From bedside to bench to bedside: role of renin-angiotensin-aldosterone system in remodeling of resistance arteries in hypertension

SPECIAL MEDICAL EDITORIALSFrom bedside to bench to bedside: role of renin-angiotensin-aldosterone system in remodeling of resistance arteries in hypertensionErnesto L. Schiffrin, and Rhian M. TouyzErnesto L. Schiffrin, and Rhian M. TouyzPublished Online:01 Aug 2004https://doi.org/10.1152/ajpheart.00262.2004MoreSectionsPDF (268 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations resistance arteries are vessels of ∼100–300 μm in lumen diameter that are an important site of resistance to blood flow. Small artery-dependent increased peripheral resistance may participate in development and complications of hypertension. The degree of remodeling of small arteries has prognostic significance over a 10-yr period, with worse prognosis for hypertensive subjects with greater remodeling. In almost all hypertensive subjects, a reduction in lumen and an increase in the media-to-lumen ratio are found, without increase in its media cross section, as a result of rearrangement of smooth muscle cells and increased collagen and fibronectin. Approximately 60% of hypertensive patients exhibit endothelial dysfunction already in stage 1 hypertension. Study of human vascular smooth muscle cells and of vessels from experimental animals has demonstrated that ANG II, aldosterone, and endothelin exert remodeling effects in large measure by activation of NADPH oxidase, and to lesser degree by stimulating xanthine oxidase and mitochondrial reactive oxygen species generation. Stimulation of angiotensin type 1 (AT1) receptors (AT1R) leads to increased reactive oxygen species in part via activation of nonreceptor tyrosine kinases such as c-src, and of PKC and phospholipase D, and thereby contributes to endothelial dysfunction by inactivating nitric oxide (NO). Treatment of hypertensive patients with angiotensin-converting enzyme (ACE) inhibitors and angiotensin receptor blockers, but not β-blockers, corrects small artery structure and endothelial dysfunction, which may favorably affect outcome in the long term, beyond the 3–5 yr of randomized clinical trials during which most antihypertensives affect outcome similarly as long as blood pressure is well controlled.RESISTANCE ARTERIES IN HYPERTENSION: FROM CLINIC TO LABORATORYA major objective of translational research from bench to bedside is to test in humans novel therapeutic modalities developed through experimentation. However, because our understanding of many human diseases, such as hypertension, is still very limited, there is a critical need for bedside to bench research. We and other researchers have demonstrated that in human hypertension, resistance arteries undergo remodeling and altered reactivity. What remains uncertain, however, is how these processes occur and whether they are primary causative events or secondary adaptive phenomena. To answer some of these questions, it is imperative to extend bedside observations and findings to the bench, where molecular and cellular processes underlying vascular changes in hypertension can be studied in isolated vessels and cells in in vitro conditions. Ideally, one would like to examine cells directly involved in the pathological process under investigation. In hypertension, this includes vascular smooth muscle cells in resistance arteries. However, because of ethical and practical constraints, this is not always possible, and most studies investigating subcellular processes in hypertension are conducted, in large part, in cells from animal models, where hypertension is spontaneous or induced experimentally. When human cells are examined, they are usually derived from large arteries and veins obtained during surgery, from postmortem specimens, or from immortalized cell lines. Because these conditions do not control for blood pressure status and other variables of subjects from whom the cells are derived, it is inordinately difficult to extrapolate findings from such cell models to what may be happening in vascular cells during the development of hypertension. Because circulating blood cells are easily accessible from humans, many investigators have studied cellular events and signaling cascades in platelets, erythrocytes, and leukocytes as markers of events in vascular cells. However, this model is suboptimal because circulating cells are devoid of a supporting extracellular matrix and adventitia and have very different morphological and functional phenotypes compared with vascular smooth muscle cells.Over the past 10 yr, we successfully developed a system for the study of isolated vessels and vascular smooth muscle cells derived from human small arteries obtained from gluteal biopsies of subcutaneous tissue. This unique approach provides multiple benefits. First, small arteries and vascular cells derived from these are obtained from healthy individuals and well-characterized hypertensive patients, with or without treatment. Second, the vascular smooth muscle cells are derived from resistance arteries, which contribute to blood pressure regulation, elevation of peripheral resistance and development of hypertension. Third, low-passaged cells that maintain their morphological and functional properties are studied. With the use of this approach, implementing “bedside to bench” research is realized, gaining a fuller understanding of cellular processes and signaling pathways contributing to