bluuebunny/crossref_metadata_2025_split huggingface dataset:
LUTEOLIN REDUCES CARDIAC DYSFUNCTIONS AND MITOCHONDRIAL OXIDATIVE STRESS IN STREPTOZOTOCIN‐INDUCED DIABETIC RATS
Authors: Su Fang , Liu Shu‐Qin , Chen Ying , Chen Fang‐Xia , Wang Hui‐Ping , Xia Qiang 
Published: June 2021

AimTo investigate the effects of luteolin on cardiac functions and mitochondrial oxidative stress in streptozotocin (STZ)‐induced diabetic rats.MethodsMale Sprague‐Dawley rats were randomly divided into a normal control group, a luteolin control group, a diabetic group, and diabetic groups orally administered with a low dose (10 mg/kg/d) or a high dose of luteolin (100 mg/kg/d) for eight weeks. The body weight, blood glucose, cardiac functions, left ventricular weight, myocardial collagen, and reactive oxygen species (ROS) levels were assayed. The cardiac mitochondrial ROS level, superoxide dismutase (SOD) activity and the mitochondrial swelling were measured.ResultsTreatment with luteolin had no effect on the blood glucose but reduced the losing of body weight in diabetic rats. High dose of luteolin markedly reduced the ratio of ventricular weight and body weight, increased the left ventricular develop pressure, and decreased the left ventricular end diastolic pressure in diabetic rats. The myocardial levels of ROS and collagen, the cardiac mitochondrial ROS level, and the mitochondrial swelling in diabetic rats were all markedly reduced by high dose of luteolin. Furthermore, high dose of luteolin significantly increased the mitochondrial SOD activity in diabetic rat hearts.ConclusionTreatment with luteolin for 8 weeks markedly improves the cardiac function, which may be related to reducing mitochondrial oxidative stress and mitochondrial swelling, in diabetic rats.This work was supported by grants from the National Natural Science Foundation of Zhejiang Province ( Y206179 ).
Isolation of microvascular endothelial cells from tissues using counterflow elutriation
Authors: James Megan L. , Brock Robert W. 
Published: June 2021

The purpose of this study was to optimize the isolation of pure and viable microvascular endothelial cell (EC) populations from a variety of murine tissues. While several methods are reported in the literature, many of these are lengthy and result in low yields. Although counterflow elutriation has been used to isolate a variety of cell types, we are unaware of its use for the isolation of ECs from tissues other than from the liver. Therefore, we proposed to evaluate the conditions necessary for separation of ECs from various tissues. Previously described procedures for isolating liver sinusoidal ECs via counterflow elutriation were optimized using our equipment. Additionally, methods for isolating ECs from aortae and kidney were developed and optimized. All cells were isolated from mice overexpressing endothelial‐specific GFP (Tg(TIE2GFP)287Sato/J). Flow cytometry was used to determine purity of isolated EC populations using staining of GFP, CD31 and DiI(3,3′‐dioctadecylindocarbocyanine)‐labeled acetylated LDL. The methods described herein result in EC populations of at least 98% purity. These cells have been successfully used for downstream applications, including molecular biological assays and cell culture. Counterflow elutriation can be employed to obtain high‐yield, high‐purity EC populations from a variety of murine tissues.This work was funded, in part, by NHLBI T32 HL090610.
Replicative senescence may be a suitable model for assessing in vivo endothelial cell oxidative stress and inflammation with aging in humans
Authors: Rippe Catarina , Blimline Mark , Nelson Keri , Seals Douglas R 
Published: June 2021

