference in the SK group (+1.2%). Despite the controlled nature of this study, consumption of the entire dose was lower among women consuming the FB.

The Oportunidades program financed this study and holds the rites to the data presented.
KIST‐A164A Increases a Quinone Reductase Potentially by Inhibiting COX‐2‐Dependent Inflammatory Signaling
Authors: Lee Saet Byoul , Kwon Hak‐Cheol , Yang Hyun Ok , Kang Kyungsu , Nho Chu Won
Published: June 2021

The up‐regulation of phase II detoxification genes is believed to play an important role in cancer prevention. Cyclooxygenase‐2 (COX‐2) is often up‐regulated in human colorectal cancers, and it is considered as one of major inflammatory factors causing carcinogenesis. However, the relationship between the induction of phase II enzyme and COX‐2 activation in terms of cancer prevention is unknown. We have identified a polyacetylene compound (KIST‐A164A) from Aster koraiensis as a major principle for a significant quinone reductase (QR) induction via transcriptional activation. We also found that KIST‐A164A significantly inhibited COX‐2 mRNA expression levels and PGE2 production in a dose‐dependent manner. Further molecular study revealed that co‐treatment with KIST‐A164A and acetylsalicylic acid, an inhibitor of COX‐2, increased the expression of QR mRNA and protein levels, which was coincidently enhancing QR activity as well as antioxidant response element (ARE) activation in Caco‐2 cells. The ARE activation was increased 2‐fold over control when the cells were treated with 50 μM KIST‐A164A in the presence of JNK1/2 inhibitor, SP600125. Taken together, we present here that KIST‐A164A increases QR through ARE and it could be related to inhibition of COX‐2‐dependent inflammatory signaling.
Effects Of Polystyrene Particle Density on Type II Pneumocytes
Authors: Benghuzzi Ham A , Cameron Joseph A , Tucci Michelle A
Published: June 2021

Inhalation of small particles is associated with respiratory diseases. The aim of the present study was to investigate the role of ultrafine particle number on lung cells and to describe the effects due to phagocytosis of particles by the cells. A549 cells were treated with 1000, 5000, and 10,000 polystyrene particles and incubated at 37„aC for periods of 24, 48, and 72 hours. Cellular viability and damage were determined on the treated cells and compared with untreated controls. The lung cells were also observed microscopically to assess cell damage. Lung cells exposed for 24 hours with 1,000 and 5,000 particles showed the greatest increase in cell number. Cellular protein levels were similar for all groups (p> 0.05) for the duration of the study. MDA levels for treated groups at 24 and 72 hours were not statistically different from the control groups. Cells treated at 48 hours with 5,000 and 10,000 particles showed increased in the MDA levels above low particle number and control treated groups. Morphological evaluation of the cells revealed increased inclusions with increasing dose. The results from this study indicate the ability of A549 cells to respond to a challenge with ultrafine particles. The concentrations tested caused an initial stimulation cell proliferation at 24 hours followed by increased damage at 48 hours. Future studies will focus on the inflammatory products formed by ingestion of the ultrafine particles.
Direct visualization of adenosine A2A and dopamine D2L receptor oligomers in a neuronal cell model
Authors: Vidi Pierre‐Alexandre , Chemel Benjamin R. , Watts Val J.
Published: June 2021

Dopamine D2 and adenosine A2A receptors are co‐expressed in GABAergic neurons in the basal ganglia where they antagonistically regulate adenylyl cyclase activity. Interactions between D2 and A2A are thought to play an important role in drug addiction and in the pharmacotherapy of Parkinson's disease. Molecular mechanisms may involve the formation of receptor oligomers signaling to alternative pathways. D2 and A2A have been shown to form homo‐ and heterooligomers both in recombinant cells and in native brain tissues. However, the stoichiometry of these complexes, as well as the regulation or their formation (notably by ligands) remain to be characterized.

