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Tiago Tomazini Gonçalves,1 Carolina M Lazaro,1 Fernanda G De Mateo,1 Marcela CB Campos,1 Jackeline GB Mezencio,1 Mario A Claudino,1 Patrícia de O Carvalho,1 R Clinton Webb,2 Fernanda BM Priviero1,2 1Laboratory of Multidisciplinary Research, Universidade São Francisco, Bragança Paulista, São Paulo, Brazil; 2Department of Physiology, Medical College of Georgia, Augusta University, Augusta, GA, USA Objective: The aim of the study was to evaluate the effects of supplementation with glucosyl hesperidin (GH), with or without physical training, on body weight, fat depot, glucose and plasma lipids, oxidative status and vascular function of rats fed with high-fat diet (HFD). Methods: After weaning, male Wistar rats were fed with an HFD plus fructose for 12 weeks and started receiving oral antioxidant supplementation and/or physical training after the fourth week of diet for eight further weeks. Body weight, epididymal and retroperitoneal fat, plasma glucose and lipids, oxidative status and mesenteric artery reactivity were evaluated. Results: Rats fed with HFD presented higher body weight gain and fat accumulation compared to control rats, while GH supplementation did not influence these parameters. Physical training reduced the body weight gain and fat accumulation and modulated the oxidative status by increasing superoxide dismutase activity and total antioxidant capacity and reducing lipid peroxidation. GH alone decreased lipid peroxidation. However, when given to exercised rats, it impaired the response elicited by physical training. HFD caused endothelial dysfunction, and neither GH nor physical exercise prevented it. Potency of sodium nitroprusside was increased in exercised animals but not in GH-supplemented rats. Conclusion: Physical exercise partially decreased the body fat accumulation, decreased plasma levels of glucose and lipids and improved general oxidative status and endothelium-independent relaxation in mesenteric arteries of rats fed with HFD. GH exhibited benefits only in the oxidative status. However, GH given in association with physical exercise did not cause further changes in addition to those promoted by physical exercise. On the contrary, in exercised animals, GH prevented those changes elicited by physical training in plasma glucose and lipids, oxidative status and endothelium-independent relaxation. Keywords: high-fat diet, physical activity, oxidative stress, antioxidants, obesity, physical exercise, supplementation, flavonoids, reactive oxygen species

Background & Aims: The treatment of cardiovascular diseases (CVD) could greatly benefit from using nitric oxide (NO) donors. This study aimed to investigate the mechanisms of action of NONO2P that contribute to the observed responses in the mesenteric artery. The hypothesis was that NONO2P would have similar pharmacological actions to sodium nitroprusside (SNP) and NO., Methods: Male Wistar rats were euthanized to isolate the superior mesenteric artery for isometric tension recordings. NO levels were measured using the DAF-FM/DA dye, and cyclic guanosine monophosphate (cGMP) levels were determined using a cGMP-ELISA Kit., Results: NONO2P presented a similar maximum efficacy to SNP. The free radical of NO (NO • ) scavengers (PTIO; 100 μM and hydroxocobalamin; 30 μM) and nitroxyl anion (NO - ) scavenger (L-cysteine; 3 mM) decreased relaxations promoted by NONO2P. The presence of the specific soluble guanylyl cyclase (sGC) inhibitor (ODQ; 10 μM) nearly abolished the vasorelaxation. The cGMP-dependent protein kinase (PKG) inhibition (KT5823; 1 μM) attenuated the NONO2P relaxant effect. The vasorelaxant response was significantly attenuated by blocking inward rectifying K + channels (K ir ), voltage-operated K + channels (K V ), and large conductance Ca 2+ -activated K + channels (BK Ca ). NONO2P-induced relaxation was attenuated by cyclopiazonic acid (10 μM), indicating that sarcoplasmic reticulum Ca2+-ATPase (SERCA) activation is involved in this relaxation. Moreover, NONO2P increased NO levels in endothelial cells and cGMP production., Conclusions: NONO2P induces vasorelaxation with the same magnitude as SNP, releasing NO • and NO - . Its vasorelaxant effect involves sGC, PKG, K + channels opening, and SERCA activation, suggesting its potential as a therapeutic option for CVD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cytokines play a crucial role in the structure and function of blood vessels in hypertension. Hypertension damages blood vessels by mechanisms linked to shear forces, activation of the renin-angiotensin-aldosterone and sympathetic nervous systems, oxidative stress, and a proinflammatory milieu that lead to the generation of neoantigens and damage-associated molecular patterns, ultimately triggering the release of numerous cytokines. Damage-associated molecular patterns are recognized by PRRs (pattern recognition receptors) and activate inflammatory mechanisms in endothelial cells, smooth muscle cells, perivascular nerves, and perivascular adipose tissue. Activated vascular cells also release cytokines and express factors that attract macrophages, dendritic cells, and lymphocytes to the blood vessels. Activated and differentiated T cells into Th1, Th17, and Th22 in secondary lymphoid organs migrate to the vessels, releasing specific cytokines that further contribute to vascular dysfunction and remodeling. This chronic inflammation alters the profile of endothelial and smooth muscle cells, making them dysfunctional. Here, we provide an overview of how cytokines contribute to hypertension by impacting the vasculature. Furthermore, we explore clinical perspectives about the modulation of cytokines as a potential therapeutic intervention to specifically target hypertension-linked vascular dysfunction., Competing Interests: Disclosures None.

