bluuebunny/crossref_metadata_2025 huggingface dataset:
Sphingosine 1‐Phosphate (S1P) Attenuates Combined Acute Alcohol Intoxication and Hemorrhagic‐Shock/Resuscitation‐Induced Microvascular Hyperpermeability in Rat Mesentery
Authors: Doggett Travis Matthew , Alves Natascha G , Yuan Sarah Y , Breslin Jerome W
Published: November 2023

Alcohol intoxication‐related injuries remain a significant clinical problem involving at least 38 million individuals and producing and estimated $223.5 billion health care cost burden in the US. In the emergency room, intoxicated trauma patients present with increased injury severity and are more hypotensive, a predictor of poor outcomes. We have previously demonstrated that acute alcohol intoxication causes microvascular hyperpermeability in vivo, and also exacerbates hemorrhagic shock/resuscitation‐ (HSR)‐induced microvascular leakage. In the current study, we tested the hypothesis that the barrier‐enhancing bioactive lipid Sphingosine 1‐Phosphate (S1P) can reverse the microvascular hyperpermeability and hypotension following combined acute alcohol intoxication and HSR. Carotid artery, jugular vein, and indwelling gastric catheters were surgically placed in anesthetized Sprague Dawley rats. Following 6–7 days recovery, conscious, unrestrained rats received 2.5 g/kg alcohol via the intragastric catheter, followed by HSR (fixed pressure of 40 mm Hg for 1 h, followed by Lactated Ringers (LR), 40% i.v. bolus + 2x total blood volume removed infusion over 1 h). During resuscitation, the rats received 0.003, 0.03, or 0.1 mg/kg S1P within the LR fluid. Following the 1‐h resuscitation phase, the rats were anesthetized for intravital microscopy of the mesenteric microcirculation. FITC‐Albumin served as a tracer to determine extravasation, quantified by measuring integrated optical intensity (IOI) of the extravascular areas adjacent to postcapillary venules. The results show that by the end of the 1‐h fluid resuscitation period, the group receiving LR supplemented with 0.1 mg/kg S1P displayed significantly restored MAP compared to Alcohol/HSR (89.1 ± 3.6 mm Hg for Alcohol/HSR + 0.1 mg/kg S1P vs. 75.2 ± 3.1 mm Hg for Alcohol/HSR, p<0.05). This trend persisted during the subsequent intravital microscopy phase of the experiment (86.3 ± 3.3 mm Hg Alcohol/HSR + 0.1 mg/kg S1P vs. 68.3 ± 4.3 mm Hg Alcohol/HSR, p<0.05). In addition, 0.1 mg/kg S1P reduced the alcohol/HSR‐induced microvascular hyperpermeability by 50% (542.3 ± 58.4 IOI units for Alcohol/HSR + 0.1 mg/kg S1P vs. 1061.7 ± 142.1 IOI units for Alcohol/HSR, p<0.001). These data show that fluid resuscitation augmented with 0.1 mg/kg S1P ameliorates alcohol/HSR‐induced hypotension and microvascular hyperpermeability more effectively than standard fluid resuscitation. Overall, the data show promise for using fluid resuscitation augmented with S1P for improving circulation in traumatic injury patients.

Support or Funding Information

Supported by NIH grants R01HL098215 and R21AA020049, and the ABMRF/Foundation for Alcohol Research.
Optimization of the Hemodynamic of the Kidney and Preservation of Glycocalyx During Kidney Transplantations ‐ A Combination of Fluid Therapy and Low Dose Norepinephrine
Authors: Eriksen Jonathan Kunisch , Moeslund Niels , Nielsen Lise Have , Petersen Jens Aage Kølsen , Jespersen Bente , Birn Henrik
Published: November 2023

Background

Delayed Graft Function (DGF) is a common complication to kidney‐transplanted patients receiving a kidney from brain dead donors. The present study is designed to evaluate the influence of fluid therapy on kidney function and the preservation of the glycocalyx, during kidney transplantation using kidneys from brain dead donors.

Method

A porcine prospective randomized animal study is performed. The recipients are randomized to either high volume fluid therapy or individualized goal directed fluid therapy (IGDT). The pigs are also randomized to either no Nor‐epinephrine (NE) or continuous low dose NE‐infusion.

Brain death is achieved by inflating a 60cc urinary catheter inserted in the epidural space until the intracranial pressure exceeds the systolic pressure. After 4 hours of brain death the kidneys are perfused with “Bridge to life®” and removed and stored cooled at 4°C for at least 18 hours.

The recipients are bilaterally nephrectomised and each gets one kidney. The recipient pigs are observed for 10 hours post‐reperfusion. Blood and urine samples are collected during this observation period. Biopsies are taken from the donor kidneys at baseline and after 3 hours of brain death, and in the recipient immediately after reperfusion and 10 hours post reperfusion.

The kidney function will be evaluated using measured GFR (mGFR) and by studying the histology of the kidney using light microscopy.

We plan to evaluate the glycocalyx degradation by analyzing blood for Syndecan‐1 and Heparan Sufalte using ELISA. Further to evaluate the histology, the biopsies will be studied in microscope, along with immunofluorescence evaluation of Syndecan‐1 for a semi‐quantitative measurement for glycocalyx.

Results

As this is an ongoing study no results have yet been obtained.

Conclusion

We hope to show a significantly better kidney function and glycocalyx preservation in the IGDT group, and even better in the group receiving NE.

