bluuebunny/crossref_metadata_2025_split huggingface dataset:
Anti‐oxidative and anti‐inflammatory role of adipose stem cells in reversing pulmonary hypertension and associated cardiac remodeling (1090.9)
Authors: Rathinasabapathy Anandharajan , Shenoy Vinayak , Bruce Erin , Espejo Andrew , Nair Anand , Francis Joseph , Raizada Mohan , Katovich Michael 
Published: June 2021

Pulmonary Hypertension (PH) is a debilitating cardiopulmonary disorder, often resulting in right heart failure. Existing therapy does not curtail the rate of mortality. Here, the protective role of adipose stem cell (ADSC) in attenuating PH was studied. Monocrotaline (MCT) was injected into 8‐weeks old male SD rats to induce PH. Post 2 weeks of MCT challenge, when the RVSP (Right Ventricular Systolic Pressure) was significantly elevated, ADSC were administered. After 4 weeks of MCT induction, RVSP (30.09±1.51 vs 86.71±4.21 mmHg), & RV Hypertrophy (RVH; 0.23±0.01 vs 0.60±0.03) were elevated. ADSC therapy significantly reduced RVSP (55.94±7.35) and RVH (0.34±0.04). Video of echocardiography demonstrated that RV end‐diastolic area, (0.12±0.01 vs 0.33±0.03 cm2) and RV ejection fraction (61.62±0.93 vs 29.47±3.76 %) was significantly dysregulated in PH. Further, the pulmonary blood flow (0.88±0.04 vs 0.58±0.06 m/s) and AT/ET (0.37±0.01 vs 0.18±0.02) was altered by MCT treatment. ADSC reverted all of these cardiac remodeling events. RT‐PCR demonstrated that the markers of fibrosis (TGFβ), pro‐inflammatory cytokines (TLR4, TNFα, IL1β) and oxidative stress (NOX2) were up regulated 4 to 5 fold and anti‐inflammatory cytokine (IL10) was down regulated 6 fold in PH animals. ADSC therapy normalized all of these parameters. Collectively, our data demonstrate that the anti‐oxidant and anti‐inflammatory role of ADSC arrested PH and associated cardiac remodeling and suggest that ADSC intervention could be a potential therapeutic alternative for the treatment of PH.Grant Funding Source: HL056921 and HL102033
Proteomic profiling of quiescent and activated airway epithelial NKCC1 (1095.9)
Authors: Liedtke Carole , Litman Paul , Gokulrangan Giridharan 
Published: June 2021

Protein scaffolds at the amino‐ or carboxyl termini have specific roles in NKCC1 activation and/or membrane expression: Activation of epithelial NKCC1 transport activity also requires posttranslational modification (PTM). The goal of this study is to identify cargo proteins associated with the protein scaffolds and to assess PTM of NKCC1 using proteomic/nano LC‐MS/MS methodology for more accurate modeling of NKCC1 regulation. NKCC1 was immunoprecipitated from Calu‐3 epithelial cells after treatment to modify NKCC1 activity and subjected to 1D gel electrophoresis. Protein bands were excised for in‐gel trypsin digestion followed by LS‐MS/MS analysis. Phosphoenrichment of in‐gel digests was used to identify sites of phosphorylation of NKCC1. We identified proteins associated with plasma membrane, endocytosis, exocytosis, intracellular signaling, structural proteins, endoplasmic reticulum, cytosol, and ubiquitination. We found that activated (okadaic acid treatment) NKCC1 mass decreased and inactive (siSPAK treatment) NKCC1 mass increased. Two unique sites of ubiquitination of NKCC1 were identified. Treatment with hyperosmotic sucrose for 4 min induced robust phosphorylation of NKCC1 in the C‐terminus that decreased after 15 min. We conclude that targeted proteomic analysis using LS‐MS/MS will generate more detailed information about NKCC1 and its associated protein scaffolds.Grant Funding Source: Supported by a grant from the NIH, RO1‐HL‐58598, and by the Clinical and Translational Science Colla
Calcium signaling in the choroid plexus epithelium (1097.11)
Authors: Longden Thomas , Bonev Adrian , Nelson Mark 
Published: June 2021

