Endothelium-dependent Vasodilator Responses Research Articles

TNF-TNFRI interaction impairs endothelial calcium signaling and elevates blood pressure in obesity

Endothelial dysfunction in small arteries plays a pivotal role in obesity-induced hypertension. Recent investigations have unveiled the detrimental impact of peroxynitrite, a reactive nitrogen species, on endothelium calcium signaling and vasodilation. Vascular inflammation has emerged as a crucial contributor to the pathogenesis of cardiovascular disorders. In this regard, the inflammatory cytokine, tumor necrosis factor (TNF), has been linked to peroxynitrite formation and endothelial dysfunction. Although TNF signaling in cardiovascular disorders is mainly attributed to immune cells, recent studies suggest that endothelial cells (ECs) and smooth muscle cells (SMCs) also produce TNF. We hypothesized that TNF derived from the vascular wall acts locally to inhibit endothelium-dependent vasodilation in obesity. Male C57BL6J mice fed a high-fat diet (HFD, 60% kcal fat) for 14 weeks showed higher blood pressures that normal chow diet (NCD)-fed mice. Flow cytometry and mRNA expression studies showed an increase in TNF levels in freshly isolated SMCs from small mesenteric arteries (MAs) in HFD mice compared to NCD mice. However, TNF levels in ECs were not different between HFD or NCD mice. Control and EC-specific TNF knockout (TNFECKO) mice showed a similar increase in blood pressure after HFD feeding. However, HFD feeding failed to elevate blood pressure in SMC-specific TNF knockout (TNFSMCKO) mice, suggesting a critical role for SMC TNF in blood pressure elevation in HFD mice. Both TNFSMCKO and TNFECKO showed a similar increase in body weight after HFD feeding when compared to control mice, suggesting that differences in weight gain did not contribute to lower blood pressure in obese TNFSMCKO mice. The pro-inflammatory effects of TNF are primarily attributed to TNF receptor-I (TNFRI) signaling. Therefore, we postulated that TNF released from SMCs acts locally on EC TNFRI to impair endothelium-dependent vasodilation. EC-specific TNFRI deletion (TNFRIECKO) reduced blood pressure, whereas SMC-specific TNFRI deletion (TNFRISMCKO) had no effect on blood pressure after HFD feeding. These findings collectively underline the critical role of SMC-derived TNF and endothelial TNFRI in driving blood pressure elevation in diet-induced obesity. Consistent with these data, endothelium-dependent vasodilation in response to acetylcholine markedly improved in obese TNFSMCKO and TNFRIECKO mice compared to obese control mice. Previous studies showed that decreased Ca2+ influx through endothelial TRPV4 (transient receptor potential vanilloid 4) channels contributes to endothelia dysfunction and blood pressure elevation in obese mice. The activity of TRPV4 channels was improved in ECs from HFD-fed TNFSMCKO and TNFRIECKO mice. Additionally, the levels of endothelial peroxynitrite were diminished in obese TNFSMCKO and TNFRIECKO mice, ameliorating the peroxynitrite-dependent impairment of endothelial function. Furthermore, treatment with a selective TNFRI signaling inhibitor (R7050, 10 mg/kg, intraperitoneally, once daily for 14 days) restored blood pressure to normal levels in HFD-fed obese mice but had no effect on the blood pressure in NCD-fed mice. R7050 treatment also restored endothelium-dependent dilation of MAs and the activity of TRPV4 channels in ECs. Collectively, our findings identify a critical role of local TNF-TNFRI signaling in the vascular wall in obesity-induced impairment of endothelial function and blood pressure elevation. The National Institutes of Health to SKS (HL146914, HL142808, and HL147555). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Enhanced Microvascular Adaptation to Acute Physical Stress and Reduced Oxidative Stress in Male Athletes Who Consumed Chicken Eggs Enriched with n-3 Polyunsaturated Fatty Acids and Antioxidants-Randomized Clinical Trial.

This randomized interventional study aimed to determine the effects of n-3 polyunsaturated fatty acids, selenium, vitamin E, and lutein supplementation in the form of enriched chicken egg consumption on microvascular endothelium-dependent vasodilation, oxidative stress, and microvascular response to an acute strenuous training session (ASTS) in competitive athletes. Thirty-one male athletes were assigned to a control (n = 17) or a Nutri4 group (n = 14) who consumed three regular or enriched chicken eggs per day, respectively, for 3 weeks. Significantly enhanced endothelium-dependent responses to vascular occlusion (PORH) and iontophoresis of acetylcholine (AChID) were observed in the Nutri4 group but not in the control group after egg consumption. Formation of peroxynitrite and hydrogen peroxide in peripheral blood mononuclear cells, as well as serum concentration of 8-iso prostaglandin F2α, decreased in the Nutri4 group while remaining unchanged in controls. PORH and AChID were reduced post-ASTS compared with pre-ASTS, both before and after the diets, in both groups. However, the range of PORH responsiveness to ASTS (ΔPORH) increased after consumption of enriched eggs. These results suggest that consumption of enriched chicken eggs has a beneficial effect on microvascular endothelium-dependent vasodilation and the reduction of oxidative stress levels in competitive athletes. Also, microvascular adaptation to the ASTS was improved after consumption of Nutri4 eggs.