Support or Funding Information

This Study is funded by the Novo Nordic fund.
Blunted Endothelial Dependent Vasodilation in the Cutaneous Microvasculature of College‐Aged African Americans
Authors: Patik Jordan C , Hurr Chansol , Christmas Kevin M. , Brothers R. Matthew
Published: November 2023

African Americans (AA) have elevated risk for hypertension, insulin resistance, and cardiovascular disease compared to their Caucasian American (CA) counterparts. Previously, we have observed attenuated vasodilation in the cutaneous vasculature during local heating in young, healthy AA compared to CA. Impaired endothelium‐dependent vasodilation in response to acetylcholine (ACh) is present in other at‐risk populations, including aging, obesity, smokers, and hypercholesterolemics. Therefore, the aim of the current study was to test the hypothesis that AA have a blunted vasodilatory response to the ACh analog methacholine (MCh). Twenty‐three (12 AA, 11 CA) volunteers free of overt disease participated in the study. AA and CA groups were matched for age (21.5±1 vs 23.9±1 years, P=0.30), BMI (25±1 vs 23.8±1, P=0.31), and sex (AA: 6M/6F, CA:6M/5F). MCh was delivered via a sterile microdialysis fiber inserted in the dermal layer of the left forearm. MCh doses of 10−6, 10−5, 10−4, 10−3, 10−2, and 1M (mixed in lactated Ringer's) were infused into the skin for 6 min each dose at a rate of 2μL/min. Cutaneous red blood cell flux was measured via laser Doppler fluximetry (LDF) continuously and averaged over the last min of each dose. Local heaters were set to 33°C throughout the dose‐response protocol. Brachial blood pressure was taken during the last min of each dose on the contralateral arm. Cutaneous vascular conductance (CVC) at each dose was calculated as LDF/MAP and then normalized to each individual's maximal CVC during combined 44° heating and 28mM sodium nitroprusside infusion (CVCmax). Using normalized data, four parameter logistic dose response curves for each group were compared with an extra sum of squares F‐test with maximum constrained to 100 (%CVCmax). Including all parameters, AA and CA dose response curves were different (P<0.001). Log EC50 for AA was at a higher dose (−2.814±0.26) compared to CA (−4.434±0.19, P<0.001). Hillslope was not different between groups (P=0.33). Baseline values were also not different (AA: 10.37±5.26 %CVCmax vs CA: 15.02±4.27 %CVCmax, P=0.54). The results of this study indicate that a significantly greater dose of MCh is required to reach 50% of maximal vasodilation in healthy, young AA compared to CA. This pharmacological evidence supports earlier findings suggestive of impaired endothelium‐dependent microvascular function in AA.

Support or Funding Information

Institutional Start‐up funds from the University of Texas at Austin
Nox1 Deletion Rescues Microvascular Dysfunction in Obese Mice
Authors: Thompson Jennifer Anne , Mintz James , Fulton David J , Stepp David W
Published: November 2023

Objective

To investigate the role of Nox1 in the development of microvascular dysfunction in obesity.

Methods

Four genotypes were generated by breeding Nox1 knock‐out (KO) mice with obese db/db mice: (HdbWnox1); lean Nox1 KO (HdbKnox1); obese (KdbWnox1); double KO (KdbKnox1). At 12 weeks of age, body composition was determined by NMR spectroscopy and fasting levels of lipids, glucose and HbA1c were measured. Glucose tolerance tests in conscious mice were generated after IP injection of a glucose load (2g/kg body weight). Mesenteric arteries were isolated and endothelium‐dependent responses to acetylcholine (Ach) (10−12–10−3 M) were assessed in the presence or absence of L‐NAME (inhibitor of nitric oxide synthase) or Tempol (superoxide dismutase mimetic).

Results

Body weight and % body fat were increased in obese (KW, KK) vs. lean (HW, HK) mice (p < 0.01), while there were no differences in KW vs. KK (% body fat, HW: 16.3 ± 2.5 %; KW: 61.2 ± 5.2 %; HK: 14.2 ± 1.2 %; KK: 50.3 ± 5.0 %). Likewise, fasting plasma glucose, HbA1c, insulin and triglyceride levels were elevated in obese mice (p < 0.01), with no differences between KW and KK. The area under the curve for glucose tolerance tests was increased by 67% in HW vs. KW and 60% in HK vs. KK (p < 0.05). KK and KW mice also exhibited similar reductions in oxygen consumption and carbon dioxide production (p < 0.01). Endothelium‐dependent responses to Ach were blunted by 48% in obese (KW) vs. lean (HW) (p < 0.01), yet similar between obese mice with Nox1 deletion (KK) and lean controls (HK). L‐NAME incubation reduced Ach responses in HW, HK and KK animals (p < 0.01), but had no influence in KW. Tempol had no impact on HW, HK and KK, but improved Ach responses in KW mice.

Conclusions

Impaired endothelial function in the microvasculature of obese db/db mice appears to be related to vascular superoxide production. Deletion of Nox1 improved microvascular endothelial function in obese db/db mice without influencing the degree of adiposity, hyperglycemia or dyslipidemia. Therefore, Nox1 may be an effective target for the prevention of vascular disease in the metabolic syndrome.

Support or Funding Information

DWS is supported by AHA‐GIA and NHLR01 98744; JAT supported by AHA 15POST25730006
Post‐exercise changes in endothelin‐1 in military veterans with respiratory complaints
Authors: Salcedo Pablo , Jiao Xilu , Chen Yang , Klein Jacquelyn C , Ndirangu Duncan , Condon Michael R , Falvo Michael J
Published: November 2023

Post‐9/11 veterans frequently report respiratory symptoms following deployment to Iraq and Afghanistan, which may be related to deployment‐related exposures. Work from our clinic has observed evidence of pulmonary vascular dysfunction (i.e., normal spirometry and reduced lung diffusing capacity [DLCO])as a common abnormality in this population. The goal of the present study was to evaluate changes in endothelin‐1 (ET‐1), a key vascular regulator, before and after a maximal cardiopulmonary exercise test and determine its relationship with measures of pulmonary gas‐exchange. Seven post‐9/11 veterans(34.7±4.1 years) with a median deployment length of 13 months and dyspnea onexertion (5.6±1.2 Borg Scale), completed a modified Bruce protocol on amotorized treadmill. Pre and post‐exercise venous plasma concentrations of ET‐1 were obtained using sensitive ELISA. Pulmonary gas‐exchange was assessed during exercise (VE/VCO2 slope) and at rest (DLCO). Baseline concentrations of ET‐1(1.04±0.2 pg/mL) were similar to post‐exercise levels (0.93±0.3; p=0.32), with three individuals demonstrating an exercise‐induced increase. The change in ET‐1 (−10.3±23.4%) was associated with exercise ventilatory efficiency (VE/VCO2 slope; r =0.77; p = 0.04), but was not associated with DLCO. However, we observed a mild reduction in DLCO (<85% predicted) in 4 veterans and normal DLCO (>85%) in the remaining group of individuals. Atrend for higher concentration of ET‐1 was observed (1.48±0.5 vs. 0.93±0.03pg/mL; p=0.08) in those with low DLCO (78.8±0.04%) versus normal DLCO (93.9±0.03%). The present study demonstrated that exercise‐induced changes in ET‐1 were associated with inefficient exercise ventilation, and individuals with reduced gas‐exchange at rest exhibit higher plasma levels of ET‐1. Future studies may consider investigating ET‐1 and pulmonary gas‐exchange in deployed veterans presenting with unexplained dyspnea.

