Extracellular vesicles (EVs) derived from human bone marrow mesenchymal stromal cells (MSCs) promote the regeneration of kidneys in different animal models of acute kidney injury (AKI) in a manner comparable with the cells of origin. However, due to the heterogeneity observed in the EVs isolated from MSCs, it is unclear which population is responsible for the proregenerative effects. We therefore evaluated the effect of various EV populations separated by differential ultracentrifugation (10K population enriched with microvesicles and 100K population enriched with exosomes) on AKI recovery. Only the exosomal-enriched population induced an improvement of renal function and morphology comparable with that of the total EV population. Interestingly, the 100K EVs exerted a proproliferative effect on murine tubular epithelial cells, both in vitro and in vivo. Analysis of the molecular content from the different EV populations revealed a distinct profile. The 100K population, for instance, was enriched in specific mRNAs (CCNB1, CDK8, CDC6) reported to influence cell cycle entry and progression; miRNAs involved in regulating proliferative/antiapoptotic pathways and growth factors (hepatocyte growth factor and insulin-like growth factor-1) that could explain the effect of renal tubular cell proliferation. On the other hand, the EV population enriched in microvesicles (10K) was unable to induce renal regeneration and had a molecular profile with lower expression of proproliferative molecules. In conclusion, the different molecular composition of exosome- and microvesicle-enriched populations may explain the regenerative effect of EVs observed in AKI.

Background— Recent clinical studies have suggested a major protective role for the antioxidant enzyme glutathione peroxidase-1 (GPx1) in diabetes-associated atherosclerosis. We induced diabetes in mice deficient for both GPx1 and apolipoprotein E (ApoE) to determine whether this is merely an association or whether GPx1 has a direct effect on diabetes-associated atherosclerosis. Methods and Results— ApoE-deficient (ApoE −/− ) and ApoE/GPx1 double-knockout (ApoE −/− GPx1 −/− ) mice were made diabetic with streptozotocin and aortic lesion formation, and atherogenic pathways were assessed after 10 and 20 weeks of diabetes. Aortic proinflammatory and profibrotic markers were determined by both quantitative reverse-transcription polymerase chain reaction analysis after 10 weeks of diabetes and immunohistochemical analysis after 10 and 20 weeks of diabetes. Sham-injected nondiabetic counterparts served as controls. Atherosclerotic lesions within the aortic sinus region, as well as arch, thoracic, and abdominal lesions, were significantly increased in diabetic ApoE −/− GPx1 −/− aortas compared with diabetic ApoE −/− aortas. This increase was accompanied by increased macrophages, α-smooth muscle actin, receptors for advanced glycation end products, and various proinflammatory (vascular cell adhesion molecule-1) and profibrotic (vascular endothelial growth factor and connective tissue growth factor) markers. Quantitative reverse-transcription polymerase chain reaction analysis showed increased expression of receptors for advanced glycation end products (RAGE), vascular cell adhesion molecule-1, vascular endothelial growth factor, and connective tissue growth factor. Nitrotyrosine levels were significantly increased in diabetic ApoE −/− GPx1 −/− mouse aortas. These findings were observed despite upregulation of other antioxidants. Conclusions— Lack of functional GPx1 accelerates diabetes-associated atherosclerosis via upregulation of proinflammatory and profibrotic pathways in ApoE −/− mice. Our study provides evidence of a protective role for GPx1 and establishes GPx1 as an important antiatherogenic therapeutic target in patients with or at risk of diabetic macrovascular disease.

