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Inducible overexpression of sFlt-1 in podocytes ameliorates glomerulopathy in diabetic mice.

Ku CH, White KE, Dei Cas A, Hayward A, Webster Z, Bilous R, Marshall S, Viberti G, Gnudi L - Diabetes (2008)

Bottom Line: Urine free VEGF-A was decreased by 50%, and cortex VEGF-A expression was upregulated by 30% (P < 0.04).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).Podocyte-specific sFlt-1 overexpression ameliorates diabetic glomerular injury, implicating VEGF-A in the pathogenesis of this complication.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Division, King's College London School of Medicine, Guy's Hospital, King's College London, London, UK.

ABSTRACT

Objective: Podocyte-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 methods: We 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.

Results: Nondiabetic 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.

Conclusions: Podocyte-specific sFlt-1 overexpression ameliorates diabetic glomerular injury, implicating VEGF-A in the pathogenesis of this complication.

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Renal cortex nephrin and TGFβ1 expression in podocyte sFlt-1 overexpressing nondiabetic control and diabetic mice after 10 weeks of treatment with VEH or DOX. Renal cortex nephrin (A) and TGFβ1 (B) expression were upregulated, respectively, by ∼4- and 1.5-fold in VEH-treated diabetic vs. VEH-treated nondiabetic control mice (*P < 0.007 for c-VEH vs. d-VEH). DOX administration did not affect nephrin expression in either diabetic or nondiabetic control mice. Conversely, DOX administration was paralleled by downregulation of TGFβ1 expression levels in diabetic mice (B) (#P = 0.001 for d-VEH vs. d-DOX). Representative Western immunoblotting and quantitation of multiple experiments expressed as % change over nondiabetic VEH-treated mice are shown for both nephrin and TGFβ1 (n = 4–8/group).
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f6: Renal cortex nephrin and TGFβ1 expression in podocyte sFlt-1 overexpressing nondiabetic control and diabetic mice after 10 weeks of treatment with VEH or DOX. Renal cortex nephrin (A) and TGFβ1 (B) expression were upregulated, respectively, by ∼4- and 1.5-fold in VEH-treated diabetic vs. VEH-treated nondiabetic control mice (*P < 0.007 for c-VEH vs. d-VEH). DOX administration did not affect nephrin expression in either diabetic or nondiabetic control mice. Conversely, DOX administration was paralleled by downregulation of TGFβ1 expression levels in diabetic mice (B) (#P = 0.001 for d-VEH vs. d-DOX). Representative Western immunoblotting and quantitation of multiple experiments expressed as % change over nondiabetic VEH-treated mice are shown for both nephrin and TGFβ1 (n = 4–8/group).

Mentions: Analysis of nephrin protein expression in total renal cortex protein lysate resulted in two bands at 196 and 200 kDa representing different degrees of nephrin glycosylation (Fig. 6A) (22). Diabetic VEH-administered animals had a higher level of nephrin expression than nondiabetic control VEH-administered mice (P = 0.002). Treatment with DOX did not alter the expression of nephrin in the diabetic and the nondiabetic control mice.


Inducible overexpression of sFlt-1 in podocytes ameliorates glomerulopathy in diabetic mice.

Ku CH, White KE, Dei Cas A, Hayward A, Webster Z, Bilous R, Marshall S, Viberti G, Gnudi L - Diabetes (2008)

Renal cortex nephrin and TGFβ1 expression in podocyte sFlt-1 overexpressing nondiabetic control and diabetic mice after 10 weeks of treatment with VEH or DOX. Renal cortex nephrin (A) and TGFβ1 (B) expression were upregulated, respectively, by ∼4- and 1.5-fold in VEH-treated diabetic vs. VEH-treated nondiabetic control mice (*P < 0.007 for c-VEH vs. d-VEH). DOX administration did not affect nephrin expression in either diabetic or nondiabetic control mice. Conversely, DOX administration was paralleled by downregulation of TGFβ1 expression levels in diabetic mice (B) (#P = 0.001 for d-VEH vs. d-DOX). Representative Western immunoblotting and quantitation of multiple experiments expressed as % change over nondiabetic VEH-treated mice are shown for both nephrin and TGFβ1 (n = 4–8/group).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2551695&req=5

f6: Renal cortex nephrin and TGFβ1 expression in podocyte sFlt-1 overexpressing nondiabetic control and diabetic mice after 10 weeks of treatment with VEH or DOX. Renal cortex nephrin (A) and TGFβ1 (B) expression were upregulated, respectively, by ∼4- and 1.5-fold in VEH-treated diabetic vs. VEH-treated nondiabetic control mice (*P < 0.007 for c-VEH vs. d-VEH). DOX administration did not affect nephrin expression in either diabetic or nondiabetic control mice. Conversely, DOX administration was paralleled by downregulation of TGFβ1 expression levels in diabetic mice (B) (#P = 0.001 for d-VEH vs. d-DOX). Representative Western immunoblotting and quantitation of multiple experiments expressed as % change over nondiabetic VEH-treated mice are shown for both nephrin and TGFβ1 (n = 4–8/group).
Mentions: Analysis of nephrin protein expression in total renal cortex protein lysate resulted in two bands at 196 and 200 kDa representing different degrees of nephrin glycosylation (Fig. 6A) (22). Diabetic VEH-administered animals had a higher level of nephrin expression than nondiabetic control VEH-administered mice (P = 0.002). Treatment with DOX did not alter the expression of nephrin in the diabetic and the nondiabetic control mice.

Bottom Line: Urine free VEGF-A was decreased by 50%, and cortex VEGF-A expression was upregulated by 30% (P < 0.04).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).Podocyte-specific sFlt-1 overexpression ameliorates diabetic glomerular injury, implicating VEGF-A in the pathogenesis of this complication.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Division, King's College London School of Medicine, Guy's Hospital, King's College London, London, UK.

ABSTRACT

Objective: Podocyte-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 methods: We 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.

Results: Nondiabetic 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.

Conclusions: Podocyte-specific sFlt-1 overexpression ameliorates diabetic glomerular injury, implicating VEGF-A in the pathogenesis of this complication.

Show MeSH
Related in: MedlinePlus