Cytomegalovirus (CMV) is a member of the β-herpesviruses and is ubiquitous, infecting 50%-99% of the human population depending on ethnic and socioeconomic conditions. CMV establishes lifelong, latent infections in their host. Spontaneous reactivation of CMV is usually asymptomatic, but reactivation events in immunocompromised or immunosuppressed individuals can lead to severe morbidity and mortality. Moreover, herpesvirus infections have been associated with several cardiovascular and post-transplant diseases (stroke, atherosclerosis, post-transplant vasculopathy, and hypertension). Herpesviruses, including CMV, encode viral G-protein-coupled receptors (vGPCRs) that alter the host cell by hijacking signaling pathways that play important roles in the viral life cycle and these cardiovascular diseases. In this brief review, we discuss the pharmacology and signaling properties of these vGPCRs, and their contribution to hypertension. Overall, these vGPCRs can be considered attractive targets moving forward in the development of novel hypertensive therapies., (© The Author(s) 2023. Published by Oxford University Press on behalf of American Journal of Hypertension, Ltd. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: There is a great increase in uterine arterial blood flow during normal pregnancy, which is a result of the cardiovascular changes that occur in pregnancy to adapt the maternal vascular system to meet the increased metabolic needs of both the mother and the fetus. The cardiovascular changes include an increase in cardiac output and more importantly, dilation of the maternal uterine arteries. However, the exact mechanism for the vasodilation is not fully known. Piezo1 mechanosensitive channels are highly expressed in endothelial and vascular smooth muscle cells of small-diameter arteries and play a role in structural remodeling. In this study, we hypothesize that the mechanosensitive Piezo1 channel plays a role in the dilation of the uterine artery (UA) during pregnancy. Methods: For this, 14-week-old pseudopregnant and virgin Sprague Dawley rats were used. In isolated segments of UA and mesenteric resistance arteries (MRA) mounted in a wire myograph, we investigated the effects of chemical activation of Piezo1, using Yoda 1. The mechanism of Yoda 1 induced relaxation was assessed by incubating the vessels with either vehicle or some inhibitors or in the presence of a potassium-free physiological salt solution (K + -free PSS). Results: Our results show that concentration-dependent relaxation responses to Yoda 1 are greater in the UA of the pseudo-pregnant rats than in those from the virgin rats while no differences between groups were observed in the MRAs. In both vascular beds, either in virgin or in pseudopregnant, relaxation to Yoda 1 was at least in part nitric oxide dependent. Discussion: Piezo1 channel mediates nitric oxide dependent relaxation, and this channel seems to contribute to the greater dilation that occurs in the uterine arteries of pseudo-pregnant rats., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Arishe, McKenzie, Dela Justina, Dos Anjos Moraes, Webb and Priviero.)

Piezo1 channel is a sensor for shear-stress in the vasculature. Piezo1 activation induces vasodilation, and its deficiency contributes to vascular disorders, such as hypertension. In this study, we aimed t o determine whether Piezo1 channel has a functional role in the dilation of pudendal arteries and corpus cavernosum (CC). For this, male Wistar rats were used, and the relaxation of the pudendal artery and CC was obtained using the Piezo1 activator, Yoda1, in the presence and absence of Dooku (Yoda1 antagonist), GsMTx4 (non-selective mechanosensory channel inhibitor) and L-NAME (nitric oxide synthase inhibitor). In the CC, Yoda1 was also tested in the presence of indomethacin (non-selective COX inhibitor) and tetraethylammonium (TEA, non-selective potassium channel inhibitor). The expression of Piezo1 was confirmed by Western blotting. Our data show that Piezo1 activation leads to the relaxation of the pudendal artery and CC as the chemical activator of Piezo1, Yoda1, relaxed the pudendal artery (47%) and CC (41%). This response was impaired by L-NAME and abolished by Dooku and GsMTx4 in the pudendal artery only. Indomethacin and TEA did not affect the relaxation induced by Yoda1 in the CC. Limited tools to explore this channel prevent further investigation of its underlying mechanisms of action. In conclusion, our data demonstrate that Piezo1 is expressed and induced the relaxation of the pudendal artery and CC. Further studies are necessary to determine its role in penile erection and if erectile dysfunction is associated with Piezo1 deficiency., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Dela Justina, de Freitas, Arishe, Giachini, Webb and Priviero.)