Choroid plexus epithelial cells (ECs) express a diverse array of pumps and ion channels that facilitate the production of cerebrospinal fluid in the brain ventricles. Our understanding of the molecular players involved in EC function is advancing, but to date little is known about EC calcium (Ca2+) signaling. We thus sought to determine the nature of Ca2+ signaling in native preparations of rat choroid plexus loaded with the Ca2+ indicator Fluo‐4, using high speed confocal imaging of intracellular Ca2+ events.We observed three major forms of Ca2+ events in ECs: 1) Uni‐ or multicellular propagating waves. 2) Brief, localized, high amplitude events reminiscent of IP3‐mediated Ca2+ release in endothelial cells, which we term Ca2+ plexars.1 3) Low amplitude quantal events reminiscent of TRPV4 sparklets in endothelial cells.2Ca2+ activity in ECs was dependent on extracellular Ca2+, as its removal abolished all signals. Similarly, blockade of the sarco/endoplasmic reticulum ATPase terminated EC Ca2+ signaling. At some sites, Ca2+ plexars initiated sparklet activity.We conclude that Ca2+ signaling in choroid plexus ECs is dependent on extracellular Ca2+ entry, which causes the release of Ca2+ from intracellular stores. Stores‐mediated plexar events also couple to sparklet events and may lead to regenerative Ca2+ release.1. Ledoux et al. (2008) PNAS 9: 1397‐1403 2. Sonkusare et al. (2012) Science 336 6081: 597‐601Grant Funding Source: Supported by National Institutes of Health and the American Heart Association.
Effect of resveratrol on the erectile machinery is due to the underlying mechanisms of the alpha 1‐adrenoceptor activation (1097.2)
Authors: Restini Carolina , Domingos Aline , Pereira Bruna , Vieira David 
Published: June 2021

The anococcygeus muscle is part of the erectile machinery in male rodents whose innervation is mainly due the autonomic system. Activation of alpha1‐adreneceptor triggers the production of reactive oxygen species (ROS). Resveratrol (RESV) is an antioxidant compound. Hypothesis: RESV may reduce the contractile responses stimulated by alpha‐1‐adrenergic agonist, phenylephrine (PE). It was evaluated the effect of RESV on the contractile response to PE on the rat anococcygeus muscle. Male Wistar rats (n=5‐7) were killed by cervical dislocation (Ethics Committee Approval: 019/2012), the anococcygeus muscle was isolated and suspended in an organ chamber bath to measure isometric contractions. Preparations were stimulated with PE (1 nmol/L to 100 mmol/L) before and after 20 min incubation with RESV or RESV plus different drugs: prazosin (Pz: 10‐8 mol/L); 2‐APB (10‐4 mol/L); GF109203X (0.5x10‐5mol/L) or atorvastatin (ATV: 10‐4mol/L). Analysis: Student paired t‐test or one‐way ANOVA, Newman‐Keuls post‐test. RESV (10‐4 mol/L) reduced maximum contraction (MC) stimulated with PE (to 69.3±1.1%). Pz (alpha1‐adrenoceptor antagonist) + RESV reduced this MC (to 55.8±4.7%, p<0.0001). 2‐APB (IP3 receptor antagonist) was effective on reducing MC (to 28.8±1.19, p<0,001). RESV+2‐APB also reduced ME (to 5.3±1.4%, p<0.0001). pD2 values were reduced under RESV+2‐APB incubation. Incubation with GF109203X (PKC inhibitor) was effective on reducing MC (to 40.5±7.3%, p<0.0001). pD2 values were also altered after GF109203X incubation. ATV (NADPH oxidase inhibitor) +RESV reduced MC (56.9±7.7%, p<0.0001). The signaling triggered through stimulation of PE on the alpha 1‐adrenoceptors are involved in the production of free radicals, which were sensitive to RESV.Grant Funding Source: Supported by CNPq/PIBIC, UNAERP
Inhibition of Rhbg‐mediated NH3 and NH4+ transport by DIDS (1098.2)
Authors: Caner Tolga , Abdulnour‐Nakhoul Solange , Brown Karen , Hamm L. , Nakhoul Nazih 
Published: June 2021