Oxidative stress and inflammation develop in the vascular endothelium with aging in rodents and humans and cause dysfunction. In vitro culture of endothelial cells to senescence may be a useful model of endothelial aging in humans, but this has not been established. Human aortic endothelial cells (HAEC) were analyzed by western blotting at early (passage 7–10) and senescent (passage 22–24) stages and compared with endothelial cells obtained in vivo from antecubital veins of healthy young (18–35 yr, n=26) and older (55–71 yr, n=26) adult males and analyzed using quantitative immunofluorescent staining. Nitrotyrosine, a marker of oxidative stress, was increased (P<0.05) with age both in vivo (1.6±0.2‐fold) and in vitro (1.9±0.1‐fold). The oxidant enzyme NADPH oxidase increased with age in vivo (1.6±0.1‐fold, P<0.05) and tended to increase in vitro (1.5±0.1‐fold, P=0.14). The antioxidant enzymes catalase and copper zinc superoxide dismutase (SOD) did not differ with age either in vivo or in vitro, whereas manganese SOD was selectively increased in senescent HAECs (1.6‐fold, P<0.05). The inflammatory proteins tumor necrosis factor α, nuclear factor κ B and monocyte chemoattractant protein‐1 increased (all P<0.05) with age both in vivo (1.6±0.2‐, 1.6±0.2‐, 1.4±0.2‐fold) and in vitro (2.4±0.5‐, 1.7±0.3‐, 1.6±0.1‐fold). In vitro culture of vascular EC to senescence may be a useful model of in vivo EC aging in humans for assessing oxidative stress and protein expression of oxidant and antioxidant enzymes and inflammatory molecules.NIH AG013038 and the Swedish Medical Research Council
Caveolin‐1 negatively regulates a metalloprotease‐dependent EGF receptor transactivation by angiotensin II
Authors: Takaguri Akira , Shirai Heigoro , Hinoki Akinari , Rizzo Victor , Eguchi Satoru 
Published: June 2021

ObjectiveThe EGF receptor (EGFR) transactivation pathway via an ADAM17‐dependent ligand shedding is the key signaling step by which angiotensin II (AngII) induces vascular smooth muscle cell (VSMC) remodeling. However, the molecular mechanism of the ADAM17 regulation remains largely unclear. We hypothesized that caveolin‐1 (Cav1), the major structural protein of a membrane microdomain, caveolae, is involved in the regulation.Methods & ResultsPhosphorylation and expression of the signaling proteins were evaluated in VSMCs prepared from rat aorta. Infection of adenovirus encoding Cav1 markedly inhibited AngII‐induced EGFR ligand shedding, EGFR transactivation, hypertrophy and migration, but not intracellular Ca2+ elevation. Interestingly, ADAM17 localized in Cav1‐rich lipid raft fractions of VSMCs, and that AngII induced a fraction shift of ADAM 17 from low to high density in the fractions. In addition, methyl‐beta‐cyclodextrin, a reagent to disrupt the caveolae structure, stimulates the EGFR ligand shedding and subsequent EGFR activation in VSMCs.ConclusionCaveolae provides an indispensable base environment for activation of AngII signal transduction linked to vascular remodeling. Moreover, our data indicate that dissociation of Cav1 from ADAM17 is a crucial mechanism by which AngII induces ADAM17 activation.
Atorvastatin Attenuates Sox9 Joint and Cardiac Calcification in ApoE null mice
Authors: Park Jeffrey , Flores Amy , Basu Nivedita , Krueger Bryan , Rajamannan Nalini M. 
Published: June 2021

BackgroundDegenerative joint disease is responsible for high morbidity in patients. The mechanisms of osteoarthritis and cardiac calcification are under investigation. We hypothesize that lipids may play a role via chondrogenesis.MethodsWe tested lipids with and without statins in the ApoE mouse to determine the Sox9 expression in the joints and valves. ApoE−/− mice (n=60). Group I (n=20) normal diet, Group II (n=20) 0.25% chol diet (w/w), and Group III (n=20) 0.25% (w/w) chol diet+atorv for the development of abnormal matrix synthesis in the joints over 24 weeks. The aortic valve (AVA) was examined for Bone formation and was assessed by microCT. Aortic Valve Disease was assessed by echo.ResultsThe chol diet induced bone in the AV and thickened knee joints with an increase in Sox9 and Cyclin as compared to the Control(>3‐fold(p<05). Atorvastatin attenuated all markers. Echo demonstrated a mild increase in the peak jet velocity. Masson Trichrome demonstrated a marked increase in the matrix synthesis.ConclusionWe demonstrated that chondrogenic mediated bone formation in the hypercholesterolemic aortic valves and abnormal matrix synthesis in the knee joints is attenuated with atorvastatin.
Life‐long caloric restriction confers pronounced AMPK‐dependent cardioprotection
Authors: Edwards Andrew G , Donato Anthony J , Lesniewski Lisa A , Gioscia Rachel A , Seals Douglas R , Moore Russell L 
Published: June 2021

