bluuebunny/crossref_metadata_2025_split huggingface dataset:
The antioxidative detection of Prdx6 to cell membrane lipid peroxidation of lung type II epithelial cells by DPPP (693.3)
Authors: Wang Yan , Manevich Yefim , Fisher Aron 
Published: June 2021

Peroxiredoxin 6 (Prdx6) is a peroxidase antioxidant enzyme, we observed the protection of Prdx6 to cell membrane lipid peroxidation of lung alveolar epithelial typeIIcells (AEC II) by using a novel fluorescent probe Diphenyl‐1‐pyrenylphosphine (DPPP). Hydrogen peroxide (H2O2) can diffuse across cell membrane, so the dose of 500 µM H2O2 was used to AEC II exposure with labeled DPPP at 400nmol/m for 1 to2 h, the AECIIwere isolated from wild type (WT) and Prdx6 transgenic overexpressor (Tg) mice and observed under fluorescence microscopy and PTI spectrofluorometer. We found that AECIIwith H2O2 exposure induced time‐related fluorescence signal increasing in cell membrane under fluorescence microscopy, and the AECIIfrom Prdx6 Tg mice showed about 36% less cell membrane lipid peroxidation compared to WT group; under PTI spectrofluorometer study, the decreased cell membrane phospholipid peroxidation in Tg Prdx6 AECIIconsistently presented about 60% decreasing remarkably in 2h‐ H2O2 exposure and about 30% in 1h‐ H2O2 exposure compared to WT group. Thus, transgenic overexpression of Prdx6 in AECII defends from H2O2‐induced cell membrane lipid peroxidation.Grant Funding Source: Heart and Lung Society
The protective role of peroxidase 6 against peroxidative stress in pulmonary microvascular endothelial cells includes its ability to repair peroxidized cell membranes (693.4)
Authors: Li Haitao , Feinstein Sheldon , Fisher Aron 
Published: June 2021

Peroxiredoxin 6 (Prdx6) is known to play a critical role in antioxidant defense of lung and other organs. Our aim in this research was to investigate whether this protective function of Prdx6 includes an ability to repair peroxidized cell membranes. Mouse pulmonary microvascular endothelial cells (PMVECs) were treated with tert‐butyl hydroperoxide (tBOOH) and lipid peroxidation was evaluated by measurement of thiobarbituric acid reactive substances (TBARS). PMVECs from Pxdx6 null mice showed much higher levels of lipid peroxidation upon tBOOH (50‐300 μM) exposure compared with wild‐type (WT).To determine whether Prdx6 is necessary for repair, PMVECs were treated with tBOOH (300 μM) for 24 h and then evaluated at 0,2,4 and 6 h after removal of the oxidant. TBARS in WT PMVECs returned to the levels of unexposed control level in 2 h. On the contrary, TBARS in PMVECs from Pxdx6 null mice were unchanged from their elevated levels during 6 h observation after cessation of tBOOH exposure. These results indicate that Prdx6 plays a critical role in protecting PMVECs against peroxidative stress both by preventing lipid peroxidation during the oxidant exposure and through its ability to repair peroxidized cell membrane during the subsequent recovery period following removal of the oxidative stress.Grant Funding Source: Supported by 5T32HL007748‐20, RO1‐HL 102011, and PO1‐HL19737
Differential effect of apoptosis‐associated tyrosine kinase on K‐Cl cotransporter activity in C6 glioblastoma cells (694.2)
Authors: Gagnon Kenneth 
Published: June 2021

