Reversal of renal dysfunction by targeted administration of VEGF into the stenotic kidney: a novel potential therapeutic approach

Renal microvascular (MV) damage and loss contribute to the progression of renal injury in renovascular disease (RVD). Whether a targeted intervention in renal microcirculation could reverse renal damage is unknown. We hypothesized that intrarenal vascular endothelial growth factor (VEGF) therapy will reverse renal dysfunction and decrease renal injury in experimental RVD. Unilateral renal artery stenosis (RAS) was induced in 14 pigs, as a surrogate of chronic RVD. Six weeks later, renal blood flow (RBF) and glomerular filtration rate (GFR) were quantified in vivo in the stenotic kidney using multidetector computed tomography (CT). Then, intrarenal rhVEGF-165 or vehicle was randomly administered into the stenotic kidneys (n = 7/group), they were observed for 4 additional wk, in vivo studies were repeated, and then renal MV density was quantified by 3D micro-CT, and expression of angiogenic factors and fibrosis was determined. RBF and GFR, MV density, and renal expression of VEGF and downstream mediators such as p-ERK 1/2, Akt, and eNOS were significantly reduced after 6 and at 10 wk of untreated RAS compared with normal controls. Remarkably, administration of VEGF at 6 wk normalized RBF (from 393.6 ± 50.3 to 607.0 ± 45.33 ml/min, P < 0.05 vs. RAS) and GFR (from 43.4 ± 3.4 to 66.6 ± 10.3 ml/min, P < 0.05 vs. RAS) at 10 wk, accompanied by increased angiogenic signaling, augmented renal MV density, and attenuated renal scarring. This study shows promising therapeutic effects of a targeted renal intervention, using an established clinically relevant large-animal model of chronic RAS. It also implies that disruption of renal MV integrity and function plays a pivotal role in the progression of renal injury in the stenotic kidney. Furthermore, it shows a high level of plasticity of renal microvessels to a single-dose VEGF-targeted intervention after established renal injury, supporting promising renoprotective effects of a novel potential therapeutic intervention to treat chronic RVD.     

Early superoxide scavenging accelerates renal microvascular rarefaction and damage in the stenotic kidney

Renal artery stenosis (RAS), the main cause of chronic renovascular disease (RVD), is associated with significant oxidative stress. Chronic RVD induces renal injury partly by promoting renal microvascular (MV) damage and blunting MV repair in the stenotic kidney. We tested the hypothesis that superoxide anion plays a pivotal role in MV dysfunction, reduction of MV density, and progression of renal injury in the stenotic kidney. RAS was induced in 14 domestic pigs and observed for 6 wk. Seven RAS pigs were chronically treated with the superoxide dismutase mimetic tempol (RAS+T) to reduce oxidative stress. Single-kidney hemodynamics and function were quantified in vivo using multidetector computer tomography (CT) and renal MV density was quantified ex vivo using micro-CT. Expression of angiogenic, inflammatory, and apoptotic factors was measured in renal tissue, and renal apoptosis and fibrosis were quantified in tissue sections. The degree of RAS and blood pressure were similarly increased in RAS and RAS+T. Renal blood flow (RBF) and glomerular filtration rate (GFR) were reduced in the stenotic kidney (280.1 ± 36.8 and 34.2 ± 3.1 ml/min, P < 0.05 vs. control). RAS+T kidneys showed preserved GFR (58.5 ± 6.3 ml/min, P = not significant vs. control) but a similar decreases in RBF (293.6 ± 85.2 ml/min) and further decreases in MV density compared with RAS. These changes were accompanied by blunted angiogenic signaling and increased apoptosis and fibrosis in the stenotic kidney of RAS+T compared with RAS. The current study shows that tempol administration provided limited protection to the stenotic kidney. Despite preserved GFR, renal perfusion was not improved by tempol, and MV density was further reduced compared with untreated RAS, associated with increased renal apoptosis and fibrosis. These results suggest that a tight balance of the renal redox status is necessary for a normal MV repair response to injury, at least at the early stage of RVD, and raise caution regarding antioxidant strategies in RAS.     