Rhbg is a non‐erythroid Rh glycoprotein expressed at the basolateral membrane of renal collecting duct. In previous studies, we demonstrated that Rhbg transported both NH4+ and NH3. As such, the Rh transporter is unique in being able to transport both the ionic and the gaseous components of ammonia. Other studies have shown that DIDS inhibited CO2 transport by AQP1. This suggested that DIDS might also inhibit transport of other gases such as NH3 by Rh proteins. This study was conducted to test this hypothesis. To do so we used ion‐selective microelectrodes and two‐electrode voltage clamp to measure changes in surface pH (pHs) and whole cell currents (I) induced by NH3/NH4+. All experiments were conducted on Xenopus oocytes expressing Rhbg. Our results indicate that in oocytes expressing Rhbg, exposure to 5mM NH4Cl caused a decrease in pHs by 0.85±0.12 and an inward current of ‐58±10 nA. The decrease in pHs is caused by NH3 influx whereas the inward current is due to electrogenic NH4+ influx. In the presence of DIDS, NH4Cl caused a significantly smaller decrease in pHs (0.57±0.07) and current (‐29±5 nA). Similarly, exposing Rhbg‐oocytes to 5mM MA/MA+ (a substitute to NH3/NH4+) caused a decrease in pHs of 0.27±0.04 and an inward current of ‐29±6 nA. In the presence of DIDS, the MA/MA+ induced changes in pHs (0.17±0.03) and ΔI (‐11±2 nA) were also inhibited. DIDS had no effect on NH3/NH4+ transport in H2O‐injected oocytes.Our data indicate that 1) DIDS inhibits transport of NH3 & MA by Rhbg without affecting endogenous NH3 & MA transport. 2) DIDS also inhibits the electrogenic transport of NH4+ & MA+ by Rhbg. 3) This is the first report to show inhibition of both gas (NH3) and ionic (NH4+) transport by Rhbg.Grant Funding Source: Supported by VA Merit grant
Suppression of physical activity and non‐resting energy expenditure during calorie restriction: role in individual differences in adaptive thermogenesis (1101.8)
Authors: Mukherjee Sromona , Demuth Emily , Spriggs Addison , Britton Steven , Koch Lauren , Novak Colleen 
Published: June 2021

Weight loss requires negative energy balance, which can induce adaptive thermogenesis‐the reduction of energy expenditure (EE) beyond that accounted for by the weight lost. Adaptive thermogenesis varies between individuals. Here, we measured EE and physical activity (PA) before and after 21 days of 50% calorie restriction (CR) in female rats with lean and obesity‐prone phenotypes‐rats strains selectively bred for high and low intrinsic aerobic capacity (HCR and LCR, respectively). Use of female rats minimized potentially confounding effects of group differences in body size. We found that CR decreased EE more than was predicted by loss of weight and lean mass, demonstrating adaptive thermogenesis. Both groups showed similar suppression in resting EE (HCR, 39%; LCR, 34%). The CR‐induced suppression in non‐resting EE, which includes activity EE, was significantly greater in HCR (37%) than in LCR (24%). CR also significantly suppressed PA in HCR but not LCR, but PA was still higher in HCR than in LCR even after CR. Weight loss did not differ between groups. These results suggest that individual differences in CR‐induced adaptive thermogenesis may be accounted for by variation in aerobic capacity. Moreover, it is likely activity EE, not resting or basal metabolism, accounts for individual variation in adaptive thermogenesis.Grant Funding Source: Supported by NIH R01NS055859 and NIH R15DK097644 to CMN
Instrument‐assisted soft tissue manipulation has no effect on the alpha7beta1 integrin pathway (1102.1)
Authors: Graham Zachary , Vardiman John , Siedlik Jake , Deckert Jake , Gallagher Philip 
Published: June 2021