vascular remodeling in hypertension. Our long-term goal is to identify putative genes and/or proteins fundamentally involved in hypertensive vascular pathology that could be used as targets for manipulation in the prevention and management of human hypertension. Ultimately and ideally, our findings will echo back from “bench to bedside” to test novel therapeutic strategies developed through experimentation. We will recapitulate our own scientific itinerary from the bedside and studies of human small artery remodeling to the bench and the use of cells derived from these human small arteries as well as cells from vessels from experimental animals, and with the insights gained in the latter, back to the bedside, with studies of the action of agents that block these mechanisms, in particular the inhibitors of the renin-angiotensin system, and their effect on vascular remodeling in hypertensive humans.STRUCTURAL CHANGES IN RESISTANCE ARTERIES IN HUMAN ESSENTIAL HYPERTENSIONIncreased peripheral resistance is the hallmark of essential hypertension (94) (Fig. 1). Increased peripheral resistance results primarily from the increased energy dissipation that occurs when blood flows through small resistance arteries with reduced lumen. Resistance arteries have a lumen of 100–300 μm (11) and play an important role in the development of hypertension (136). Small artery remodeling may also lead to complications of hypertension (137), including myocardial ischemia (13, 61, 78), stroke (22), and renal failure (79). Recently it was demonstrated that increased media-to-lumen ratio is associated with increased cardiovascular events over a 10-yr period of follow-up of 150 subjects, which included normotensive controls, essential hypertensive individuals, and subjects with hyperaldosteronism, pheochromocytoma, renovascular hypertension, and diabetes (124).Fig. 1.Blood pressure is the product of cardiac output and total peripheral resistance. In general, increased peripheral resistance is the major factor contributing to elevated pressure in essential hypertension. Flow diagram demonstrates vascular mechanisms contributing to increased peripheral resistance, a hallmark of hypertension. Vasoactive agents, mechanical factors and oxidative stress interact to influence vascular structure and function. ANG II, angiotensin II; ET-1, endothelin-1; ↑, increase; −, no change.Download figureDownload PowerPointResistance arteries from normotensive and hypertensive patients, and from persons with diabetes and dyslipidemia, have been mainly investigated by obtaining biopsies of gluteal subcutaneous tissue, followed by dissection of small arteries. This technique, which was first introduced by Heagerty and Mulvany (1), is invasive but well tolerated and allows studies of well-characterized hypertensive subjects, individuals with dyslipidemia, and persons with diabetes, etc. The first finding with this technique has been that in essential hypertension small arteries do not exhibit hypertrophy, but rather “eutrophic remodeling” (62, 107, 140–144, 146), in which the outer diameter and lumen of these vessels are smaller, the media-to-lumen ratio is greater, but the cross-sectional area of the media is not different from that of age and sex-matched normotensive subjects (62, 107, 142). The increased media-to-lumen ratio (105, 143, 144) is the most reproducible parameter in longitudinal and cross-sectional studies (135). In small arteries from patients with essential hypertension there does not appear to be any smooth muscle cell hypertrophy or hyperplasia in the media (81) whereas there may be in secondary hypertension (125–127). Smooth muscle cells are rearranged around a smaller lumen, and this is accompanied by increased extracellular matrix deposition (71, 72, 74). The restructuring of smooth muscle cells may be the result of increased vasoconstriction (6) or of increased apoptosis in the periphery of the vessel with enhanced growth toward the lumen (73) and could also entail motility changes in smooth muscle cells contributing to this cellular rearrangement. Recently, we found that whereas structural remodeling of small arteries is found in all hypertensive patients, impairment of endothelial function is found in ∼60% and left ventricular hypertrophy in 45% of hypertensive patients (115) with stage I hypertension (according to the recent Joint National Committee VII classification) (21). We have concluded that altered structure of small arteries may be the first manifestation of target organ damage in hypertensive humans before the appearance of microalbuminuria, thickening of the intima media of the carotid arteries, or development of cardiac hypertrophy. It still remains to be established whether remodeling of resistance arteries precedes the development of hypertension or is a consequence of elevated blood pressure. In patients with severe hypertension, secondary hypertension, and in acromegalic patients (103, 122, 125–127), small arteries undergo hypertrophic remodeling, where media growth encroaches on the lumen to increase the media-to-lumen ratio and media cross-sectional area.Role of ANG II in Vascular Structural Changes in HypertensionVascular smooth muscle cells are dynamic, multifunctional cells that contribute to arterial remodeling through numerous processes, including cell growth (hyperplasia and hypertrophy), apoptosis, elongation of cells, reorganization of cells and/or altered extracellular matrix composition (8, 