Acute daily exposure to ambient air pollution impairs endothelium-dependent vasodilator responses in young, healthy individuals.

Exposure to ambient air pollution influences cardiovascular (CV) morbidity and mortality. The differential effects of changing particulate or gaseous air pollution on endothelial function in young healthy individuals remain unclear. The aim of this study was to evaluate the relationships between exposures to different pollutants and vascular function in a group of 39 young (33±11 years old) subjects with low CV risk. Flow-mediated dilatation (FMD) and nitroglycerin-mediated dilatation (NMD) were performed, when air pollution reached highest levels (heating period) and repeated in a subgroup of 18 participants a few months later (just before the heating period starts). Daily mean concentrations of PM2.5 and PM10 were inversely correlated with FMD, and this relationship remained significant after adjusting for factors known to affect vascular dysfunction. Endothelial function did not differ between the two time points studied. However, we observed a strong inverse association between the change in the concentration of particulate matter (deltaPM2.5 and deltaPM10) and the change in FMD (deltaFMD) between the two visits (R= -0.65, p= 0.02; R= -0.64, p= 0.02, respectively). In summary, we provide evidence that the concentration of PM2.5 and PM10, but not SO2, NO, NO2, CO, or O3 is associated with impaired endothelial function in young, healthy individuals.

Imaging photoplethysmography quantifies endothelial dysfunction in patients with risk factors for cardiovascular complications

The endothelium plays an important role in the pathogenesis of cardiovascular pathology, so that revealing its dysfunction can predict outcomes of the disease. However, the correct assessment of the endothelial function remains an unsolved problem. Registration of endothelium dependent vasodilation in response to nitric oxide synthesis using available noninvasive technologies does not show sufficient specificity and sensitivity, although it helps to detect its differences in population studies. In this pilot study, we compared the dynamics of microcirculation of the forearm skin and its spectral parameters during mild heating (up to 42 °С) between groups of smokers (n = 17) and non-smokers (n = 21) using imaging photoplethysmography synchronized with electrocardiography. The study included healthy men with similar anthropometric characteristics at the age of 51 ± 8 years and 48 ± 13 years, respectively. During the comparative analysis, no intergroup difference was found in the perfusion increase in either the axon- or endothelial-related phases of the reaction to local hyperemia. However, it was revealed that the power spectral density in the frequency band 0.01–0.02 Hz is significantly smaller in smokers than in non-smokers only during the phase of endothelium induced vasodilation: 4.0 ± 1.1 versus 5.3 ± 1.2 rel.un., respectively (P = 0.005). At the same time, no difference in the spectral power in the same band was observed during the axon-mediated phase, thus indicating high sensitivity and selectivity for assessment of the nitric oxide dependent vascular reactivity. The obtained results can be used for noninvasive evaluation of other cardiometabolic risk factors for early prediction and timely prevention of adverse outcomes by optimizing and intensifying treatment.

Abstract 484: Antidiabetic Agent, Canagliflozin, Is A Potent Vasodilator Of The Coronary Microvasculature

Objective: Sodium-glucose cotransporter-2 (SGLT-2) inhibitors are cardioprotective even in the absence of preexisting diabetic microvascular disease. Our group has shown SGLT-2 inhibitor, canagliflozin, increases myocardial perfusion without an attendant increase in angiogenesis. We hypothesize canagliflozin accentuates vasodilation of coronary microvasculature to increase perfusion. Methods: Coronary resistance arterioles (174 ± 60μm in diameter) isolated from Sprague-Dawley rats either receiving daily canagliflozin (15mg/kg, oral gavage) for four weeks (n = 4) or no treatment (n = 4) were studied in vitro in a pressurized (40mmHg), no-flow state using video microscopy. Following preconstriction of microvessels with thromboxane analog U46619 or endothelin-1, concentration response curves to vasodilatory adenosine 5’-diphosphate (ADP), acetylcholine (ACh), and sodium nitroprusside (SNP) were completed. Additionally, nitric-oxide-dependent ADP-induced vasodilation of microvessels isolated from treated rats was measured in the presence of N ω -Nitro-L-arginine methyl ester (L-NAME) 10μM. Results: Canagliflozin treatment was associated with a significant increase in endothelium-dependent vasodilatory response to ADP (p = 0.039) (Figure 1). Pretreatment with L-NAME, an inhibitor of nitric oxide synthase, eliminated this vasodilatory response. Compared to ADP, vasodilatory response to ACh was muted (p = 0.552). Endothelium-independent vasodilatory response to SNP, a nitric oxide donor, strongly trended toward an increase in the treated group (p = 0.099). Conclusions: Canagliflozin improves microvascular vasodilation in an endothelium-dependent and endothelium-independent fashion not yet described. These findings implicate both vascular endothelium and arteriolar smooth muscle cells in altered vasomotor physiology and may, in part, account for the observed increase in myocardial blood flow with canagliflozin treatment.

Hyperoxaluria Induces Endothelial Dysfunction in Preglomerular Arteries: Involvement of Oxidative Stress.