Molecular pathways involved in injury-repair and ADPKD progression

The major hallmark of Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the formation of many fluid-filled cysts in the kidneys, which ultimately impairs the normal renal structure and function, leading to end-stage renal disease (ESRD). A large body of evidence suggests that injury-repair mechanisms are part of ADPKD progression. Once cysts have been formed, proliferation and fluid secretion contribute to the cyst size increase, which eventually causes stress on the surrounding tissue resulting in local injury and fibrosis. In addition, renal injury can cause or accelerate cyst formation.In this review, we will describe the various mechanisms activated during renal injury and tissue repair and show how they largely overlap with the molecular mechanisms activated during PKD progression. In particular, we will discuss molecular mechanisms such as proliferation, inflammation, cell differentiation, cytokines and growth factors secretion, which are activated following the renal injury to allow the remodelling of the tissue and a proper organ repair. We will also underline how, in a context of PKD-related gene mutations, aberrant or chronic activation of these developmental pathways and repair/remodelling mechanisms results in exacerbation of the disease.     

The role of TNF-alpha in diabetic nephropathy: pathogenic and therapeutic implications

Diabetes mellitus and its complications are a public health problem. Diabetic nephropathy has become the main cause of renal failure, and furthermore is associated with a dramatic increase in cardiovascular risk. Unfortunately, the mechanisms leading to the development and progression of renal injury in diabetes are not yet fully known. There is now evidence that activated innate immunity and inflammation are relevant factors in the pathogenesis of diabetes. Furthermore, different inflammatory molecules, including pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), play a critical role in the development of microvascular diabetic complications, including nephropathy. This review discusses the role of TNF-alpha as a pathogenic factor in renal injury, focusing on diabetic nephropathy, and describes potential treatment strategies based on modulation of TNF-alpha activity.     

Oxidative Stress and Modification of Renal Vascular Permeability Are Associated with Acute Kidney Injury during P. berghei ANKA Infection

Malaria associated-acute kidney injury (AKI) is associated with 45% of mortality in adult patients hospitalized with severe form of the disease. However, the causes that lead to a framework of malaria-associated AKI are still poorly characterized. Some clinical studies speculate that oxidative stress products, a characteristic of Plasmodium infection, as well as proinflammatory response induced by the parasite are involved in its pathophysiology. Therefore, we aimed to investigate the development of malaria-associated AKI during infection by P. berghei ANKA, with special attention to the role played by the inflammatory response and the involvement of oxidative stress. For that, we took advantage of an experimental model of severe malaria that showed significant changes in the renal pathophysiology to investigate the role of malaria infection in the renal microvascular permeability and tissue injury. Therefore, BALB/c mice were infected with P. berghei ANKA. To assess renal function, creatinine, blood urea nitrogen, and ratio of proteinuria and creatininuria were evaluated. The products of oxidative stress, as well as cytokine profile were quantified in plasma and renal tissue. The change of renal microvascular permeability, tissue hypoxia and cellular apoptosis were also evaluated. Parasite infection resulted in renal dysfunction. Furthermore, we observed increased expression of adhesion molecule, proinflammatory cytokines and products of oxidative stress, associated with a decrease mRNA expression of HO-1 in kidney tissue of infected mice. The measurement of lipoprotein oxidizability also showed a significant increase in plasma of infected animals. Together, our findings support the idea that products of oxidative stress, as well as the immune response against the parasite are crucial to changes in kidney architecture and microvascular endothelial permeability of BALB/c mice infected with P. berghei ANKA.     

The role of microvesicles in tissue repair

Microvesicles (MVs) are released by almost all cells in resting and activated conditions. First described several years ago, it is only recently that their mechanisms of action are being elucidated, and their potential role in health and disease is drawing increasing attention. The main function of MVs is signaling through specific interactions with target cells and the transferring of gene products. Gaining further insights into the molecular specificity of MVs has allowed identification of the cellular source and may provide new diagnostic tools in the future. Indeed, an increasing body of evidence indicates that MVs are capable of mediating tissue repair in models of acute kidney and liver injury. In this review, we will discuss the mechanisms through which MVs from stem cells may act on target cells and may modify the response to injury. Furthermore, MVs from inflammatory cells are suspected to be involved in various diseases, such as cardiovascular and renal diseases, pathological pregnancy, tumors and sepsis. MVs are no doubt also involved in modulating immunity, and future studies will clarify their functional role in negatively modulating the cell response. Their role in physiological and pathological processes is increasingly appreciated. Depending on the cell source and the condition, MVs may be either beneficial or detrimental to the host. The recognition of their pathogenetic role may suggest new approaches to future therapies.     

