bluuebunny/crossref_metadata_2025 huggingface dataset:
The Impact of High Mobility Group Box 1 Protein (HMGB1) on Renal Ischemia‐Reperfusion Injury in Male and Female Spontaneously Hypertensive Rats (SHR)
Authors: Crislip G. Ryan , Tipton Ashlee J. , Sullivan Jennifer C.
Published: June 2021

HMGB1 is an endogenous molecule released by dying cells which contributes to renal ischemia‐reperfusion injury (IRI). We recently published that male SHR have more pro‐inflammatory immune cells in the kidney. The goal of this study was to test the hypothesis that female SHR have less HMGB1 than males resulting in a lower degree of renal IRI. HMGB1 levels were measured via ELISA in kidneys of 12 week old male and female SHR (N=5). Additional age matched rats were pre‐treated with control or HMGB1 neutralizing antibody (300 μg/rat via IP; N=4) 1 hour before occluding renal arteries for 1 hour. Renal blood flow velocity (RBV) was measured via ultrasound before and 24 hours after reperfusion. Kidneys were isolated for mRNA extraction and histology. HMGB1 levels were greater in the renal cortex of female vs. male SHR (0.33±0.05 vs. 0.16±0.02 ng/mg; p=0.01). Medullary HMGB1 levels were comparable between sexes. Pre‐treatment with anti‐HMGB1 did not alter RBV compared to controls (% decrease of male SHR: 34±2 to 35±5, NS; female SHR: 35±6 to 22±6, NS) or expression of renal VCAM1 or ICAM1 in either sex. Using histological analysis we determined that anti‐HMGB1 decreased CD3+ T cell infiltration in both sexes, verifying a role for HMGB1 in inflammation following IRI. In conclusion, despite greater renal HMGB1 levels in female SHR, there were no sex differences in the functional role of HMGB1 inhibition on renal IRI or inflammation.
Vasopressin infusion increases intravesical pressure in Wistar rats.
Authors: Cafarchio Eduardo Mazuco , Silva Luiz Augusto , Colombari Eduardo , Ogihara Cristiana Akemi , Almeida Roberto Lopes , Sato Monica Akemi
Published: June 2021

Objective

This study aimed to investigate the effect of vasopressin on intravesical pressure, renal blood flow and arterial pressure in Wistar rats.

Methods

Adult female Wistar rats (~250 g) were anesthetized with 2% halothane in 100% O2 and the femoral artery and vein were cannulated for mean arterial pressure (MAP) and heart rate (HR) recordings, and drug infusion, respectively. A miniaturized Doppler flow probe was placed around the left renal artery and the renal conductance (RC) was calculated as the ratio of Doppler shift and MAP. The urinary bladder was cannulated for measurement of the intravesical pressure (IP). Saline (vehicle) or 0.25, 0.5 and 1 ng/mL of vasopressin were infused in a ratio of 0.1 mL/min/5 min in rats with bilateral urether ligature. Data are as mean±SE and were submitted to unpaired Student t‐test. Significance level was set at p<0.05. Vasopressin increased the IP (70±30, 107±34, and 105±39% at 0.25, 0.5 and 1 ng/mL, respectively, N=6) compared to saline (1.8±0.5%, N=6). The RC was reduced (−43±18, −26±2, and − 34±12%) by 0.25, 0.5 and 1 ng/mL of vasopressin, respectively, compared to saline (−2±0.02%). No significant changes were observed in MAP (13±15, 7±16, and 6±8 mmHg) after 0.25, 0.5 and 1 ng/mL of vasopressin, respectively, or saline (2±2 mmHg). No changes were also observed on HR (9±21, 7±22, and −5±17 bpm) after 0.25, 0.5 and 1 ng/mL of vasopressin, respectively, or saline (−10±9 bpm).

Conclusions

Infusions of subpressor doses of vasopressin evoked renal vasoconstriction and increased the intravesical pressure in rats with bilateral urether ligature, suggesting that vasopressin can likely increase the contractility of the detrusor muscle regardless the effects on renal blood flow.

Supported by: FAPESP and PIBIC‐CNPq.
A rat model of doxorubicin‐induced late‐onset cardiotoxicity
Authors: Gibson Noah M , Lien Chia‐Ying , Jensen Brock T , Hydock David S , Hayward Reid
Published: June 2021

Doxorubicin (DOX) is one of the most effective and commonly used chemotherapeutic agents in childhood cancer treatments. However, the primary side effect of juvenile DOX exposure is a late‐onset cardiotoxicity that can be manifested decades after treatment. To study late‐onset cardiotoxicity, our lab developed a rat model that expresses cardiac dysfunction in adulthood following juvenile DOX exposure. Twenty‐five day old female Sprague‐Dawley rats were randomly selected to receive a cumulative dose of 14 mg DOX/kg body weight or equivalent saline (SAL) in serial injections of 2/mg/kg/day for 7 days. At 16 weeks of age echocardiography and Langendorff preparations were performed to assess cardiac function and geometry. Significant differences were observed in the following cardiac parameters: left ventricular developed pressure (SAL 94±19 mmHg, DOX 66±7 mmHg), fractional shortening (SAL 62±4%, DOX 52±10%), myocardial performance index (SAL 0.77±0.03, DOX 1.02±0.32), aortic valve maximal flow velocity (SAL 92±4 cm/sec, DOX 67±16 cm/sec) and left ventricular mass (SAL 558±79 g, DOX 375±74 g). DOX treated animals also exhibited a blunted growth curve as evidenced by a lower body mass at 16 weeks of age (SAL 346±43 g, DOX 241±45 g). This model of juvenile DOX treatment may allow for the study of mechanisms associated with late‐onset cardiotoxicity and interventions to protect against it.
Physiologic cyclic strain stimulates heme oxygenase‐1 gene expression in endothelial cells: role in cell survival and proliferation
Authors: Durante William , Liu Xiao‐ming , Peyton Kelly J.
Published: June 2021