We have used bimolecular fluorescence complementation (BiFC) for direct visualization of A2A homodimers and A2A ‐ D2 heterodimers in a neuronal cell model. Initial studies revealed that the receptor‐BiFC fusion proteins remained functional. Preliminary experiments using multicolor BiFC indicated that prolonged exposure to D2 antagonist spiperone promotes A2A ‐ D2 dimerization whereas D2 agonist quinpirole treatment upregulates A2A homodimerization. In addition, studies combining BiFC with FRET provided evidence for the formation of higher‐order A2A D2 oligomers.

This project was funded by Purdue University and NIMH grant award MH060397 to VJW.
Cerebral microvascular dysfunction in angiotensin II‐induced hypertension
Authors: Vital Shantel A. , Russell Janice , Granger D. Neil
Published: June 2021

Inflammation has been implicated in the pathogenesis of hypertension. Although hypertension is a major risk factor for ischemic stroke, and inflammation contributes to the pathogenesis of stroke, relatively little is known about the inflammatory effects of arterial hypertension in the cerebral microcirculation. The objective of this study was to determine whether angiotensin II (AngII)‐induced hypertension promotes blood cell‐endothelial cell adhesion in cerebral venules. Leukocyte and platelet adhesion were monitored in cerebral venules of C57Bl/6 mice via a cranial window. Hypertension was induced by subcutaneous implantation of an AngII‐loaded Alzet pump for a 2 wk period. Mice were placed on either a normal (ND) or high salt diet (HSD). AngII infusion induced a dose‐dependent (400–2000 ng/kg/min) increase in arterial pressure, which was higher in HSD mice. Mice on either ND or HSD exhibited an increased adhesion of both leukocytes and platelets at 600 ng/kg/min. Higher AngII doses did not elicit further increases in blood cell adhesion in ND mice, but further dose‐dependent increases were noted HSD mice. Blood brain barrier (BBB) permeability was increased by AngII in both ND and HSD mice. These findings indicate that the cerebral microcirculation assumes an inflammatory and pro‐thrombogenic phenotype in AngII‐induced hypertension, which may account for the accompanying BBB dysfunction. (HL26441)
AICAR preconditioning prevents postischemic leukocyte rolling and adhesion:Role of KATP channels and heme oxygenase
Authors: Gaskin Frederick Spencer , Kamada Kazuhiro , Yusof Mozow , Durante William , Gross Garrett , Korthuis Ronald
Published: June 2021

We have shown that eNOS activation plays a critical role as a trigger for the anti‐inflammatory state that is induced by activation of 5′‐adenosine monophosphate‐activated protein kinase (AMPK) with AICAR (AICAR‐PC), such that postcapillary venules fail to support increased leukocyte‐endothelial cell adhesive interactions (LEI) on exposure to ischemia/reperfusion (I/R) 24 hrs later. The aim of the present study was to determine if ATP‐sensitive potassium channels (KATP) and heme oxygenase (HO) are effectors of AICAR‐PC. We also examined the role of KATP as triggers of AICAR‐PC. To address these issues, C57BL/6J mice were treated with either 5‐hydroxydecanoate (5‐HD, mitochondrial KATP (mKATP) inhibitor) or tin protoporphyrin (SnPP, HO inhibitor) during I/R 24 hrs after AICAR‐PC. I/R induced a marked increase in LEI relative to sham control mice, responses that were prevented by AICAR‐PC 24 hrs prior to I/R. The anti‐adhesive effects of AICAR‐PC were abolished by 5‐HD or SnPP but not an inactive PP treatment during I/R. The HO inducer hemin mimicked the anti‐inflammatory actions of AICAR‐PC. Interestingly, AICAR‐PC in the presence of KATP inhibition failed to prevent its anti‐adhesive actions. Our results indicate that AMPK agonists produce an anti‐inflammatory phenotype in postcapillary venules that is mediated by mKATP‐ and HO‐dependent mechanisms during I/R. Supported by AA14945, DK 43785, and HL73414.
Resveratrol attenuates mitochondrial ROS generation in endothelial cells treated with cigarette smoke extract
Authors: Ungvari Zoltan , Mukhopadhyay Parta , Labinskyy Nazar , Podlutsky Andrej , Pacher Pal , Csiszar Anna
Published: June 2021