Instrument assisted soft tissue mobilization (IASTM) is a manual therapy technique used by clinicians to treat soft tissue injuries or aid in recovery. However, the mechanism of IASTM has yet to elucidated. A potential mechanism that IASTM may work is through mechanoreceptors of the sarcolemma. The alpha7beta1 integrin is a mechanoreceptor that relays membrane stretch signals to the cytoplasm through focal adhesion kinase (FAK), which can then relay the signal through cSrc and ERK1/2. In this study IASTM was administered using a standardized protocol on the gastrocnemius of 11 male subjects. Muscle biopsies were taken pre‐treatment and 24, 48, and 72 hours post‐treatment and analyzed via Western immunoblotting. We saw no change in the expression of the alpha7beta1 integrin or changes in expression of total or phosphorylated FAK, cSrc or ERK1/2 at any point of the study. We conclude that IASTM has no effect on the alpha7beta1 signaling pathway at 24‐72 hours post‐treatment.Grant Funding Source: Supported by a General Research Grant from the University of Kansas awarded to J.P. Vardiman
The effects of KATP channels on skeletal muscle fatigue and recovery are dependent on muscle stimulus frequency and fibre type (1102.9)
Authors: Charter Mackenzie , Murrant Coral 
Published: June 2021

ATP sensitive potassium (KATP) channels are hypothesized to protect against ATP depletion during maximal muscle contraction by decreasing force production, however their function in submaximal contractions and recovery is unknown. We sought to test whether KATP channels altered the force of contraction during fatigue and recovery of mouse slow (soleus; SOL) and fast (extensor digitorum longus; EDL) twitch muscle, in vitro, at maximal and submaximal stimulus frequencies. We fatigued SOL (60Hz, 60 contractions per minute (CPM) and 20Hz, 60CPM) and EDL (100Hz, 60CPM and 40Hz, 60CPM) for 5 minutes in the absence or presence of a KATP channel inhibitor (10‐5M glibenclamide; GLIB) or a KATP channel opener (10‐5M pinacidil; PIN) and then observed recovery (SOL: 60Hz 0.6 CPM and 20Hz, 0.6CPM; EDL:100Hz 0.6CPM and 40Hz 0.6 CPM). GLIB had no effect on fatigue or recovery of SOL or EDL at maximum stimulus frequencies but significantly attenuated force in SOL (12.2%) and EDL (6.9%) at submaximal frequencies. GLIB significantly enhanced force during recovery of SOL by 22.5% at submaximal stimulus frequencies. PIN did not affect fatigue of SOL or EDL at maximal stimulus frequencies or the recovery of SOL but significantly attenuated force of EDL during recovery by 8.9%. PIN had no effect on fatigue or recovery during submaximal contractions. Thus, the effect of KATP channels is dependent on both fibre type and stimulus frequency.Grant Funding Source: Supported by NSERC, Canada
Aerobic training prevents the impaired vascular reactivity of fructose‐treated rats (1106.22)
Authors: Medeiros Renata , Gaique Thaiane , Bernardes Thais , Motta Nadia , Brito Fernanda , Bertoldi Julia , Santos Caroline , Oliveira Karen , Nóbrega Antonio 
Published: June 2021