Expression of the potassium‐chloride (K‐Cl) cotransporter is linked to the growth and spread of gynecological cancers. Immunofluorescent localization of three K‐Cl cotransporter isoforms (KCC1, KCC3, and KCC4) to migrating extensions of rat glioblastoma (C6) cells suggests their potential role in the enhanced motility of these tumors. RT‐PCR amplification of mRNA from C6 cells validated the expression of kinases and phosphatases known to control cation‐chloride cotransporter activity. Xenopus laevis oocytes co‐injected with KCC3 and apoptosis‐associated tyrosine kinase (AATYK1) cRNA showed isosmotic K+ influx similar to hyposmotically‐activated K+ influx, whereas no change in isosmotic K+ influx was observed with co‐injection of KCC1 and AATYK1 cRNA. This differential effect of AATYK1 on two KCC isoforms prompted testing of the potential of AATYK1 as a novel target for affecting glioblastoma motility. Genistein, a general tyrosine kinase inhibitor, failed to reduce the hyposmotically‐stimulated K+ influx of C6 cells or inhibit their motility in a scratch wound assay indicating that the catalytic activity of the tyrosine kinase is not involved. Molecular identification of both serine/threonine kinase and phosphatase binding sites on AATYK1 suggests that it may scaffold other K‐Cl cotransporter regulatory proteins and thus represent a novel target for anti‐cancer therapeutics.Grant Funding Source: Supported by a Special Projects Grant from the University of Saskatchewan College of Medicine
Characteristics of collective cell migration in human keratinocytes at subatmospheric pressure (694.5)
Authors: Hsu Chih‐Chin , Chow Shu‐Er 
Published: June 2021

A negative pressure incubator incorporating the electric cell‐substrate impedance sensing (ECIS) technique has been built to quantify monolayer keratinocytes movement in serum free media at ambient pressure (AP) and at the subatmospheric pressure of 635 mmHg (SP). Monolayer cell motion at the two pressures was continuously recorded by ECIS in the frequency range of 22.5‐64 kHz. Cells in both pressures had similar viability. Greater migration distance was found in cells at SP. We underwent fluorescent staining and protein analysis for E‐cadherin, phosphorylated ‐catenin (p‐catenin), and acetylated ‐tubulin in cells at the both pressures serially at 0, 3, 6, 9, and 12 hours after wounding the monolayer cells. A significant decrease of cell‐substratum conductance (AP: SP= ~5 cm0.5: ~3 cm0.5) and increase of membrane capacitance (AP: SP= ~4F/cm2: ~8F/cm2) were observed early in the monolayer cell movement at SP. Increased cell‐substratum distance and membrane ruffling observed in scanning electron microscopic (EM) images may be respectively responsible for these measurements. Long and stable microtubules may also contribute to the cell deformation. In cells treated with SP, a progressive drop of intercellular resistance from 1.2 to 0.8 cm2 and a significant change were observed 6 hours after wound closure. The gradual loss of cell junctions associated with decreased adherent junction proteins was identified in serial fluorescent stains and transmission EM images in cells at SP. Characteristics of collective cell migration at SP include early release of cell‐substratum adhesion, quick formation of membrane ruffling, and an ensued decrease of cell junction protein expressions.Grant Funding Source: Chang Gung Medical Research Programs
P2X4 receptors mediate atheroprotective gene expression in an in vitro model of steady flow in vascular endothelial cells (696.6)
Authors: Sathanoori Ramasri , Olde Björn , Rosi Federica , Müller Christa , Erlinge David 
Published: June 2021

Vascular endothelial cells are in direct contact with blood flow and are exposed to fluid shear stress. The role of P2X4 receptors in shear stress‐mediated mechanotransduction is well documented. Studies have shown endothelial cells to release ATP in response to steady flow that in turn modulates cellular functions via P2 receptors in the vasculature. However, the molecular mechanisms governing ATP‐mediated P2 receptor signaling downstream of steady flow in endothelial cells remains largely unknown. Although steady flow‐induced KLF2 expression implicated in atheroprotection is well studied, whether ATP regulates KLF2 remains unanswered and is the aim of this study.Human umbilical vein endothelial cells (HUVECs) were either exposed to steady flow at 12 dynes/cm2 or to exogenous administration of a non‐hydrolysable analog of ATP (ATPγS) in the presence of the P2X4 antagonist (PSB‐12253) and P2X4 siRNA. We validated the in vitro orbital shaker model for steady flow showing cell alignment in the direction of flow and upregulation of KLF2 and NOS3. Further, addition of ATPγS to static cultures led to an increase in KLF2 mRNA as early as 3h. Exposing HUVECs to apyrase and PSB‐12253 reduced KLF2 and NOS3 expression under steady flow. Furthermore, the knockdown of P2X4 confirmed its role in regulating KLF2. These results suggest that ATP is involved in regulating KLF2 in part via the P2X4 receptor.
miR‐23b regulates CDK‐activating kinase complex through cyclin H repression to modulate endothelial transcription and growth under flow (696.7)
Authors: Wang Kuei‐Chun , Li Yi‐Shuan , Chien Shu 
Published: June 2021