52, 72, 81, 125, 127) (Fig. 2). Hyperplasia refers to an increase in vascular smooth muscle cell number associated with DNA synthesis and is stimulated by ANG II (55, 127, 130). Hypertrophy, a reversible process, refers to increased cell size due to increased protein synthesis and/or increased intracellular cell water volume (68). ANG II stimulates hypertrophy by stimulating protein synthesis and by inducing activation of transmembrane transport systems, which influence transmembrane movement of ions and water. In experimental models of hypertension, hyperplasia, and hypertrophy have been demonstrated to contribute, to varying degrees, to vascular remodeling (3, 32, 106, 153). We demonstrated that ANG II stimulates both hyperplasia and hypertrophy and that in human vascular smooth muscle cells from resistance arteries of essential hypertensive patients, these growth responses are enhanced (175, 176).Fig. 2.Molecular and cellular mechanisms whereby ANG II influences vascular structure in hypertension. ANG II binds to the ANG type 1 (AT1) receptor (AT1R), leading to activation of receptor tyrosine kinases, such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR) and insulin-like growth factor-1 receptor (IGF-1R), and nonreceptor tyrosine kinases, such as c-Src. In addition, ANG II:AT1R binding induces activation of NAD(P)H oxidase resulting in intracellular generation of reactive oxygen species (ROS), which influence redox-sensitive signaling molecules, such as mitogen-activated protein (MAP) kinases (p38MAP kinase, JNK, ERK1/2, and ERK5), transcription factors [NF-κB, AP-1, and hypoxia-inducible factor (HIF-1)] and matrix metalloproteinases (MMP). ANG II may downregulate (indicated by minus sign) peroxisome proliferator-activated receptors (PPARs), which have anti-inflammatory effects, thus enhancing vascular inflammation. These signaling events regulate vascular smooth muscle cell growth, extracellular matrix (ECM) protein production and inflammatory responses. Under pathological conditions, altered ANG II signaling leads to altered growth, fibrosis, and inflammation, which contribute to structural remodeling in hypertension. PAI, plasminogen activator inhibitor; RXR, retinoid X receptor.Download figureDownload PowerPointIn eutrophic remodeling, characteristic of vessels in mild essential hypertension, apoptosis and vascular fibrosis may also be important (72–74). Apoptosis, gene-regulated cell death, is involved in the fine tuning of media growth and is increased in some vascular beds (36, 151) and decreased in others (31) in hypertensive rats. The exact role of apoptosis in arterial remodeling remains unclear, and it is unknown whether apoptosis is a growth-associated compensatory and adaptive process or a primary event. However, an imbalance between growth and apoptosis could be important (128, 190). Many studies (27, 151, 166) have suggested that ACE inhibitors and AT1R blockers could contribute to regression of vascular wall growth through activation of proapoptotic pathways. Apoptosis may also play a role in microvascular rarefaction, which has been implicated in the development of hypertension (56). Furthermore, there is evidence that detachment of vascular smooth muscle cells and endothelial cells (anoikis) may also contribute to vascular dysfunction and remodeling in hypertension (98, 159).Vascular fibrosis involves the accumulation of extracellular matrix proteins, such as collagen, elastin, fibrillin, fibronectin, and proteoglycans, in the vascular media. Increased collagen deposition in the vascular media has been demonstrated in experimental hypertension and in subcutaneous resistance arteries from essential hypertensive patients (74, 120, 130, 164). Increased collagen I and III mRNA and enhanced collagen protein synthesis by fibroblasts have also been demonstrated in patients with essential hypertension (28). Experimental studies indicate that collagen accumulation may be related, in part, to increased ANG II-stimulated synthesis (43, 130, 177). In addition to inducing production, ANG II regulates collagen degradation by attenuating interstitial matrix metalloproteinase (MMP) activity and by enhancing tissue inhibitor of metalloproteinase-1 production (16, 189). A similar role may be played by endothelin-1 in blood vessels and the heart (4). In patients with untreated hypertension, serum levels of MMP-1 (88) and MMP-9 (91) have been reduced with normalization after 1 yr of lisinopril treatment (88). These data further support a role for ANG II in profibrotic processes in human hypertension.Increasingly it has been appreciated that evidence of an inflammatory reaction may be recorded in association with the hypertensive process (7, 17, 73, 91, 165). It is unclear whether angiotensin II, endothelin-1, or aldosterone, agents that participate in the pathophysiology of hypertension and that induce inflammation in the heart, the vasculature, and the kidney (12, 82, 129, 171, 193) or blood pressure elevation by itself, through effects on adhesion molecules, chemokines, or endothelin-1 induced by cyclic mechanical strain (18, 195, 201) are associated with the inflammatory response that is found in hypertension. As well, the intriguing possibility that inflammation may activate the renin-angiotensin system and contribute to vascular remodeling and hypertension has been raised (194). Recently, it has been demonstrated that the activators of nuclear receptors, such as the peroxisome