Fructose intake is associated with cardiometabolic disorders, but few studies investigated its role on vascular reactivity and the potential effects of exercise training. Thus, we assessed the effect of fructose intake on vascular reactivity in trained rats. Therefore male Wistar rats (n=7/group) received tap water (control group, C) or Fructose 10% (F group) ad libitum for two weeks. Then, rats underwent treadmill exercise for eight weeks (FT group). Caloric intake was higher in fructose groups compared with C group (P<0.01), although we found no change in body weight and in lipid profile, except the higher HDL in FT group (P<0.01). Training increased serum NO (C=296.5±42.8; F=325.8±51.6; FT=718.4±151.2; P<0.01). Regarding body composition, F increased body fat content compared with C and FT groups (P=0.03), whereas the FT group showed higher protein content (P<0.01). Finally, vascular vasoconstrictor reactivity was increased by fructose since F group required lower concentrations of phenylephrine to promote 50% of the maximum contractile effect when compared to C and FT groups (C=1.31‐6±2.4‐7; F=2.43‐7±6.5‐8; FT= 7.99‐7±1.5‐7 M; P<0,01). Vasodilator response to acetylcholine was also impaired in F group and was preserved by exercise training (C= 3.13‐7±1.9‐7; F=3.64‐6±9.6‐7; FT=3.13‐7±1.9‐7M; P<0.01). Thus, aerobic exercise prevented fructose‐impaired vascular reactivity in rats.Grant Funding Source: CNPq, FAPERJ, CAPES, FINEP
Glucolipotoxicity‐induced p21 suppresses pro‐survival signaling in pancreatic β cells (1108.9)
Authors: Hernandez Angelina , Colvin E. , Fueger Patrick 
Published: June 2021

Type 2 diabetes manifests from peripheral insulin resistance and a loss of functional β cell mass due to decreased β cell function, survival, or proliferation. The glucolipotoxic environment in the diabetic milieu activates a stress response in β cells, resulting in cell death and decreased survival through various mechanisms. One key player implicated in β cell dysfunction is FOXO‐1, a transcription factor phosphorylated and inhibited by the pro‐survival kinase Akt. Interestingly, palmitate in combination with high glucose induced expression of p21, a cell cycle inhibitor that activates the intrinsic apoptotic pathway and reduces Akt cell survival signaling. The molecular mechanism by which p21 activates apoptosis in β cells during metabolic stress is still unknown. We hypothesized that p21‐mediated suppression of Akt leads to FOXO‐1 activation, ultimately resulting in the expression of pro‐apoptotic factors. To investigate the molecular events that p21 acts through to increase death and reduce cell survival, we adenovirally overexpressed p21 in isolated islets and 832/13 cells. p21 overexpression increased phosphorylation of the stress‐activated protein kinase JNK. In contrast, FOXO‐1 phosphorylation was suppressed, indicating activation of FOXO‐1. We are actively investigating whether a constitutively‐active Akt will reverse this p21‐mediated phenomena of FOXO‐1 activation. Therefore, we propose that p21‐mediated inactivation of Akt leads to cell death via JNK and FOXO1 activation. Our findings suggest a molecular mechanism by which metabolic stress‐ or glucolipotoxicity‐induced p21 expression leads to β cell death‐a mechanism that could potentially be targeted to protect β cell mass.Grant Funding Source: Supported by NIH grant DK078732 and William Townsend Porter Fellowship from APS
A long form of the alpha subunit of the human epithelial Na+ channel forms hyperactivated channels (1109.12)
Authors: Berman Jonathan , Brand Cristin , Awayda Mouhamed 
Published: June 2021