Endothelial cells (ECs) are constantly subjected to cyclic strain that arises from the periodic change in vessel wall diameter as a result of pulsatile blood flow. Application of physiologic levels of cyclic strain regulates EC proliferation and apoptosis; however, the underlying mechanisms are not known. Since heme oxygenase‐1 (HO‐1) is a potent modulator of EC function, the present study investigated whether HO‐1 contributes to the biological actions of cyclic strain. Administration of physiologic cyclic strain (6% at 1 hertz) to human aortic ECs stimulated an increase in HO‐1 mRNA, protein, and activity that was prevented by the antioxidant N‐acetyl‐ L‐cysteine. Cyclic strain also evoked an increase in HO‐1 promoter activity that was blocked by mutating the antioxidant response element in the promoter or by overexpressing dominant‐negative Nrf2. Finally, application of cyclic strain inhibited EC apoptosis and proliferation. Although pharmacological inhibition or genetic deletion of HO‐1 reversed the anti‐apoptotic effect, it enhanced the anti‐proliferative action of cyclic strain. In conclusion, the present study demonstrates that physiologic cyclic strain stimulates HO‐1 gene expression via an oxidative pathway that involves Nrf2, and that induction of HO‐1 promotes the anti‐apoptotic action of cyclic strain while counteracting its anti‐proliferative effect. Supported by NIH grant HL59976.
S100A4 is a key regulator of matrix metalloproteinases expression in vascular smooth muscle cells
Authors: Lemaitre Vincent , Weir Jessica‐Anne , Miura Hiroto
Published: June 2021

Introduction

Matrix metalloproteinases (MMPs) are critical in vascular smooth muscle cell (VSMC) dedifferentiation, proliferation, and migration in atherosclerosis. S100A4, a Ca2+‐ binding protein, is expressed in atheromatous and restenotic VSMCs, and regulates MMP expression in tumors. We hypothesized that S100A4 modulates MMP expression in VSMCs.

Methods

VSMCs from wildtype (WT) and S100a4−/− (KO) mouse aortas were treated with platelet‐derived growth factor (PDGF)‐BB (20 ng/ml) and/or angiotensin II (AII; 100 nM) for 24 hours. Supernatants were used for gelatin‐zymography and total RNA for real‐time PCR analysis.

Results

Culture supernatants from KO cells exhibited lower MMP‐2 activity compared to WT cells in all groups (non‐treated, PDGF, AII, and PDGF+AII). MMP‐2 and ‐9 activities were elevated in WT cells after treatment with PDGF and AII, respectively, but were significantly lower in KO cells, which also showed decreased Mmp‐14 mRNA expression (average delta Ct, WT vs. KO: 2.6±0.3 vs. 4.2±0.2, non‐treated cells; 4.5±0.03 vs. 5.7±0.1 with PDGF; 2.9±0.1 vs. 4.1±0.2 with AII, p<0.05; n=3). Western blot showed decreased phosphorylated ERK, an activator of MMPs, in PDGF‐treated KO cells compared to WT.

Conclusion

S100A4 plays a key role in the regulation of MMP‐2, ‐9, and ‐14 produced in activated VSMCs.
Evidence for pleiotropic cardioprotection by the bradycardic agent ivabradine
Authors: Kleinbongard Petra , Heusch Gerd
Published: June 2021

Background

In prior studies in pigs, ivabradine reduced infarct size even when given only at reperfusion and in the absence of heart rate reduction (atrial pacing).

Methods

Ventricular cardiomyocytes were isolated enzymatically from C57Bl/6J mouse hearts (n=15), aliquoted and incubated in Tyrode buffer ± ivabradine (3 μM) for 30 min, respectively. Simulated ischemia ± ivabradine was induced (hypoxia, pH 6.5, 310 mOsm/l, 60 min) and followed by reperfusion ± ivabradine (normoxia, pH 7.4, 250 mOsm/l, 5 min). Viability (trypan blue exclusion) was quantified in >; 200 cells/per preparation. In a subset of hearts (n=5), cardiomyocytes were loaded with CM‐H2DCFDA (5 μM) as an indicator for reactive oxygen species (ROS) formation. Intracellular fluorescence was detected before and after simulated ischemia ± ivabradine.

Results

Viability was comparable at baseline (control, 73±4%; ivabradine, 73±2%), and it remained relatively stable with 60 min normoxia (53±2%). Simulated I/R reduced cardiomyocyte viability (9±3%), whereas it was better preserved with ivabradine (27±2%, p=0.026 versus I/R). Preliminary data indicate a reduction of ischemia‐induced ROS formation (3.1±0.4 a.u.) by ivabradine (2.0±0.4 a.u.).

Conclusion

Ivabradine improves ventricular cardiomyocyte viability during simulated I/R, possibly by reduction of ROS formation.
Hemodynamic and bioenergetics of hearts from obese adult mice overfed during early life
Authors: Vieira Anatalia Kutianski , de Mattos Ana Marcondes , Neves Fabiana Alves , Bernardo Amélia Faustino , Cortez Erika Costa , de Souza Larissa Ramos , Miranda Glauciane Lacerda , de Melo Soares dos Santos Vivian , de Souza Rodrigues‐Cunha Alessandra Cordeiro , Garcia‐Souza Érica Patrícia , Sichieri Rosely , Moura Anibal Sanchez
Published: June 2021

In this study we examined heart hemodynamic and metabolism of adult obese mice overfed during lactation. Heart left ventricle from obese (OG) and control (CG) groups were studied. Analysis of key proteins of cardiomyocyte metabolism and bioenergetics were associated to heart hemodynamic by the Langendorff System and high‐resolution respirometry by Oroboros 2k‐Oxygraph during Baseline (CGBL, OGBL) and Ischemia/Reperfusion (CGIR, OGIR) conditions. OG showed a significant increase in total body mass (p<0,0001), epididymal fat (p=0,0018), retroperitoneal fat (p=0,0046) and glycemia (p=0,0471) when compared with CG. Hemodynamic showed lower Max dp/dt (p=0,0007) and heart frequency (BPM) (p=0,0178) in OGIR when compared to OGBL; higher Tau (relaxation time) (p=0,001) in OGIR compared to CGIR and OGLB. Respiratory control ratio (RCR) of isolated heart fibers of OGBL was decreased to fatty acid and carbohydrate oxidation, OGIR only to fatty acid. There were no difference in O©ü consumption of OGBL and OGIR. Hearts from OGBL and OGIR presented lower content of phosphoAKT and IR when compared to CG. We suggested that early life overnutrition induces permanent alterations of cardiac activity and that this process is associated with proteins of the insulin signaling cascade and mitochondrial energy metabolism.