Cardiovascular diseases (CVD) are the number 1 cause of death in the United States and hypertension is a highly prevalent risk factor for CVD. It is estimated that up to 50 % of the hypertensive trait is genetically inherited while the other 50 % is determined by modifiable factors involving lifestyle, behaviors, and the environment. Interestingly, the hypertensive trait is induced or inhibited by epigenetic modifications modulated by modifiable factors. This review focused on the underlying mechanisms of stress, sleep deprivation, obesity and sedentarism as key players for epigenetic modifications contributing to the development of the hypertensive trait and, on the other hand, how epigenetic modifications induced by physical exercise and healthier habits may contribute to overturn and prevent the inheritance of hypertension trait. Furthermore, adversities during gestation and perinatal life also increase the risk for hypertension and CVD later in life, which can perpetuate the inheritance of the hypertensive trait whereas healthier habits during gestation and lactation may counteract fetal programming to improve the cardiovascular health of the progeny. Therefore, it is promising that a healthier lifestyle causes long-lasting epigenetic modifications and is transmitted to the next generation, strengthening the fight against the inheritance of hypertension., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Aims: Vascular dysfunction is a clinical hallmark of diabetes. While various pathways drive vascular alterations in diabetes, many gaps persist in understanding this process. Heat-shock protein 70 (HSP70) has a long-recognized role in diabetes, but the contributions of HSP70 to the diabetic vasculature remain largely unknown., Main Methods: We determined the systemic and local (aorta) levels of HSP70 in control (CTL) and streptozotocin (STZ)-induced diabetic rats. Functional studies were conducted in a wire myograph in the presence or absence of a pharmacological inhibitor for HSP70 (VER155008). Calcium (Ca 2+ ) dynamics was indirectly evaluated as a function of change in force development in vehicle and VER-treated vessels, as well as in the presence of inhibitors for voltage-dependent and -independent plasmalemmal Ca 2+ channels. Furthermore, mimicking the extracellular diabetic environment, we exposed aortic rings to serum from CTL and STZ-induced animals, which contains higher levels of HSP70, as well as to purified recombinant HSP70. Then, we performed functional studies following the modulation of Toll-like receptor 4 (TLR4) and its co-adaptor MD2, which interact with HSP70., Key Findings: HSP70 plays a dual role in diabetes-induced vascular dysfunction: intracellular (i)HSP70 affects Ca 2+ handling mechanisms, and extracellular (e)HSP70 modulates the TLR4-MD2 complex., Significance: These newly discovered roles of HSP70 push forward the field of vascular biology and open research avenues for other diseased states associated with altered vascular responses., Competing Interests: Declaration of competing interest Authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022. Published by Elsevier Inc.)

This study was designed to connect aortic stiffness to vascular contraction in young male and female Wistar rats. We hypothesized that female animals display reduced intrinsic media-layer stiffness, which associates with improved vascular function. Atomic force microscopy (AFM)-based nanoindentation analysis was used to derive stiffness (Young's modulus) in biaxially (i.e., longitudinal and circumferential) unloaded aortic rings. Reactivity studies compatible with uniaxial loading (i.e., circumferential) were used to assess vascular responses to a selective α1 adrenergic receptor agonist in the presence or absence of extracellular calcium. Elastin and collagen levels were indirectly evaluated with fluorescence microscopy and a picrosirius red staining kit, respectively. We report that male and female Wistar rats display similar AFM-derived aortic media-layer stiffness, even though female animals withstand higher aortic intima-media thickness-to-diameter ratio than males. Female animals also present reduced phenylephrine-induced aortic force development in concentration-response and time-force curves. Specifically, we observed impaired force displacement in both parts of the contraction curve (Aphasic and Atonic) in experiments conducted with and without extracellular calcium. Additionally, collagen levels were lower in female animals without significant elastin content and fragmentation changes. In summary, sex-related functional differences in isolated aortas appear to be related to dissimilarities in the dynamics of vascular reactivity and extracellular matrix composition rather than a direct response to a shift in intrinsic media-layer stiffness.

Aging impairs the expression of HSP70, an emergent player in vascular biology. However, it is unknown if age-related alterations in HSP70 are linked to a decline in arterial function. In this study, we test the hypothesis that the contributions of HSP70 to vascular contraction are diminished in middle-aged animals. We determined the basal levels of HSP70 in the aorta of young and middle-aged Sprague Dawley male rats using Western blotting. Functional studies were performed in a wire myograph system. Force development in response to phenylephrine was assessed in the presence or absence of extracellular calcium (Ca 2+ ), and in aortic rings treated or non-treated with an HSP70 inhibitor. Fluorescent probes were used to evaluate vascular oxidative stress and nitric oxide levels. We report that middle-aged rats have significantly lower levels of HSP70. Blockade of HSP70 attenuated vascular phasic and tonic contraction in isolated aortas. It appears that a functional HSP70 is required for proper Ca 2+ handling as inhibition of this protein led to reduced force-displacement in response to Ca 2+ dynamics. Furthermore, middle-aged aortic rings exposed to the HSP70 inhibitor display higher reactive oxygen species levels without changes in nitric oxide. In summary, we show that middle-aged animals have lower levels of HSP70 in aortas, which associates with an age-related decline in vascular responses to α-1 adrenergic stimulation.