proliferator-activated receptors (PPARs), that are well-known hypolipidemic agents (the fibrates, PPAR α-agonists), or insulin sensitizers (glitazones, PPAR-γ agonists), downregulate the cardiac (37) and vascular inflammatory response in experimental animals (34, 35, 69) and serum markers of inflammation in humans (60). Furthermore, these inflammatory markers have been found to predict the risk of developing hypertension (42, 150). Chronic subclinical inflammation has been associated with the insulin resistance syndrome (45) that may precede the development of hypertension (57). Thus PPARs and vasoactive substances may be endogenous modulators of the inflammatory process that plays a role in structural changes that occur in the vasculature in hypertension, and inflammation may contribute to accelerate vascular damage in cardiovascular disease including hypertension. In fact, ANG II has been shown to downregulate PPARs, an effect mediated through activation of NF-κB (167). Blockade of the action of these agents or activation of PPARs may contribute to slow down cardiovascular damage in hypertension.Vascular inflammation is characterized by recruitment of monocytes and lymphocytes into the subendothelial space, production of chemotactic cytokines, increased expression of adhesion molecules, reactive smooth muscle cell proliferation, and altered extracellular matrix production and degradation. These processes, together with lipid oxidation, are proatherogenic, particularly in damaged arteries in hypertension. ANG II has significant proinflammatory actions in the vascular wall, inducing the production of reactive oxygen species, such as superoxide (O2−·) and H2O2, cytokines, adhesion molecules, and activation of redox-sensitive inflammatory genes (156). Vascular O2−· and H2O2 function intracellularly as second messengers, influencing redox-sensitive signaling molecules that regulate vascular smooth muscle cell contraction/dilation, cell growth (hypertrophy and hyperplasia), apoptosis, and extracellular matrix protein content (172, 191, 203). ANG II also modulates expression of proinflammatory molecules in the vessel wall, such as VCAM-1, ICAM, and E-selectin expression through redox-dependent pathways (23, 82, 119). In vascular smooth muscle cells, ANG II stimulates VCAM-1 production, chemokine monocyte chemotactic protein-1, and the cytokine IL-6 (116), which stimulate the recruitment of mononuclear leukocytes into the vessel media. Many of these factors are increased in plasma from hypertensive patients and it has been suggested that elevated circulating levels of cytokines and chemokines may reflect vascular inflammation and target organ damage in hypertensive patients (66, 96). To support the role of endogenous ANG II in vascular inflammation, AT1R blockers have been shown to reduce serum levels of VCAM-1, TNF-α, and O2−· in patients with early atherosclerosis (110).FUNCTIONAL ABNORMALITIES OF RESISTANCE ARTERIES IN HYPERTENSIONEnhanced myogenic tone has been reported in experimental hypertension (76), and it has been suggested that this is a primary event in hypertension that may lead to the chronic vascular changes described above. Most vasoconstrictor agents such as endothelin-1 and vasopressin as well as norepinephrine appear to elicit normal or diminished constrictor responses in clinical hypertension (1, 62, 141, 142). Whether there is amplification of vasoconstrictor responses by the structural or mechanical reduction of lumen diameter according to the law of Laplace (47, 48, 80, 199, 200) is controversial (75). However, ANG II appears to frequently elicit enhanced vasoconstrictor responses in essential hypertension (142). This effect may be direct or indirect through increased sympathetic activity (29, 117). Enhanced ANG II-induced vascular contraction is attributed to increased cytoplasmic free Ca2+ concentration ([Ca2+]i) and is probably due to postreceptor signaling changes (179, 180) (Fig. 3). Recent studies (20) demonstrate that increased sensitivity of the contractile machinery through RhoA/Rho kinase-dependent pathways may also contribute to increased vascular contractility in hypertension.Fig. 3.Molecular and cellular mechanisms whereby ANG II influences vascular function in hypertension. ANG II increases intracellular free Ca2+ concentration ([Ca2+]i) by stimulating Ca2+ influx and through mobilization from sarcoplasmic reticular (SR) stores, the latter in response to inositol 1,4,5-trisphosphate produced by activation of PLC. Increased activity of the RhoA/Rho kinase Ca2+-sensitizing pathway and PKC-dependent pathways (dashed line) also influence contractility. ANG II-induced NAD(P)H oxidase-mediated generation of superoxide (O2−·) quenches endothelial-derived nitric oxide (NO), resulting in decreased cGMP production and increased peroxynitrite (ONOO−) formation, with consequently altered vasodilation. In hypertension, altered ANG II signaling leads to increased contraction, decreased dilation, and increased vascular tone. DAG, diacylglycerol; MLC, myosin light chain; GEF, guanine nucleotide exchange factor; p, phosphorylated; G, G protein; ↑, increase; ↓, decrease.Download figureDownload PowerPointThe endothelium is critically involved in modulating vascular relaxation through release of endothelial-derived NO, stimulation of vascular smooth muscle cell soluble guanylate cyclase, and the subsequent increase in intracellular cGMP. 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