Urolithiasis is a worldwide problem and a risk factor for kidney injury. Oxidative stress-associated renal endothelial dysfunction secondary to urolithiasis could be a key pathogenic factor, similar to obesity and diabetes-related nephropathy. The aim of the present study was to characterize urolithiasis-related endothelial dysfunction in a hyperoxaluria rat model of renal lithiasis. Experimental approach: Endothelial dysfunction was assessed in preglomerular arteries isolated from control rats and in which 0.75% ethylene glycol was administered in drinking water. Renal interlobar arteries were mounted in microvascular myographs for functional studies; superoxide generation was measured by chemiluminescence and mRNA and protein expression by RT-PCR and immunofluorescence, respectively. Selective inhibitors were used to study the influence of the different ROS sources, xanthine oxidase, COX-2, Nox1, Nox2 and Nox4. Inflammatory vascular response was also studied by measuring the RNAm expression of NF-κB, MCP-1 and TNFα by RT-PCR. Results: Endothelium-dependent vasodilator responses were impaired in the preglomerular arteries of the hyperoxaluric group along with higher superoxide generation in the renal cortex and vascular inflammation developed by MCP-1 and promoted by NF-κB. The xanthine oxidase inhibitor allopurinol restored the endothelial relaxations and returned superoxide generation to basal values. Nox1 and Nox2 mRNA were up-regulated in arteries from the hyperoxaluric group, and Nox1 and Nox2 selective inhibitors also restored the impaired vasodilator responses and normalized NADPH oxidase-dependent higher superoxide values of renal cortex from the hyperoxaluric group. Conclusions: The current data support that hyperoxaluria induces oxidative stress-mediated endothelial dysfunction and inflammatory response in renal preglomerular arteries which is promoted by the xanthine oxidase, Nox1 and Nox2 pathways.

MATERNAL SALT INTAKE AND MICROVASCULAR ENDOTHELIAL FUNCTION DURING THE THIRD TRIMESTER OF HEALTHY PREGNANCY

Objective: Even in healthy individuals, high-salt (HS) loading leads to endothelial dysfunction, a hallmark of cardiometabolic diseases, independently of the changes in blood pressure (BP). Still, the effect of the amount of daily salt intake during healthy pregnancy on maternal endothelium-dependent vascular reactivity has not yet been investigated. Thus, this study aimed to assess the effect of the amount of daily salt intake during third trimester of pregnancy on maternal microvascular reactivity to stimuli in healthy pregnant women. Design and method: The present study was designed as a cross-sectional study in which all the data for each pregnant woman were obtained at one-time point between 37 and 40 weeks of gestation. Daily salt intake was estimated based on 24-h urinary sodium excretion using appropriate formula [1-g salt (NaCl) = 393.4 mg Na = 17.1 mmol Na]. Microvascular endothelium-dependent vasodilation in response to vascular occlusion (PORH), iontophoresis of acetylcholine (AChID) and local heating (HEAT), as well as endothelium-independent vasodilation in response to iontophoresis of sodium nitroprusside (SNP) was assessed by Laser Doppler flowmetry (LDF). Results: Preliminary results of the present study involve data obtained from 27 healthy pregnant women. According to the amount of daily salt intake, pregnant women were divided in low-salt (LS, < 5 g of salt/day, N = 4), normal-to-high salt (NHS, 5.0–7.5 g of salt/day, N = 12) and high-salt (HS, > 7.5 g of salt/day, N = 11) group. All pregnant women were normotensive, and BP values did not differ between groups. HEAT was significantly lower in HS compared to LS and NHS group, while PORH and AChID tended to be lower in HS compared to LS and NHS group, but without statistical significance. SNPID did not significantly differ between the groups. There was weak to moderate negative correlation between daily salt intake and functional markers of microvascular endothelium-dependent vasodilation (PORH, AChID and HEAT). Conclusions: The amount of daily salt intake significantly affects maternal endothelium-dependent microvascular reactivity during third trimester of healthy pregnancy, i.e. the increased salt intake significantly reduces the endothelium-dependent dilatation of the skin microcirculation, especially responses mediated by nitric-oxide (NO).

FUNCTIONAL FOOD ENRICHED IN N-3 POLYUNSATURATED FATTY ACIDS, SELENIUM, LUTEIN AND VITAMIN E ENHANCES MICROVASCULAR ENDOTHELIAL FUNCTION OF COMPETITIVE ATHLETES

