Background-Atherosclerotic renovascular disease may augment deterioration of renal function and ischemic nephropathy compared with other causes of renal artery stenosis (RAS), but the underlying mechanisms remain unclear. This study was designed to test the hypothesis that concurrent early atherosclerosis and hypoperfusion might have greater early deleterious effects on the function and structure of the stenotic kidney. Methods and Results-Regional renal hemodynamics and function at baseline and during vasoactive challenge (acetylcholine or sodium nitroprusside) were quantified in vivo in pigs by electron-beam computed tomography after a 12-week normal (nϭ7) or hypercholesterolemic (HC, nϭ7) diet, RAS (nϭ6), or concurrent HC and a similar degree of RAS (HCϩRAS, nϭ7). Flash-frozen renal tissue was studied ex vivo. Basal cortical perfusion and single-kidney glomerular filtration rate (GFR) were decreased similarly in the stenotic RAS and HCϩRAS kidneys, but tubular fluid reabsorption was markedly impaired only in HCϩRAS. Perfusion responses to challenge were similarly blunted in the experimental groups. Stimulated GFR increased in normal, HC, and RAS (38.3Ϯ3.6%, 36.4Ϯ7.6%, and 60.4Ϯ9.3%, respectively, PϽ0.05), but not in HCϩRAS (6.5Ϯ15.1%). These functional abnormalities in HCϩRAS were accompanied by augmented perivascular, tubulointerstitial, and glomerular fibrosclerosis, inflammation, systemic and tissue oxidative stress, and tubular expression of nuclear factor-B and inducible nitric oxide synthase. Conclusions-Early chronic HCϩRAS imposes distinct detrimental effects on renal function and structure in vivo and in vitro, evident primarily in the tubular and glomerular compartments. Increased oxidative stress may be involved in the proinflammatory and progrowth changes observed in the stenotic HCϩRAS kidney, which might potentially facilitate the clinically observed progression to end-stage renal disease.

Background-Endothelial progenitor cells (EPCs) promote neovascularization and endothelial repair. Renal artery stenosis (RAS) may impair renal function by inducing intrarenal microvascular injury and remodeling. We investigated whether replenishment with EPCs would protect the renal microcirculation in chronic experimental renovascular disease. Methods and Results-Single-kidney hemodynamics and function were assessed with the use of multidetector computed tomography in vivo in pigs with RAS, pigs with RAS 4 weeks after intrarenal infusion of autologous EPCs, and controls. Renal microvascular remodeling and angiogenic pathways were investigated ex vivo with the use of micro-computed tomography, histology, and Western blotting. EPCs increased renal expression of angiogenic factors, stimulated proliferation and maturation of new vessels, and attenuated renal microvascular remodeling and fibrosis in RAS. Furthermore, EPCs normalized the blunted renal microvascular and filtration function. Conclusions-The present study shows that a single intrarenal infusion of autologous EPCs preserved microvascular architecture and function and decreased microvascular remodeling in experimental chronic RAS. It is likely that restoration of the angiogenic cascade by autologous EPCs involved not only generation of new vessels but also acceleration of their maturation and stabilization. This contributed to preserving the blood supply, hemodynamics, and function of the RAS kidney, supporting EPCs as a promising therapeutic intervention for preserving the kidney in renovascular disease. (Circulation. 2009;119:547-557.)Key Words: blood flow Ⅲ kidney Ⅲ progenitor cells Ⅲ renal artery stenosis Ⅲ hypertension, renal E ndothelial progenitor cells (EPCs) mobilized endogenously in response to ischemia play a crucial role in augmenting neovascularization of ischemic tissues and endothelial replacement after vascular injury. Replenishment of such cells may limit vascular injury through reconstitution of the luminal barrier and cellular secretion of paracrine factors, providing a novel therapeutic option. 1,2 Indeed, growing experimental and clinical evidence underscores the critical role that circulating cells play in healing the endothelium when the intrinsic system is unable to adequately support tissue repair. Targeted delivery of EPCs has been shown to improve the function of the infarcted myocardium, 3 decrease hindlimb ischemia, 4,5 rescue the kidney from acute ischemia injury, 6 and participate in glomerular endothelial repair in glomerulonephritis. 7 Clinical Perspective p 557Ischemic nephropathy secondary to renal artery stenosis (RAS) represents an important cause of renovascular disease and hypertension that may induce renal injury and lead to end-stage renal disease. The presence of renovascular disease also constitutes an independent predictor for increased morbidity and mortality in cardiovascular disease and cardiac events. 8 We have shown previously that the kidney exposed to chronic RAS shows significant functional deteriorati...