Support or Funding Information

VA RR&D 1I21RX001079‐01
Endothelial hyper‐permeability and cardiomyocyte dysfunction following Coronary Artery Bypass Grafting
Authors: Velten Markus , Duerr Georg Daniel , Hamiko Marwan , Frede Stilla , Rogers Lynette K , Baumgarten Georg , Hoeft Andreas , Hilbert Tobias
Published: November 2023

Objective

Ischemia Reperfusion Injury (IRI) is contributing to morbidity and mortality in various cardiac diseases including coronary artery bypass graft surgery (CABG). We hypothesized that CABG induced IRI is resulting in capillary leakage due to Angiopoietins (ANG) and cardiomyocytes dysfunction affecting perioperative morbidity after CABG.

Approach

After institutional revenue board approval and informed consent, serum samples (n = 28) were collected pre CABG surgery, 1, 6, and 24 h after aortic de‐clamping. ANG1, ANG2, sTIE2, HMGB1, sRAGE and IL‐6 serum concentrations were analyzed by ELISA. Endothelial cells (HPMECs) and primary murine cardiomyocytes were incubated with human serum to assess the impact of IRI on endothelial permeability and myocardial contractility after CABG.

Results

CABG induced an early and sustained increase of ratio of ANG2/ANG1 (7‐fold change in ANG2/ANG1 ratio after 24 h compared to pre‐surgery). These changes correlated with elevated serum lactate levels, postoperative fluid balance, as well as the duration of mechanical ventilation. Permeability of ECs significantly increased after incubation with post‐surgery patient serum showing a marked shift of ANG2/ANG1 balance (18‐fold) compared to serum with a less pronounced increase (6‐fold). Furthermore, CABG resulted in increased IL‐6, HMGB1, and sRAGE serum content. Pre‐incubation with serum containing high levels of IL‐6 amplified the ANG2 secretion by HPMECs however this was not influenced by blocking IL‐6. Moreover, contractility of murine cardiomyocytes was significantly reduced after incubation with human serum 1h after aortic de‐clamping. These changes correlated with increased HMGB1 and sRAGE levels.

Conclusion

CABG induced IRI altered the relative levels of serum ANG2 to ANG1, increased HMBG1 and sRAGE concentrations, resulting in endothelial hyper‐permeability and impaired cardiomyocytes contractility. This influences perioperative morbidity, leading to changes in clinical surrogate parameters of tissue hypo‐perfusion, vascular leakage, and pulmonary dysfunction.
Thermoregulatory and Ventilatory Responses in Humans with a Patent Foramen Ovale During Passive Cooling While Immersed in 20 °C Water
Authors: Davis James T , Hay Madeline W , Hardin Alyssa M , Lovering Andrew T
Published: November 2023

Approximately 35% of the general healthy population has a patent foramen ovale (PFO). People without a PFO (PFO−) have a lower esophageal temperature (Tesoph) under resting conditions and during exercise. Additionally, while breathing super cooled (−2.0 °C) air at the end of a 10‐minute exercise bout (2:30 stages at 25, 50, 75 and 90% of VO2MAX), PFO− subjects did not increase Tesoph as much as subjects with a PFO (PFO+). These data suggest blunted respiratory system cooling in PFO+ subjects. Taken together, these data suggest differences between PFO− and PFO+ subjects in thermoregulatory and ventilatory responses at rest and during exercise. However, to our knowledge it has not been prospectively determined if a PFO has any role on thermoregulatory and ventilatory responses to passive cooling.

Fourteen males matched for height, weight and age participated in this study (9 PFO−; age: 26 ± 6 yrs, height: 180.1 ± 7.2 cm, weight: 83.3 ± 12.3 kg, BSA: 2.0 ± 0.2 m2; 5 PFO+; age: 28 ± 6 yrs, height: 175.8 ± 7.3 cm, weight: 77.7 ± 6.0 kg, BSA: 2.0 ± 0.1 m2). Subjects came to the lab at the same time each morning and laid in a reclined position, immersed to the nipple line in a water bath (19.6 ± 0.7 °C) until 1) 60 minutes elapsed, 2) Tesoph decreased to 35.5 °C, 3) sustained shivering occurred (oxygen uptake [VO2] > 25% of resting values for 5 consecutive minutes), or 4) subjects asked to get out of the water.

We measured Tesoph, VO2 and minute ventilation (VE) every 5 minutes. At rest before immersion PFO− subjects had a lower Tesoph than PFO+ subjects (PFO−: 36.5 ± 0.2, PFO+: 36.8 ± 0.2, p = .04). At the end of immersion PFO− subjects also had a lower Tesoph than PFO+ subjects (PFO− : 36.0 ± 0.4, PFO+: 36.5 ± 0.3, p = .04). There were no differences in VO2 or VE at any time point. Furthermore, after the initial increase in Tesoph upon immersion, PFO− subjects decreased Tesoph throughout the remainder of the trial; while PFO+ subjects drop in Tesoph plateaued after 50 minutes of immersion. These data suggest that PFO− subjects are unable to retain heat as effectively as PFO+ subjects during passive cooling. This may be due to the possibility that PFO− subjects have greater respiratory system heat loss than PFO+ subjects.