Objective: A substantial proportion of competitive athletes do not reach dietary goals for various macro- and micronutrients, important in creating a favorable oxidative environment, which is one of a principal determinant of normal endothelial function. The present study aimed to determine the effect of functional food consumption enriched in four nutrients (Nutri4) on microvascular endothelium-dependent and endothelium-independent vasodilation of skin microcirculation in competitive athletes. Design and method: Thirty-one competitive athletes were divided to Nutri4 group (N = 17) who ate three n-3 PUFAs, selenium, lutein, and vitamin E enriched hen eggs/daily for 3 weeks (mg per egg: n-3 PUFAs 342, selenium 0.0191, lutein 0.616, vitamin E 1.098), and Control group (N = 14) who ate three regular hens’ eggs/daily (mg per egg: n-3 PUFAs 146, selenium 0.0183, lutein 0.110, vitamin E 0.595). Blood pressure (BP), body mass index (BMI), body composition and serum lipid profile were measured before and after respective dietary protocol. Microvascular endothelium-dependent vasodilation in response to post-occlusive reactive hyperemia (PORH) and iontophoresis of acetylcholine (AChID), and endothelium-independent vasodilation in response to iontophoresis of sodium nitroprusside (SNP) was assessed by Laser Doppler flowmetry (LDF). Results: Mean arterial pressure (MAP) significantly decreased, and BMI, body composition and serum lipid profile remained unchanged following dietary protocol in Nutri4 group, while all variables remained unchanged in Control group. Microvascular PORH (R-O% before 162.6 ± 40.1 vs. after 185.5 ± 63.8, P = 0.050) and AChID (flow increase before 15.6 ± 4.7 vs. after 18.8 ± 5.0, P = 0.045) was significantly increased, while SNPID remained unchanged following dietary protocol in Nutri4 group (flow increase before 15.8 ± 4.9 vs. 14.0 ± 4.2, P = 0.209). In contrast, none was significantly changed in Control group. Conclusions: Consumption of n-3 PUFA, selenium, lutein, and vitamin E enriched functional foods improves peripheral endothelium-dependent microvascular reactivity in competitive athletes, independently of changes in BP and blood lipid concentrations. Importantly, consumption of large amounts of regular hen eggs (3/day for 3 weeks) did not have a detrimental effect on vascular function, BP and serum lipid profiles in athletes.

Stratification by Race Reveals Disparate Vascular Toxicity in Response to Anti‐Cancer Therapies

IntroductionCardiovascular toxicity is the primary reason for halting anti‐cancer therapy (CTx) or limiting the dose, and cardiovascular side effects of CTx represent a major clinical problem with implications for survival and quality of life. Ongoing work in our laboratory suggests that CTx induces microvascular endothelial dysfunction, which is a well‐established predictor of future CV events. CV disease in general and CTx‐induced adverse events are significantly more prevalent in Black patients (B) relative to non‐Hispanic White patients (NHW). Despite clinical evidence of these health disparities, little to no data exist to explain the underlying causes. Therefore, we tested the novel hypothesis that CTx‐induced microvascular dysfunction is augmented in B, which may explain the increase in cardiotoxic events in this population compared to NHW.MethodsFresh human tissues were obtained as de‐identified surgical discard specimens. Peripheral adipose arterioles (100–200 µm) from healthy B and NHW were incubated ex vivowith the CTx agents, doxorubicin (Dox), trastuzumab (TZM), or paclitaxel (PTX) for 16–20 h and used for isolated arteriole preparations. Endothelium‐dependent vasodilation in response to flow (Flow‐Mediated Dilation, FMD; 5–100 cmH2O pressure gradient) or acetylcholine (ACh; 10−9 to 10−5 M) and smooth muscle‐dependent dilation to papaverine (100 µM) were evaluated. Data are presented as % max. diameter ± SEM, N. Statistical significance was established with two‐way ANOVA RM.ResultsUnder control conditions, there was no difference in microvascular FMD in healthy B (ACh: 89.8±4.0%, n=5; 85.8±1.8%, n=6) vs. NHW (ACh: 86.2±3.2%, n=6; 78.7±5.9%, n=8). Ex vivo exposure to Dox impaired endothelial function in arterioles from both B and NHW (ACh: 45.0±10.5%, n=4 and 34.2±9.8%, n=7; FMD: Figure 1A). TZM impaired endothelial function to a greater extent in arterioles from B (ACh: 36.1±7.1%, n=3; FMD: Figure 1B) compared to NHW (ACh: 44.4±8.6%, n=8; FMD: Figure 1B). Interestingly, PTX did not affect endothelial function in NHW (ACh: 91.5±1.6%, n=4; FMD: Figure 1C), but impaired endothelial function in arterioles from B (ACh: 54.7±12.9%, n=3; FMD: Figure 1C). Max dilator capacity to the endothelium‐independent dilator papaverine was not changed in either group (not shown).ConclusionOur findings reveal disparate effects of CTx agents on microvascular endothelial function from B and NHW patients. This raises the possibility that microvascular dysfunction may contribute to the known disparities in adverse CV clinical outcomes of cancer patients and potentially other CV pathologies (e.g., disparities in adverse events associated with PTX‐eluting stents.) Future work should differentiate between biological/genetic vs. social/environmental contribution to observed microvascular responses to CTx.

Monocrotaline‐induced Pulmonary Hypertension Impairs Diaphragm Vasomotor Function

