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


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.
Ku
Ching-Hsin
CH
Cardiovascular Division, King's College London School of Medicine, Guy's Hospital, King's College London, London, UK.
White
Kathryn E
KE
Dei Cas
Alessandra
A
Hayward
Anthea
A
Webster
Zoe
Z
Bilous
Rudy
R
Marshall
Sally
S
Viberti
Giancarlo
G
Gnudi
Luigi
L
D000818:Animals; D015153:Blotting, Western; D048909:Diabetes Complications / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D003920:Diabetes Mellitus / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D003928:Diabetic Nephropathies / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D004318:Doxycycline / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007678:Kidney Glomerulus / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D008854:Microscopy, Electron; D050199:Podocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
D008565:Membrane Proteins; D042461:Vascular Endothelial Growth Factor A; C113267:nephrin; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D004318:Doxycycline