Supported by FAPERJ (E‐26/110.578/2009)
Regenerative cerebral arteriogenesis is induced by GM‐CSF and inhibited by anti‐inflammatory cardiovascular medications
Authors: Persson Anja Bondke , Duelsner André , Gatzke Nora , Buschmann Ivo R.
Published: June 2021

Restoration of blood flow in case of stenotic brain‐supplying arteries relies on restoration of arterial inflow, achieved by an outgrowth of preexistent arterioles (arteriogenesis) and depends on monocyte migration into the vesel wall. It was assumed that a stimulation of monocytic function induces collateral growth, while anti‐inflammatory substances might interfere negatively.

In two animal models of cerebral hypoperfusion (3‐vessel occlusion/rats, common carotid artery occlusion/mice), administration of the pro‐arteriogenic Granulocyte Colony‐ Stimulating Factor (40ug/kg) augmented angiographic collateral diameters and restored cerebrovascular reserve capacities (CVRC) (20+ 8%, cont. −14+14%). Acetylsalicylic acid (ASA), used in patients at risk for arterial stenosis, interfered with (a) CVRC restoration (1+4%), (b) collateral vessel growth and (c) monocyte migration, while Clopidogrel did not. Current data indicate similar effects for other anti‐inflammatory drugs and other vascular beds, which may explain why their effects on outcomes may fall behind effects on cardiovascular risk factors.

Anti‐inflammatory drugs should be used with care in patients with vascular occlusions until large‐scale clinical data on antiarteriogenic effects have been obtained.
Effect of combined aerobic and resistance exercise training on platelet adhesiveness under shear flow in sedentary men
Authors: Chen Yi‐Ching , Wang Jong‐Shyan
Published: June 2021

Aerobic or resistance exercise may modulate the risk of vascular thrombotic events. This study investigated how combined aerobic and resistance exercise training influenced platelet adhesiveness under various shear flows. Twelve healthy sedentary males were trained on a bicycle ergometer (60% VO2max for 30 minutes) for 3 days per week and an isokinetic dynamometer (alternating 60°–180°/s for 6 to 10 sessions) for 2 days per week for 6 weeks. A novel microfluidic chamber combined with fluorescence instrument, which provided shear stress values covering the entire physiological range of human circulation, was used to assess platelet adhesiveness in whole blood. The experimental results demonstrated that a higher degree of platelets adhered to fibronectin‐coated surface revealed at 5 dyne/cm2 compared to 1 dyne/cm2 of shear stress. Strenuous, acute exercise significantly increased the platelet adhesiveness under 1 and 5 dyne/cm2 of shear stresses. However, combined aerobic and resistance exercise training suppressed the extents of shear‐mediated platelet adhesiveness at rest and immediately strenuous exercise. Hence, these findings may provide an effective strategy for developing a suitable exercise regimen that prevents early thrombotic events, further retarding the progression of cardiovascular disease.
Physiological and Biomechanical Analysis of a Foot‐Tapping Device
Authors: Brilla Lorrie , Suprak David N , Callahan Rory
Published: June 2021

Fidgeting may be effective to increase daily energy expenditure. The purpose of this study was to describe the effects of foot tapping using a variable resistance device in subjects in different age ranges with no lower extremity limitations. The subjects were young, 19–34 years (M: 7, F: 7), middle aged, 35–54 years (M: 4, F: 3), and older, >;55 years (M: 4, F: 4) adults. Movement factors of the limb were measured with accelerometry, goniometry, and electromyography and whole body metabolic cost was assessed. Subjects performed eight 5‐minute conditions (toe and heel, low resistance versus higher resistance, and speed, 100 bpm and 140 bpm) with each separated by a recovery period. Data were analyzed using repeated measures ANOVA at an alpha level of 0.05. There were no statistical differences in either mean or peak shank acceleration, tibialis anterior or gastrocnemius average Integrated EMG, ankle range of motion, or metabolic cost for younger and middle aged subjects. For older adults, METs were statistically different in Toe‐Low Resistance‐100 (TL100) versus Heel‐High Resistance‐140 (HH140), TL100 at 1.06±0.17 versus HH140 1.27±0.28. The TL100 versus HH140 comparison was at the extreme of the conditions, lowest contrasted with highest. However, the slight increase in the mean values of 0.19 METs probably would not have a consequential effect on metabolic cost. At the average weight of the older adult subjects tested, approximately two kilocalories per minute would be expended.
Effects of resistance interval and constant training on muscular hemodynamic kinetics in sedentary men
Authors: Chiang Cheng‐Ling , Wang Jong‐Shyan
Published: June 2021

This study investigated the effects of resistance interval (RIT) and constant (RCT) training on muscular hemodynamic/metabolic responses to isokinetic exercise. Twelve sedentary males were randomly divided into RIT and RCT groups. The subjects were trained at alternating 60°–180°/s (RIT, n=6) or constant 120°/s (RCT, n=6) in knee extension/flexion for 6 to 10 sessions for 2 days/week for 6 weeks. A muscle fatigue test was performed at isokinetic contraction with 180°/s for 30 repetitions. A near‐infrared spectroscopy was applied to assess dynamic changes of [HHb] in vastus lateralis. The kinetics formula is: HHb(t)=HHb(b)+Ai(1−e−(t−TDi)/τi)+Ap(1−e−(t−TDp)/τp)+As(1−e−(t−TDs)/τs).HHb(b) is rest baseline value, and the subscripts i, p and s refer to initial, primary, and slow components, respectively. A is the asymptotic amplitude for the exponential term; τ is the time constant; and TD is time delays. The TDp and τp were summed to provide an indication of the overall dynamics of the initial and primary component (MRT). The results indicated that RIT significantly reduced the values of TDp, τp, Ap, and MRT during isokinetic contraction. However, no significant changes in muscular hemodynamic kinetics occurred following RCT. Hence, we conclude that RIT effectively improves hemodynamic/metablobic efficiencies in contracting skeletal muscles compared to RCT.
Total/HDL cholesterol ratio predicts impaired NO‐dependent vasodilation in human skin
Authors: Dahmus Jessica D. , Craighead Daniel H. , Kenney W. Larry , Alexander Lacy M.
Published: June 2021

