Effect Of Astrocyte-derived Vegf Under Conditions Of Hypoxia Induction On Brain Microvascular Endothelial Cells

ObjectiveA co-culture model of brain microvascular endothelial cells(BMEC)and astrocytes (Ast) was established to study the effect of astrocyte-derived Vascular endothelial growth factor (VEGF) on BMEC.Methods(1) Astrocytes and Brain microvascular endothelial cells (BMEC) were isolated and purified from Wistar rats cerebral cortex and were identified by the expression of glial fibrillary acidic protein(GFAP) andⅧfactor related antigen respectively.(2) The purified Ast was was developed in hypoxia chamber for 30 minutes and then the culture was returned to normoxia conditions to develop. The expression level of VEGF mRNA in Ast was detected with semi-quantitative RT-PCR and compared with the control culture.(3) To build Co-culture model between Ast and BMEC through Transwell. The effect of astrocyte-derived VEGF on migration and LDH leakage rate of BMEC was detected.Result(1) The purity of cultured Ast and BMEC was 95%,90% respectively and satisfied our needs.(2) The expression of VEGF was at a low level under normoxia conditions in Ast(3) Up-regulation of VEGF was occur during hypoxia and reoxgyenization and the expression level raised to the highest at 8h after normoxia and was significantly improved compared with control group(P<0.01)(4) Astrocyte-derived VEGF could stimulate migration and LDH leakage of BMEC, administration of 4μg/ml sFlt could obviously inhibit this process(P<0.01).Conclusion Chronic ischemia cerebrovascular disease has been associated with changes in neurovascular behavior, mediated, in part, by induction of vascular endothelial growth factor (VEGF).Our study demonstrate that hypoxia can obviously up-regulate VEGF in cerebral astrocytes. Due to the special anatomical relation among astrocytes,neurons and brain microvascular, on the one hand, VEGF by Ast binds to the receptors in vascular endothelial cells and causes proliferation and migration of vascular endothelial cells and induces angiogenesis, reducing the degeneration of neurons. On the other hand, VEGF could be released to the space of cells and play as a neurotrophic factor role on neurons directly. At the same time, we notice that VEGF by Ast could improve brain microvascular permeablity (tested by LDH leakage), it will induce cerebral edema.These results clearly indicate that VEGF could play a critical role in ischemia cerebrovascular disease...