Vascular fibrosis in aging and hypertension: molecular mechanisms and clinical implications

Aging is the primary risk factor underlying hypertension and incident cardiovascular disease. With aging, the vasculature undergoes structural and functional changes characterized by endothelial dysfunction, wall thickening, reduced distensibility, and arterial stiffening. Vascular stiffness results from fibrosis and extracellular matrix (ECM) remodelling, processes that are associated with aging and are amplified by hypertension. Some recently characterized molecular mechanisms underlying these processes include increased expression and activation of matrix metalloproteinases, activation of transforming growth factor-β1/SMAD signalling, upregulation of galectin-3, and activation of proinflammatory and profibrotic signalling pathways. These events can be induced by vasoactive agents, such as angiotensin II, endothelin-1, and aldosterone, which are increased in the vasculature during aging and hypertension. Complex interplay between the “aging process” and prohypertensive factors results in accelerated vascular remodelling and fibrosis and increased arterial stiffness, which is typically observed in hypertension. Because the vascular phenotype in a young hypertensive individual resembles that of an elderly otherwise healthy individual, the notion of “early” or “premature” vascular aging is now often used to describe hypertension-associated vascular disease. We review the vascular phenotype in aging and hypertension, focusing on arterial stiffness and vascular remodelling. We also highlight the clinical implications of these processes and discuss some novel molecular mechanisms of fibrosis and ECM reorganization

Genomic and non-genomic effects of androgens in the cardiovascular system: clinical implications

The principle steroidal androgens are testosterone and its metabolite 5α-dihydrotestosterone (DHT), which is converted from testosterone by the enzyme 5α-reductase. Through the classic pathway with androgens crossing the plasma membrane and binding to the androgen receptor (AR) or via mechanisms independent of the ligand-dependent transactivation function of nuclear receptors, testosterone induces genomic and non-genomic effects respectively. AR is widely distributed in several tissues, including vascular endothelial and smooth muscle cells. Androgens are essential for many developmental and physiological processes, especially in male reproductive tissues. It is now clear that androgens have multiple actions besides sex differentiation and sexual maturation and that many physiological systems are influenced by androgens, including regulation of cardiovascular function [nitric oxide (NO) release, Ca2+ mobilization, vascular apoptosis, hypertrophy, calcification, senescence and reactive oxygen species (ROS) generation]. This review focuses on evidence indicating that interplay between genomic and non-genomic actions of testosterone may influence cardiovascular function

Vascular smooth muscle contraction in hypertension

Hypertension is a major risk factor for many common chronic diseases, such as heart failure, myocardial infarction, stroke, vascular dementia and chronic kidney disease. Pathophysiological mechanisms contributing to the development of hypertension include increased vascular resistance, determined in large part by reduced vascular diameter due to increased vascular contraction and arterial remodelling. These processes are regulated by complex interacting systems such as the renin angiotensin aldosterone system (RAAS), sympathetic nervous system, immune activation and oxidative stress, which influence vascular smooth muscle function. Vascular smooth muscle cells are highly plastic and in pathological conditions undergo phenotypic changes from a contractile to a proliferative state. Vascular smooth muscle contraction is triggered by an increase in intracellular free calcium concentration ([Ca2+]i), promoting actin-myosin cross-bridge formation. Growing evidence indicates that contraction is also regulated by calcium-independent mechanisms involving RhoA-Rho kinase (ROCK), protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) signaling, reactive oxygen species and reorganization of the actin cytoskeleton. Activation of immune/inflammatory pathways and noncoding RNAs are also emerging as important regulators of vascular function. Vascular smooth muscle cell [Ca2+]i, not only determines the contractile state but also influences activity of many calcium-dependent transcription factors and proteins thereby impacting the cellular phenotype and function. Perturbations in vascular smooth muscle cell signaling and altered function influence vascular reactivity and tone, important determinants of vascular resistance and blood pressure. Here we discuss mechanisms regulating vascular reactivity and contraction in physiological and pathophysiological conditions and highlight some new advances in the field, focusing specifically on hypertension