Previously we have shown that water soluble components of cigarette smoke (CS) up‐regulate NAD(P)H oxidase‐dependent ROS production in endothelial cells. In the present study we tested the effect of CS on mitochondrial ROS production in cultured coronary arterial endothelial cells (CAECs) and determined whether the sirtuin activator resveratrol (RES) protects CAECs against the deleterious effects of CS. Mitochondrial O2.‐ production was assessed in CS extract treated HCAECs by flow cytometry and confocal microscopy using MitoSOX Red. CS extract (0.04 to 4 ug/mL) elicited dose‐dependent increases in mitochondrial O2.‐ generation which were prevented by resveratrol pre‐treatment (1–10 uM; for 24 h). At higher concentrations CSE also elicited apoptosis in CAECs (as reflected by the increased cellular annexin V staining and caspase 3 activity), which was also prevented by resveratrol. Thus, RES at nutritionally relevant concentrations attenuates CS‐induced mitochondrial oxidative stress and protects endothelial cells from apoptotic cell death.
Regulation of Basal and Beta‐Adrenergic‐Stimulated Ventricular Cardiomyocyte Contraction Rate by Distinct Populations of Chloride Channels
Authors: Sellers Zachary M. , Xiang Kevin , Best Philip M.
Published: June 2021

A variety of Cl− channels have been identified in ventricular cardiomyocytes and are presumed to be important in modulation of cardiac action potential duration. In the current study we examined if modulation of Cl− channel activity affects ventricular cardiomyocyte contraction rate. Ventricular myocytes were isolated from neonatal mice and cultured until they formed a functional syncytium. Contraction rates were captured by video imaging and analyzed in the presence and absence of beta‐adrenergic receptor stimulation by isoproteronol (ISO). Cl− channel activity was inhibited by various Cl− channel inhibitors. Glibenclamide, DPC, and CFTRinh‐172 significantly, but transiently, inhibited baseline contraction rate. All three drugs significantly inhibited prolonged increases in ISO‐stimulated cardiomyocyte contraction rate. 9‐AC elicited a prolonged inhibitory effect on baseline contraction rate, but failed to significantly alter ISO‐stimulated contraction rate. Tamoxifen transiently increased baseline contraction rate, but significantly inhibited ISO‐stimulated increases in contraction rate. Inhibitors of CFTR, calcium‐activated, and swelling‐activated Cl− channels modulate ventricular cardiomyocyte contraction rate, but do so in distinct ways. Our results indicate an important role for CFTR in the regulation of cardiomyocyte contraction rate. Supported by a GIA from the AHA.
Apoptotic susceptibility, muscle and mitochondrial perturbations in skeletal muscle of p53 wild‐type (WT) and knockout (KO) mice
Authors: Saleem Ayesha , Hood David A.
Published: June 2021

Evidence indicates that p53 plays a role in modulating apoptosis, cell‐cycle arrest and aerobic metabolism. Our goal was to evaluate the role of p53 in regulating basal and exercise‐induced apoptosis in skeletal muscle. p53 KO mice had impaired whole muscle cytochrome c oxidase (COX) activity and reduced intermyofibrillar (IMF) state 3 respiration. The rate of reactive oxygen species (ROS) production during state 3 was ∼75% higher in subsarcolemmal (SS) mitochondria and ∼ 2‐fold higher in IMF mitochondria in KO mice. Despite this, KO mice had a 46% lower maximum speed (Vmax) of mitochondrial permeability transition pore (mtPTP) opening and a 19% greater time to Vmax in SS mitochondria. There were no changes in pore kinetics in the IMF mitochondria. However, a ∼2‐fold decrease in basal cytochrome c release in the IMF mitochondria in p53 KO mice was detected. Surprisingly, muscle weight / gram of body weight was 1.7‐fold higher in KO, compared to the WT animals. p53 KO mice that were endurance trained for eight weeks on a running wheel ran ∼5‐fold lower distances than the WT mice, and exhibited a trend towards decreased ROS production in the SS mitochondria. Muscle fatigue, evaluated using in situ muscle contraction, was also greater in KO animals. These data indicate that lack of p53 impairs mitochondrial function, endurance capacity and reduces apoptotic potential in skeletal muscle.
P2X Receptor Blockade Inhibits Autoregulation of Renal Blood Flow In Vivo
Authors: Osmond David Alan , Inscho Edward W
Published: June 2021