The human cutaneous circulation is a representative vascular bed to assess preclinical systemic microvascular dysfunction. A standardized local heating protocol (0.5°C/30sec) was performed to determine risk factors associated with eNOS‐mediated NO‐dependent vasodilation, as measured by the 42°C plateau in skin blood flow (SkBF). Laser speckle contrast imaging (LSCI), a state‐of‐the‐art method for imaging a larger skin surface area compared to single‐point laser‐Doppler flowmetry was used to measure SkBF. We sought to determine the cardiovascular risk factors most predictive of preclinical impairments in cutaneous microvascular dysfunction in healthy human subjects across a broad range of age, blood pressure, and serum cholesterol (CHOL) concentrations. Subjects included 48 men and women (age: 19–88 yr, body mass index: 19.7–36.2 kg/m2, systolic pressure: 90–150 mmHg, diastolic pressure: 50–96 mmHg, total serum CHOL: 121–321 mg/dl, total/HDL ratio: 1.79–7.47, LDL: 54–199 mg/dl, HbA1C: 5.1–5.9, and fasting blood glucose: 80–114 mg/dl). A stepwise regression analysis was performed to determine the best predictor(s) of NO‐dependent vasodilation. The total/HDL ratio predicted impairments in the NO‐dependent plateau (R=0.27, p = 0.01) response to local heating. Microvascular dysfunction as measured by attenuated eNOS‐mediated vasodilation is associated with adverse cholesterol ratios.

R01 AG07004, HL093238, HL089301
Knockdown of tyrosine hydroxylase in the nucleus of the solitary tract (NTS) reduces elevated blood pressure during chronic intermittent hypoxia (CIH)
Authors: Bathina Chandra Sekhar , Rajulapati Anuradha , Franzke Michelle , Yamamoto Kenta , Mifflin Steve
Published: June 2021

We tested the hypothesis that tyrosine hydroxylase (TH) knockdown in NTS reduces cardiovascular responses to CIH, a model of the arterial hypoxemia observed during sleep apnea. Adult male Sprague‐Dawley rats were implanted with radiotelemetry transmitters and AAV constructs with a GFP reporter having either short hairpin RNA for TH (shRNA) or scrambled virus (scrambled) were injected into caudal NTS. Virus injected rats were exposed to 7 days CIH (alternating 6 min periods of 10% O2 and 4 min of 21% O2 from 8am to 4pm; from 4pm to 8am rats were exposed to 21% O2). CIH increased MAP and HR during the day in both the scrambled (n= 14) and shRNA (n=13) groups (all p < 0.001). During the night MAP and HR remained elevated in the scrambled rats (both p<0.001) but not in the shRNA group. The number of TH‐immunoreactive neurons per section was reduced by 20% in sections with GFP fluorescence; shRNA 28±1 vs. scrambled 35±1 (p=.005) without altering the number of dopamine â‐hydroxylase (DBH) – immunoreactive neurons; shRNA 45±3 cells/section vs. scrambled 45±3 cells/section. Western blot analysis showed reductions in TH protein levels of 30% and 10% in caudal and sub‐postremal NTS respectively. Exposure to CIH increased MAP beyond the period of exposure to CIH. Knockdown of TH in the NTS reduces this persistent increase in MAP. This indicates that NTS A2 neurons play a role in the cardiovascular responses to CIH. Supported by HL‐088052
Chronic Depressive Symptoms and Gender: Impact on Conduit Vascular Outcomes
Authors: Stanley Shyla C , Butcher Joshua T , Frisbee Stephanie J , D'Audiffret Alexandre C , Frisbee Jefferson C
Published: June 2021

Chronic depressive symptoms have become recognized as independent risk factors for vascular disease, we investigated the impact of chronic stress/depressive symptoms on vascular outcomes between genders using an establish model from behavioral research. Male and female mice were exposed to 8 weeks of unpredictable chronic mild stress (UCMS; e.g., wet bedding, predator sound/smell, random disruption of light/dark cycle) and indexes of depressive behavior (e.g., coat status, grooming, mobility) were exacerbated in females vs. males. However, dilator responses to methacholine were more strongly impaired in aortic rings of UCMS males than females (p<0.05). NOS inhibition was without effect in UCMS males, while blunting reactivity in UCMS females (p<0.05). COX inhibition impaired dilator responses of UCMS males, but only affected UCMS females at higher concentrations of methacholine. Combined NOS/COX blockade abolished methacholine reactivity. Endothelium‐independent dilator responses were not different across all groups. These results suggest that UCMS impairs vascular function, the severity of which is gender‐dependent. The severity of induced depressive symptoms was not indicative of poorer vascular outcomes. In conclusion, while female mice exhibited increased susceptibility to UCMS‐induced depressive symptoms, they maintained superior vascular function versus male counterparts.
Regulation of antioxidant enzyme expression in response to exercise in skeletal muscles of dyslipidemic mice
Authors: Nguyen Albert Quyen , Thorin Eric
Published: June 2021