INTRODUCTIONPulmonary hypertension (PH) is a chronic and progressive disease characterized by high arterial blood pressure and compensatory vascular remodeling in the pulmonary circulation. Mounting evidence suggests that the effects of PH extend beyond the pulmonary circulation. In humans and animals, PH induces diaphragm contractile dysfunction and diaphragm muscle atrophy. While the mechanistic bases for the PH‐induced diaphragm dysfunction is relatively unknown, alterations in the vasomotor control of the diaphragm resistance vasculature (i.e. arterioles) may play a key role. Therefore, we tested the hypothesis that PH increases the vasoconstrictor responsiveness and impairs the vasorelaxation of diaphragm arterioles.METHODSAdult female Sprague‐Dawley rats (n=8) were divided into healthy control (HC; n=2), ovariectomized (OVX; n=3), and PH + OVX (n=3). PH + OVX animals were given a single intraperitoneal injection of monocrotaline (MCT; 50mg/kg) to induce PH. Disease progression was then monitored via echocardiography. Once animals developed PH (~3 weeks post injection), the diaphragm was removed and first‐order arterioles (1A) from the medial costal diaphragm were isolated, cannulated, and pressurized at 90 cmH2O. After a 60 min equilibration period and spontaneous tone development, vasoconstrictor responses to potassium chloride (KCl) (10‐100 mM) and endothelium‐dependent and ‐independent vasorelaxation responses to cumulative doses of acetylcholine (ACh) and sodium nitroprusside (SNP) (10‐9 to 10‐4 M) were determined.RESULTSSpontaneous tone (%) development was not different across all three groups (HC: 36 ± 9%; OVX: 33 ± 9%; PH + OVX: 39 ± 2%; P>0.05). Compared to control vessels, maximal percent vasoconstriction with KCl was decreased with OVX (67 ± 2 versus 50 ± 1%; P<0.05). Maximal vasoconstriction (%) was reduced with PH + OVX (36 ± 4%) compared to vessels from both healthy control and OVX groups (P<0.05). Ovariectomy had no effect on endothelium‐dependent vasodilation responses to ACh versus intact control animals (P>0.05). The maximal ACh‐mediated vasodilation with PH + OVX (29 ± 9%) was decreased versus both healthy (45 ± 6%; P<0.05) and OVX animals (41 ± 4%; P<0.05). Across all three groups, there were no differences in percent relaxation with SNP (P>0.05).CONCLUSIONThese findings suggest that estrogen plays a role in the maintenance of K+ channel activity and/or the contribution of K+ channels to diaphragm vasomotor tone, and that PH further disrupts K+ channel function in diaphragm arterioles. In addition, the absence of estrogen in OVX animals had no effect on endothelial function whereas PH + OVX impaired endothelium‐dependent vasorelaxation of diaphragm resistance vessels. These findings suggest that PH, at least in the rodent model utilized herein, elicits endothelial dysfunction in the diaphragm vasculature and provide a potential mechanism for the impaired diaphragm contractile dysfunction in PH patients.

Simvastatin Improves Microcirculatory Function in Nonalcoholic Fatty Liver Disease and Downregulates Oxidative and ALE-RAGE Stress.

Increased reactive oxidative stress, lipid peroxidation, inflammation, and fibrosis, which contribute to tissue damage and development and progression of nonalcoholic liver disease (NAFLD), play important roles in microcirculatory disorders. We investigated the effect of the modulatory properties of simvastatin (SV) on the liver and adipose tissue microcirculation as well as metabolic and oxidative stress parameters, including the advanced lipoxidation end product–receptors of advanced glycation end products (ALE-RAGE) pathway. SV was administered to an NAFLD model constructed using a high-fat–high-carbohydrate diet (HFHC). HFHC caused metabolic changes indicative of nonalcoholic steatohepatitis; treatment with SV protected the mice from developing NAFLD. SV prevented microcirculatory dysfunction in HFHC-fed mice, as evidenced by decreased leukocyte recruitment to hepatic and fat microcirculation, decreased hepatic stellate cell activation, and improved hepatic capillary network architecture and density. SV restored basal microvascular blood flow in the liver and adipose tissue and restored the endothelium-dependent vasodilatory response of adipose tissue to acetylcholine. SV treatment restored antioxidant enzyme activity and decreased lipid peroxidation, ALE-RAGE pathway activation, steatosis, fibrosis, and inflammatory parameters. Thus, SV may improve microcirculatory function in NAFLD by downregulating oxidative and ALE-RAGE stress and improving steatosis, fibrosis, and inflammatory parameters.

Vasoprotective Endothelial Effects of Chronic Cannabidiol Treatment and Its Influence on the Endocannabinoid System in Rats with Primary and Secondary Hypertension.

Our study aimed to examine the endothelium (vascular)-protecting effects of chronic cannabidiol (CBD) administration (10 mg/kg once daily for 2 weeks) in aortas and small mesenteric (G3) arteries isolated from deoxycorticosterone-induced hypertensive (DOCA-salt) rats and spontaneously hypertensive rats (SHR). CBD reduced hypertrophy and improved the endothelium-dependent vasodilation in response to acetylcholine in the aortas and G3 of DOCA-salt rats and SHR. The enhancement of vasorelaxation was prevented by the inhibition of nitric oxide (NO) with L-NAME and/or the inhibition of cyclooxygenase (COX) with indomethacin in the aortas and G3 of DOCA-salt and SHR, respectively. The mechanism of the CBD-mediated improvement of endothelial function in hypertensive vessels depends on the vessel diameter and may be associated with its NO-, the intermediate-conductance calcium-activated potassium channel- or NO-, COX-, the intermediate and the small-conductance calcium-activated potassium channels-dependent effect in aortas and G3, respectively. CBD increased the vascular expression of the cannabinoid CB1 and CB2 receptors and aortic levels of endocannabinoids with vasorelaxant properties e.g., anandamide, 2-arachidonoylglycerol and palmitoyl ethanolamide in aortas of DOCA-salt and/or SHR. In conclusion, CBD treatment has vasoprotective effects in hypertensive rats, in a vessel-size- and hypertension-model-independent manner, at least partly via inducing local vascular changes in the endocannabinoid system.