P2X1 receptor activation is implicated in afferent arteriolar autoregulatory behavior in vitro. Experiments were performed to test the hypothesis that P2X receptor blockade with pyridoxal phosphate‐6‐azo‐benzene‐2,4‐disulfonic acid (PPADS, 20μmoles/kg) inhibits renal autoregulation in vivo. An occluder, placed around the aorta, was used to adjust renal arterial pressure in 10mmHg decrements. Autoregulatory profiles of renal blood flow (RBF) were measured before and during P2 receptor blockade using an ultrasonic flow probe placed around the left renal artery. Baseline RBF was similar between control and PPADS treated groups, averaging 8.3±0.7 and 8.4±0.6 ml/min, respectively. Under control conditions, the change in RBF averaged 0.7±0.9% (0.0±0.1 ml/min; n=6) at arterial pressures between 100 and 120 mmHg. Conversely, RBF decreased by 10.5±1.4% (0.8±0.2 ml/min; n=5; P<0.05) over the same pressure range, during PPADS treatment. Bolus i.v. infusions of a P2X1 receptor selective agonist, α, β‐methylene ATP (αβ‐mATP), reduced RBF dose‐dependently. 500ng of αβ‐mATP increased mean arterial pressure (MAP) by 8.2±0.7mmHg and reduced RBF by 50% from 6.1±0.4 to 3.0±0.7ml/min. During PPADS treatment, the effects of αβ‐mATP (500ng) on MAP and RBF were eliminated. These data support the hypothesis that P2X1 receptor activation is essential for initiating autoregulation in vivo.
Both cell volume and calmodulin contribute to the regulation of macula densa nitric oxide
Authors: Komlosi Peter , Zhang Zhi‐Ren , Steele Stacy , Chu Wen‐Feng , Bell Phillip Darwin
Published: June 2021

Elevations in luminal sodium chloride concentration ([NaCl]L) alter macula densa (MD) intracellular calcium concentration and result in the release of nitric oxide (NO). Also, concomitant elevations in [NaCl]L and luminal osmolality (osmL) lead to shrinkage of macula densa cells, while increases in [NaCl]L at constant osmL produce cell swelling. Currently, there is little information concerning the intracellular signaling event that leads to the activation of MD NO synthase. Current studies were performed to determine whether changes in MD cell volume and/or calmodulin‐dependent signaling activate MD NO synthase. Thick ascending limb‐glomerulus preparations were isolated and perfused from rabbits, and MD NO release was detected with fluorescence microscopy using DAF‐FM. Elevations in [NaCl]L (from 0 to 150 mmol/L) at constant osmL produced an increase in DAF‐FM fluorescence of 5±0.5%, while concomitant elevations in [NaCl]L and osmL led to an increase of 12±0.7%. Luminal application of furosemide, a blocker of Na+:2Cl‐:K+ cotransport, reversed [NaCl]L‐dependent NO release (‐0.04±0.2%). Also, application of the calmodulin‐inhibitor calmidazolium blocked [NaCl]L‐dependent NO release (1.8±1.8%). Thus, [NaCl]L‐dependent NO production by MD cells may be modulated by changes in cell volume and requires activation of calmodulin signaling.
Tubuloglomerular feedback (TGF) is decreased in COX‐1 knockout mice after chronic angiotensin II (Ang II) infusion
Authors: Araujo Magali , Welch William J
Published: June 2021