Support or Funding Information

2015 Eugene and Clarissa Evonuk Memorial Graduate Fellowship
The role of nitric oxide (NO) in the expression of the human intermediate and small conductance calcium‐activated potassium channels, KCa3.1 and KCa2.3
Authors: Jones Heather Marie , Hussain Yusaf M
Published: November 2023

Modulation of the calcium‐activate potassium channels that play a role in vascular smooth relaxation may lead to changes in blood pressure by altering vessel tone. Nitric oxide (NO) is an intermediate molecule released from vascular endothelial cells which also will regulate vascular tone. This project hypothesized that exogenous NO will increase expression of calcium‐activated potassium channels to modulate vascular smooth muscle hyperpolarization and relaxation. Additionally, inhibition of endogenous NO will decrease expression of these channels. Total protein expression levels of the intermediate conductance calcium‐activated potassium channel, KCa3.1, and the small conductance calcium‐activated potassium channel, KCa2.3 were examined in this study. Cells endogenously expressing KCa3.1 and KCa2.3 were treated with increasing concentrations (1.0, 2.5 and 10.0 μM) of NO donors and a nitric oxide synthase inhibitor for varying incubation periods (4.0, 8.0 and 12.0 hours). The NO donors used were sodium nitroprusside (SNP) and n‐nitrosoglutathione (GSNO) and the nitric oxide synthase inhibitor used was Nω‐Nitro‐L‐Arginine Methyl Ester Hydrochloride (L‐NAME). Total protein from the treated cells was quantified and equal protein was run on SDS‐PAGE and protein bands were quantified and normalized to b‐actin. Concurrent, non‐treated cells were grown and run as a comparison at each time point. Western blots were scanned and bands intensity used as an indication of expression and quantified using ImageJ. Results showed that SNP significantly increased expression of KCa3.1 at all concentrations of the donor and between the 4 and 8 hour time intervals. The expression of KCa2.3 was also increased in the presence of SNP. Inhibition of KCa3.1 expression by L‐NAME was seen at a concentration of 2.5 μM at 8 hours (p = 0.032) and there was no significant inhibition of the KCa2.3 with L‐NAME. This study demonstrated a potential therapeutic effect that can be obtained in hypertensive patients by applying SNP and GSNO to increase expression of the KCa3.1 and KCa2.3 channels, which will play a role in increased endothelial derived hyperpolarization of vascular smooth muscle.
Sex Differences in the Expression of Cardiac miR‐29 Family microRNAs in Diabetic Male and Female Rats and its Correlation with Increased Risk for Coronary Artery Disease in Diabetic Females
Authors: Lum‐Naihe Kelly , Pulakat Lakshmi
Published: November 2023

Diabetic females are at increased risk of heart failure (HF) and coronary artery disease (CAD) than age‐matched diabetic males and non‐diabetic females. Factors contributing to this sex difference in diabetic heart disease are not understood. We have previously shown that progression of diabetes mellitus (DM) in male rats correlates with increased expression of cardiac miR‐29 family of microRNAs. The miR‐29 family is composed of three members who share an identical seed sequence, however, they exhibit different roles in vivo: miR‐29a is a biomarker of hypertrophy and fibrosis, miR‐29b is involved in aneurysm formation in Marfan's syndrome, and miR‐29c is a marker of hyperglycemia. Here we investigated whether sex differences exist in the expression of cardiac miR‐29 family in 5‐month old Zucker lean (ZL) and Zucker diabetic fatty (ZDF) rats, and examined whether overexpression of the miR‐29 family miRNAs is detrimental to human male and female coronary artery vascular smooth muscle cells (hCAVSMCs). Cardiac miR‐29 expression was assessed by qRT‐PCR and cardiac structure was examined by histological analysis. Expression of miR‐29a, b and c were elevated in both diabetic female (ZDF‐F) and male (ZDF‐M) rats when compared to lean female (ZL‐F) and male (ZL‐M) rats (p‐values for ZDF‐F vs. ZL‐F: miR‐29a, 0.054, miR‐29b, 0.03, miR‐29c, 0.02; for ZDF‐M vs. ZL‐M: miR‐29a, 0.0001, miR‐29b, 0.0015, miR‐29c, 0.0001), however, relative to lean controls, miR‐29a and miR‐29c were increased to a greater extent in ZDF‐M (~10.3‐ and 26.3‐fold greater, respectively) than in ZDF‐F (~4.8‐ and 8.8‐fold, respectively). In contrast, in ZDF‐F, miR‐29b was ~18.1‐fold greater than ZL‐F, while in ZDF‐M, miR‐29b was ~4.7‐fold higher than ZL‐M. Examination of cardiac sections stained with H&E or Masson's trichrome revealed structural damage in ZDF‐F, which was not observed in any other group. Transfection of primary cultures of male and female hCAVSMCS with miR‐29 mimics individually showed that 48‐hour post‐transfection, loss of cell membrane integrity, as assessed by propidium iodide (PI) exclusion assay, was highest in cells transfected with miR‐29b compared to those transfected with miR‐29a or miR‐29c. TUNEL assay further confirmed this detrimental effect of miR‐29b in hCAVSMCs. Since miR‐29b expression was higher in heart tissues of ZDF‐F compared to all other groups, and miR‐29b was most effective in inducing cell death compared to other miR‐29 family miRNAs in human CAVSMCs, we conclude that the sex difference in miR‐29b expression may underlie higher risk for CAD in diabetic females.

Support or Funding Information

LSME, UM‐Columbia(LP), NIH 1R01HL118376‐01(LP)
Increasing Muscle Mass Improves Cavernosal Function in Obesity
Authors: MicKinac Nathan K. , Larion Sebastian , Mintz James D. , Fulton David J. , Stepp David W.
Published: November 2023

Obesity is a known risk factor for erectile dysfunction but the causal mechanisms remain poorly understood. Moreover, exercise is known to improve erectile function but the components of exercise (weight loss, aerobic capacity, or muscle mass) that contribute are unclear. The purpose of our study was to investigate whether increased muscle mass in an obese mouse model improves the erectile function of the corpus cavernosum.