The flow regulation of microRNAs (miRNAs) in endothelial cells (ECs) plays a significant role in vascular homeostasis and diseases. We used a combination of an in vitro cell perfusion system and in vivo studies on experimental animals to elucidate the molecular mechanism by which the pulsatile shear flow (PS)‐induced miR‐23b exerts anti‐proliferative effects on ECs. Our results demonstrated that PS induces the transcription factor KLF2 to promote miR‐23b biosynthesis; the increase in miR‐23b then represses cyclin H (CCNH) to impair the activity and integrity of CDK‐activating kinase complex (CAK). The inhibitory effect of miR‐23b on CAK further suppresses cell cycle progression via reducing the basal transcription and phosphorylation of CDK2/4. While PS regulates the miR‐23b/CAK pathway to exert anti‐proliferative effects on ECs, oscillatory shear flow (OS) has little effect on the miR‐23b/CAK pathway and does not cause EC growth arrest. Such flow pattern‐dependent phenomena are validated with an in vivo model on rat carotid artery. Local delivery of miR‐23b precursors to rat carotid artery significantly reduces the EC proliferative phenotype induced by flow disturbance. Our findings unveil a novel mechanism by which hemodynamic forces modulate endothelial homeostasis through miR‐23b and demonstrate the potential of developing miR‐23b‐based therapeutics for cardiovascular diseases.Grant Funding Source: Supported by NIH HL106579 and HL108375
Integration of background genome variation to dissect complex cardiovascular phenotypes (697.5)
Authors: Dwinell Melinda , Lazar Jozef , Shimoyama Mary 
Published: June 2021

The genetic underpinnings of cardiovascular (CV) disease are far more complex than originally imagined. Human genome wide association studies (GWAS) have identified candidate genes, but to date have predicted only a small fraction of the genetic basis. Some of the “missing heritability” is likely due to the interaction of multiple genes and alleles contributing to the disease. Phenotypic and genotypic characterization of two complete comsomic rat panels provides the foundation to analyze the influence of genetic background on CV traits. Phenotyping of PhysGen consomic strains, developed by introgressing Brown Norway (BN) chromosomes onto the Salt‐Sensitive (SS) or Fawn Hooded Hypertensive (FHH) backgrounds, identified consomic strains in introgressed BN chromosomes alter phenotypes on only one genomic background or under only one environmental condition. Introgression of BN chromosome 1 differentially regulates left ventricle infarct size following ischemia on SS and FHH backgrounds. HIF pathway genes on chromosome 1 are nearly identical between SS and FHH, although 12 genes on background chromosomes contain non‐synonymous SNPs (2 in SS only, 6 in FHH only, and 4 in both compared to BN), potentially altering the HIF pathway and CV phenotypes in response to chronic hypoxia. Additional in silico analysis, using gene, QTL and phenotype annotations, in conjunction with strain‐specific sequence variation, enables virtual variant profiles for complex phenotypes or disease pathways to be created for each consomic strain.Grant Funding Source: Supported by R01HL094271 and R01HL064541
Acute ingestion of dietary nitrate increases muscle blood flow via local vasodilation during exercise in young adults (705.9)
Authors: Richards Jennifer , Racine Matthew , Hearon Christopher , Patel Rakesh , Dinenno Frank 
Published: June 2021