Prostaglandins (PGs) are important modulators of renal hemodynamics. PGE2 and PGI2 are the major PGs in the renal cortex of mice and acute infusion of Ang II induces a COX‐2 dependent increase of these PGs. Prostaglandin E synthase (PGES) is coexpressed with COX‐2 in the macula densa (MD). We previously showed that MD COX‐2 modulates TGF responses in the rat. Genetic deletion of COX‐1 is associated with renal vasoconstriction, decreased renal and urinary PGs and attenuated ang II‐induced hypertension. However, the effects of lower levels of Ang II in the absence of COX‐1 are not known. The present study was designed to evaluate the effects of slow pressure doses of Ang II (400 ng/kg/min) on TGF in COX‐1 knockout mice. Four groups of mice were studied: 1) COX‐1 wild type (WT) ; 2) COX‐1 knockout (KO); 3) WT infused with Ang II for 14 days; 4) COX‐1 KO infused with Ang II for 14 days. Chronic Ang II infusion increased significantly TGF in WT mice (WT: 9.3±0.7 vs WT+AngII: 12.2±1.7 mmHg, p<0.001). However, chronic ang II decreased TGF in COX‐1 KO mice (KO: 11.0±1.2 vs KO+AngII: 8.3±0.6 mmHg, p<0.01). Ang II infusion increased MAP similarly in both groups (WT: 93±2 vs KO: 91±3 mmHg, NS, n=6 mice). These data demonstrate the effects of MD COX‐2 production of PGs in the absence of COX‐1 and their influence in TGF. During chronic Ang II infusion, TGF is increased in WT mice, but reduced in COX‐1 KO mice. This suggests that COX‐1 is stimulated by Ang II and produces vasoconstricting PGs. In the absence of COX‐1, COX‐2 produces primarily vasodilating PGs in response to Ang II, resulting in a lower TGF. These data further demonstrate that PGs generated by both COX isoforms regulate TGF.
Frequency and distribution of CGG/GCC repeats of FRAXA and FRAXE genes in South Indian non‐specific MR cases
Authors: Katikala Lavanya , Guruju Mallikarjuna , Thelma B. K. , Ramesh Om Sai , Sujatha M. , Anandaraj MPJS
Published: June 2021

Fragile X syndrome is one of the X‐linked disorders associated with moderate to severe mental retardation. Fragile X A syndrome (FRAXA) and fragile X E syndromes (FRAXE) are caused by trinucleotide expansions of CGG and GCC repeat expansion at the 5′ UTR of FMR1 and FMR2 genes respectively. Based on the number of the CGG repeats, individuals are classified into normal (5–60), permutated (60–200 repeats) and full mutated individuals (>200 repeats). We have screened 335 individuals from south India with MR of unknown etiology. All the individuals are clinically evaluated by a 15 item check list, molecular diagnosis was carried out using PCR and sizing of the amplicons were carried out by polyacrylamide gel electrophoresis. Of the 335 cases screened 12 unrelated cases positive for FRAXA. No cases of FRAXE have been identified. All Positive cases were confirmed by southern blotting using stb12.3 probe. Our study is the first study to report the frequency of FRAXE in south Indian population. We found the frequency of FRAXA to be 3.5 % and FRAXE to be 0%. Radioactive PCR is best to exactly estimate the allele sizes thereby finding the allelic variants in the population. We found 25 FRAXA and 20 FRAXE allelic variants in our population. The most frequent FRAXA allele size was 29 CGG repeats and that of FRAXE was 15 GCC repeats in South Indian non‐specific MR cases.
Lysophosphatidic acid receptor subtype‐specific regulation of human oral fibroblast healing responses
Authors: Cerutis D. Roselyn , George JoJu , Headen Karmel V. , Brady Stephanie A. , Schulte Nancy A. , Toews Myron L. , Parrish Lawrence C. , McVaney Timothy P. , Wilwerding Terry M. , Mattson John S.
Published: June 2021