Lean mice heterozygous for db, the leptin receptor, were bred to mice lacking myostatin, a myokine the limits muscle growth. Four groups were studied. Lean controls, lean myostatin‐null mice, obese control and obese myostatin‐null mice. All mice were phenotyped using metabolic cages and analysis of plasma chemistry. Responses of cavernosal strips were assessed in vitro in response to phenylephrine and acetylcholine.

Obese mice compared to lean were characterized by a two‐fold increase in body weight, marked adiposity, low oxygen consumption, muscle atrophy, dyslipidemia and hyperglycemia. Deletion of myostatin in lean mice markedly increased muscle mass (35%, p<0.05) but otherwise produced no phenotypic alterations. Deletion of myostatin increased muscle mass to a similar degree as lean mice and improved glucose control in obese mice with no effect on lipids, body weight or body composition.

Despite similar cavernosal mass, obese mice showed increased vasoconstrictor force generation (which would restrain erection) to phenylephrine when compared to the lean controls (102%, p<0.05). Deletion of myostatin had no effect in lean mice but reduced constriction to normal values in obese mice. Vasodilation to acetylcholine tended to be reduced in obese mice but no significant differences emerged among the four groups.

Overall, these results support the hypothesis that increased muscle tone can improve erectile function in obese mice. This effect, potentially due to improved glucose tolerance by the larger muscle mass, may explain to some degree how exercise improves erectile function in obesity.

Support or Funding Information

Work supported by NHLBI 092446 and 124773 and the Dean's Summer Research Program
Helium induces Caveolin Secretion and decreases Permeability in Human Umbilical Vein Endothelial Cells
Authors: Landau Maximilian Alexander , Smit Kirsten Fernande , Warmbrunn Moritz , Preckel Benedikt , Hollmann Markus Werner , Patel Hemal H , Albrecht Martin , Weber Nina Claudia
Published: November 2023

Caveolins are structural proteins essential for the formation of caveolae, small lipid enriched membrane invaginations. Caveolins have been shown to be involved in anesthetic‐induced cardioprotection and play an important role in signal transduction in various cell types. The noble gas helium, a potent cardioprotectant, leads to decreased protein levels of caveolin‐1 and caveolin‐3 in mice hearts, whereas higher levels of these caveolins were found in the serum of mice. These data suggest that caveolins are secreted into the bloodstream after helium inhalation. As actin filaments are an integral part of the cytoskeleton and are located in close connection to caveolins in the plasma membrane we hypothesized that helium might affect the cytoskeleton and thereby induce secretion of caveolins. To investigate this hypothesis we used human umbilical vein endothelial cells (HUVEC), as the endothelium is considered the first target organ that gets in contact with circulating blood.

HUVEC were isolated from fresh umbilical cords, pooled and grown upon a confluent monolayer in passage 3–5. Cells were exposed for 20 minutes to either helium (5% CO2, 25% O2, 70% He) or control gas (5% CO2, 25% O2, 70% N2) in a specialized gas chamber. Cells and supernatant were collected after 0 minutes, 2, 4, 6, 12 and 24 hours for Western blot analysis, RT‐PCR, immunofluorescence staining and permeability measurements.

In the helium group, protein analysis showed a significantly lower expression of caveolin‐1 at 6 hours in cell lysate (13.35±1.30 in controls vs. 8.02±1.29 in helium treated cells, average light intensity (AVI), p<0.05) and a higher expression in supernatant starting at 6h (1.42±0.25 in controls vs. 8.26±4.5 in helium treated cells, AVI, p<0.05) Actin staining demonstrated a significant increase of the border/inner ratio of the filaments at 12 hours after helium exposure. Cell permeability measurement revealed a significantly lower flow‐through of FITC‐BSA at 6 (p<0.05) and 12 hours (p<0.05) in the helium group. RT‐PCR of Cav‐1 did not show significant differences.

These findings suggest that caveolin‐1 is secreted after helium exposure in vitro and that the actin cytoskeleton filaments are regulated by helium. This mechanism might finally lead to a decreased permeability of the endothelium after helium inhalation.

Support or Funding Information

This project was in part funded by the IARS (International Anesthesia Research Society) and the SCA (Society of Cardiovascular Anesthesiologists).
Ventilatory Efficiency and Oscillatory Breathing As Clinical Markers of Surgical Outcomes in Heart Failure
Authors: Jesse Nicholas , Raichlin Eugenia , Schultz Harold
Published: November 2023

Background

Ventilatory efficiency (Ve/VCO2) and oscillatory breathing (OB) during exercise have been shown to be important predictors of long‐term survival in patients with chronic systolic heart failure (CHF). However, the status of Ve/VCO2 and OB in patients after undergoing heart transplant (HT) or left ventricular assist device (LVAD) implant is less clear. The aim of this study was to assess the outcomes of HT or LVAD implantation on Ve/VCO2 and OB. We hypothesized that Ve/VCO2 and OB are improved (i.e. decreases in Ve/VCO2 slope and breathing interval variability) after HT and LVAD and that these changes correlate with improved exercise tolerance (6 minute walk).

Methods

Cardiopulmonary exercise test (CPET) data were retroactively analyzed for Ve/VCO2 slope and OB (coefficient of variability of breath interval) during progressive treadmill exercise from eight CHF patients: HT (n=4), and LVAD (n=4). Sample size was restricted due to insufficient CPET data in most HT/LVAD patient records at the time of analysis.

Results

We found that Ve/VCO2 was reduced in three patients in each group after the intervention [LVAD: 38 ± 6.1(SD) before vs. 34 ± 3 after (6 % improvement) n =4] [HT: 45 ± 17 before vs. 35 ± 6 after (12 % improvement) n= 4]. The reduction in Ve/VCO2 in the three patients from each group correlated with a reduction in exercise OB. One patient in each group exhibited increased Ve/VCO2 and OB after the intervention. Overall, the direction and magnitude of the change in Ve/VCO2 correlated well with changes in OB for all patients (r2 = 0.70). There also was a correlation between improved VE/CO2 or OB and improved exercise tolerance (increased distance walked in 6 min).