Gut Microbiota Has a Crucial Role in the Development of Hypertension and Vascular Dysfunction in Toll-like Receptor 7-Driven Lupus Autoimmunity.

Our group has investigated the involvement of gut microbiota in hypertension in a murine model of systemic lupus erythematosus induced by Toll-like receptor (TLR)-7 activation. Female BALB/c mice were randomly assigned to four experimental groups: an untreated control (CTR), a group treated with the TLR7 agonist imiquimod (IMQ), IMQ-treated with vancomycin, and IMQ-treated with a cocktail of broad-spectrum antibiotics. We carried out faecal microbiota transplant (FMT) from donor CTR or IMQ mice to recipient IMQ or CTR animals, respectively. Vancomycin inhibited the increase in blood pressure; improved kidney injury, endothelial function, and oxidative stress; and reduced T helper (Th)17 infiltration in aortas from IMQ-treated mice. The rise in blood pressure and vascular complications present in IMQ mice were also observed in the CTR mice recipients of IMQ microbiota. Reduced relative populations of Sutterella and Anaerovibrio were associated with high blood pressure in our animals, which were increased after stool transplantation of healthy microbiota to IMQ mice. The reduced endothelium-dependent vasodilator responses to acetylcholine induced by IMQ microbiota were normalized after interleukin-17 neutralization. In conclusion, gut microbiota plays a role in the TLR7-driven increase in Th17 cell, endothelial dysfunction, vascular inflammation, and hypertension. The vascular changes induced by IMQ microbiota were initiated by Th17 infiltrating the vasculature.

Probiotics Prevent Hypertension in a Murine Model of Systemic Lupus Erythematosus Induced by Toll-Like Receptor 7 Activation.

Our group tested the effects of Lactobacillus fermentum CECT5716 (LC40) and/or Bifidobacterium breve CECT7263 (BFM) in the prevention of gut dysbiosis, hypertension and endothelial dysfunction in a pharmacologically-induced model of systemic lupus erythematosus (SLE). We treated eight-week-old BALB/cByJRj mice without (Ctrl) or with the agonist of TLR-7 Imiquimod (IMQ) for 8 weeks. Concomitantly, LC40 (109 CFU/mL) and BFM (109 CFU/mL) were administered through oral gavage once a day. IMQ induced intestinal dysbiosis consisting of a decrease in the α-diversity measured with Chao-richness and numbers of species. LC40 and BFM did not restore these parameters. The three-dimensional principal component analysis of bacterial taxa in stool samples presented perfect clustering between Ctrl and IMQ groups. Clusters corresponding to LC40 and BFM were more akin to IMQ. BFM and LC40 were detected colonizing the gut microbiota of mice treated respectively. LC40 and BFM decreased plasma double-stranded DNA autoantibodies, and B cells in spleen, which were increased in the IMQ group. Also, LC40 and BFM treatments activated TLR9, reduced T cells activation, and Th17 polarization in mesenteric lymph nodes. Aortae from IMQ mice displayed a decreased endothelium-dependent vasodilator response to acetylcholine linked to pro-inflammatory and pro-oxidative status, which were normalized by both BFM and LC40. In conclusion, we demonstrate for the first time that the chronic treatment with LC40 or BFM prevented hypertension and endothelial dysfunction in a mouse lupus model induced by TLR-7 activation.

Regular transient limb ischemia protects endothelial function against hypercholesterolemic damage in rabbits.

Regular transient limb ischemia (RTLI) can prevent atherosclerosis in hypercholesterolemic rabbits. As endothelial dysfunction is the initial factor leading to atherosclerosis, we investigated the effect of RTLI on endothelial function in hypercholesterolemic rabbits. We randomly allocated 15 New Zealand white rabbits to three groups, five animals per group: the hypercholesterolemic group (Group H), the sham RTLI group (Group S), and the RTLI group (Group L). All rabbits received hypercholesterolemic fodder daily. No intervention was performed on the rabbits in Group H. Rabbits in Group S were kept in hutches, with a deflated cuff applied to their left hind limb for 60 min every day. For rabbits in Group L, RTLI (six cycles of 5-min ischemia and 5-min reperfusion of the left hind limb) was applied once daily for 12 weeks. At the end of week 12, a segment of the abdominal aorta was isolated from each rabbit for in vitro measurement of the endothelium-dependent vasodilation (EDV) response to different concentrations of acetylcholine and the endothelium-independent vasodilation (EIV) response to sodium nitroprusside. The EDV response was significantly higher in Group L than in Groups S and H (p < 0.05), with no significant difference between Groups S and H (p > 0.05). There was no difference in the EIV response among the three groups. RTLI could improve the EDV response, protecting endothelial function against hypercholesterolemic damage.

Gut microbiota contributes to the development of hypertension in a genetic mouse model of systemic lupus erythematosus.

