The Change Of TLR4 Expression In The Oxidation-damaged VECs Regulated By DFMG And Its Effects On The Proliferation And Migration Of VSMCs

Objective :To observe whether the change of TLR4 expression in the oxidation-damaged vascular endothelial cells(VECs) regulated by 7-Difluoromethyl-5,4’-dimethoxygenistein(DFMG) can influence the proliferation and migration of vascular smooth muscle cells(VSMCs).Method:(1)The VECs-VSMCs coculture model was established, and LPC was used for inducing the injury of VECs. CCK-8 assay and Transwell-migration method was used to determine the proliferation and migration of VSMCs.ELISA was used to analyze the amount of interleukin 6(IL-6) and human tumor necrosis factor-α(TNF-α) in the VECs supernatant, and Western blot and Quantitative real-time PCR was used to detect TLR4 protein and gene expression of VECs in the injured VECs-VSMCs coculture model.(2)VECs were pre-incubated with DFMG at different concentrations and different times, and CCK-8 assay was used again to determine the proliferation of VSMCs in the injured VECs-VSMCs coculture model. Then,VECs is processed by DFMG, TLR4 overexpression or silencing, the proliferation and migration of VSMCs in the injured VECs-VSMCs coculture model was analysed by CCK-8 assay and Transwell-migration method.Results:(1) LPC-induced injured VECs caused the proliferation and migration of VSMCs.(2)The amount of IL-6 and TNF-α in the VECs supernatant increased with the LPC concentration in the injured VECs-VSMCs coculture model, and the30μM group became the model of the following experiment.(3)The injured VECs-VSMCs coculture group had a higher TLR4 protein and gene expression in VECs than the normal VECs-VSMCs coculture group.(4)The proliferation of VSMCs in the injured VECs-VSMCs coculture model was influenced by DFMG on VECs in a time- and concentration- dependent manner, and 1μM and 12 h was selected as the best treatment concentration and time of DFMG.(5)TLR4 over expression has a similar role with LPC, which can promote the proliferation and migration of VSMCs in co-culture model, while DFMG and TLR4 silencing have similar effects, which can inhibit the proliferation and migration of VSMCs in co-culture model.Conclusion:(1) LPC-induced injured VECs caused the proliferation and migration of VSMCs.(2)The change of TLR4 expression in the injured VECs regulated by DFMG might influence the proliferation and migration of VSMCs.