Sexual dysfunction (SD) is widely reported by cancer survivors. However, this is an issue underestimated by doctors and the contribution of anticancer therapies for the development of SD in cancer survivors is understudied and poorly understood. Sexual function involves the activation of a neurovascular system that leads to penile erection in males and clitoral engorgement in females. Anticancer therapies can cause damage to the neurovascular circuit responsible for normal sexual function and thus, individual or combined therapies could play a role in the development of SD in all types of cancer survivors and not only those affected by genital cancers. In this review, the pathophysiology of SD and possible mechanisms underlying SD induced by anticancer therapies will be discussed. The effects of chemotherapy, radiotherapy and surgical interventions on the vasculature and nerves as well as their effects on sex hormones and inflammatory processes could link the biological effects of these interventions with SD. In conclusion, this review reports evidence that, despite psychological aspects and the disease itself, anticancer therapies are able to induce direct and indirect effects in males and females that could lead to SD in cancer survivors even after the end of the treatment., Competing Interests: Conflict of interest Authors declare no conflict of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

People living with hypertension have a higher risk of developing heart diseases, and hypertension remains a top cause of mortality. In hypertension, some detrimental changes occur in the arterial wall, which include physiological and biochemical changes. Furthermore, this disease is characterized by turbulent blood flow, increased fluid shear stress, remodeling of the blood vessels, and endothelial dysfunction. As a complex disease, hypertension is thought to be caused by an array of factors, its etiology consisting of both environmental and genetic factors. The Mosaic Theory of hypertension states that many factors, including genetics, environment, adaptive, neural, mechanical, and hormonal perturbations are intertwined, leading to increases in blood pressure. Long-term efforts by several investigators have provided invaluable insight into the physiological mechanisms responsible for the pathogenesis of hypertension, and these include increased activity of the sympathetic nervous system, overactivation of the renin-angiotensin-aldosterone system (RAAS), dysfunction of the vascular endothelium, impaired platelet function, thrombogenesis, vascular smooth muscle and cardiac hypertrophy, and altered angiogenesis. Exosomes are extracellular vesicles released by all cells and carry nucleic acids, proteins, lipids, and metabolites into the extracellular environment. They play a role in intercellular communication and are involved in the pathophysiology of diseases. Since the discovery of exosomes in the 1980s, numerous studies have been carried out to understand the biogenesis, composition, and function of exosomes. In this review, we will discuss the role of exosomes as intercellular messengers in hypertension.

The virus responsible for the coronavirus disease of 2019 (COVID-19) is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Evidences suggest that COVID-19 could trigger cardiovascular complications in apparently healthy patients. Coronaviruses are enveloped positive-strand RNA viruses acting as a pathogen-associated molecular pattern (PAMP)/ danger-associated molecular patterns (DAMP). Interestingly, Toll-like receptor (TLR) 3 recognize both PAMPs DAMPs and is activated by viral double-stranded RNA (dsRNA) leading to activation of TIR receptor domain-containing adaptor inducing IFN-β (TRIF) dependent pathway. New evidence has shown a link between virus dsRNA and increased BP. Hence, we hypothesize that COVID-19 infection may be over activating the TLR3 through dsRNA, evoking further damage to the patients, leading to vascular inflammation and increased blood pressure, favoring the development of several cardiovascular complications, including hypertension., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Cardiovascular disease (CVD) is still the leading cause of illness and death in the Western world. Cardiovascular aging is a progressive modification occurring in cardiac and vascular morphology and physiology where increased endothelial dysfunction and arterial stiffness are observed, generally accompanied by increased systolic blood pressure and augmented pulse pressure. The effects of biological sex on cardiovascular pathophysiology have long been known. The incidence of hypertension is higher in men, and it increases in postmenopausal women. Premenopausal women are protected from CVD compared with age-matched men and this protective effect is lost with menopause, suggesting that sex-hormones influence blood pressure regulation. In parallel, the heart progressively remodels over the course of life and the pattern of cardiac remodeling also differs between the sexes. Lower autonomic tone, reduced baroreceptor response, and greater vascular function are observed in premenopausal women than men of similar age. However, postmenopausal women have stiffer arteries than their male counterparts. The biological mechanisms responsible for sex-related differences observed in cardiovascular aging are being unraveled over the last several decades. This review focuses on molecular mechanisms underlying the sex-differences of CVD in aging., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Dela Justina, Miguez, Priviero, Sullivan, Giachini and Webb.)

