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Expression profiles of podocytes exposed to high glucose reveal new insights into early diabetic glomerulopathy.

ABSTRACT: Podocyte injury has been suggested to have a pivotal role in the pathogenesis of diabetic glomerulopathy. To glean insights into molecular mechanisms underlying diabetic podocyte injury, we generated temporal global gene transcript profiles of podocytes exposed to high glucose for a time interval of 1 or 2 weeks using microarrays. A number of genes were altered at both 1 and 2 weeks of glucose exposure compared with controls grown under normal glucose. These included extracellular matrix modulators, cell cycle regulators, extracellular transduction signals and membrane transport proteins. Novel genes that were altered at both 1 and 2 weeks of high-glucose exposure included neutrophil gelatinase-associated lipocalin (LCN2 or NGAL, decreased by 3.2-fold at 1 week and by 7.2-fold at 2 weeks), endothelial lipase (EL, increased by 3.6-fold at 1 week and 3.9-fold at 2 week) and UDP-glucuronosyltransferase 8 (UGT8, increased by 3.9-fold at 1 week and 5.0-fold at 2 weeks). To further validate these results, we used real-time PCR from independent podocyte cultures, immunohistochemistry in renal biopsies and immunoblotting on urine specimens from diabetic patients. A more detailed time course revealed changes in LCN2 and EL mRNA levels as early as 6 hours and in UGT8 mRNA level at 12 hours post high-glucose exposure. EL immunohistochemistry on human tissues showed markedly increased expression in glomeruli, and immunoblotting readily detected EL in a subset of urine samples from diabetic nephropathy patients. In addition to previously implicated roles of these genes in ischemic or oxidative stress, our results further support their importance in hyperglycemic podocyte stress and possibly diabetic glomerulopathy pathogenesis and diagnosis in humans.

SUBMITTER: Jain S

PROVIDER: S-EPMC3068212 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Expression profiles of podocytes exposed to high glucose reveal new insights into early diabetic glomerulopathy.

Jain Sanjay S De Petris Laura L Hoshi Masato M Akilesh Shreeram S Chatterjee Rajshekhar R Liapis Helen H

Laboratory investigation; a journal of technical methods and pathology 20101122 4

Podocyte injury has been suggested to have a pivotal role in the pathogenesis of diabetic glomerulopathy. To glean insights into molecular mechanisms underlying diabetic podocyte injury, we generated temporal global gene transcript profiles of podocytes exposed to high glucose for a time interval of 1 or 2 weeks using microarrays. A number of genes were altered at both 1 and 2 weeks of glucose exposure compared with controls grown under normal glucose. These included extracellular matrix modulat ...[more]

PMID: 21102505

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Project description:ObjectivePodocyte-specific, doxycycline (DOX)-inducible overexpression of soluble vascular endothelial growth factor (VEGF) receptor-1 (sFlt-1) in adult mice was used to investigate the role of the VEGF-A/VEGF receptor (VEGFR) system in diabetic glomerulopathy.Research design and methodsWe studied nondiabetic and diabetic transgenic mice and wild-type controls treated with vehicle (VEH) or DOX for 10 weeks. Glycemia was measured by a glucose-oxidase method and blood pressure by a noninvasive technique. sFlt-1, VEGF-A, VEGFR2, and nephrin protein expression in renal cortex were determined by Western immunoblotting; urine sFlt-1, urine free VEGF-A, and albuminuria by enzyme-linked immunosorbent assay; glomerular ultrastructure by electron microscopy; and VEGFR1 and VEGFR2 cellular localization with Immunogold techniques.ResultsNondiabetic DOX-treated transgenic mice showed a twofold increase in cortex sFlt-1 expression and a fourfold increase in sFlt-1 urine excretion (P < 0.001). Urine free VEGF-A was decreased by 50%, and cortex VEGF-A expression was upregulated by 30% (P < 0.04). VEGFR2 expression was unchanged, whereas its activation was reduced in DOX-treated transgenic mice (P < 0.02). Albuminuria and glomerular morphology were similar among groups. DOX-treated transgenic diabetic mice showed a 60% increase in 24-h urine sFlt-1 excretion and an approximately 70% decrease in urine free VEGF-A compared with VEH-treated diabetic mice (P < 0.04) and had lower urine albumin excretion at 10 weeks than VEH-treated diabetic (d) mice: d-VEH vs. d-DOX, geometric mean (95% CI), 117.5 (69-199) vs. 43 (26.8-69) mug/24 h (P = 0.003). Diabetes-induced mesangial expansion, glomerular basement membrane thickening, podocyte foot-process fusion, and transforming growth factor-beta1 expression were ameliorated in DOX-treated diabetic animals (P < 0.05). Diabetes-induced VEGF-A and nephrin expression were not affected in DOX-treated mice.ConclusionsPodocyte-specific sFlt-1 overexpression ameliorates diabetic glomerular injury, implicating VEGF-A in the pathogenesis of this complication.

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Expression profiles of podocytes exposed to high glucose reveal new insights into early diabetic glomerulopathy.

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Expression profiles of podocytes exposed to high glucose reveal new insights into early diabetic glomerulopathy.

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