The Alpha subunit of ENaC is critical to the ability to form a channel. Alpha‐like subunits have been described and have been shown to affect channel activity and/or regulation. A new isoform of the alpha subunit containing an extended N‐terminus resulting in a 728 a.a. subunit was recently identified. We examined the electrophysiological properties of channels formed by this isoform. Channels formed by α728βγ exhibited much higher activity than those formed by α669βγ in the Xenopus oocyte expression system. These differences were much larger under conditions which elevated the intracellular [Na+]. α728βγ also exhibited reduced current/voltage rectification. These differences occurred despite similar single channel conductances (~5.5 pS). The role of intracellular Na+ and Na+ self‐inhibition in these differences was examined. We find that α728βγ sustained much higher activity under elevated [Na+] than α669βγ and that intracellular and acute increase of [Na+] by injection of Na2SO4 led to further increase of conductance. α728βγ channels were further activated by extracellular cleavage by trypsin or subtilisin; with 2 notable differences: 1) fold activation was smaller than that observed with α669βγ, and 2) the large baseline activity of α728βγ channels was observed in the absence of detectable cleavage at the plasma membrane. These data indicate that α728 forms the structural basis of an ENaC with differing activity and regulation than α669.Grant Funding Source: National Institute of Diabetes and Digestive and Kidney Diseases Grant DK‐55626
Glial cell line‐derived neurotrophic factor (GDNF) promotes barrier maturation and wound healing in intestinal epithelial cells in vitro (1111.1)
Authors: Meir Michael , Bergauer Lisa , Flemming Sven , Germer Christoph‐Thomas , Schlegel Nicolas 
Published: June 2021

Recent data suggest that neurotrophic factors that derive from the enteric nervous system are involved in intestinal epithelial barrier regulation. In this context the glial cell line‐derived neurotrophic factor (GDNF) was shown to affect gut barrier properties in vivo directly or indirectly by yet undefined processes. Here we investigated whether GDNF directly affects intestinal epithelial barrier functions. Permeability (PE) was estimated by measurements of 4 kD FITC‐dextran flux across differentiated and across immature Caco2 monolayers. Recombinant human GDNF (100 ng/ml) was applied for 24h to test possible effects on epithelial barrier functions. GDNF application resulted in significant epithelial barrier stabilisation only in Caco monolayers with immature barrier functions. This effect was paralleled by a more linearized and augmented staining pattern of tight junction proteins claudin1 and occludin at the cell borders. Barrier maturation by GDNF correlated with augmented cAMP‐levels in immature Caco2 cells. Moreover, wound healing assays showed a significantly faster closure of the wounded areas following application of GDNF.These data show that GDNF directly affects intestinal epithelial cells and promotes epithelial wound healing and barrier maturation. This effect may be mediated by augmented cAMP levels following GDNF application.Grant Funding Source: Supported by DFG
MLCK regulation of claudin 3 expression contributes to IL1β mediated weakening of colonic epithelial tight junctions (1113.1)
Authors: Haines Ricci , Beard Richard , Yuan Sarah , Wu Mack 
Published: June 2021

Interleukin 1β (IL1β) is an inflammatory cytokine well recognized for its role in autoinflammatory diseases including multiple sclerosis and ulcerative colitis, both associated with increased endothelial and epithelial barrier dysfunction respectively. Epithelial barrier integrity is maintained, in part, by expression of tight junction proteins such as occludin, zonula occludens, and claudin family of proteins. The major claudins involved in colonic epithelial barrier function are claudin 2, claudin 3, and claudin 4. In addition to structural components of tight junctions, kinases play a role in mediating signaling events that can affect gene expression. Recently, myosin light chain kinase (MLCK) has been shown to play a role in IL1β mediated abrogation of epithelial tight junctions, however the manner in which MLCK contributes to this dysfunction is not known. Our studies show that IL1β downregulated claudin 3, a sealing claudin, in young adult mouse epithelial (YAMC) and CACO 2 cells. In addition, we show that the MLCK inhibitor ML7 attenuated IL1β induced claudin 3 downregulation, and improved barrier function as measured by electric cell‐substrate impedance sensing (ECIS). Moreover, the role of MLCK mediated downregulation of claudin 3 was further characterized by immunocytochemistry. These results suggest that MLCK plays a critical gene regulatory role in addition to its known place in epithelial cell contractility in response to inflammation.Grant Funding Source: Supported by VA Merit Review 5101BX000799, the NIH grants HL96640, HL61507 and HL70752