Background: LPA1–5 receptors (LPARs) bind lysophosphatidic acid (LPA), a G protein‐coupled lipid mediator involved in wound healing. We have published that LPA regulates the growth and migration of human gingival (GF) and periodontal ligament (PDLF) fibroblasts, and that they express LPA1–3; however, which of these subtype(s) mediate stimulation of MAPK, elevation of intracellular calcium, or collagen gel contraction is not known. Methods: Serum‐starved cells in 24‐well plates were stimulated with LPA or subtype‐specific agonists [NAEPA (LPA1); FAP‐12 (LPA2); OMPT (LPA3)] for 5 min, and activation of p44/p42 MAPK was assessed by SDS‐PAGE and WB. Serum‐starved cells in 96‐well plates were loaded with Fluo‐3 and elevation of intracellular calcium after stimulation with LPA or subtype‐specific agonists was measured using the FlexStation II (Molecular Devices). Cells in collagen gels were stimulated with LPA or subtype‐specific agonists, and gel areas were measured over time using Sorcerer (Optomax). Results: For GF and PDLF, LPA1–3 agonists stimulate MAPK, but LPA1 and LPA3 mediate elevation of intracellular calcium and collagen gel contraction. The responses mediated through LPA1 and LPA3 were inhibited by Ki16425, an LPA⅓ antagonist. Conclusion: LPA signals through different LPAR subtypes to control specific wound healing‐related responses of GF and PDLF. Support: NIH/NIDCR 1 R15 DE016855‐02 (D.R.C).
Anandamide biosynthesis catalyzed by the sequential actions of a phospholipase (ABH4) and phosphodiesterase (GDE1) on the precursor N‐acyl phosphatidylethanolamines
Authors: Simon Gabriel Moore , Cravatt Benjamin F
Published: June 2021

Anandamide is a member of the N‐acyl ethanolamine (NAE) family of signaling lipids and an endogenous ligand of cannabinoid receptors that is implicated in many physiological processes including pain, inflammation, appetite, and cognition. The biosynthetic precursor for NAEs are N‐acyl phosphatidyl ethanolamines (NAPEs) but the precise enzymatic steps leading to release of NAEs from NAPEs are unclear. Three pathways have been proposed: (1) direct release of NAE from NAPE by a D‐type phospholipase (NAPE‐PLD), (2) phospholipase C‐mediated cleavage of NAPE generating phospho‐NAE followed by action of a phosphatase and (3) double O‐deacylation of NAPE followed by cleavage of the phosphodiester bond of the resulting GP‐NAE. Evidence is presented in support of the third pathway: In mice containing a targeted deletion of the NAPE‐PLD gene, AEA levels were unchanged in brain and enzymatic conversion of NAPE to NAE absolutely required cleavage of the O‐acyl chains. Here we present evidence that a B‐type phospholipase (ABH4) and a metal‐dependent phosphodiesterase (GDE1) act on NAPEs and GP‐NAEs, respectively, to generate anandamide via the deacylation pathway.
Investigating the anti‐hypertriglyceridemic effect of Stearoyl‐CoA Desaturase 1 deficiency under liver X receptor activation
Authors: Chu Kiki , Ntambi James M
Published: June 2021