The role of microvesicles in tissue repair

Microvesicles (MVs) are released by almost all cells in resting and activated conditions. First described several years ago, it is only recently that their mechanisms of action are being elucidated, and their potential role in health and disease is drawing increasing attention. The main function of MVs is signaling through specific interactions with target cells and the transferring of gene products. Gaining further insights into the molecular specificity of MVs has allowed identification of the cellular source and may provide new diagnostic tools in the future. Indeed, an increasing body of evidence indicates that MVs are capable of mediating tissue repair in models of acute kidney and liver injury. In this review, we will discuss the mechanisms through which MVs from stem cells may act on target cells and may modify the response to injury. Furthermore, MVs from inflammatory cells are suspected to be involved in various diseases, such as cardiovascular and renal diseases, pathological pregnancy, tumors and sepsis. MVs are no doubt also involved in modulating immunity, and future studies will clarify their functional role in negatively modulating the cell response. Their role in physiological and pathological processes is increasingly appreciated. Depending on the cell source and the condition, MVs may be either beneficial or detrimental to the host. The recognition of their pathogenetic role may suggest new approaches to future therapies.     

The interface of mechanical loading and biological variables as they pertain to the development of tendinosis

Different tendons are designed to withstand different mechanical loads in their individual environments. Variable physiologic loading ranges and correspondingly different injury thresholds lead to tendon heterogeneity. Also, tendon heterogeneity is evident when examining how different tendons regulate their response to changes in mechanical loading (over- and under-loading). The response of tendons to changes in mechanical loading plays an important role in the induction and progression of tendinosis which is tendon degeneration without inflammation. Tendon overuse injury is likely related to abnormal mechanical loading that deviates from normal mechanical loading in magnitude, frequency, duration and/or direction. Mechanical loading that results in tendon overuse injury can initiate a repair process but, after failed initial repair, non-resolving chronic attempted repair appears to lead to a "smoldering" fibrogenesis. Contributions of regulatory components, including minor components in the "nerve-mast cell-myofibroblast axis", are key features in the development and progression of tendinosis. Hormonal and genetic factors may also influence risk for tendinosis. Further understanding of how tendinosis induction is related to mechanical use/overuse, how tendinosis progression is related to abnormal regulation of attempted repair, and how induction and/or progression are modulated by other risk factors may lead to interventions that mitigate risk and enhance functional repair.     

NLRP3 Inflammasome Mediates Albumin-induced Renal Tubular Injury through Impaired Mitochondrial Function

Proteinuria serves as a direct causative factor of renal tubular cell injury and is highly associated with the progression of chronic kidney disease via uncertain mechanisms. Recently, evidence demonstrated that both NLRP3 inflammasome and mitochondria are involved in the chronic kidney disease progression. The present study was undertaken to examine the role of NLRP3 inflammasome/mitochondria axis in albumin-induced renal tubular injury. In patients with proteinuria, NLRP3 was significantly up-regulated in tubular epithelial cells and was positively correlated with the severity of proteinuria. In agreement with these results, albumin remarkably activated NLRP3 inflammasome in both in vitro renal tubular cells and in vivo kidneys in parallel with significant epithelial cell phenotypic alteration and cell apoptosis. Genetic disruption of NLRP3 inflammasome remarkably attenuated albumin-induced cell apoptosis and phenotypic changes under both in vitro and in vivo conditions. In addition, albumin treatment resulted in a significant mitochondrial abnormality as evidenced by the impaired function and morphology, which was markedly reversed by invalidation of NLRP3/caspase-1 signaling pathway. Interestingly, protection of mitochondria function by Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP) or cyclosporin A (CsA) robustly attenuated albumin-induced injury in mouse proximal tubular cells. Collectively, these findings demonstrated a pathogenic role of NLRP3 inflammasome/caspase-1/mitochondria axis in mediating albumin-induced renal tubular injury. The discovery of this novel axis provides some potential targets for the treatment of proteinuria-associated renal injury.     