AIMSTo determine whether myocardial resistance to ischemia/reperfusion (I/R) injury is enhanced in the senescent heart after life‐long caloric restriction, and whether AMP‐activated protein kinase (AMPK) plays a role in that phenotype.METHODS AND RESULTSHearts from ad libitum fed (AL) and life‐long calorically restricted (LCR, 40% restriction) B6D2F1 mice were examined by echocardiography, and 25/90 minute global I/R. AMPK inhibition (AraA) was applied to both groups (AL+AraA, LCR+AraA) prior to I/R. LCR hearts were smaller (AL, 270 ± 7 mg vs. LCR, 185 ± 5 mg, p < 0.001), and had thinner posterior walls (PWTd: AL, 1.07 ± 0.05 mm vs. LCR, 0.84 ± 0.05 mm, p < 0.05), but similar in vivo luminal dimension (LVIDd; LVIDs: AL, 3.8 ± 0.14; 2.0 ± 0.19 mm vs. LCR, 3.5 ± 0.18; 1.8 ± 0.17 mm, both p > 0.2). LCR hearts were protected from infarction (Infarct Size: AL, 28 ± 4 % vs. LCR, 10 ± 1 %, p < 0.01) and post‐ischemic functional deficit (LVDP recovery: AL, 65 ± 8 % vs. LCR, 93 ± 7 %, p < 0.01), and both effects were abolished by AraA (Infarct size: LCR+AraA, 22 ± 4 %; LVDP recovery: LCR+AraA, 82 ± 9 %, both p vs. AL > 0.1). LCR dramatically increased AMPKα phosphorylation (AL, 1.30 ± 0.03 vs. LCR, 1.55 ± 0.03, ratio units, p < 0.001), but not expression (AL, 1.21 ± 0.07 vs. LCR, 1.19 ± 0.01, ratio units, p = 0.8).CONCLUSIONSLife‐long caloric restriction induces powerful AMPK‐dependent I/R resistance, and involves elevated AMPKα phosphorylation.
Effects of exercise training on cardiac mitochondrial apoptosis in ovariectomized rats
Authors: Hsu Chih‐Chao , Ou Hsiu‐Chung , Lee Shin‐Da 
Published: June 2021

Cardiac apoptosis is found in ovariectomized rats; however, the effects of exercise training on cardiac apoptosis is still unclear in menopausal or bilateral oophorectomied women was available. The purpose of this study was to evaluate the therapeutic effects of exercise training on cardiac mitochondrial apoptotic pathways.MethodsEight sham‐operated rats and eight ovariectomized rats at 5–6 months of age were served as negative (sham) and positive (OVX) control. Additional 8 ovariectomized rats underwent treadmill running exercise 1 hour daily for 3 month. After exercise training or sedentary status, the excised hearts were measured by positive TUNEL assays, Hematoxylin‐eosin staining and Western Blotting.ResultsExercise training significantly prevented the increasing of whole heart weight and OVX‐induced cardiac cell apoptosis. In addition, exercise training decrease the OVX‐induced tBid, Bad, Bax, Bak, cytochrome c overexpression, and caspase‐9 and caspase‐3 activation.ConclusionsExercise training suppressed ovariectomy‐induced cardiac apoptosis through inhibiting mitochondrial apoptotic pathways in rat models. The findings may provide one of possible therapeutic approaches for preventing cardiac apoptosis in menopausal or bilateral oophorectomied women.
Ion transport homeostasis in Alpha‐1 AMPK KO mice
Authors: Lazo Yoskaly , Baile Goretti , Martínez Laura , Bernal Marisa , Viollet Benoit , Gimenez Ignacio 
Published: June 2021