Stearoyl‐CoA desaturase (SCD) is the rate‐limiting enzyme necessary for the biosynthesis of monounsaturated fatty acids. Previously, we showed that 129SvEv mice with targeted disruption of the SCD1 gene (SCD1−/−) exhibit reduced hepatic triglyceride (TG) synthesis and reduced levels of TGs in very low‐density lipoprotein (VLDL) fractions. Furthermore, in recent studies we showed that SCD1 deficiency prevented the hypertriglyceridemic effect and reduced hepatic TG accumulation upon liver X receptor (LXR) activation with a synthetic agonist T0901317. To investigate the anti‐hypertriglyceridemic effect of SCD1 deficiency under LXR activation, hepatic TG production rate was determined by measuring the appearance of plasma TG under conditions in which TG hydrolysis by lipoprotein lipase (LPL) is inhibited using Poloxamer 407. Both wild‐type and SCD1−/− mice showed the same level of increase in the rate of hepatic TG production under LXR activation, suggesting that the ability of SCD1−/− mice to secrete TG rich‐VLDL particles is not impaired. In addition, induction of hepatic LPL gene expression upon LXR activation was significantly higher in the SCD1−/− mice compared to wild‐type counterpart. These results suggest that the lack of increase in plasma TG upon LXR activation in SCD1−/− mice may be due to increase in the clearance of TG rich‐VLDL particles via an increase in the hydrolysis of VLDL‐TG by LPL.
Galectin‐3 Promotes Formation of Lamellipodia in Corneal Epithelial Cells by Interacting with α3β1 Integrins
Authors: Saravanan Chandrassegar , Liu Fu‐Tong , Gipson Ilene , Panjwani Noorjahan
Published: June 2021

Recent studies in our laboratory have shown that galectin‐3, a β‐galactoside binding lectin, mediates re‐epithelialization of corneal wounds. In an effort to characterize the molecular mechanism by which galectin‐3 stimulates the cell migration, the goals of the present study are to: (i) characterize the lectin‐induced migratory phenotypes and Rac1 activation in corneal epithelial cells in vitro, and (ii) determine whether carbohydrate‐based interactions between galectin‐3 and integrins play a role in the lectin‐induced lamellipodia formation in corneal epithelium. Specifically, exogenous galectin‐3 but not galectin‐1, and ‐8 induced formation of lamellipodia in human corneal epithelial cells in a β‐lactose‐inhibitable manner and increased Rac1 activity. α3β1, α6β1, α6β4 and & αv‐integrins and laminin‐5 were identified as major galectin‐3‐binding proteins in corneal epithelial cells by affinity chromatography of corneal epithelial cell lysates on a galectin‐3‐Sepharose column, followed by MALDI‐TOF MS analysis. Furthermore, preincubation of cells with anti‐α3 integrin function‐blocking antibody (P1B5) significantly inhibited the galectin‐3‐induced lamellipodia. These data suggest that galectin‐3 may influence re‐epithelialization of corneal wounds by initiating the formation of lamellipodia, at least in part, through its interaction with α3β1 integrins.
Modulation of the Nicotinic Acetylcholine Receptor Function by Cholesterol
Authors: Hoyo‐Rivera Natalie Del , Baez‐Pagan Carlos A. , Quesada Orestes , Lasalde‐Dominicci Jose A.
Published: June 2021

The nicotinic acetylcholine receptor (nAChR), located in the cell membranes of neurons and muscle cells, mediates the transmission of nerve impulses across cholinergic synapses. The nAChR is also found in the electric organs of electric rays (e.g. Torpedo californica). Cholesterol is a key lipid for maintaining the correct functionality of membrane proteins and has been found to alter the nAChR function. We were thus interested to probe the changes in the functionality of different nAChRs when expressed in cell membranes with modified cholesterol to phospholipid ratios (C/P). In this study, we examined the effect of increasing the C/P of Xenopus laevis oocytes expressing the neuronal α‐7, muscle‐type or Torpedo californica nAChRs in the functioning of the nAChR. Using the two‐electrode voltage clamp technique it was found that the neuronal ‐7 and Torpedo nAChRs are significantly more sensitive to small increases in C/P than the muscle‐type nAChR. This study clearly illustrates that a physiologically relevant increase in membrane cholesterol concentration renders a significant fraction of the neuronal ‐7 and Torpedo nAChRs “inactivated” whereas the muscle‐type nAChR tends to resist this functional inhibition. This work was supported by NIH grants RO1GM56371‐11, NCRR 1S0RR13705 and S06‐GM08102‐27 to JAL‐D. NDH‐R and CAB‐P were supported by the NIH‐MBRS RISE Grant R25GM61151.