Objective-Atherosclerotic renovascular disease (ARVD) aggravates renal scarring more than other causes of renal artery stenosis (RAS), but the underlying pathogenic mechanisms of this potential profibrotic effect remain unclear. We tested the hypothesis that coexistence of atherosclerosis and RAS interferes with renal tissue remodeling. Methods and Results-Single-kidney hemodynamics and function were quantified in vivo with electron-beam computed tomography in 3 groups of pigs (nϭ7 each): normal pigs, pigs 12 weeks after induction of unilateral RAS (RAS group), and pigs with similar-degree RAS fed a 12-week 2% hypercholesterolemic diet (HCϩRAS, simulating early ARVD). Kidneys were studied ex vivo by Western blotting and immunohistochemistry. Renal volume, renal blood flow, and glomerular filtration rate were similarly decreased in RAS and HCϩRAS ischemic kidneys, accompanied by similar increased expression of profibrotic factors like transforming growth factor-␤, tissue inhibitor of metalloproteinase-1, and plasminogen activator inhibitor-1. Nevertheless, HCϩRAS kidneys showed increased intrarenal fibrosis compared with RAS-only kidneys. Furthermore, expression of nuclear factor-B was increased, expression of extracellular (matrix metalloproteinase-2) and intracellular (ubiquitin) protein degradation systems was decreased, and apoptosis was blunted. Conclusions-Diet-induced HC superimposed on RAS accelerates the development of fibrosis in the stenotic kidney by amplifying profibrotic mechanisms and disrupting tissue remodeling. These alterations might contribute to renal disease progression in ARVD and might account for the increased propensity for end-stage renal disease.

Objective-Mechanisms of renal injury distal to renal artery stenosis (RAS) remain unclear. We tested the hypothesis that it involves microvascular remodeling consequent to increased oxidative stress. Methods and Results-Three groups of pigs (nϭ6 each) were studied after 12 weeks of RAS, RASϩantioxidant supplementation (100 IU/kg vitamin E and 1 g vitamin C daily), or controls. The spatial density and tortuousity of renal microvessels (Ͻ500 m) were tomographically determined by 3D microcomputed tomography. The in situ production of superoxide anion and the expression of vascular endothelial growth factor (VEGF), its receptor VEGFR-2, hypoxia-inducible-factor (HIF)-1␣, von Hippel-Lindau (VHL) protein, and NAD(P)H oxidase (p47phox and p67phox subunits) were determined in cortical tissue. RAS and RASϩantioxidant groups had similar degrees of stenosis and hypertension. The RAS group showed a decrease in spatial density of cortical microvessels, which was normalized in the RASϩantioxidant group, as was arteriolar tortuousity. RAS kidneys also showed tissue fibrosis (by trichrome and Sirius red staining), increased superoxide anion abundance, NAD(P)H oxidase, VHL protein, and HIF-1␣ mRNA expression. In contrast, expression of HIF-1␣, VEGF, and VEGFR-2 protein was downregulated. These were all significantly improved by antioxidant intervention. Conclusions-Increased oxidative stress in the stenotic kidney alters growth factor activity and plays an important role in renal microvascular remodeling, which can be prevented by chronic antioxidant intervention. (Arterioscler Thromb

This study underscores the importance of the renal microcirculation in renovascular disease. Intra-renal administration of VEGF preserved renal MV architecture and function of the stenotic kidney, which in turn preserved renal haemodynamics and function and decreased renal fibrosis. These observations suggest that preventing renal MV loss may be a potential target for therapeutic approaches for patients with chronic renovascular disease.