Dyslipidemia combined with a sedentary lifestyle is a well‐characterized combination of risk factor for cardiovascular diseases (CVD). Although preventive physical exercise delays the onset of CVD, the underlying mechanisms are not clear. We hypothesized that exercise regulates antioxidant enzyme expression that counteracts the deleterious effects of dyslipidemia. To test this hypothesis, 3‐month old (m/o) severely dyslipidemic (LDLR−/−, hApoB+/+) mice (SD) and wild type (WT) controls were exposed or not (sedentarity, SED) for 3 months to voluntary physical training (PT, □4.5 km/day). At 6‐m/o, the endothelial function of the femoral artery was measured using a pressure myograph and the surrounding skeletal muscles were isolated to quantify antioxidant gene expression by qPCR. No differences in endothelial function were noted between groups. In contrast, GPx1 expression decreased by 25% (p<0.05) in SD mice; PT, however, increased (p<0.05) its expression in both WT (+25%) and SD (+54%) mice. Prdx3 mRNA was higher (p<0.05) in SD compared to WT by 44% and 50% in SED and PT mice. Likewise, MnSOD and Txn1 expressions were elevated in SD mice compared to the other groups (p<0.05) and that was prevented by PT. In conclusion, analysis of mRNA expression reveals that the regulation of antioxidant enzymes is sensitive to dyslipidemia and PT, unveiling the dynamic and sensitive nature of the endogenous antioxidants.
Ascorbic acid inhibits histamine‐receptor mediated sustained post‐exercise vasodilation in humans
Authors: Romero Steven A. , Kono Jordan M. , Halliwill John R.
Published: June 2021

An acute bout of aerobic exercise elicits a sustained post‐exercise vasodilation which is histamine‐receptor mediated. We tested the hypothesis that the antioxidant ascorbic acid would inhibit this histaminergic vasodilation. Four healthy men performed 1 hr unilateral dynamic knee extension at 60% of peak power on 3 separate days: 1) control, 2) intravenous ascorbic acid infusion (primer dose 0.5 g/kg, maintenance dose 0.017 g/kg), 3) combined ascorbic acid infusion and oral H1/H2 receptor antagonism (540mg fexofenadine, 300mg ranitidine). Femoral artery blood flow (Doppler ultrasound) was measured before exercise and for 2h post‐exercise. Femoral vascular conductance was calculated as flow/pressure. Pre‐exercise conductance was similar across all 3 days (P >; 0.05). Compared with pre‐exercise, conductance was elevated on the control day (Δ 58 ± 7%, P < 0.05), but was unchanged with ascorbic acid infusion (Δ 10 ± 6%, P >; 0.05) and combined ascorbic acid and histamine receptor antagonism (Δ 17 ± 5%, P >; 0.05). Post‐exercise conductance was greater on the control day compared with ascorbic acid infusion alone or with histamine receptor antagonism (both P < 0.05). Thus, ascorbic acid inhibits the histaminergic sustained post‐exercise vasodilation, suggesting that exercise‐induced oxidative stress activates a histaminergic pathway in previously exercised muscle.
The effect of short‐term exercise training and nitric oxide on the adaptation of femoral vascular conductance at the onset of contraction
Authors: Just Timothy , Jendzjowsky Nicholas G. , DeLorey Darren S.
Published: June 2021

We tested the hypothesis that short‐term mild‐ (M) and heavy‐intensity (H) exercise training (ET) would speed the adaptation of femoral vascular conductance (FVC) at the onset of contraction by a nitric oxide (NO) dependent mechanism. Sprague‐Dawley rats (n=18) were randomized to sedentary (S), M (20m/min5% grade) or H (40m/min5% grade) ET groups and trained 5d/wkfor 4 wks with equal ET volume. Rats were anesthetised and instrumented for measurement of arterial blood pressure and femoral artery blood flow. FVC was calculated. The triceps surae muscle group was stimulated to contract at 30% and 60% of maximal contractile force (MCF) before and after NO synthase blockade (L‐NAME, 5mg/kg IV). FVC data were fit with a mono‐exponential model. The time constant (τ) for FVC was not different (p>;0.05) between groups at 30% (S = 18 ± 13s; M = 15 ± 5s; H = 13 ± 14s) or 60% MCF (S = 24 ± 17s; M = 16 ± 4s; H = 19 ± 4s). L‐NAME increased (p<0.05) τ in all groups at 30% MCF (S = 25 ± 12s; M = 21 ± 6s; H = 27 ± 13s), but did not alter (p>;0.05) τ at 60% MCF (S = 25 ± 9s; M = 22 ± 9s; H = 20 ± 3s). The primary findings from this study were that: 1) short‐term ET did not alter the adaptation of FVC at the onset of contraction; and 2) regardless of training status, NO contributed to the regulation of FVC kinetics at the onset of moderate‐intensity contraction (30% MCF), whereas NO was not required for vasodilation at the onset of heavy‐intensity contraction (60% MCF). NSERC, Canada.
Role of purinergic neurotransmission in different brainstem CO2‐chemoreceptor regions
Authors: Wenker Ian Christopher , Sobrinho Cleyton R , Takakura Anna C , Moreira Thiago S , Mulkey Daniel K
Published: June 2021

Central chemoreception is the mechanism by which specific brainstem regions, such as the solitary tract (NTS), medullary rapheand retrotrapezoid nucleus (RTN), regulate breathing in response to changes in tissue CO2/pH. In the RTN, the mechanism of chemoreception involves modulation of chemosensitive neurons by CO2‐evoked astrocytic ATP release. However, it is not clear whether purinergic signaling influences CO2‐responsiveness of other putative chemoreceptors. This study aimed to determine whether neuronal activity and CO2‐responsivness of NTS and raphe chemoreceptors are influenced by purinergic signaling in vitro and in vivo. In brain slices, cell‐attached recordings show that CO2‐sensitive neurons in the NTS, but not the raphe, respond to focal ATP application with ~3 fold increase in firing rate. However, bath application of a non‐specific P2‐receptor blocker (PPADS) had no effect on baseline activity or CO2‐responsivness of NTS nor raphe neurons. In anaesthetized adult rats, injection of PPADS into either the caudal NTS or medullary raphe had no effect on baseline phrenic nerve activity or the ventilatory response to CO2. Further, immunohistochemical analysis of an astrocyte marker indicates that the RTN has a higher density of astrocytes compared to the NTS or medullary raphe. Together, these results suggest that purinergic signaling is a unique feature of RTN chemoreception.
Reactive oxygen species (ROS) and purinergic signaling contribute to the thyroid hormone (TH)‐induced vasodilation
Authors: Fürstenau Basso Cristina R. , Bechara Luiz R. G. , Belló‐Klein Adriane , Brum Patrícia C. , Barreto‐Chaves Maria Luiza M
Published: June 2021