Renal epithelial injury and fibrosis

Chronic kidney disease at a certain advanced stage inevitably progresses to end stage renal failure characterized by the progressing loss of nephrons accompanied by the increasing appearance of fibrotic tissue, called renal fibrosis. The urgent question is whether renal fibrosis is a response to injury or if fibrosis acquires a self-sustaining progressive potential that actively contributes to the deterioration of the kidney. The present review distinguishes between renal fibrosis subsequent to a glomerular injury and fibrosis subsequent to a primary tubular injury. Glomerular diseases enter a progressing course after encroaching onto the tubule leading to what is generally called “tubulointerstitial fibrosis”. The progression of the injury at the level of the tubulointerstitium appears to be fully dependent on the progression of the disease in the corresponding glomerulus. Primary tubular injuries have a very good chance of recovery. If they develop a local fibrotic process, this seems to be supportive for recovery. Cases in which recovery fails appear to secondarily initiate a glomerular disease accounting for a glomerulus-dependent vicious cycle to progression. Even if most researchers think of renal fibrosis as a process promoting the progression of the disease this review points out that the available structural evidence speaks in favour of a protective role of fibrosis supporting recovery after acute tubular injury or, under progressing circumstances, providing a firm three-dimensional framework that permits still intact or partially damaged nephrons to survive. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease.     

Endothelin-A receptor blockade slows the progression of renal injury in experimental renovascular disease

Endothelin (ET)-1, a potent renal vasoconstrictor with mitogenic properties, is upregulated by ischemia and has been shown to induce renal injury via the ET-A receptor. The potential role of ET-A blockade in chronic renovascular disease (RVD) has not, to our knowledge, been previously reported. We hypothesized that chronic ET-A receptor blockade would preserve renal hemodynamics and slow the progression of injury of the stenotic kidney in experimental RVD. Renal artery stenosis, a major cause of chronic RVD, was induced in 14 pigs and observed for 6 wk. In half of the pigs, chronic ET-A blockade was initiated (RVD+ET-A, 0.75 mg·kg(-1)·day(-1)) at the onset of RVD. Single-kidney renal blood flow, glomerular filtration rate, and perfusion were quantified in vivo after 6 wk using multidetector computer tomography. Renal microvascular density was quantified ex vivo using three-dimensional microcomputer tomography, and growth factors, inflammation, apoptosis, and fibrosis were determined in renal tissue. The degree of stenosis and increase in blood pressure were similar in RVD and RVD+ET-A pigs. Renal hemodynamics, function, and microvascular density were decreased in the stenotic kidney but preserved by ET-A blockade, accompanied by increased renal expression of vascular endothelial growth factor, hepatocyte growth factor, and downstream mediators such as phosphorilated-Akt, angiopoietins, and endothelial nitric oxide synthase. ET-A blockade also reduced renal apoptosis, inflammation, and glomerulosclerosis. This study shows that ET-A blockade slows the progression of renal injury in experimental RVD and preserves renal hemodynamics, function, and microvascular density in the stenotic kidney. These results support a role for ET-1/ET-A as a potential therapeutic target in chronic RVD.     

Mechanisms of non-canonical activation of ataxia telangiectasia mutated

ATM is a master regulator of the cellular response to DNA damage. The classical mechanism of ATM activation involves its monomerization in response to DNA double-strand breaks, resulting in ATM-dependent phosphorylation of more than a thousand substrates required for cell cycle progression, DNA repair, and apoptosis. Here, new experimental evidence for non-canonical mechanisms of ATM activation in response to stimuli distinct from DNA double-strand breaks is discussed. It includes cytoskeletal changes, chromatin modifications, RNA–DNA hybrids, and DNA single-strand breaks. Noncanonical ATM activation may be important for the pathology of the multisystemic disease Ataxia Telangiectasia.     

Unfolded protein response at the cross roads of tumourigenesis,oxygen sensing and drug resistance in clear cell renal cell carcinoma

Clear cell renal cell carcinoma (ccRCC) is the most common form of kidney cancer. Despite therapeutic advances, long term survival in patients diagnosed with advanced disease is low. Efforts to understand the mechanisms promoting disease progression will likely produce novel therapeutic targets. The unfolded protein response (UPR) is activated when unfolded protein accumulates in the endoplasmic reticulum (ER) upon cellular stress. Constitutive UPR activation is a characteristic of many malignancies. We discuss the accumulating evidence that describes a role for the UPR in ccRCC. Studies focused on UPR signalling may provide compelling avenues for therapeutic intervention in the future.     