Dietary nitrate (NO3‐) supplementation with beetroot juice (BR) can reduce the O2 cost of submaximal exercise and increase exercise tolerance. NO3‐ is converted to nitrite and can be reduced to the vasodilator nitric oxide amid a low O2 environment and BR is observed to increase hindlimb blood flow in rats during treadmill exercise. We tested the hypothesis that BR increases forearm blood flow (FBF) via local vasodilation during handgrip exercise in young adults (n=7; 24±2 years). FBF (Doppler ultrasound) and blood pressure (finometer) were measured at rest and during graded handgrip exercise at 5, 15, and 25% maximal voluntary contraction (MVC) lasting 4 minutes each. During 25% MVC exercise, systemic hypoxia (80%SaO2) began and exercise continued for 3 additional minutes. Subjects then ingested concentrated BR (12.6 mmol nitrate) and repeated the exercise bout 2h post consumption. Compared to control, BR significantly increased FBF at 15% (183±29 vs. 165±21 ml/min), 25% (331±40 vs.293±43 ml/min), and 25% + hypoxia (393 ±59 vs. 357±49 ml/min; all P<0.05), and this was due to increases in vascular conductance. Further, the increase in FBF from control was greatest at 25% MVC in normoxia and hypoxia (Δ~35‐40 ml/min) compared with 15% (Δ~18 ml/min) and 5% (Δ~5 ml/min). These findings may have important implications for aging and diseased populations that have impaired muscle perfusion and exercise intolerance.Grant Funding Source: Supported by HL095573
Influence of 5‐HT1A agonist administration associated with E2 replacement on neuroendocrine in ovariectomized rats (707.11)
Authors: Fonseca Fabricia , Mecawi André , Araujo Iracema , Monteiro Lívia , Pereira Gislaine , Rodrigues José , Reis Luís 
Published: June 2021

We intend to examine the effect of repeated use of an agonist of 5‐HT1A receptor in sodium appetitive response in ovariectomized (OVX) rats and investigate the effect of estrogen therapy. Female Wistar rats (n=6‐14) were ovariectomized and formed groups as follow: OVX and OVX + E2 (estradiol cypionate, 80µg/Kg, SC, 14 days) separated in groups of 8‐OH‐DPAT (5‐HT1A receptor agonist, 250µg/Kg, IP, 7 days) or vehicle (0.9% NaCl) in baseline and sodium depletion (furosemide 20mg/kg, SC) protocols. The animals were submitted to following analyses: (i) plasma Na+ (ii) radioimmunoassay to measure oxytocin (OT), vasopressin (AVP), angiotensin II (ANG II) plasma levels. The E2 increased Na+ (P<0.01) plasma levels in OVX‐E2‐8‐OH‐DPAT compared to the OVX‐E2‐OH‐DPAT group in sodium depletion condition. Plasma OT increased (P<0.001) in OVX‐E2‐8‐OH‐DPAT group compared to OVX‐E2 group in baseline condition. OT plasma levels also significantly increased in OVX‐E2 group compared to OVX rats (P<0.01). AVP plasma concentration increased in OVX‐E2‐DPAT group in relation to OVX‐E2 (P<0.001) group following volume depletion. Plasma concentration of ANG II decreased in OVX group compared to OVX‐E2 (P<0.01) group in basal condition. Plasma concentration of ANG II decreased in OVX group compared to OVX‐E2 (P<0.01) group in basal condition. Thus, these results suggest that repeated administration of 5‐HT1A agonist associated with E2 replacement differentially trigger mechanisms for neuroendocrine in electrolyte homeostasis.Grant Funding Source: FAPERJ, CNPq and CAPES
A module implemented in Excel® for predicting fluid requirements in complex thermal environments (707.7)
Authors: Gonzalez Richard , Kenefick Robert , Charkoudian Nisha , Cheuvront Samuel , Muza Stephen 
Published: June 2021