The participation of ROS production and nucleotides signaling in the TH‐induced vasodilation was investigated. T3 was added to VSMC primary culture or administered to rats to evaluate the expression of Nox1 and 4, P2Y1,2, and 6. Redox status was assessed by the measurement of glutathiones (Arbor Assays, MI, USA). Results show that T3 (10−8 M, 24 h) increased Nox1 protein and Nox4 mRNA expression in VSMC (n=3). Besides, the expression of P2Y1, 2 and 6 mRNA seem to be decreased in T3‐treated VSMC (10−8 and 10−7 M, 24 h). Experimental hyperthyroidism was confirmed by increased cardiac mass (heart weight (mg)/tibia's length (mm): 33.17 ± 3.34, n=15 vs 27.49 ± 2.24, n=14 in control, p<0.05) and non‐detectable levels of total T4. Nox4 protein expression is increased in T3‐treated animals (7 μg/100 g body weight, i.p., 14 days), n=4. Moreover, oxidized glutathione (GSSG) is significantly diminished in hyperthyroid animals (4.15 ± 0.60, n=8 vs 1.98 ± 0.30, n=9 in control group, p<0.05) while the ratio reduced (GSH)/oxidized glutathione is significantly higher in T3 group (6.71 ± 1.4, n=8 vs 15.64 ± 3.5 in control animals, n=9, p<0.05), suggesting a more preserved antioxidant status in these animals. The present data disclose the participation of ROS production and purinergic signaling in the TH‐induced vasodilation.
Hydrogen peroxide (H2O2) is not increased in angiotensin II (AngII)‐stimulated neurons overexpressing superoxide dismutase (SOD)
Authors: Case Adam J , Cruz Rodrigo Franco , Zimmerman Matthew C
Published: June 2021

Previous studies have shown that SOD overexpression in neurons inhibits AngII intra‐neuronal signaling; thus, suggesting that superoxide radicals (O2•−) are key signaling intermediates driving AngII‐induced neuronal activation. However, considering H2O2 is a direct product of O2•− dismutation by SOD, an alternative interpretation is that increased H2O2 mediates the inhibition of AngII signaling in SOD overexpressing neurons. We aimed to determine if H2O2 levels are indeed increased in SOD overexpressing neurons stimulated with AngII. To query this, we used a neuronal cell culture model expressing HyPer, a H2O2‐sensitive fluorescent protein. To determine the sensitivity of HyPer to H2O2, neurons were stimulated with increasing concentrations of H2O2 (1–100 μM) and changes in HyPer fluorescence were observed by real‐time confocal imaging. Robust increases in HyPer fluorescence were detected after treatment of ≥5 μM, but not after 1 μM H2O2. In contrast, no changes in HyPer fluorescence were observed after AngII (100 nM) stimulation in control or SOD overexpressing neurons. These data suggest that AngII does not increase H2O2 levels in SOD overexpressing neurons, or does so at concentrations below the level of HyPer detection. These data further support the hypothesis that SOD overexpression inhibits AngII intra‐neuronal signaling by decreasing O2•− levels, but not by increasing H2O2 levels. AHA 0930204N.
EtOH consumption contributes to a ‘pro‐injury’ environment in the alcoholic lung.
Authors: Eaton Amity F , Brown Lou Ann , Helms My N.
Published: June 2021

Chronic alcohol consumption increases ICU morbidity and mortality due to acute respiratory distress syndrome (ARDS). One reason is that alcohol regulates alveolar epithelial sodium channels (ENaC) via oxidant injury. We used mice to mimic acute and chronic alcohol consumption and tested whether ethanol (EtOH) would affect lung fluid clearance by altering lung ENaC activity. Chronic ethanol animals were tracheally instilled and imaged in vivo to observe lung fluid clearance. Lung fluid clearance showed a latent increase in clearance rates in mice receiving EtOH. Ethanol mice given an acute tracheal instillation of LPS and imaged for four hours showed lung fluid clearance early compared to control mice. But, ethanol mice with overnight exposure to LPS, showed reduced alveolar fluid clearance like the alveolar flooding seen in ARDS. Together, these data suggest that chronic EtOH consumption leads to altered regulation of ENaC, contributing to a ‘pro‐injury’ environment in the alcoholic lung. Funding support: R00HL09226 awarded to MNH; Children's Center for Development in Lung Biology Grant awarded to MNH.
Beneficial Effects of Angiotensin (1–7) On Dysfunctional CD34+ Endothelial Progenitor Cells (EPC) from Patients with Heart Failure
Authors: Hazra Sugata , Cole‐Jeffrey Colleen T. , Pepine Carl J , Katovich Michael J , Grant Maria B , Raizada Mohan K
Published: June 2021

Endothelial dysfunction is well established in heart failure (HF) and thus HF individuals can be benefitted from autologous cell therapy. In HF patients, the numbers of circulating EPCs vary with NYHA stage. We hypothesized that CD34+ EPCs are dysfunctional in heart failure (HF) patients which can be corrected by angiotensin 1–7 (Ang‐(1–7)). Peripheral blood was collected from control (n=10) and NYHA class II‐IV HF patients (n=15). CD34+ cells were enumerated by flow cytometry and were assessed for migratory potential, NO generation, ROS levels with and without Ang1–7. HF patients showed 63% reduction in number of CD34+cells, 60% reduction in BFU‐E, 74% increase in CFU‐GEMM colonies, reduced expression of ACE2, MAS1 and IL‐10 whereas increased expression of TNF‐α and IL‐6 compared to controls. Migratory potential of CD34+ cells in HF patients were reduced by 77%. Additionally, CD34+ cells from class III donors had 44% reduction in NO/ROS ratio due to increased ROS and reduced NO. Ang‐(1–7) treatment improved their migratory response, reduced ROS and increased NO which resulted in a 54% increase in NO/ROS ratio. These data support that CD34+ cells isolated from HF individuals are dysfunctional, with a reduction in number, colony forming ability, migratory potential, NO generation and increased oxidative stress and these dysfunctions can be corrected by Ang‐(1–7), which suggests therapeutic potential of Ang‐(1–7).