Persistent kidney dysfunction in swine renal artery stenosis correlates with outer cortical microvascular remodeling

Percutaneous transluminal renal stenting (PTRS) does not consistently improve renal function in patients with atherosclerotic renovascular disease, but the mechanisms underlying irreversible kidney injury have not been fully elucidated. We hypothesized that renal dysfunction after PTRS is linked to ongoing renal microvascular (MV) remodeling. Pigs were studied after 10 wk of atherosclerosis and renal artery stenosis (ARAS), ARAS treated with PTRS 4 wk earlier, and normal controls (n = 10 each). Renal blood flow (RBF) and glomerular filtration rate (GFR) were studied using multidetector computer tomography. Renal microvascular architecture (micro-CT), angiogenic activity, oxidative stress, and fibrosis were evaluated ex vivo. Four weeks after PTRS, blood pressure was normalized. However, GFR and RBF remained similarly decreased in untreated ARAS and ARAS+PTRS (P < 0.05 vs. normal). MV rarefaction was unaltered after revascularization, and the spatial density of outer cortical microvessels correlated with residual GFR. Interstitial fibrosis and altered expression of proangiogenic and profibrotic factors persisted after PTRS. Tubulointerstitial injury in ARAS persisted 4 wk after mechanically successful PTRS, and vessel loss correlated with residual renal dysfunction. MV loss and fibrosis in swine ARAS might account for persistent renal dysfunction after PTRS and underscore the need to assess renal parenchymal disease before revascularization.     

Adjunctive mesenchymal stem/stromal cells augment microvascular function in poststenotic kidneys treated with low-energy shockwave therapy

Effective therapeutic strategies are needed to preserve renal function in patients with atherosclerotic renal artery stenosis (ARAS). Low-energy shockwave therapy (SW) and adipose tissue-derived mesenchymal stem/stromal cells (MSCs) both stimulate angiogenesis repair of stenotic kidney injury. This study tested the hypothesis that intrarenal delivery of adipose tissue-derived MSCs would enhance the capability of SW to preserve stenotic kidney function and structure. Twenty-two pigs were studied after 16 weeks of ARAS, ARAS treated with a SW regimen (bi-weekly for 3 weeks) with or without subsequent intrarenal delivery of adipose tissue-derived MSCs and controls. Four weeks after treatment, single-kidney renal blood flow (RBF) before and after infusion of acetylcholine, glomerular filtration rate (GFR), and oxygenation were assessed in vivo and the renal microcirculation, fibrosis, and oxidative stress ex vivo. Mean arterial pressure remained higher in ARAS, ARAS + SW, and ARAS + SW + MSC compared with normal. Both SW and SW + MSC similarly elevated the decreased stenotic kidney GFR and RBF observed in ARAS to normal levels. Yet, SW + MSC significantly improved RBF response to acetylcholine in ARAS, and attenuated capillary loss and oxidative stress more than SW alone. Density of larger microvessels was similarly increased by both interventions. Therefore, although significant changes in functional outcomes were not observed in a short period of time, adjunct MSCs enhanced pro-angiogenic effect of SW to improve renal microvascular outcomes, suggesting this as an effective stratege for long-term management of renovascular disease.     

Contribution of apoptotic cell death to renal injury

Cell number abnormalities are frequent in renal diseases, and range from the hypercellularity of postinfectious glomerulonephritis to the cell depletion of chronic renal atrophy. Recent research has shown that apoptosis and its regulatory mechanisms contribute to cell number regulation in the kidney. The role of apoptosis ranges from induction to repair and progression of renal injury. Death ligands and receptors, such as TNF and FasL, proapoptotic and antiapoptotic Bcl-2 family members and caspases have all been shown to participate in apoptosis regulation in the course of renal injury. These proteins represent potential therapeutic targets, which should be further explored.     