Military and public health reports have expressed a need for improved sweating rate prediction models (msw,pred) that calculate hourly and daily water needs. Using both meta‐analysis and experimental studies (150 volunteers and >1000 cases), we developed an energy balance module within the Microsoft Excel® spreadsheet package. The module incorporates previous legacy heat balance data and predicts msw,pred . The core model inputs environmental parameters, metabolic (Pandolf equation), clothing coefficients, and calculates net effective radiation fluxes, sensible and latent heat fluxes, and outputs evaporative heat loss to estimate fluid requirements. The altitude elevation (<3000 m) can be specified in the model. An output file estimating hourly and daily water requirements and the energy fluxes is generated. The main advantages of the module are: (1) it requires only a laptop/smartphone computation running standard Microsoft Windows®, enabling it to be used at a desktop or in the field; and (2), it can be adapted quickly to different environmental scenarios and meteorological data formats. Model calculations of msw,pred compared with measured data showed a predictive accuracy ranging from 95% for the best to 75% for the worst prediction in complex thermal environments with solar loads. The module is a useful tool allowing guidance in military, public health, occupational and sports medicine settingsGrant Funding Source: Supported by:Knowledge Preservation Program, USAMRMC
Hydration status when increasing daily fluid intake with various beverages (707.8)
Authors: Tucker Matthew , Ganio Matthew , Adams J. D. , Brown Lemuel , Ridings Christian , Burchfield Jenna , Robinson Blake , McDermott Jamie , Moyen Nicole , Schreiber Brett , Washington Tyrone , Bermudez Andrea , Bennett Meredith , Buyckx Maxime 
Published: June 2021

Relatively little is known about how hydration status is affected when increasing fluid ingestion with various beverages. Purpose: To investigate the hydration status of healthy, free‐living males when given varying volumes of different beverage types. Methods: 36 males (age 24±5 y; mass 75.1±11.4 kg) were randomly divided into 4 treatment groups for 2 weeks of testing: water only, water+cola, water+diet cola, or water+cola+diet cola+orange juice. Volumes of fluid were split evenly between beverages within each 24‐h treatment period. 24‐h fluid ingestion (including fluid in food) for weeks 1 and 2 was 35 and 40 ml/kg body mass, respectively. Serum osmolality (Osmserum), total body water (TBW) via bioelectrical impedance, 24‐h Osmurine and volume (Volurine) were analyzed after 24‐h of intervention. Results: Total beverage consumption independent of treatment, increased from week 1 to 2 (1721±384 to 2096±437 ml; P <.05). Independent of week there were no hydration differences between treatments (P >.05). Independent of treatment increasing fluid intake did not change TBW (43.7±5.5 vs 43.5±5.2 kg), Osmserum (292±4 vs 293±5 mOsm), Osmurine (599±221 vs 561±217 mOsm) or Volurine (1526±624 vs 1621±651 ml) (P >.05). Conclusion: Regardless of fluid volume or beverage type consumed, 24‐h hydration status did not differ. This suggests that fluid contained in various beverages is equally effective in hydrating the body.Grant Funding Source: Funded by The Coca‐Cola Company
Hypoxia induced hERG trafficking defect leads to apoptosis: involvement of NADPH oxidase generated ROS and Hsp90 (710.2)
Authors: Vaddi Damodara , Piao Lin , Khan Shakil , Wang Ning , Prabhakar Nanduri , Nanduri Jayasri 
Published: June 2021

The alpha subunit of the voltage gated human ether‐go‐go‐related (hERG) potassium channel regulates cell excitability in a broad range of cell lines. hERG channels are also expressed in a variety of cancer cells and control cell proliferation and apoptosis. Sustained hypoxia (SH) alters gating properties of hERG currents in neuroblastoma (SH‐SY5Y) cells. In the present study, we examined the molecular mechanisms underlying SH altered hERG currents in SH‐SY5Y cells. Patch‐clamp studies revealed that 4days of SH decreased hERG tail currents by 40‐50%. SH down regulated the surface expression of the fully glycosylated form (fg) of hERG protein with accumulation of the endoplasmic reticulum (cg) form. These effects were stimulus dependent and reversible. Accumulation of cg form by SH was due to impaired trafficking of hERG protein. Prolonged association with molecular chaperone Hsp90 resulting in complex oligomeric insoluble aggregates contribute to SH induced ER retention and defective trafficking. SH increased NADPHoxidase‐2 (Nox2)‐mediated ROS generation. Silencing Nox2 prevented SH‐induced degradation of fg and accumulation of cg forms. HERG downregulation by hypoxia resulted in reduced cell proliferation, altered cell cycle distribution and increase in apoptotic cells and all these effects were prevented by genetic silencing of Nox2.Grant Funding Source: HL‐90554
Pulmonary hypertension is present in a rodent model of bronchopulmonary dysplasia at 21 days but not 6 months of age (715.1)
Authors: Kleinertz Aaron , Conhaim Robert , Shuster Taylor , Mahan Sidney , Eldridge Marlowe , Bates Melissa 
Published: June 2021