Heat-shock protein 70 (HSP70) contributes to cellular calcium (Ca 2+ ) handling mechanisms during receptor-mediated vascular contraction. Interestingly, previous studies have independently reported sex-related differences in HSP70 expression and Ca 2+ dynamics. Still, it is unknown if sex, as a variable, plays a role in the impact that HSP70 has upon vascular contraction. To narrow this gap, we investigated if differences exist in the expression levels of HSP70 in the aorta, and if targeting this protein contributes to sex disparity in vascular responses. We report that, compared with male animals, female rats present a reduction in the basal levels of HSP70. More compelling, we found that the blockade of HSP70 has a greater impact on phenylephrine-induced phasic and tonic vascular contraction in female animals. In fact, it seems that the inhibition of HSP70 significantly affects vascular Ca 2+ handling mechanisms in females, which could be associated with the fact that these animals have impaired HSP70 expression. Corroborating this idea, we uncovered that the higher sensitivity of female rats to HSP70 inhibition does not involve an increase in NO-dependent vasodilation nor a decrease in vascular oxidative stress. In summary, our findings reveal a novel mechanism associated with sex-specific differences in vascular responses to α-1 adrenergic stimulation, which might contribute to unraveling the network of intertwined pathways conferring female protection to (cardio)vascular diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 de Oliveira, Priviero, Webb and Nunes.)

Cells respond to stress by activating a variety of defense signaling pathways, including cell survival and cell death pathways. Although cell survival signaling helps the cell to recover from acute insults, cell death or senescence pathways induced by chronic insults can lead to unresolved pathologies. Arterial hypertension results from chronic physiological maladaptation against various stressors represented by abnormal circulating or local neurohormonal factors, mechanical stress, intracellular accumulation of toxic molecules, and dysfunctional organelles. Hypertension and aging share common mechanisms that mediate or prolong chronic cell stress, such as endoplasmic reticulum stress and accumulation of protein aggregates, oxidative stress, metabolic mitochondrial stress, DNA damage, stress-induced senescence, and proinflammatory processes. This review discusses common adaptive signaling mechanisms against these stresses including unfolded protein responses, antioxidant response element signaling, autophagy, mitophagy, and mitochondrial fission/fusion, STING (signaling effector stimulator of interferon genes)-mediated responses, and activation of pattern recognition receptors. The main molecular mechanisms by which the vasculature copes with hypertensive and aging stressors are presented and recent advancements in stress-adaptive signaling mechanisms as well as potential therapeutic targets are discussed.

Background: Erectile dysfunction (ED) has been shown to be related with inflammatory markers in humans. Chronic infusion of TNF-α caused ED in mice while TNF-α knockout mice exhibited improvement in the relaxation of the corpus cavernosum (CC)., Aim: Since obesity triggers an inflammatory process, we aimed to investigate the hypothesis that in obesity, Toll-like receptor 9 (TLR9) activation leads to increased TNF-α levels and impairment in CC reactivity., Methods: Four-week old male C57BL6 (WT) and TLR9 mutant (TLR9 MUT ) mice were fed a standard chow or high fat diet (HFD) for 12 weeks. Body weight and nonfasting blood glucose were analyzed. Contractile and relaxation responses of the CC were evaluated by electrical field stimulation and concentration response curves to phenylephrine and acetylcholine. Protein expression of nNOS, TNF-α, TNF-R1, TLR9 and MyD88 were measured by western blot. Plasma levels of TNF-α were measured by ELISA., Outcome: In obesity, impaired cavernosal relaxation is associated with the activation of the innate immune system, by increasing the production of TNF-α through the activation of TLR9 in the macrophages., Results: After 12 weeks of HFD both WT and TLR9 MUT mice had increased body weight and nonfasting blood glucose compared to standard chow. In the CC, acetylcholine-induced relaxation was not changed. A trend to increased contraction to phenylephrine and KCl was seen in WT HFD only. electrical field stimulation-induced relaxation of the CC was decreased in WT HFD as well as nNOS expression in the CC of WT HFD, but not in TLR9 MUT HFD. In the CC, protein expression of TLR9 and MyD88 was similar in all groups. While circulating levels of TNF-α presented only a trend to increase in mice fed HFD, the CC expression of TNF-α was increased only in WT HFD mice., Clinical Translation: The innate immune system can be a target for the treatment of erectile complications in obesity., Strengths and Limitations: This is the first study demonstrating that activation of TLR9 expressed in macrophages leads to impaired cavernosal relaxation. The main limitation of the study is the lack of understanding about the source/expression of the macrophages in the cavernous tissue. Further, herein, the experiments were performed only in isolated cavernous tissue (in vitro), thus the lack of knowledge on how the TLR9 modulates the in vivo response of the erectile tissue is another limitation of this study., Conclusion: Our findings indicate that CC dysfunction observed in obesity is at least in part mediated by the production of TNF-α upon activation of TLR9 expressed in the macrophages. Priviero F, Calmasini F, Dela Justina V, et al. Macrophage-Specific Toll Like Receptor 9 (TLR9) Causes Corpus Cavernosum Dysfunction in Mice Fed a High Fat Diet. J Sex Med 2021;18:723-731., (Copyright © 2021 International Society for Sexual Medicine. Published by Elsevier Inc. All rights reserved.)