AMP‐activated kinase (AMPK) regulates cell energy balance by modulating ATP synthesis and consumption. In epithelia, AMPK interacts with and regulate ion transport systems, including CFTR, ENaC and NKCC. We studied renal function and measured blood pressure in mice with genetic deletion of the alpha‐1 catalytic subunit of AMPK (a1KO mice; C57Bl6J background). When fed a standard diet (0.3%NaCl) a1KO mice showed mild antidiuresis (WT=0.053 vs. KO=0.038 mL/d/g bw). No significant differences in glomerular filtration rate, sodium or potassium handling were found. When fed low Na+ (0.02%) or high salt (4%NaCl) diets, a1KO mice achieved ion and water balance. Lack of aversion for the high salt diet in a1KO mice suggested increased sodium appetite. Aldosterone urinary excretion was consistently higher in a1KO mice (WT=0.028 vs. KO=0.037 ng/d/g bw). Telemetric 24h blood pressure records showed significantly lower blood pressure in a1KO mice, especially nocturnal diastolic pressure (WT=93.5 vs. KO=86.1 mmHg). We found that a1KO mice lose water and sodium in their feces compared to wild‐type mice. In the kidney, we observed up‐regulation of the alpha‐2 catalytic subunit, which may provide functional compensation for the loss of alpha‐1 activity. Thus, kidneys from a1KO mice apparently have normal function and may be actually compensating for extra renal water and ion losses.MICIIN Spain BFU2007/62119
Differential regulation of NaPi2a and NaPi2c by parathyroid hormone
Authors: Okamura Kayo , Giral Hector , Caldas Yupanqui , Sutherland Eileen , Millard Andrew , Levi Moshe , Blaine Judith 
Published: June 2021

Parathyroid hormone (PTH) is a critical regulator of renal phosphate (Pi) homeostasis. In the kidney Pi reabsorption is regulated by the sodium phosphate cotransporters NaPi2a and NaPi2c. It has previously been shown that PTH‐induced removal of NaPi2c from the renal proximal tubule brush border membrane (BBM) is approximately 3 times slower than that of NaPi2a. The reason for this is unknown. Using a novel application of total internal reflection fluorescence microscopy (TIR‐FM) to the apical membrane of renal proximal tubule cells we have found that a dynamic actin cytoskeleton and the unconventional myosin motor myosin VI are required for PTH induced removal of NaPi2a from the BBM. Here we demonstrate that a dynamic actin cytoskeleton is required for NaPi2c removal from the BBM in response to PTH. In addition we investigate the role of myosin VI and membrane lipid dynamics in PTH‐induced trafficking of NaPi2c. Differences in the use of motor proteins or clustering of the cotransporters within the BBM may account for the differences in the kinetics of the response of NaPi2a and NaPi2c to PTH.Supported by NIH K08 DK080989‐01A1.
Going to extremities: is variation in human limb length and proportions a paradox?
Authors: Auerbach Benjamin Miller 
Published: June 2021

Group comparisons indicate thermoregulation, nutrition, and locomotor efficiency as factors affecting human limb length and proportional variation. Humans from warmer climates have longer limbs relative to torso length and higher intralimb intermembral indices, likely reflecting thermoregulatory necessity; distal segments have higher surface area‐to‐mass relative than proximal segments, and longer limbs dissipate more heat. Despite this pattern, paradoxically, limb proportions and limb lengths were shown to not correlate in previous research. Subsequent analysis concluded that limb proportions are adaptively constrained and do not closely reflect limb elongation.Using a global sample of 3680 pre‐industrial skeletons, this study reexamines this incongruity. Coefficients of variation show that variance in proximal relative to distal elements is not universal among all human groups, though distal elements generally have greater variance. Scaling analyses indicate longer limbs have disproportionately longer distal elements and disproportionately shorter proximal elements, following thermoregulatory expectations. Different allometric relationships for proximate and distal elements may obscure these patterns when their lengths are expressed as ratios. Variation in these patterns among groups, and the effects of mechanics and nutrition, are further considered.
Renal epithelial Na channel protein expression is reduced in H,K‐ATPase knockout mice
Authors: Greenlee Megan Michelle , Lynch I. Jeanette , Gumz Michelle L. , Cain Brian D. , Wingo Charles S. 
Published: June 2021