Conclusions

Ve/VCO2 and OB were improved after LVAD or HT implant in the majority of CHF patients tested and correlated with improved exercise tolerance. Patients will be followed to determine long‐term survivability. Further analysis as additional CPET data becomes available will give a more complete picture of the efficacy of these measurements as diagnostically relevant markers of improved ventilatory function and long‐term outcomes in LVAD and HT patients.

Support or Funding Information

NIH PO1‐HL62222
Doxorubicin Decreases Cyclooxygense‐2 Levels in Heart, Brain, and Pial Arteries from Ovariectomized Female Rats
Authors: Gonzales Rayna , Raman Puneet , Vijayavel Nirmal , Kerrigan Colleen , Hale Taben , Carroll Chad , Dickinson Jared , Angadi Sidartha
Published: November 2023

Doxorubicin (DOX) is a highly effective chemotherapy agent. Its use is hampered however owing to severe dose‐dependent cardiovascular toxicity in cancer survivors. Multiple mechanisms have been implicated in the pathogenesis of DOX cardiotoxicity, one of which involves inflammation mediated by activation of the NFκB/TLR4/COX‐2 pathway in the heart. Knowledge regarding the toxic effects of DOX‐induced inflammation in other organ systems such as the brain has not been thoroughly investigated. Therefore, we explored the inflammatory potential of DOX by assessing TLR4 and COX‐2 levels in cortex and pial arteries isolated from ovariectomized (OVX) female Sprague‐Dawley rats. We hypothesized that DOX would promote inflammation by increasing COX‐2 expression along with expression of its upstream innate immune receptor, TLR4, both of which are under the transcriptional regulation of NFκB. OVX rats were treated with three, bi‐weekly, i.p. injections of DOX (4 mg/kg; cumulative dose 12 mg/kg) or vehicle (saline) and euthanized 5 days after the last dose. Tissues were isolated, snap frozen, homogenized, and analyzed for COX‐2, TLR4 and NFκBp65 levels using standard western blotting. Although COX‐2 is typically considered an inducible enzyme, it has been shown to be expressed under basal conditions. In cortex and pial arteries from vehicle‐treated rats, measurable levels of COX‐2 and TLR4 protein were detected. However, contrary to our hypothesis, levels of COX‐2 and TLR4 were decreased following DOX. In pial arteries, DOX elicited a reduction in both the COX‐2 72 kDa band 74 kDa band with the greatest reduction observed in the 72 kDa. The 72 kDa band corresponds to the partially glycosylated inactive form of enzyme, while the 74 kDa band has been shown to represent the fully glycosylated active form. Cytosolic levels of NFκBp65 levels were detected in brain and pial vessel lysate, however levels were not altered by DOX. Similar to brain and pial arteries, basal expression of COX‐2 and TLR4 were detected in left ventricle and DOX treatment attenuated expression. In conclusion, although others have suggested the involvement of the NFκB pathway during the development and progression of DOX‐induced cardiomyopathy, our studies demonstrate a possible novel action for the anticancer agent implicating anti‐inflammatory mechanisms, particularly in female cohorts with low circulating levels of gonadal hormones.

Support or Funding Information

Arizona State University Intramural Funds (SA and JD), Midwestern University Intramural Funds (CC), Sarver Heart Center (RG)
Chronic inhibition of endoplasmic reticulum (ER) stress attenuates cardiovascular but not appetite responses to MC3/4R antagonism in SHR
Authors: do Carmo Jussara Marcia , da Silva Alexandre A. , Wang Zhen , Hall John E.
Published: November 2023

We previously demonstrated that activation of the central nervous system (CNS) melanocortin system plays a key role in modulating sympathetic nerve activity (SNA) and blood pressure (BP) in spontaneous hypertensive rats (SHR). Previous studies also suggest that endoplasmic reticulum (ER) stress may contribute to cardiovascular pathophysiology and hypertension in SHR. However, whether there is a major interaction of ER stress and melanocortin‐4 receptor (MC4R) activation in contributing to hypertension and appetite regulation in SHR is still unknown. In this study we examined the role of ER stress alone or in combination with MC3/4R antagonism in controlling BP and metabolic functions in SHR. Male SHR (n=7) were implanted with BP telemetry transmitters and an intracerebroventricular (ICV) cannula was inserted into the lateral ventricle at 12 weeks of age. After 10 days of recovery, food intake, BP and heart rate (HR) were measured 24‐hrs/day. After stable baseline measurements for 5 days, tauroursodeoxycholic (TUDCA, 100 mg/kg, IP) was injected daily for 22 consecutive days to inhibit ER stress, which was followed by a 10‐day combined treatment with the MC3/4R antagonist (SHU‐9119, 1 nmol/h, ICV) for 10 days. Chronic ER stress inhibition alone did not alter food intake (16±1 vs. 14±3g), body weight (293±4 vs. 301±4 g), MAP (166±2 vs. 169±3 mmHg) or HR (335±6 vs. 338±2 bpm). Combined treatment with MC3/4R antagonist significantly increased food intake by 120% and body weight by 19%, while reducing BP by 9±2 mmHg and HR by 28±5 bpm when compared to −25 mmHg and −45 bpm with MC3/4R antagonist alone. No changes were observed in blood glucose levels. These results indicate that chronic inhibition of ER stress did not alter appetite, HR or BP, but attenuated BP and HR responses to MC3/4R antagonist in SHR.

Support or Funding Information

(NHLBI‐PO1HL51971, NIGMS‐P20GM104357 and AHA SDG5680016)
Severe Sleep Apnea Is Associated with Telomere Lengthening
Authors: Polonis Katarzyna , Singh Prachi , Becari Christiane , Covassin Naima , Druliner Brooke , Johnson Ruth , Boardman Lisa , Somers Virend K
Published: November 2023

Background

Obstructive sleep apnea (OSA) increases the risk for metabolic and cardiovascular disease. Telomere shortening has been linked to hypertension, diabetes mellitus, and cardiovascular diseases such as stroke and myocardial infarction. Because these conditions are highly prevalent in sleep apnea, we hypothesized that telomere length (TL) would be reduced in patients with OSA.