Hypertension is an important cardiovascular risk factor that is prevalent in the systemic lupus erythematosus patient population. Here, we have investigated whether intestinal microbiota is involved in hypertension in a genetic mouse model of systemic lupus erythematosus. Twenty-six-week-old female NZW/LacJ (control) and NZBWF1 (F1 hybrid of New Zealand Black and New Zealand White strains; systemic lupus erythematosus) mice were treated for 6 weeks with a broad-spectrum antibiotic mixture or with vancomycin. Faecal microbiota transplantation was performed from donor systemic lupus erythematosus group to recipient to germ-depleted or germ-free mice. Antibiotic treatment inhibited the development of hypertension and renal injury, improved endothelial dysfunction and vascular oxidative stress, and decreased aortic Th17 infiltration in NZBWF1 mice. High BP and vascular complications found in systemic lupus erythematosus mice, but not autoimmunity, kidney inflammation and endotoxemia, were reproduced by the transfer of gut microbiota from systemic lupus erythematosus donors to germ-free or germ-depleted mice. Increased proportions of Bacteroides were linked with high BP in these mice. The reduced endothelium-dependent vasodilator responses to acetylcholine and the high BP induced by microbiota from hypertensive systemic lupus erythematosus mice were inhibited after IL-17 neutralization. Changes in T-cell populations, endothelial function, vascular inflammation and hypertension driven by a genetic systemic lupus erythematosus background can be modified by antibiotic-induced changes in gut microbiota. The vascular changes induced by hypertensive systemic lupus erythematosus microbiota were mediated by Th17 infiltration in the vasculature.

Restoration of Vitamin D Levels Improves Endothelial Function and Increases TASK-Like K+ Currents in Pulmonary Arterial Hypertension Associated with Vitamin D Deficiency.

Background: Vitamin D (vitD) deficiency is highly prevalent in patients with pulmonary arterial hypertension (PAH). Moreover, PAH-patients with lower levels of vitD have worse prognosis. We hypothesize that recovering optimal levels of vitD in an animal model of PAH previously depleted of vitD improves the hemodynamics, the endothelial dysfunction and the ionic remodeling. Methods: Male Wistar rats were fed a vitD-free diet for five weeks and then received a single dose of Su5416 (20 mg/Kg) and were exposed to vitD-free diet and chronic hypoxia (10% O2) for three weeks to induce PAH. Following this, vitD deficient rats with PAH were housed in room air and randomly divided into two groups: (a) continued on vitD-free diet or (b) received an oral dose of 100,000 IU/Kg of vitD plus standard diet for three weeks. Hemodynamics, pulmonary vascular remodeling, pulmonary arterial contractility, and K+ currents were analyzed. Results: Recovering optimal levels of vitD improved endothelial function, measured by an increase in the endothelium-dependent vasodilator response to acetylcholine. It also increased the activity of TASK-1 potassium channels. However, vitD supplementation did not reduce pulmonary pressure and did not ameliorate pulmonary vascular remodeling and right ventricle hypertrophy. Conclusions: Altogether, these data suggest that in animals with PAH and severe deficit of vitD, restoring vitD levels to an optimal range partially improves some pathophysiological features of PAH.

Cholesterol Disrupts H 2 S‐Mediated Vasodilation in Large Arteries

Small resistance size arteries are key players in vascular resistance and overall blood pressure control. We previously demonstrated that within endothelial cells (EC), H2S-mediated vasodilation involves activation of transient receptor potential cation channel subfamily V member 4 (TRPV4)-dependent Ca2+ influx to activate nearby large-conductance Ca2+-activated potassium (BK) channel in resistance-size arteries. Comparison of vasodilatory responses in large and small (resistance-size) arteries show that small arteries dilate at lower concentrations of H2S donors vs large arteries. The importance of endogenous H2S in endothelium-dependent vasodilatory responses in small resistance size arteries highlights a role for H2S in regulating blood pressure and flow. However, little is known of the differences in H2S signaling in large and small arteries and how these differences modulate vasodilation. Based on previous reports that cell membrane cholesterol negatively regulates TRPV4 mobility and BK activity, we hypothesized that H2S-mediated vasodilation is disrupted by elevated EC membrane cholesterol in large arteries. We first verified EC plasma membrane cholesterol content was higher in large vs small arteries by immunofluorescence using the cholesterol stain, filipin III (20 µg/ml) in EC labeled with the glycocalyx marker tomato lectin (20 µg/ml) and the nuclear stain SYTOX green (large 145.20 ± 13.67, small: 59.85 ± 5.50, p = 0.0004, n = 5 animals/group). Next, we examined the effect of cholesterol depletion using methyl β cyclodextrin (MBCD, 100 μM), on dilatory responses to H2S in large (300-380 μm) and small (60-130 μm) mesenteric arteries using the H2S donor, NaHS (1, 10 or 100 μM). In large arteries, MBCD pretreatment significantly enhanced H2S-mediated vasodilation (10 μM +vehicle: 0.72% ± 2.17, +MBCD: 9.71% ± 2.710, p = 0.026), (100 μM +vehicle: 2.25% ± 5.74, +MBCD: 20.55% ± 7.687, p < 0.0001) n = 5 animals/group. Although NaHS dilated small arteries more than large arteries, there was no effect of cholesterol depletion in small arteries (1 μM +vehicle: 16.25% ± 2.36, +MBCD: 16.29% ± 4.44, NS), (10 μM +vehicle: 33.22% ± 7.10, +MBCD: 39.00% ± 5.85, NS), (100 μM +vehicle: 59.84% ±5.98, +MBCD: 63.18% ± 4.639, NS) n = 3 animals/group. Additionally, immunofluorescence studies of EC TRPV4 expression showed no difference between large and small arteries (large 39.6 ± 12.9, small 41.8 ± 14.1, NS) n =3 animals/group. These studies suggest that membrane cholesterol disrupts H2S-mediated vasodilation and contributes to the relative differences in sensitivity to H2S-mediated vasodilation in large and small arteries.

