Research of Molecular Mechanism of Puerarin on Regulating the VEGF/Ang/Tie2 Pathway to Improve Retinal Neovascularization Under Hypoxic Conditions Based on Molecular Docking
Objective Based on molecular docking technology and vitro experimental verification,this study aims to investigate the effect of puerarin on the proliferation,migration and tube of rat retinal microvascular endothelial cells (rRMECs) under hypoxic-induced conditions induced by cobalt chloride (CoCl2),and the regulatory effect of VEGF/Ang/Tie2 pathway during retinal neovascularization (RNV) formation. Methods The CCK-8 method was used to screen out the suitable concentration of CoCl2 modeling and the concentration of puerarin intervention in rRMECs. The experiments were divided into control group,hypoxia model group,puerarin high-dose group,puerarin low-dose group. The effects of puerarin on the proliferation,migration and tube formation of rRMECs were observed by cell proliferation,scratch experiment and tubular formation experiment. At the same time,the binding potential of puerarin on RNV protein targets was evaluated by molecular docking technology,and the prediction results were verified by RT-qPCR. Results CCK-8 results showed that under hypoxic conditions,200 μmol/L CoCl2 can significantly restrain cell viability and is the best concentration. In addition,10 and 40 μmol/L puerarin was in the low-and high-dose groups. In vitro studies have found that puerarin can reduce the proliferation rate,mobility,number of duct-forming nodes and number of intersections of rRMECs under hypoxia conditions. The results of molecular docking showed that puerarin had good affinity with key proteins in the VEGF/Ang/Tie2 pathway. In addition,RT-qPCR verification results showed that puerarin could significantly reduce the expression of Ang2,HIF-1 D,VEGF-A,VE-PTP and other genes in rRMECs under hypoxia conditions,and upregulate the expression of Ang1,Tie2 and VE-cadherin genes,and had a certain dose-effect relationship. Conclusion Puerarin can inhibit the proliferation,migration and ducting of retinal microvascular endothelial cells under hypoxia conditions,and has a certain inhibitory RNV effect,and its mechanism of action may be related to the regulation of VEGF/Ang/Tie2 pathway by puerarin.
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