In preterm infants, mechanical ventilation with supplemental O2 can cause bronchopulmonary dysplasia (BPD), characterized by dysmorphic pulmonary vasculature and simplified alveoli. Although vascular cross‐sectional area is diminished, contributing to right ventricular hypertension (RVH), it is unknown whether this improves with age. We hypothesized that RVH would continue into adulthood in a rat model of BPD. Rats were exposed to 21% or 95% O2 from birth to postnatal day 8. At 21 days or 6 months of age, a fluid‐filled catheter was placed in the RV to measure RV pressure and inject 2x108 4μm fluorescent spheres. At the conclusion of the experiment, lungs were harvested, air dried, and imaged using laser scanning confocal microscopy to investigate capillary density. At 21 days, rats exposed to 95% O2 demonstrated RVH (31±5 mmHg), which was absent in both 21 day controls (16±2 mmHg, p=0.028) and 6 month old rats exposed to 95% O2 (20±10 mmHg, p=0.014). Interestingly, the density and distribution of spheres was not different between 6 month old rats exposed to 95% or 21%, suggesting that the normalization of right heart pressure by 6 months is the result of improved alveolar capillary density. Our findings indicate that children and infants with BPD may be at high risk for RVH, but that this resolves with age.Grant Funding Source: Supported by NIH 5R01HL115061
14‐3‐3 Isoforms bind directly exon B of the 5’UTR of human surfactant protein A2 mRNA (717.5)
Authors: Noutsios Georgios , Ghatas Paul , Bennett Stephanie , Tsotakos Nikolaos , Floros Joanna 
Published: June 2021

Human surfactant protein A, an innate immunity molecule is encoded by two genes SFTPA1 (SP‐A1) and SFTPA2 (SP‐A2). The 5’ untranslated (5’UTR) splice variant of SP‐A2 (ABD), but not of SP‐A1 (AD), contains exon B (eB). eB is an enhancer for transcription and translation, and contains cis regulatory elements. Specific trans‐acting factors including 14‐3‐3, bind eB. The 14‐3‐3 protein family contains 7 isoforms some of which (ζ/δ, β/α, τ/θ, ε, γ, η) were found by mass spectroscopy to be present in eB electromobility assays (REMSA) (AJPLC MP 304:L722, 2013). Here, we investigated whether 14‐3‐3 binds directly to eB using three different approaches. i) REMSAs were performed, with either NCI‐H441 cell extracts or purified 14‐3‐3 isoforms (β, η, τ, ζ) expressed in E. coli, incubated with wild type eB‐, and random‐ RNAs. Formed complexes were analyzed with mass spectroscopy, ii) Surface Plasmon Resonance analysis (Biacore), and iii) eB RNA pull down assays. We found by REMSA that 14‐3‐3 isoforms β, τ and η but not ζ bind directly eB; these do not bind SP‐A1 (AD) that does not contain eB. Moreover, when isoform 14‐3‐3 η was tested with Biacore and pull down assays a direct binding to eB RNA was observed as with REMSA. In conclusion the 14‐3‐3 protein family affects differential regulation of SP‐A1 and SP‐A2 by binding directly to SP‐A2 mRNA.Grant Funding Source: Supported by NIH HL34788