The importance of the epithelial Na+ channel (ENaC) to Na+ reabsorption is highlighted by genetic gain of function mutations in ENaC that lead to severe hypertension (Liddle's Syndrome). Spicer et al (Am J Phys, 2001) demonstrated that colonic ENaC activity was reduced in HKα2−/− mice, suggesting that H+,K+‐ATPases are required for complete ENaC function. Given the importance of functional ENaC to renal Na+ transport, we investigated whether double knockout of H+,K+‐ATPase α subunits in mice (HKα1,2−/−) affected ENaC expression. Real time quantitative PCR was used to evaluate the mRNA expression of α‐, β‐, and γ‐ENaC subunits in cortex and medulla from wild type (WT) and HKα1,2−/− mice. Cortical α‐ and β‐ENaC mRNA expression was slightly decreased (~20–25%) in HKα1,2−/− mice compared to WT (p=0.10 and 0.08, respectively). Importantly, we observed a 45% reduction in medullary α‐ENaC protein expression (n=3, p<0.05) in HKα1,2−/− mice vs WT, as determined by densitometry of immunoblots. In summary, these data suggest that renal H+,K+‐ATPases are needed for appropriate expression of ENaC subunits and are important for renal Na+ adaptation.Studies were funded by NIH RO1‐DK‐049750 to C.S.W and 5T32DK007518‐22 to M.M.G.
Identification of epithelial Na+ channel (ENaC) inter‐subunit Cl‐ coordinating residues suggests a trimeric alpha, gamma, beta channel architecture
Authors: Collier Daniel M , Snyder Peter M 
Published: June 2021

The extracellular domain of the epithelial Na+ channel (ENaC) is exposed to a wide range of anion concentrations in the kidney. We have previously demonstrated that extracellular Cl− inhibits ENaC activity. To identify residues involved in Cl− inhibition, we mutated residues in the extracellular domain of α‐, β‐ and γENaC that are homologous to the Cl− binding site in ASIC1 and tested the effect of Cl− on the activity of ENaC expressed in Xenopus oocytes. We identified two functionally significant Cl− sites in αβγENaC. One site is formed by residues in the thumb domain of αENaC and palm domain of βENaC. Mutation of residues in this domain decreased Cl− inhibition and decreased Na+ self‐inhibition. The second site is formed by residues at the interface of the thumb domain of βENaC and the palm domain of γENaC. Mutation of these residues also decreased Cl− inhibition, yet had no effect on Na+ self‐inhibition. In contrast, mutations in the thumb domain of γENaC and palm of αENaC had minimal effect on Cl− inhibition or Na+ self‐inhibition. The data demonstrate that Cl− inhibits ENaC activity by two discrete Na+ dependent and Na+ independent mechanisms. Furthermore, our results suggest that ENaC adopts an αγβ rather than αβγ confirmation, providing additional insight into the functional significance of α, β and γENaC subunit interactions.PMS is supported by NIH HL072256
RAGE and ERM proteins in EMT of human alveolar epithelial cells
Authors: Buckley Stephen T , Medina Carlos , Ehrhardt Carsten 
Published: June 2021