Methods

Genomic DNA was isolated from circulating leukocytes in 176 subjects (42.8 ± 13.8 years, 22.7 % women) who had undergone overnight polysomnographic evaluation. Of these individuals, 92 subjects had an Apnea Hypopnea Index (AHI) >5 events/hour and were classified as OSA patients. TL was measured using a quantitative PCR where a telomere to single copy gene ratio is determined and represented as number of base pairs (bp). The effect of OSA on TL was determined by comparing a control group (no OSA) (AHI<5, n=92) with patients with mild OSA (≥ 5 AHI <15, n=25), and those with moderate to severe OSA (AHI ≥15, n=59) in unadjusted and age‐adjusted models.

Results

Compared to control subjects, mild and moderate to severe OSA subjects were older: 48.9 ± 11.2 vs. 36.0 ± 11.2 years (p<0.001) and 50.8 ± 13.1 vs. 36.0 ± 11.2 years (p<0.001), respectively. Statistical models examining the effects of OSA severity on TL showed a J‐shaped curve : longer TL in moderate to severe OSA (4867 ± 221 bp) and controls (4820 ± 160 bp), and the shortest TL in mild OSA (4742 ± 155 bp). The mean TL in moderate to severe OSA was significantly longer than in mild OSA (p=0.0048). These effects were more pronounced after adjustment for age, when we observed significant differences in mean TL between moderate to severe OSA and mild OSA (p=0.0036) and between moderate to severe OSA and the control group (p=0.0379) (Fig. 1). However, the mean TL between control subjects and subjects with mild OSA was not different.

Conclusion

Moderate to severe OSA patients had longer TL compared with mild OSA and control group. This association was independent of age. Even though OSA is associated with increased cardiovascular risk, telomere lengthening in subjects with moderate to severe OSA suggests the presence of protective compensatory mechanisms to prevent telomere shortening in OSA. On the other hand, the existence of longer telomere in subjects with moderate and severe OSA may also indicate triggering of molecular mechanisms which may contribute to cancer development. Further studies are needed to explain the role of hypoxia in telomere dynamics in OSA, and the interactions between telomere length and increased longevity reported in older patients with OSA.

Support or Funding Information

NIH R01 grant (#HL65176); Singh P. is supported by AHA scientist development grant (11SDG7260046). Becari C. is supported by grant from CNPq‐Brazil (# 203802/2014‐4).
Chronic Insulin‐clamp Causes Postprandial Diuresis in Sprague Dawley Rats
Authors: Irsik Debra L. , Alaisami Rabei , Washington Ashley R. , Brands Michael W.
Published: November 2023

Understanding the physiologic role of insulin in salt and volume homeostasis is important for interpreting its contribution to disease states such as diabetes or metabolic syndrome. This study tested the hypothesis that the rise in plasma insulin that occurs postprandially is required to minimize renal sodium (Na+) and volume losses. Rats were instrumented with chronic artery and vein catheters, housed in metabolic cages, and connected to Instech hydraulic swivels. Na+ intake was clamped chronically at normal by feeding Na+ deficient diet and infusing saline 24 hr/day. Measurements of 24 hr Na+ and volume excretions and 4‐ and 8‐hr postprandial Na+ and volume excretions were made under control conditions (CTL) and during insulin‐clamped conditions (I‐C) to prevent the postprandial insulin surge. The chronic I‐C was accomplished by administering STZ and subsequently infusing insulin 24 hr/day to clamp insulin at normal levels that maintained normal fasting blood glucose. Twenty‐four hr urine volume and Na+ excretion were greater in the I‐C vs. CTL rats (33±2 vs. 17±1 ml/day and 3.8±0.1 vs. 2.9±0.2 mmol/day, respectively; n=5–9; p<0.05) despite lower fasting blood glucose levels (93±9 vs. 129±5 mg/dl). Postprandial measurements were made after feeding a calorie‐dense gel to rats that had been fasted overnight. Postprandial urine volume was greater in I‐C rats after 4 hrs (5.5±1.6 vs. 2.7±0.3 ml, n=4–8; p<0.05) and 8 hrs (7.0 vs. 2.8 ml), and postprandial GFR was lower in the I‐C rats (1.8±0.4 vs. 2.2±0.4 ml/min), consistent with decreased tubular reabsorption. Postprandial blood glucose increased more in the I‐C rats (307±80 vs. 141±11 mg/dl, n=4–8; p<0.05). These data suggest that the postprandial surge in insulin is necessary to prevent renal salt and volume losses, because rats that were unable to increase insulin after a meal exhibited increased diuresis. Future studies will be needed to quantify the relative roles of insulin‐mediated glucose disposal vs. insulin‐mediated renal tubular Na+ reabsorption.

Support or Funding Information

HL056259 and HL056259‐S1
High Salt Activation of Collecting Duct‐Derived Nitric Oxide Synthase Suppresses the Renin‐Angiotensin‐Aldosterone (RAAS) System
Authors: Pollock Jennifer S , Pollock David M , Dugas Courtney , Hyndman Kelly A
Published: November 2023