Vascular smooth muscle contraction in hypertension

Hypertension is a major risk factor for many common chronic diseases, such as heart failure, myocardial infarction, stroke, vascular dementia, and chronic kidney disease. Pathophysiological mechanisms contributing to the development of hypertension include increased vascular resistance, determined in large part by reduced vascular diameter due to increased vascular contraction and arterial remodelling. These processes are regulated by complex-interacting systems such as the renin-angiotensin-aldosterone system, sympathetic nervous system, immune activation, and oxidative stress, which influence vascular smooth muscle function. Vascular smooth muscle cells are highly plastic and in pathological conditions undergo phenotypic changes from a contractile to a proliferative state. Vascular smooth muscle contraction is triggered by an increase in intracellular free calcium concentration ([Ca2+]i), promoting actin–myosin cross-bridge formation. Growing evidence indicates that contraction is also regulated by calcium-independent mechanisms involving RhoA-Rho kinase, protein Kinase C and mitogen-activated protein kinase signalling, reactive oxygen species, and reorganization of the actin cytoskeleton. Activation of immune/inflammatory pathways and non-coding RNAs are also emerging as important regulators of vascular function. Vascular smooth muscle cell [Ca2+]i not only determines the contractile state but also influences activity of many calcium-dependent transcription factors and proteins thereby impacting the cellular phenotype and function. Perturbations in vascular smooth muscle cell signalling and altered function influence vascular reactivity and tone, important determinants of vascular resistance and blood pressure. Here, we discuss mechanisms regulating vascular reactivity and contraction in physiological and pathophysiological conditions and highlight some new advances in the field, focusing specifically on hypertension

Chemerin receptor blockade improves vascular function in diabetic obese mice via redox-sensitive- and Akt-dependent pathways

Chemerin and its G protein-coupled receptor [chemerin receptor 23 (ChemR23)] have been associated with endothelial dysfunction, inflammation, and insulin resistance. However, the role of chemerin on insulin signaling in the vasculature is still unknown. We aimed to determine whether chemerin reduces vascular insulin signaling and whether there is interplay between chemerin/ChemR23, insulin resistance, and vascular complications associated with type 2 diabetes (T2D). Molecular and vascular mechanisms were probed in mesenteric arteries and cultured vascular smooth muscle cells (VSMCs) from C57BL/6J, nondiabetic lean db/m, and diabetic obese db/db mice as well as in human microvascular endothelial cells (HMECs). Chemerin decreased insulin-induced vasodilatation in C57BL/6J mice, an effect prevented by CCX832 (ChemR23 antagonist) treatment. In VSMCs, chemerin, via oxidative stress- and ChemR23-dependent mechanisms, decreased insulin-induced Akt phosphorylation, glucose transporter 4 translocation to the membrane, and glucose uptake. In HMECs, chemerin decreased insulin-activated nitric oxide signaling. AMP-activated protein kinase phosphorylation was reduced by chemerin in both HMECs and VSMCs. CCX832 treatment of db/db mice decreased body weight, insulin, and glucose levels as well as vascular oxidative stress. CCX832 also partially restored vascular insulin responses in db/db and high-fat diet-fed mice. Our novel in vivo findings highlight chemerin/ChemR23 as a promising therapeutic target to limit insulin resistance and vascular complications associated with obesity-related diabetes

Vasoprotective effects of NOX4 are mediated via polymerase and transient receptor potential melastatin 2 cation channels in endothelial cells

Background: NOX4 activation has been implicated to have vasoprotective and blood pressure (BP)-lowering effects. Molecular mechanisms underlying this are unclear, but NOX4-induced regulation of the redox-sensitive Ca 2+ channel TRPM2 and effects on endothelial nitric oxide synthase (eNOS)-nitric oxide signalling may be important. Method: Wild-type and LinA3, renin-expressing hypertensive mice, were crossed with NOX4 knockout mice. Vascular function was measured by myography. Generation of superoxide (O 2- ) and hydrogen peroxide (H 2 O 2 ) were assessed by lucigenin and amplex red, respectively, and Ca 2+ influx by Cal-520 fluorescence in rat aortic endothelial cells (RAEC). Results: BP was increased in NOX4KO, LinA3 and LinA3/NOX4KO mice. This was associated with endothelial dysfunction and vascular remodelling, with exaggerated effects in NOX4KO groups. The TRPM2 activator, ADPR, improved vascular relaxation in LinA3/NOX4KO mice, an effect recapitulated by H 2 O 2 . Inhibition of PARP and TRPM2 with olaparib and 2-APB, respectively, recapitulated endothelial dysfunction in NOX4KO. In endothelial cells, Ang II increased H 2 O 2 generation and Ca 2+ influx, effects reduced by TRPM2 siRNA, TRPM2 inhibitors (8-br-cADPR, 2-APB), olaparib and GKT137831 (NOX4 inhibitor). Ang II-induced eNOS activation was blocked by NOX4 and TRPM2 siRNA, GKT137831, PEG-catalase and 8-br-cADPR. Conclusion: Our findings indicate that NOX4-induced H 2 O 2 production activates PARP/TRPM2, Ca 2+ influx, eNOS activation and nitric oxide release in endothelial cells. NOX4 deficiency impairs Ca 2+ homeostasis leading to endothelial dysfunction, an effect exacerbated in hypertension. We define a novel pathway linking endothelial NOX4/H 2 O 2 to eNOS/nitric oxide through PARP/TRPM2/Ca 2+ . This vasoprotective pathway is perturbed when NOX4 is downregulated and may have significance in conditions associated with endothelial dysfunction, including hypertension