Heat-shock protein 70 (HSP70) is a ubiquitously expressed molecular chaperone with various biological functions. Recently, we demonstrated that HSP70 is key for adequate vascular reactivity. However, the specific mechanisms targeted by HSP70 to assist in this process remain elusive. Since there is a wealth of evidence connecting HSP70 to calcium ([Formula: see text]), a master regulator of contraction, we designed this study to investigate whether blockade of HSP70 disrupts vascular contraction via impairment of [Formula: see text] handling mechanisms. We performed functional studies in aortas isolated from male Sprague Dawley rats in the presence or absence of exogenous [Formula: see text], and we determined the effects of VER155008, an inhibitor of HSP70, on [Formula: see text] handling as well as key mechanisms that regulate vascular contraction. Changes in the intracellular concentration of [Formula: see text] were measured with a biochemical assay kit. We report that blockade of HSP70 leads to [Formula: see text] mishandling in aorta stimulated with phenylephrine, decreasing both phasic and tonic contractions. Importantly, in [Formula: see text] free Krebs' solution, inhibition of HSP70 only reduced the [Formula: see text] of the phasic contraction if the protein was blocked before IP3r-mediated [Formula: see text] release, suggesting that HSP70 has a positive effect towards this receptor. Corroborating this statement, VER155008 did not potentiate an IP3r inhibitor's outcomes, even with partial blockade. In another set of experiments, the inhibition of HSP70 attenuated the amplitude of the tonic contraction independently of the moment VER155008 was added to the chamber (i.e., whether it was before or after IP3r-mediated phasic contraction). More compelling, following re-addition of [Formula: see text], VER155008 amplified the inhibitory effects of a voltage-dependent [Formula: see text] channel blocker, but not of a voltage-independent [Formula: see text] channel inhibitor, indicating that HSP70 has a positive impact on the latter. Lastly, the mechanism by which HSP70 modulates vascular contraction does not involve the [Formula: see text] sensitizer protein, Rho-kinase, nor the SERCA pump, as blockade of these proteins in the presence of VER155008 almost abolished contraction. In summary, our findings shed light on the processes targeted by HSP70 during vascular contraction and open research avenues for potential new mechanisms in vascular diseases.