Endothelium‐Independent, but Not Endothelium‐Dependent, Human Microvascular Vasodilation Differs Between Young, Healthy Females and Males

Estrogen increases nitric oxide production, but a direct comparison of endothelium-dependent and endothelium-independent mechanisms of human microvascular vasodilation between females and males has not been completed. The purpose of this study was to examine endothelium-dependent and endothelium-independent mechanisms of human cutaneous microvascular vasodilation between young, healthy females and males. Females were grouped by menstrual cycle phase to assess the influence of female sex hormones. We hypothesized that females in menstrual cycle phases of elevated estrogen would have an increased endothelium-dependent vasodilation response compared to females in phases of decreased estrogen or males. Eighteen participants (7 males; 6 females in early follicular phase, F-EF; 5 females in luteal phase, F-L) were instrumented with two microdialysis fibers in the skin of the forearm. A dose-response was completed at each site: 1) acetylcholine (ACh; 10-10 – 10-1 M) to assess endothelium-dependent vasodilation, and 2) sodium nitroprusside (SNP; 10-10 – 10-1 M) to assess endothelium-independent vasodilation. Doses were perfused for 5 minutes. Maximal vasodilation was elicited by heating the skin to 43°C and infusing 54 mM SNP. Laser-Doppler flowmetry (LDF) provided an index of skin blood flow. Cutaneous vascular conductance (CVC) was calculated (LDF/mean arterial pressure) and normalized to maximal blood flow (%CVCmax). Data are presented as mean ± SD. There was no interaction between main effects of group and dose in %CVCmax response to ACh, but there was a main effect of dose (p < 0.0001). The ACh logEC50 was similar between all groups (males: -3.8 ± 0.76M; F-EF: -2.7 ± 1.00M; F-L: -4.0 ± 0.61M). There was a significant interaction between group and dose in %CVCmax response to SNP (p = 0.04). At 10-4 M SNP, F-L (20 ± 9%CVCmax) displayed a lower response compared to males (46 ± 19%CVCmax, p = 0.004). At 10-3 M SNP, F-EF (46 ± 25%CVCmax, p = 0.038) and F-L (36 ± 15%CVCmax, p = 0.002) displayed lower responses compared to males (64 ± 16%CVCmax). The response to 10-2 M and 10-1 M SNP were not statistically different between groups. The SNP logEC50 was not different between F-EF and F-L; however, the SNP logEC50 was lower in males (-4.6 ± 0.47M) compared to F-EF (-3.3 ± 0.64M, p = 0.02) and F-L (-2.8 ± 0.36M, p < 0.0001). There was no difference between groups or phase for maximal CVC responses (p > 0.05). Endothelium-dependent mechanisms of microvascular vasodilation were similar between males and females, regardless of menstrual cycle phase. Males displayed heightened endothelium-independent vasodilation in response to lower doses of SNP, but also appeared to exhibit tachyphylaxis at the higher concentrations, compared to females.

Lead and mercury 28 day exposure at small concentrations reduces smooth muscle relaxation by decreasing cGMP

Cardiovascular diseases are among the main causes of mortality in the world. There is evidence of cardiovascular harm after exposure to low lead or mercury concentrations, but the effects of chronic exposure to the association of low doses of these toxic metals are still unknown. This work evaluated after 4 weeks, the association effects of low concentrations of lead and mercury on blood pressure and vascular resistance reactivity. Wistar rats were exposed for 28 days to lead acetate (1st dose of 4 μg/100 g and subsequent doses of 0.05 μg /100 g/day to cover daily losses) and mercury chloride (1st dose of 2.17 μg/kg and subsequent doses of 0.03 μg/kg/ day to cover daily losses) and the control group received saline, i.m. Results showed that treatment increased blood pressure and induced left ventricular hypertrophy. The mesenteric vascular reactivity to phenylephrine and the endothelium-dependent vasodilator response assessed by acetylcholine did not change. Additionally, reduced involvement of vasoconstrictor prostanoids derived from cyclooxygenase was observed in the PbHg group. By other regulatory routes, such as potassium channels, the vessel showed a greater participation of BKCa channels, and a reduction in the participation of Kv channels and SKCa channels. The endothelium-independent smooth muscle relaxation was significantly impaired by reducing cGMP, possibly through the hyperstimulation of Phosphodiesterase-5 (PDE5). Our results suggested that exposure to low doses of lead and mercury triggers this compensatory mechanism, in response to the augment of arterial pressure.