Upregulation of Nrf2 and decreased redox signaling contribute to renoprotective effects of chemerin receptor blockade in diabetic mice

Chemerin, acting through its receptor ChemR23, is an adipokine associated with inflammatory response, glucose and lipid metabolism and vascular function. Although this adipokine has been associated with the development and progression of kidney disease, it is not clear whether the chemerin/ChemR23 system plays a role in renal function in the context of diabetes. Therefore, we sought to determine whether ChemR23 receptor blockade prevents the development and/or progression of diabetic nephropathy and questioned the role of oxidative stress and Nrf2 in this process. Renal redox state and function were assessed in non-diabetic lean db/m and diabetic obese db/db mice treated with vehicle or CCX832 (ChemR23 antagonist). Renal reactive oxygen species (ROS) production, which was increased in diabetic mice, was attenuated by CCX832. This was associated with an increase in Nox 4 expression. Augmented protein oxidation in db/db mice was not observed when mice were treated with CCX832. CCX832 also abrogated impaired Nrf2 nuclear activity and associated downregulation in antioxidants expression in kidneys from db/db mice. Our in vivo findings highlight the role of the redox signaling and Nrf2 system as renoprotective players during chemerin receptor blockade in diabetic mice. The chemerin/ChemR23 system may be an important target to limit renal dysfunction associated with obesity-related diabetes

NADPH Oxidase 5 Is a Pro‐Contractile Nox Isoform and a Point of Cross‐Talk for Calcium and Redox Signaling‐Implications in Vascular Function

Background NADPH Oxidase 5 (Nox5) is a calcium‐sensitive superoxide‐generating Nox. It is present in lower forms and higher mammals, but not in rodents. Nox5 is expressed in vascular cells, but the functional significance remains elusive. Given that contraction is controlled by calcium and reactive oxygen species, both associated with Nox5, we questioned the role of Nox5 in pro‐contractile signaling and vascular function. Methods and Results Transgenic mice expressing human Nox5 in a vascular smooth muscle cell–specific manner (Nox5 mice) and Rhodnius prolixus, an arthropod model that expresses Nox5 endogenoulsy, were studied. Reactive oxygen species generation was increased systemically and in the vasculature and heart in Nox5 mice. In Nox5‐expressing mice, agonist‐induced vasoconstriction was exaggerated and endothelium‐dependent vasorelaxation was impaired. Vascular structural and mechanical properties were not influenced by Nox5. Vascular contractile responses in Nox5 mice were normalized by N‐acetylcysteine and inhibitors of calcium channels, calmodulin, and endoplasmic reticulum ryanodine receptors, but not by GKT137831 (Nox1/4 inhibitor). At the cellular level, vascular changes in Nox5 mice were associated with increased vascular smooth muscle cell [Ca2+]i, increased reactive oxygen species and nitrotyrosine levels, and hyperphosphorylation of pro‐contractile signaling molecules MLC20 (myosin light chain 20) and MYPT1 (myosin phosphatase target subunit 1). Blood pressure was similar in wild‐type and Nox5 mice. Nox5 did not amplify angiotensin II effects. In R. prolixus, gastrointestinal smooth muscle contraction was blunted by Nox5 silencing, but not by VAS2870 (Nox1/2/4 inhibitor). Conclusions Nox5 is a pro‐contractile Nox isoform important in redox‐sensitive contraction. This involves calcium‐calmodulin and endoplasmic reticulum–regulated mechanisms. Our findings define a novel function for vascular Nox5, linking calcium and reactive oxygen species to the pro‐contractile molecular machinery in vascular smooth muscle cells