OBJECTIVEMonocyte chemoattractant protein-1 (MCP-1), a chemokine binding to the CC chemokine receptor 2 (CCR2) and promoting monocyte infiltration, has been implicated in the pathogenesis of diabetic nephropathy. To assess the potential relevance of the MCP-1/CCR2 system in the pathogenesis of diabetic proteinuria, we studied in vitro if MCP-1 binding to the CCR2 receptor modulates nephrin expression in cultured podocytes. Moreover, we investigated in vivo if glomerular CCR2 expression is altered in kidney biopsies from patients with diabetic nephropathy and whether lack of MCP-1 affects proteinuria and expression of nephrin in experimental diabetes.RESEARCH DESIGN AND METHODSExpression of nephrin was assessed in human podocytes exposed to rh-MCP-1 by immunofluorescence and real-time PCR. Glomerular CCR2 expression was studied in 10 kidney sections from patients with overt nephropathy and eight control subjects by immunohistochemistry. Both wild-type and MCP-1 knockout mice were made diabetic with streptozotocin. Ten weeks after the onset of diabetes, albuminuria and expression of nephrin, synaptopodin, and zonula occludens-1 were examined by immunofluorescence and immunoblotting.RESULTSIn human podocytes, MCP-1 binding to the CCR2 receptor induced a significant reduction in nephrin both mRNA and protein expression via a Rho-dependent mechanism. The MCP-1 receptor, CCR2, was overexpressed in the glomerular podocytes of patients with overt nephropathy. In experimental diabetes, MCP-1 was overexpressed within the glomeruli and the absence of MCP-1 reduced both albuminuria and downregulation of nephrin and synaptopodin.CONCLUSIONSThese findings suggest that the MCP-1/CCR2 system may be relevant in the pathogenesis of proteinuria in diabetes.

D iabetic nephropathy is a major microvascular complication of diabetes, representing the leading cause of endstage renal disease in the Western world, and a major cause of morbidity and mortality in both type 1 and type 2 diabetic subjects. Clinical hallmarks of diabetic nephropathy include a progressive increase in urinary albumin excretion and a decline in glomerular filtration rate (GFR), which occur in association with an increase in blood pressure, ultimately leading to endstage renal failure. 1 These renal functional changes develop as a consequence of structural abnormalities, including glomerular basement membrane thickening, mesangial expansion with extracellular matrix accumulation, changes in glomerular epithelial cells (podocytes), including a decrease in number and/or density, podocyte foot process broadening and effacement, glomerulosclerosis, and tubulointerstitial fibrosis.Diabetic nephropathy occurs only in a minority of subjects with either type 1 or type 2 diabetes and seems to result from the interaction between genetic susceptibility and environmental insults, primarily metabolic and hemodynamic in origin. Over the last decade, the cellular and molecular mechanisms by which these insults translate to structural and functional abnormalities leading to diabetic nephropathy have been increasingly delineated. In particular, it has been determined that both metabolic and hemodynamic stimuli lead to the activation of key intracellular signaling pathways and transcription factors, thus triggering the production/release of cytokines, chemokines, and growth factors, which mediate and/or amplify renal damage.In the present review, we summarize molecular and cellular mechanisms that seem to be responsible for hypertensioninduced renal injury in diabetes, with particular focus on the role of increased intracapillary glomerular pressure, more recently discovered components of the renin-angiotensin system (RAS), such as angiotensin-converting enzyme (ACE) 2, and the increasing knowledge that has been gained emphasizing cross-talk between metabolic and hemodynamic pathways in amplifying diabetes-related renal injury. Impact of Hypertension on Diabetic NephropathyThe relationship between hypertension and poor vascular outcomes, including progression of renal disease, is unequivocal and independent of other confounding factors. The impact of hypertension on outcomes is exponential rather than linear. A sustained reduction in blood pressure seems to be currently the most important single intervention to slow progressive nephropathy in type 1 and type 2 diabetes. Long-term follow-up studies of initially normotensive diabetic subjects without renal disease demonstrate a blood pressure-dependent decline in GFR with blood pressure levels within the reference range. 2 Patients with a blood pressure corresponding to Ͻ130/80 mm Hg rarely develop microalbuminuria and show an annual decline in GFR close to the age-matched normal population. Diabetic patients with a blood pressure between 130/80 and 140/90 mm Hg have ...

Combined with their independent anti-atherosclerotic actions, and their important effects on dyslipidaemia and insulin resistance, PPAR agonists may be useful for the prevention of diabetic complications, including kidney disease, even in type 1 diabetes.