Hypertension is the most common chronic disease in the world, yet the precise cause of elevated blood pressure often cannot be determined. Animal models have been useful for unraveling the pathogenesis of hypertension and for testing novel therapeutic strategies. The utility of animal models for improving the understanding of the pathogenesis, prevention, and treatment of hypertension and its comorbidities depends on their validity for representing human forms of hypertension, including responses to therapy, and on the quality of studies in those models (such as reproducibility and experimental design). Important unmet needs in this field include the development of models that mimic the discrete hypertensive syndromes that now populate the clinic, resolution of ongoing controversies in the pathogenesis of hypertension, and the development of new avenues for preventing and treating hypertension and its complications. Animal models may indeed be useful for addressing these unmet needs.

We tested the hypothesis that statins would decrease renal injury in renal artery stenosis (RAS) by restoring angiogenesis and attenuating intrarenal microvascular (IMV) remodeling. Single-kidney hemodynamics and function were quantified using electron-beam-computed tomography (CT) in normocholesterolemic pigs after 12 wk of experimental RAS, RAS supplemented with simvastatin (RAS+simvastatin), and normal controls. Renal circulation was also studied in vivo using angiography and ex vivo using a unique 3D micro-CT imaging technique. Angiogenic and remodeling pathways were subsequently explored in renal tissue. Blood pressure and the degree of stenosis were similarly increased in RAS groups. Simvastatin in RAS enhanced both intrarenal angiogenesis and peri-stenosis arteriogenesis and increased the expression of angiogenic growth factors and hypoxia-inducible factor-1alpha. Furthermore, simvastatin decreased tissue-transglutaminase expression and IMV inward remodeling, restored IMV endothelial function, decreased fibrogenic activity, and improved renal function. Chronic simvastatin supplementation promoted angiogenesis in vivo, decreased ischemia-induced IMV remodeling, and improved IMV function in the stenotic kidney, independent of lipid lowering. These novel renoprotective effects suggest a role for simvastatin in preserving the ischemic kidney in chronic RAS.

Background-Hypercholesterolemia (HC) and atherosclerosis can elicit oxidative stress, coronary endothelial dysfunction, and myocardial ischemia, which may induce growth-factor expression and lead to myocardial neovascularization. We tested the hypothesis that chronic antioxidant intervention in HC would attenuate neovascularization and preserve the expression of hypoxia-inducible factor (HIF)-1␣ and vascular endothelial growth factor (VEGF). Methods and Results-Three groups of pigs (nϭ6 each) were studied after 12 weeks of normal or 2% HC diet or HCϩantioxidant supplementation (100 IU/kg vitamin E and 1 g vitamin C daily). Myocardial samples were scanned ex vivo with a novel 3D micro-CT scanner, and the spatial density and tortuosity of myocardial microvessels were determined in situ. VEGF mRNA, protein levels of VEGF and VEGF receptor-1, HIF-1␣, nitrotyrosine, and superoxide dismutase (SOD) were determined in myocardial tissue. The HC and HCϩantioxidant groups had similar increases in serum cholesterol levels. HC animals showed an increase in subendocardial spatial density of microvessels compared with normal (160.5Ϯ11.8 versus 95.3Ϯ8.2 vessels/cm 2 , PϽ0.05), which was normalized in HCϩantioxidant (92.5Ϯ20.5 vessels/cm 2 , PϽ0.05 versus HC), as was arteriolar tortuosity. In addition, HC induced upregulation of VEGF, HIF-1␣, and nitrotyrosine expression and decreased SOD expression and activity, all of which were preserved by antioxidant intervention. Conclusions-Changes