The receptor for advanced glycation end products (RAGE) purportedly plays an important role in lung homoeostasis, whilst ezrin/radixin/moesin (ERM) proteins serve as important crosslinkers between the plasma membrane and cytoskeleton. Here, we investigated the role of RAGE and ERM proteins in cytokine‐induced epithelial‐mesenchymal transition (EMT) of alveolar epithelial cells (A549).A549 cells were stimulated with TGF‐β1 and/or Cytomix [IL‐1β + TNF‐α + IFN‐γ]. Changes in expression of EMT markers were measured using immunofluorescence microscopy (IFM) and Western blot (WB). RAGE expression and localisation were evaluated by WB and IFM. Total and phosphorylated ERM levels were assessed by WB. Co‐localisation of F‐actin and ERM proteins was also investigated.When challenged with TGF‐β1 and/or Cytomix, A549 underwent EMT with a significant change in phenotype. This was coupled with altered expression of RAGE and a marked redistribution of ERM proteins from the cytoplasm to the cell periphery. F‐actin stress fibres formed following induction of EMT, were found to co‐localise with ERM proteins. Moreover, ERM proteins were found to undergo phosphorylation.Together, these results suggest that both RAGE and ERM proteins may play an important role in EMT of A549 following exposure to TGF‐β1 and/or Cytomix.STB is funded by an IRSCET postgraduate scholarship.
Estrogen induces nitric oxide production in human bronchial epithelial cells
Authors: Townsend Elizabeth A , Thompson Michael A , Cassivi Stephen D , Pabelick Christina M , Prakash Y S 
Published: June 2021

Women have a higher incidence and severity of asthma exacerbations compared to men. Asthma symptoms fluctuate with hormonal status suggesting a role for sex steroids in modulating bronchoconstriction. We have previously shown that acute estrogen decreases intracellular calcium in human airway smooth muscle thereby facilitating bronchodilation, similar to estrogen‐induced vasodilation. Akin to endothelium, airway epithelium is a source of nitric oxide (NO), a bronchodilator. We hypothesized that estrogens facilitate bronchodilation by generating NO in bronchial epithelium. In acutely dissociated human bronchial epithelial cells, 5–10 min exposure to 1 μM ACh or 17β‐estradiol (E2) (1 and 10 nM) resulted in increased DAF‐2 fluorescence (NO indicator). Both ER specific agonists THC (ERα) and DPN (ERβ) (10 nM,) induced NO production comparably. E2 effects were abrogated by the estrogen receptor (ER) antagonist ICI 182,780 (Faslodex®). eNOS phosphorylation was enhanced by E2 while E2‐induced NO production was attenuated by Ca2+ chelation (BAPTA; 20 μM, 30 min) and eNOS inhibition (L‐NAME). Overall, these data indicate a role for airway epithelium‐derived NO in the bronchodilatory effects of estrogens in human ASM.Supported by NIH grants HL090595 (CMP), HL088029 & HL56470 (YSP), and Mayo Graduate School (EAT).
Disturbed inspiratory rhythm in rat brainstem slices by seizure‐like bursting due to theophylline‐evoked GABAA receptor block
Authors: Panaitescu Bogdan , Kuribayashi Junya , Ruangkittisakul Araya , Ballanyi Klaus 
Published: June 2021

Methylxantines theophylline and caffeine can evoke seizures at clinical doses. Here we studied if this can disturb inspiratory activity and whether blockade of γ‐aminobutyric acid‐A (GABAA) receptors is involved.In newborn rat brainstem‐spinal cords, inspiratory‐related cervical nerve bursting driven by the pre‐Bötzinger Complex (preBötC) was perturbed by >0.1mM caffeine or theophylline. At millimolar levels, seizure‐like discharges occurred that resembled non‐respiratory activity due to bicuculline. In brainstem slices, seizure‐like bursting in inspiratory active hypoglossal nucleus or preBötC area was less pronounced than in the en bloc model for the same concentration. GABAA receptor activation with muscimol hyperpolarized preBötC neurons with a decrease of input resistance and blocked inspiratory rhythm. Rhythm restoration by 5–10mM theophylline in muscimol was accompanied by recovery of membrane potential and resistance.Findings suggest that methylxanthines at millimolar doses perturb preBötC bursting in the slice model via blockade of GABAA receptors, whereas disturbance of rhythm at submillimolar levels in the en bloc model may be due to another mechanism.Supported by AHFMR, CIHR, MFN‐Health