Increased O-Linked β-N-acetylglucosamine (O-GlcNAc) is observed in several pathologies, and unbalanced O-GlcNAcylation levels favor endothelial dysfunction. Whether augmented O-GlcNAc impacts the uterine artery (UA) function and how it affects the UA during pregnancy remains to be elucidated. We hypothesized that glucosamine treatment increases O-GlcNAc, leading to uterine artery dysfunction and this effect is prevented by pregnancy. Pregnant (P) and non-pregnant (NP) Wistar rats were treated with glucosamine (300 mg/kg; i.p.) for 21 days. Concentration response-curves (CRC) to acetylcholine (in the presence or absence of L-NAME) and sodium nitroprusside were performed in UAs. In NP rats, glucosamine treatment increased O-GlcNAc expression in UAs accompanied by decreased endothelium-dependent relaxation, which was abolished by L-NAME. Endothelial nitric oxide synthase (eNOS) activity and total Akt expression were decreased by glucosamine-treatment in NP rats. Further, NP rats treated with glucosamine displayed increased glycogen synthase kinase 3 beta (GSK3β) activation and O-GlcNAc-transferase (OGT) expression in the UA. P rats treated with glucosamine displayed decreased O-GlcNAc in UAs and it was accompanied by improved relaxation to acetylcholine, whereas eNOS and GSK3β activity and total Akt and OGT expression were unchanged. Sodium nitroprusside-induced relaxation was not changed in all groups, indicating that glucosamine treatment led to endothelial dysfunction in NP rats. The underlying mechanism is, at least in part, dependent on Akt/GSK3β/OGT modulation. We speculate that during pregnancy, hormonal alterations play a protective role in preventing O-GlcNAcylation-induced endothelial dysfunction in the UAs., Competing Interests: Declaration of competing interest The author(s) declare no potential conflicts of interest, financial or otherwise on this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Aims: Hyperglycemia in combination with oxidative stress plays a significant pathophysiological role in diabetic testicular dysfunction, often leading to infertility. Activation of Toll-like receptor 4 (TLR4) has been reported to mediate oxidative stress during diabetes. However, engagement of the TLR4 signaling pathway in diabetic testicular dysfunction has not been previously explored. Herein, we investigated the role of TLR4 in reactive oxygen species (ROS) production and in the phosphorylation status of ERK1/2 in primary Leydig cells exposed to high glucose and in testis isolated from diabetic rats., Main Methods: Testicular levels of TLR4 and phospho-ERK1/2 were determined by Western blotting. ROS production was detected with a fluorescent probe. Additionally, primary Leydig cells were exposed to normal (5.5 mmol/l) or elevated (33 mmol/l) glucose concentrations and treated with or without a TLR4 inhibitor, CLI095 (10 - 5  mol/l) for 24 h, followed by evaluation of TLR4 and phospho-ERK1/2 expression levels by Western blotting and immunofluorescence staining, respectively., Key Findings: We show that high glucose induces the expression of TLR4 in Leydig cells. Additionally, we demonstrate that blockade of this receptor in this cell population reduces oxidative stress and restores the levels of phospho-ERK1/2., Significance: Our findings provide new insight into TLR4 interaction with ROS and MEK/ERK pathway in Leydig cells exposed to high glucose and present a rationale for the development of new therapeutics for diabetic testicular dysfunction., Competing Interests: Declaration of competing interest The authors declare no competing interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Toll-like receptors (TLRs), such as TLR4 and 9, recognize pathogen-associated molecular pattern (PAMPs) and danger-associated molecular patterns (DAMPs) and are associated with increased blood pressure (BP). TLR3, residing in the endosomal compartment, is activated by viral double-stranded RNA (dsRNA) leading to activation of TIR receptor domain-containing adaptor inducing IFN-β (TRIF) dependent pathway. Besides foreign pathogens, the immune system responds to endogenous markers of cellular damage such as mitochondrial dsRNA (mtdsRNA). New evidence has shown a link between dsRNA and increased BP. Moreover, TLR3 activation during pregnancy was demonstrated to develop preeclampsia-like symptoms in both rats and mice. Hence, we hypothesize that the dsRNA derived from viral nucleic acids or cellular damage (mtdsRNA) will increase the inflammatory state through activation of TLR3, contributing to vascular dysfunction and increased BP. Therefore, inhibition of TLR3 could be a therapeutic target for the treatment of hypertension with potential improvement in vascular reactivity and consequently, a decrease in BP., (© 2020 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Background: Aging is a major non-modifiable risk factor for hypertension. Changes in aging are similar to those seen in hypertension in the vasculature. Also, aging increases the vascular dysfunction that occurs in hypertension. L-arginase action reduces substrate (L-arginine) availability for the formation of nitric oxide (NO). This reduces the level of NO and leads to reduced vasodilation and ultimately, vascular dysfunction. This study examines the hypothesis that age-dependent vascular dysfunction in SHRs is mediated by arginase., Methods: Young (12-14 weeks) and old (11-12 months) male Wistar and spontaneously hypertensive rats (SHR) were used. Mean arterial pressure (MAP) was measured in the rats. They were then euthanized and mesenteric resistance arteries (MRAs) and thoracic aortae were excised and placed in ice-cold physiological salt solution (PSS). Arterial segments were either snap-frozen in liquid nitrogen and stored for immunoblotting studies or cut into 2mm rings for reactivity studies. Cumulative concentration-response curves to acetylcholine (Ach: 10 -9 - 3x10 -5 M) and sodium nitroprusside (SNP: 10 -12 - 3x10 -5 M) were performed in the absence or presence (30-minute exposure) of L-arginase, 0.05U/ML (MRA) or 0.5U/ML (aorta). Vessels were pre-contracted with phenylephrine (PE; 3x10 -6 M)., Results: MAP increased during aging in the SHRs p<0.05 but not in the Wistar rats. Arginase impaired the endothelium-dependent relaxation responses of thoracic aortic and MRA arterial rings to Ach in the old Wistars and SHRs (Emax aorta: 29.42±2.19% vs 7.94±1.86%). Arginase also impaired endothelium-independent relaxation response to SNP in the old SHRs only (Emax aorta: 88.62±4.10% vs 31.45±10.61%). We also observed no differences in the serum arginase activity in the four groups of rats. On the contrary, arginase activity in the aortae of young Wistar rats was reduced compared to other groups., Conclusions: Arginase impairs both endothelium-dependent and -independent vasorelaxation responses, through the NO signaling pathway.