We reported that collecting duct nitric oxide synthase‐1 knockout (CDNOS1KO) mice display a salt‐sensitive blood pressure phenotype with reduced Na excretion in response to a high salt (HS) diet. To further delineate the relationship of CD NOS1 activation and Na homeostasis, we designed experiments to test the hypothesis that HS induced activation of CD NOS1 functions to suppress the RAAS. Adult male CDNOS1KO and flox control mice were placed on normal (NS) or HS diet for 7 days (NS: 0.4% NaCl, HS: 4% NaCl). NO production in isolated CD preparations from the CDNOS1KO mice was significantly reduced compared to control mice on NS (0.5±0.09 vs 1.0±0.03 pmol/mg pr/h, p< 0.05) and HS (1.5±0.3 vs 2.8±0.2 pmol/mg pr/h, p<0.05). HS induced urinary NOx excretion was also significantly reduced in CDNOS1KO mice by >50% compared to control. HS induced significantly higher plasma Na levels in CDNOS1KO mice (+8 mEq/L; p<0.05) compared to control mice. Plasma K levels were similar in NS and HS in both genotypes. HS significantly reduced plasma aldosterone (532.6±93.0 vs 55.6±10.4 pg/ml) and urinary aldosterone excretion in the control mice (4.2±0.7 vs 1.7±0.4 ng/day), however this reduction was abolished in CDNOS1KO mice (402.0±68 vs 276.5±55 pg/ml; 4.1±0.8 vs 3.7±0.7 ng/day). Plasma renin concentration was similar in both genotypes with HS intake yet blood pressure was significantly higher in CDNOS1KO mice, thus CDNOS1KO mice display an in appropriately high level of plasma renin with HS. Furthermore, renal renin expression was enhanced in CDNOS1KO mice compared to controls. These data demonstrate that loss of CD NOS1 facilitates sustained elevations in plasma Na levels as well as plasma and intra‐renal renin that further enhances activation of aldosterone. These findings indicate that CD NO, especially NOS1‐derived, is vital for systemic Na homeostasis under HS conditions.

Support or Funding Information

NIH P01HL95499
Median Preoptic AT1a Receptor Knockdown Does Not Diminish Drinking in Response to Peripheral Angiotensin II Administration
Authors: Shell Brent , Farmer Gef , Emeh Robert , Little Joel , Cunningham Tom
Published: November 2023

Thirst is an essential mechanism for organisms to maintain body fluid homeostasis and is therefore tightly regulated. Angiotensin II (Ang II) has long been known to stimulate thirst by both by its action peripherally as a circulating hormone, and centrally acting as a peptide neurotransmitter. Peripherally circulating Ang II can elicit thirst by stimulating the Subfornical Organ (SFO), which in turn stimulates other hypothalamic nuclei regulating body fluid homeostasis such as the Median Preoptic Nucleus (MnPO) presumably through Ang II signaling. Central Ang II administered via intracerebroventricular (ICV) infusion also induces thirst by directly stimulating regions such as the MnPO. Previous studies have shown that lesions of the MnPO block drinking responses to peripheral or ICV Ang II while lesions of the SFO decrease drinking stimulated by peripheral Ang II but not ICV Ang II. Based on these observations, it has been hypothesized that Ang II receptors in the MnPO are critical for drinking responses to centrally or peripherally administered Ang II. We tested this hypothesis by microinjecting an adeno‐associated virus with short hairpin RNA matched to the Angiotensin Type 1a receptor (AT1aR) into the MnPO and testing the rats for drinking responses elicited by peripheral or centrally administered Ang II. Sprague Dawley rats were separated into subcutaneous (SC) and ICV Ang II administration groups. The SC group was pretested for their drinking response to 2 mg/kg Ang II, and those animals that drank in response to the SC Ang II administration were utilized in the study. All animals were injected with the shAt1aR virus or a control virus on Day 0, and were allowed to recover before drinking tests on day 14 and day 18. ICV animals were microinjected with the viruses on Day 0 and instrumented with a chronic lateral ventricle cannula. These animals were tested for drinking responses to 2 ng Ang II in 1ul aCSF 14 and 18 days later. Unexpectedly, we found that knockdown of AT1aR in the MnPO did not reduce drinking in response to SC Ang II (p>.05), but did significantly reduce drinking in response to ICV Ang II (P<.05). These results suggest that AT1aRs in MnPO are necessary for drinking responses following ICV Ang II but not SC Ang II. It is possible that drinking responses to peripheral Ang II involves co‐transmitters, such as glutamate, in the MnPO that produce drinking in the absence of AT1aRs or that other Ang II receptors mediate this response.

Support or Funding Information

R01 HL119458

P01 HL088052
Blood Pressure in Old Female SHR is not Salt Sensitive and Tempol Fails to Decrease Blood Pressure during High Salt Intake
Authors: Dalmasso Carolina , Maranon Rodrigo O. , Patil Chetan N. , Reckelhoff Jane F.
Published: November 2023

Oxidative stress contributes to the development and maintenance of hypertension in male rodents; however studies in females have rarely shown a reduction in blood pressure (BP) with antioxidants. Tempol, a superoxide dismutase mimetic, decreases BP in young male spontaneously hypertensive rats (SHR), but fails to reduce BP in either young or old female SHR, despite the fact that females have similar or higher levels of oxidative stress markers. In contrast, recent experiments from our laboratory have shown that Tempol decreases BP in old female SHR treated chronically with Acetazolamide. Acetazolamide is a carbonic anhydrase inhibitor that increases sodium delivery to the distal nephron and may increase distal oxidative stress. Thus the data suggest that oxidative stress‐mediated BP control is dependent on increased sodium delivery to the distal nephron. The aim of the present work was to test the hypothesis that tempol will decrease blood pressure in old female SHR chronically given a high salt intake. Female SHR (20 months of age) were divided into two groups (n=7/group): Control + Tempol (C+T), and High Salt Diet + Tempol (HSD+T). After baseline mean arterial pressure (MAP; telemetry; 5 days), rats received low salt diet (NaCl 0.3%; C+T) or high salt diet (NaCl 8%; HSD+T). On day 22, rats in both groups (C+T and HSD+T) were given tempol (30 mg/kg/d) in the drinking water for 21 days. Baseline MAP was similar between groups (C+T: 174±7; HSD+T: 168±5 mm Hg, p=NS), and MAP was not salt sensitive (C+T: 178±5; HSD+T: 178±4 mm Hg, p=NS). Tempol treatment had no effect on MAP in either group of female SHR (C+T: 179±5; HSD+T: 178±5 mm Hg). Because tempol in acetazolamide treated female SHR reduces MAP, whereas tempol has no effect on MAP in females on high salt diet, the data suggest that in hypertensive female rats, tempol reduces MAP only when proximal sodium reabsorption is blocked.

Support or Funding Information

Supported by NIH‐R01HL66072, PO1HL51971 (JFR), 14POST18640015 (ROM).