Vascular endothelial growth factor induces nephrogenesis and vasculogenesis

The expression of vascular endothelial growth factor (VEGF) and its recepto rs Flt-1 and Flk-1 in the rat kidney was examined during ontogeny using Nor thern blot analysis and immunocytochemistry. In prevascular embryonic kidne ys (embryonic day 14 [E14]), immunoreactive Flt-1 and Flk-1 were observed i n isolated angioblasts, whereas VEGF was not detected. Angioblasts aligned forming cords before morphologically differentiating into endothelial cells , In late fetal kidneys (E19), immunoreactive VEGF was detected in glomerul ar epithelial and tubular cells, whereas Flt-1 and Flk-1 were expressed in contiguous endothelial cells. To determine whether VEGF induces endothelial cell differentiation and vascular development in the kidney, the effect of recombinant human VEGF (5 ng/ml) was examined on rat metanephric organ cul ture, a model known to recapitulate nephrogenesis in the absence of vessels . After 6 d in culture in serum-free, defined media, metanephric kidney gro wth and morphology were assessed. DNA content was higher in VEGF-treated ex plants (1.9 +/- 0.17 mu g/kidney, n = 9) than in paired control explants (1 .4 +/- 0.10 mu g/kidney, n = 9) (P < 0.05). VEGF induced proliferation of t ubular epithelial cells, as indicated by an increased number of tubules and tubular proliferating cell nuclear antigen-containing cells. VEGF induced upregulation of Flk-1 and Flt-1 expression, as assessed by Western blot ana lysis. Developing endothelial cells were identified and localized using imm unocytochemistry and electron microscopy. Flt-1, Flk-1, and angiotensin-con verting enzyme-containing cells were detected in VEGF-treated explants, whe reas control explants were negative. These studies confirmed previous repor ts indicating that the expression of VEGF and its receptors is temporally a nd spatially associated with kidney vascularization and identified angiobla sts expressing Flt-1 and Flk-1 in prevascular embryonic kidneys. The data i ndicate that VEGF expression is downregulated in standard culture condition s and that VEGF stimulates growth of embryonic kidney explants by expanding both endothelium and epithelium, resulting in vasculogenesis and enhanced tubulogenesis. These data suggest that VEGF plays a critical role in renal development by promoting endothelial eel differentiation, capillary formati on, and proliferation of tubular epithelia.