Introduction: Male and female sexual dysfunction (SD) is considered a multifactorial condition. Numerous studies have shown the involvement of inflammatory processes in this pathological condition. Sexual intercourse requires healthy and functioning vessels to supply the pelvic region in both males and females, generating penile erection and clitoral and vaginal lubrication, respectively. Cardiovascular diseases and associated risk factors may contribute negatively to pelvic blood flow, possibly through immune system activation., Aim: The study aimed to address the correlation between vascular inflammation driven by immune system activation and SD in males and females., Methods: A literature review was performed to identify articles addressing male and female SD and vascular inflammation. Key words included "male and female sexual dysfunction," "vascular inflammation," "iliac and pudendal arteries dysfunction," "genitourinary tract," and "blood flow.", Main Outcome Measures: Management of systemic and local inflammation may be a useful alternative to improve SD and reduce the risk of cardiovascular diseases in the future., Results: Increased levels of cytokines and chemokines have been detected in humans and animals with hypertension, obesity, and diabetic conditions. Chronic activation of the innate immune system, especially by pathogen- or damage-associated molecular patterns, and metabolic-related disorders may act as triggers further contributing to an increased inflammatory condition. Due to the reduced size of vessels, SD and retinal vascular impairments have been shown to be predictive factors for cardiovascular diseases. Therefore, considering that blood flow to the genitalia is essential for sexual function, endothelial dysfunction and vascular remodeling, secondary to chronic immune system activation, may be implicated in male and female vasculogenic SD., Conclusions: Several conditions appear to play a role in SD. In the present review, we have identified a role for the immune system in generating vascular and tissue impairments contributing to erectile dysfunction and female SD. Calmasini FB, Klee N, Webb RC, et al. Impact of Immune System Activation and Vascular Impairment on Male and Female Sexual Dysfunction. Sex Med Rev 2019;7:604-613., (Copyright © 2019 International Society for Sexual Medicine. Published by Elsevier Inc. All rights reserved.)

Heart failure (HF) has a strong association with the development of lower urinary tract symptoms, especially overactive bladder (OAB); although this condition remains poorly investigated. In this study, we assess the aortocaval fistula (ACF) model as a novel experimental model of micturition dysfunction, associated with HF, focused on the molecular and functional studies to evaluate the autonomic nervous system and urinary bladder remodeling. Male rats were submitted to ACF for HF induction. Echocardiography, cystometric, histomorphometry and molecular analysis, as well as concentration-response curves to carbachol and ATP and frequency-response curves to electrical field stimulation (EFS) were evaluated in Sham and HF (4- and 12-weeksendpoint) groups. Compared to SHAM, HF groups exhibited progressive increases in the left ventricle (LV) mass and fractional shortening which indicates cardiac dysfunction, although HF was characterized only after 12 weeks by the reduced ejection fraction. For micturition function, HF groups presented increased non-voiding contractions (NVC) and decreased bladder capacity; however, when comparing HF groups, these urinary parameters were significantly impaired over the weeks (12-weeks). The contractile responses induced by CCh, ATP and EFS were greater in detrusor muscle (DSM) from HF rats. mRNA expression for muscarinic receptors (M2 and M3) was higher in DSM only after 12 weeks of ACF, in addition to MMP9 and TGF-beta. Histomorphometric revealed increased urothelium thickness in both HF groups, whereas DSM thickness occurred only after 12 weeks. Thus, the ACF model induced cardiac dyfunction with progressive micturition dysfunction over the weeks, characterized by increased DSM contractile mechanisms as well as extracellular matrix remodeling in the urinary bladder, representing a useful tool to evaluate the OAB associated with HF., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Morphological and physiological changes in the vasculature have been described in the evolution and maintenance of hypertension. Hypertension-induced vascular dysfunction may present itself as a contributing, or consequential factor, to vascular remodeling caused by chronically elevated systemic arterial blood pressure. Changes in all vessel layers, from the endothelium to the perivascular adipose tissue (PVAT), have been described. This mini-review focuses on the current knowledge of the structure and function of the vessel layers, specifically muscular arteries: intima, media, adventitia, PVAT, and the cell types harbored within each vessel layer. The contributions of each cell type to vessel homeostasis and pathophysiological development of hypertension will be highlighted.

The aim of the present study was to evaluate the potency and maximal responses (E(max)) to the adenosine receptor agonists N(6)-cyclopentyladenosine (CPA), N-ethylcarboxamidoadenosine (NECA) and N(6)-(3-iodobenzyl)-5'-N-methylcarbaxamidoadenosine (IB-MECA) in right atria from trained rats. We also investigated the interaction between the training bradycardia and the sensitivity of the chronotropic response mediated by adenosine receptor stimulation. Animals were submitted to run training for 60 min, 5 days a week, over a period of 8 weeks. Mean blood pressure and heart rate were measured in conscious animals. Right atria were isolated and concentration-response curves to CPA, NECA and IB-MECA were obtained. A reduction in heart rate was found in trained rats, indicating that the training programme was successful in inducing physical conditioning. The three adenosine receptor agonists induced a concentration-dependent negative chronotropic response. The rank order of potency and E(max) for the three adenosine receptor agonists was CPA > NECA > IB-MECA. Dynamic exercise for 8 weeks did not alter the E(max) for CPA, NECA and IB-MECA. Similarly, the potencies of CPA and NECA were not affected by run training, whereas the potency of IB-MECA was reduced (6.10 +/- 0.09 vs 5.66 +/- 0.10 for sedentary and trained groups, respectively). In conclusion, run training for 8 weeks induced a desensitization of the chronotropic response to IB-MECA without changing the potency of CPA and NECA. These findings exclude the participation of adenosine receptors in the training bradycardia.