This work was supported by HL56921
Mechanosensitive activation of ATP‐sensitive K+ channels in native pulmonary epithelium depends on pannexin and connexin hemichannel mediated ATP release
Authors: Richter Katrin , Kiefer Kevin Peter , Lintges Michael , Geissert Lea , Clauss Wolfgang , Fronius Martin
Published: June 2021

Objective

The pulmonary epithelium (PE) is permanently exposed to mechanical forces (shear force, stretch). There is evidence that distention of PE leads to an activation of ATP‐sensitive K+ (KATP) channels. The aim of this study was to clarify the mechanisms of KATP channel activation and the contribution of hemichannels in particular.

Methods

A modified Ussing chamber that enabled the application of hydrostatic pressure (HP) was used to mechanically stimulate native PE derived from Xenopus laevis while the short circuit current (ISC) was recorded. Activation of KATP channels was ascertained by the application of the specific inhibitor HMR 1098. Contribution of hemichannels was investigated by application of connexin (meclofenamic acid) and pannexin (probenecide) inhibitors. Extracellular ATP was determined via a luminescence ATP detection assay.

Results

Apical application of HP led to a pressure‐induced current decrease (17.6 ± 2%, P<0.001). This effect was prevented by HMR 1098. The HP‐induced effect was also reversed by the hemichannel inhibitors. Under control conditions (without pharmaca) the HP‐induced effect was accompanied by an increased ATP release which was also inhibited by the hemichannel inhibitors.

Conclusion

KATP channel activation in response to HP is caused by ATP release through a mechanosensitive activation of connexin and pannexin hemichannels.
The relationship of muscle sympathetic nerve activity to the sympathetically‐mediated thermic effect of food in young healthy subjects
Authors: Matzek Luke James , Miles John , Jensen Michael , Levine James , Joyner Michael , Curry Timothy
Published: June 2021

The thermic effect of food (TEF) is partially mediated by β‐adrenergic receptors (βAR) and is an important component of daily energy expenditure. Greater sympathetic activity has been associated with decreased BAR responsiveness. We hypothesized that muscle sympathetic nerve activity (MSNA) would be inversely correlated with the βAR component of TEF (TEFβ). We measured MSNA (microneurography) and energy expenditure (EE) (indirect calorimetry) before and for 5 h after a mixed meal (40% daily EE). Measurements were made during systemic βAR blockade with propranolol (BB) and saline (NS) infusion on separate days in 10 healthy subjects (2 female, age 28±2 y, BMI 24.4±0.6 kg/m2). Resting MSNA was 39±3 bursts/100 heartbeats (hb). Within 1 h after the meal, MSNA significantly increased during both NS (12±3 bursts/100 hb) and BB (18±4) but was not significantly different between the two days, (P=0.375). The maximum increase in REE after the meal was 269±29 (NS) and 198±22 (BB) kcal/24 h (P<0.05). TEFβ (TEFβ = AUC TEFNS – AUC TEFBB) was 48±20% of total TEF. TEFβ was positively correlated with resting MSNA (R2=0.70, P<0.05). Contrary to our hypothesis, we found that greater levels of MSNA were associated with greater TEFβ. These results indicated that resting MSNA may be an important contributor to daily energy expenditure. Studies are needed to determine if greater MSNA is associated with a resistance to weight gain.

The project described was supported by the Mayo Foundation for Medical Education and Research and NIH Grant Numbers K23 DK82424, P30 DK50456, and UL1 RR024150.
Treatment with sulodexide restores compromised glucose tolerance in diet‐induced obese mice
Authors: Eskens Bart , Vink Hans , VanTeeffelen Jurgen
Published: June 2021

We showed in a previous study that feeding mice a high‐fat diet (HFD) resulted in an early reduction in endothelial glycocalyx barrier properties in hindlimb muscle microcirculation which was associated with increased blood glucose levels during a glucose tolerance test. In the current study we investigated whether sulodexide (SUL), an endothelial glycocalyx mimetic, could improve glucose tolerance at the early stage of diet‐induced obesity (DIO).

C57Bl/6 mice were fed a HFD for 6 weeks (n=12); SUL (0.15 mg/mL) was given in the drinking water in the last two weeks (n=5). After 6 weeks the mice were anesthetized and glucose tolerance was measured by i.p. injection of 1 g/kg glucose and the area under the curve of blood glucose (AUCgluc) and plasma insulin (AUCins) were determined for 90 minutes after glucose injection.

While baseline glucose and insulin levels were not affected by the SUL treatment, AUCgluc was significantly lower in the SUL treated mice (242 ± 43 versus 404 ±‐ 43 mmol/L * 90 min). AUCins was not affected by the SUL treatment (2108 ± 431 versus 2952 ± 512 μU/ml * 90 min).

The results of the current study suggest that preventing or diminishing endothelial glycocalyx loss may be a promising intervention for improving glucose homeostasis in obesity.

Supported by DFN 2006.00.027, and CTMM work package 01C‐104–04‐PREDICCT
ACE2 gene therapy decreases fibrosis in the pancreas of high fat diet‐fed mice
Authors: Chodavarapu Harshita , Chhabra Kavaljit , Shenoy Vinayak , Raizada Mohan K , Yue Xinping , Lazartigues Eric
Published: June 2021