Mineralocorticoid receptor blockade prevents vascular remodelling in a rodent model of type 2 diabetes mellitus

Abstract Mineralocorticoid receptors (MRs), which are activated by mineralocorticoids and glucocorticoids, actively participate in mechanisms that affect the structure and function of blood vessels. Although experimental and clinical evidence shows that vascular damage in diabetes is associated with structural alterations in large and small arteries, the role of MR in this process needs further studies. Thus, we tested the hypothesis that MR, through redox-sensitive mechanisms, plays a role in diabetes-associated vascular remodelling. Male, 12-14-weeks-old db/db mice, a model of type 2 diabetes and their non-diabetic counterpart controls (db/+) were treated with spironolactone (MR antagonist, 50 mg/kg/day) or vehicle for 6 weeks. Spironolactone treatment did not affect blood pressure, fasting glucose levels or weight gain, but increased serum potassium and total cholesterol in both, diabetic and control mice. In addition, spironolactone significantly reduced serum insulin levels, but not aldosterone levels in diabetic mice. Insulin sensitivity, evaluated by the HOMA (homoeostatic model assessment)-index, was improved in spironolactone-treated diabetic mice. Mesenteric resistance arteries from vehicle-treated db/db mice exhibited inward hypertrophic remodelling, increased number of smooth muscle cells and increased vascular stiffness. These structural changes, determined by morphometric analysis and with a myography for pressurized arteries, were prevented by spironolactone treatment. Arteries from vehicle-treated db/db mice also exhibited augmented collagen content, determined by Picrosirius Red staining and Western blotting, increased reactive oxygen species (ROS) generation, determined by dihydroethidium (DHE) fluorescence, as well as increased expression of NAD(P)H oxidases 1 and 4 and increased activity of mitogen-activated protein kinases (MAPKs). Spironolactone treatment prevented all these changes, indicating that MR importantly contributes to diabetes-associated vascular dysfunction by inducing oxidative stress and by increasing the activity of redox-sensitive proteins

NADPH oxidase 5 is a pro‐contractile Nox isoform and a point of cross‐talk for calcium and redox signaling‐implications in vascular function

Background: NADPH Oxidase 5 (Nox5) is a calcium‐sensitive superoxide‐generating Nox. It is present in lower forms and higher mammals, but not in rodents. Nox5 is expressed in vascular cells, but the functional significance remains elusive. Given that contraction is controlled by calcium and reactive oxygen species, both associated with Nox5, we questioned the role of Nox5 in pro‐contractile signaling and vascular function. Methods and Results: Transgenic mice expressing human Nox5 in a vascular smooth muscle cell–specific manner (Nox5 mice) and Rhodnius prolixus, an arthropod model that expresses Nox5 endogenoulsy, were studied. Reactive oxygen species generation was increased systemically and in the vasculature and heart in Nox5 mice. In Nox5‐expressing mice, agonist‐induced vasoconstriction was exaggerated and endothelium‐dependent vasorelaxation was impaired. Vascular structural and mechanical properties were not influenced by Nox5. Vascular contractile responses in Nox5 mice were normalized by N‐acetylcysteine and inhibitors of calcium channels, calmodulin, and endoplasmic reticulum ryanodine receptors, but not by GKT137831 (Nox1/4 inhibitor). At the cellular level, vascular changes in Nox5 mice were associated with increased vascular smooth muscle cell [Ca2+]i, increased reactive oxygen species and nitrotyrosine levels, and hyperphosphorylation of pro‐contractile signaling molecules MLC20 (myosin light chain 20) and MYPT1 (myosin phosphatase target subunit 1). Blood pressure was similar in wild‐type and Nox5 mice. Nox5 did not amplify angiotensin II effects. In R. prolixus, gastrointestinal smooth muscle contraction was blunted by Nox5 silencing, but not by VAS2870 (Nox1/2/4 inhibitor). Conclusions: Nox5 is a pro‐contractile Nox isoform important in redox‐sensitive contraction. This involves calcium‐calmodulin and endoplasmic reticulum–regulated mechanisms. Our findings define a novel function for vascular Nox5, linking calcium and reactive oxygen species to the pro‐contractile molecular machinery in vascular smooth muscle cells