The Effect Of Medical Serum Of Youguiwan On VEGF/PI3K/Akt Signaling Pathway In Rat Retinal Microvascular Endothelial Cells In Vitro Under High Glucose And Hypoxia Conditions

Objective: By the establishment of SD rat retinal microvascular endothelial cells(RRMECs)model in vitro high glucose and hypoxia condition,observing the effect of medical serum of Yougui Wan on the proliferation and migration to model cells as well as on the VEGF/ PI3-K/Akt signaling pathway,and to preliminary explore of the target and molecular mechanisms for Yougui Wan in prevention and treatment of PDR,which will provide new ideas and scientific basis for the prevention of PDR.Method:The high glucose and hypoxia model of SD rat retinal microvascular endothelial cells in vitro was established successfully with 33mmol/L glucose and 100umol/L Co Cl2 simultaneously cultured for 4 days,to simulate the proliferative diabetic retinopathy(PDR)pathological changes,and use it as a research object.blank control group,high glucose group,model group and high,middle and low dose group of medicine serum were set up according to different culture conditions.observing the effects of different dose group of medicine serum of Yougui Wan on proliferation and migration in RRMECs under high glucose and hypoxia conditions by MTT and Transwell method.By RT-PCR method and Western-blot method to detect the effects of each dose group of medicine serum on the gene and protein expression levels of VEGF,VEGFR-2,PI3 K,Akt in model cells..Result: 1.The high glucose and hypoxia model of SD rat retinal microvascular endothelial cells in vitro was established successfully with 33mmol/L glucose and 100umol/L Co Cl2 simultaneously cultured for 4 days,which can be used to simulate the pathological state of PDR.2.The effect of medical serum of Yougui Wan on proliferation in RRMECs under high glucose and hypoxia conditions :On the 1st day,the middle dose group medical serum of You gui Wan have a protective effect on cell proliferation induced by high glucose and hypoxia(p<0.003);On the 2nd day,there was no difference in the proliferation of the cells between the bl ank group,model group and the each dose group of medicine serum(p>0.003);On the 4th day,the middle and high dose groupmedical serum of Yougui Wan inhibited the cell proliferation i nduced by high glucose and hypoxia(p<0.003),The effect on the migration of RRMECs:on the 2nd day,the middle and high dose group medical serum of Yougui Wan could reduce the inhibitory effect of high glucose and hypoxia o n cell migration(p<0.003);On the 4th day,the low,middle and high dose group medical serum of Yougui Wan could inhibit the promotion of cell migration by high glucose and hypoxia(p<0.003).3.The effect of medical serum of Yougui Wan on the gene expression levels of VEGFm RNA,VEGFR-2m RNA,PI3 Km RNA,Aktm RNA in RRMECs under high glucose and hypoxia conditions :The high glucose and high glucose hypoxia can cause the expression of VEGFm RNA,VEGFR-2m RNA,PI3 Km RNA and Aktm RNA in RRMECs increased(p<0.003),while the low,middle and high dose medical serum of Yougui Wan could reduce those high exp ression(p<0.003),and the high dose group on VEGFR-2m RNA,PI3 Km RNA and Akt m RNA overexpression significantly inhibited by high glucose and hypoxia(p<0.003).The effect of the protein expression levels of VEGF,VEGFR-2,PI3 K,Akt in RRMECs :The conditions of high glucose and hypoxia could induce the enhanced expression of VEGF,VEGFR-2,PI3 K and Akt in RRMECs(p<0.003),However,The middle and high dose medical serum of Yougui Wan could reduce those high expression(p <0.003).Conclusion:The medical serum of Yougui Wan can inhibit the proliferation and migration of RRMECs under high glucose and hypoxia conditions by reducing the gene expression of VEGFm RNA,VEGFR-2m RNA,PI3 Km RNA,Aktm RNA and VEGF,VEGFR-2,PI3 K,Akt protein expression,thus inhibiting retinal neovascularization,ultimately achieve the role of prevention and treatment of PDR.