Type 2 diabetes mellitus (T2DM) is characterized by progressive loss of β‐cell function and insulin resistance. High fat diet (HFD) activates the renin‐angiotensin system (RAS) and causes hyperglycemia. RAS over‐activity has been linked to pancreatic fibrosis characterized by excessive deposition of collagen and elevated TGF‐β. We propose that Angiotensin Converting Enzyme 2 (ACE2) improves β‐cell dysfunction by attenuating Angiotensin‐II‐induced pancreatic fibrosis. To test our hypothesis, mice (n=6/group) were fed with HFD or regular diet for 16 weeks. At the 13th week, HFD‐fed mice were treated with Lenti‐mACE2 or Lenti‐GFP (intra‐pancreatic injection, 10^6 TU/ml in 100 μl saline). After 16 weeks on HFD, ACE2 gene therapy prevented the increase in fasting blood glucose levels (HFD‐ACE2: 152 ±9 vs. HFD‐GFP: 194 ±10 mg/dl P<0.05), plasma insulin levels (HFD‐ACE2: 6 ±2 vs. HFD‐GFP: 12 ±1 ng/ml P<0.05), and improved glucose tolerance (HFD‐ACE2: 52037 ±2264 vs. HFD‐GFP: 44702 ±1571 P<0.05). Quantitative RT‐PCR shows that HFD produced a 3‐fold increase in collagen I mRNA in the pancreas. ACE2 treatment significantly reduced mRNA expression of TGF‐β1 (HFD‐ACE2: 0.24 ±0.06 vs. HFD‐GFP: 1.4 ±0.3 P<0.05) and collagen I (HFD‐ACE2: 0.26 ±0.1 vs. HFD‐GFP: 1.6 ±0.09 P<0.001). In conclusion, our data demonstrate that ACE2 gene therapy could have a potential therapeutic value in T2DM by attenuating pancreatic fibrosis.
Protective effect of Hedyotis diffusa on lipopolysaccharide (LPS)‐induced liver damage
Authors: Kang Min‐Jung , Kim Jung‐In
Published: June 2021

Oxidative stress is a common mechanism of liver injury. The purpose of this study is to investigate hepatoprotective effects of Hedyotis diffusa (HD) against lipopolysaccharide (LPS)‐induced liver damage in rats. Male Sprague‐Dawley rats (BW 150–170 g, n=21) were divided in to three groups. Normal group and LPS group were fed AIN‐93G diet, whereas LPS‐HD group was offered Ain‐93G diet containing 1% ethanol extract of HD for 4 weeks. Normal group were injected saline (0.5 mL/kg BW, i.p.). LPS and LPS‐HD group were injected LPS (6 mg/kg BW, i.p.). Rats were sacrificed 4 hours after the injection of saline or LPS and blood and liver samples were collected. Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities were measured. Hepatic thiobarbituric acid reactive substance (TBARS) levels and superoxide dismutase (SOD) activities were measured. Serum AST and ALT activities of LPS‐HD group were significantly lower than those of LPS group. Consumption of Hedyotis diffusa significantly decreased hepatic TBARS levels compared with LPS group. Hepatic SOD activities were significantly increased by consumption of Hedyotis diffusa compared with LPS group. Thus we conclude that Hedyotis diffusa could exert protective effect against LPS‐induced liver damage in rats, at least partially due to antioxidant activity. This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2010–0011980).
Regulation of Human Intestinal SLC26A3 Expression by Hepatocyte Nuclear Factors
Authors: Kumar Anoop , Alakkam Anas , Borthakur Alip , Anbazhagan Arivarasu N , Saksena Seema , Gill Ravinder K , Malakooti Jaleh , Alrefai Waddah A , Dudeja Pradeep K
Published: June 2021

DRA (Down Regulated in Adenoma, SLC26A3) plays a key role in Cl− absorption in the intestine. DRA expression is decreased in congenital chloride diarrhea and IBD. Hepatocyte nuclear factor 4 α (HNF4 α) has been shown to modulate basal DRA expression in Caco2 cells whereas intestine‐specific HNF1α/HNF1β deficient mice showed diarrheal phenotype and decreased DRA expression. However, the role of individual HNFs or their combined effects on DRA expression is not known. Therefore, detailed analysis of DRA transcription was performed in response to over‐expression or silencing of HNFs. Results showed that overexpression of HNF1α, HNF1β or HNF4α in Caco‐2 cells increased DRA promoter activity by ~20, 10, and 2.5 fold, respectively (p<0.05). Similarly, their overexpression increased DRA mRNA levels (~ 4–5 fold with HNF1α, ~2–3 with HNF1β and ~1.5 with HNF4α (p<0.05). DRA mRNA expression decreased by ~50%, 35% and 30% by silencing of HNF1α, HNF1β and HNF4α, respectively. Silencing both HNF1α and HNF1β further decreased DRA expression (~70%). The attenuation of HNF1α decreased the level of HNF4α (but not HNF1β) and vice versa indicating that HNFs can modulate the expression of each other as well. ChIP assays revealed that DRA promoter harbors binding sites for HNFs at − 814/−219 bp region. These studies demonstrate that HNFs directly stimulate DRA expression and could serve as novel anti‐diarrheal targets. (Grant Support: VA/NIDDK/Gates Foundation)
Intact colonic K+ secretion in KCNMB2 knockout mice
Authors: Larsen Casper Kornbech , Praetorius Helle Anette , Leipziger Jens
Published: June 2021

Colonic K+ secretion occurs via the pump‐leak mechanism; K+ is taken up from the basolateral side via the Na+/K+ pump and NKCC1 and leaves the cells via apical K+ channels. The BK channel has been identified as the relevant K+ channel involved in K+ secretion in mammalian distal colon. It consists of 4 α‐subunits (encoded by KCNMA1) and 4 modulatory β‐subunits, of which there are 4 isoforms (KCNMB1–4). Previously, we have shown that the β2‐(KCNMB2) is the only β‐subunit expressed in the murine distal colon and that aldosterone increases the expression of both the α‐ and the β2‐subunit.

We hypothesize that upregulation of the β2‐subunit participates in aldosterone‐induced colonic K+ secretion. Here we use a global β2 subunit (KCNMB2) knockout mouse, in which we study distal colonic K+ secretion by open‐circuit recordings in Ussing chamber. The luminal Ba2+‐sensitive short circuit current was taken as measure of BK channel‐mediated electrogenic K+ secretion. Resting K+ secretion was 20.8 μA·cm−2 ± 3.8 and 28.2 μA·cm−2 ± 4.4 in WT and KO tissue, respectively (n=6). We found that apical ATP, known to activate K+ secretion via luminal P2Y receptors, increased colonic K+ secretion to −127.2 μA·cm−2 ± 9.47 (WT) and −73.4 μA·cm−2 ± 4.9 (KO) We report the surprising finding that knockout of the assumed β‐subunit (KCNMB2) of the BK channel leaves resting and ATP‐stimulated K+ secretion intact.

Supported by the Lundbeck foundation.