Congenital DNA repair deficiency results in protection against renal ischemia reperfusion injury in mice

Cockayne syndrome and other segmental progerias with inborn defects in DNA repair mechanisms are thought to be due in part to hypersensitivity to endogenous oxidative DNA damage. The accelerated aging-like symptoms of this disorder include dysmyelination within the central nervous system, progressive sensineuronal hearing loss and retinal degeneration. We tested the effects of congenital nucleotide excision DNA repair deficiency on acute oxidative stress sensitivity in vivo . Surprisingly, we found mouse models of Cockayne syndrome less susceptible than wild type animals to surgically induced renal ischemia reperfusion injury, a multifactorial injury mediated in part by oxidative damage. Renal failure-related mortality was significantly reduced in Csb^−/– mice, kidney function was improved and proliferation was significantly higher in the regenerative phase following ischemic injury. Protection from ischemic damage correlated with improved baseline glucose tolerance and insulin sensitivity and a reduced inflammatory response following injury. Protection was further associated with genetic ablation of a different Cockayne syndrome-associated gene, Csa . Our data provide the first functional in vivo evidence that congenital DNA repair deficiency can induce protection from acute stress in at least one organ. This suggests that while specific types of unrepaired endogenous DNA damage may lead to detrimental effects in certain tissues, they may at the same time elicit beneficial adaptive changes in others and thus contribute to the tissue specificity of disease symptoms.     

Role of fibroblast growth factor-binding protein in the pathogenesis of HIV-associated hemolytic uremic syndrome

A characteristic finding of childhood HIV-associated hemolytic uremic syndrome (HIV-HUS) is the presence of endothelial injury and microcystic tubular dilation, leading to a rapid progression of the renal disease. We have previously shown that a secreted fibroblast growth factor-binding protein (FGF-BP) is upregulated in kidneys from children affected with HIV-HUS and HIV nephropathy. Here, we sought to determine the potential role of FGF-BP in the pathogenesis of HIV-HUS. By immunohistochemical and in situ hybridization studies, we observed FGF-BP protein and mRNA upregulation in regenerating renal tubular epithelial cells from kidneys of HIV-Tg26 mice with late-stage renal disease, that is, associated with the development of microcystic tubular dilatation and accumulation of FGF-2. Moreover, FGF-BP increased the FGF-2-dependent growth and survival of cultured primary human renal glomerular endothelial cells and enhanced FGF-2-induced MAPK/ERK2 activation, as well as the proliferation of immortalized GM7373 endothelial cells. We propose that HIV-Tg26 mice are a clinically relevant model system to study the role of FGF-BP in the pathogenesis of HIV-associated renal diseases. Furthermore, the upregulation of FGF-BP by regenerating renal tubular epithelial cells may provide a mechanism by which the regenerative and angiogenic activity of FGF-2 in renal capillaries can be modulated in children with HIV-HUS and other renal disease.     

Low-flow vascular remodeling in the metabolic syndrome X

Peripheral microvascular dysfunction is a common affliction in patients with the metabolic syndrome X. Previous studies have described a number of vascular impairments in vasomotor control in both human patients and animal models of syndrome X, but the net effect of these impairments on microvascular structure has not been examined. The goal of the current study was to test the hypothesis that syndrome X reduces muscle perfusion and induces vascular remodeling. The obese Zucker rat was used as a model of syndrome X, and the microcirculation of the hindlimb and brain were examined. Obese Zucker rats were obese, hyperlipidemic, hyperinsulinemic, and hyperglycemic. Blood flow to the hindlimb was reduced by 59% in obese rats relative to lean rats. Skeletal muscle resistance arteries of the hindlimb microcirculation of obese rats had thinner walls, smaller lumens, and reduced distensibility. Hindlimb microvessels from obese rats also demonstrated reduced expression of vascular smooth muscle cell markers. Each of these traits is consistent with low-flow remodeling. In contrast, the cerebral microcirculation, where flow is vigorously autoregulated, showed no vascular remodeling nor were there changes in microvascular smooth muscle marker expression. Neither physical activity nor muscle mass were significantly different between lean and obese rats. Taken together, these findings suggest that syndrome X, by reducing hindlimb blood flow, induces a marked remodeling of microcirculation to favor smaller, less distensible vessels. This remodeling may result in an architectural limitation of maximum perfusion capacity and may be an important maladaption in the progression of peripheral microvascular disease.