The Inhibitory Effects Of Luteolin On Vascular Smooth Muscle Cell Proliferation,Migration And Intikal Hyperplasia By Transforming Growth Factor ? Receptor 1

Research background and purpose:Previous studies have shown that the migration of vascular smooth muscle cell(VSMC)from media to the intima and their abnormal proliferation are closely related to the occurrence and development of restenosis and related diseases.Basic medical research and clinical studies have demonstrated that Buyang Huanwu Decoction(BYHWD)is an effective prescription for the treatment of coronary atherosclerotic heart disease(CHD)and restenosis.BYHWD contains Luteolin,a flavonoid from safflower that exerts various protective effects on the cardiovascular system;however,its specific mechanism of action has not yet been fully elucidated.This study aimed to explore and verify the inhibitory effects of Luteolin on VSMC proliferation,migration,and intimal hyperplasia by examining the suppression of transforming growth factor ? receptor 1(TGFBR1)through in vitro and in vivo experiments.Further,this study aimed to clarify the therapeutic basis of BYHWD,presented as scientific evidence for the clinical application of this traditional Chinese medicine formulation used in the treatment of restenosis and related diseases.Methods:In vitro experiments:Two cell lines,rat thoracic aorta vascular smooth muscle cells(A7r5)and human aortic vascular smooth muscle cell(HASMC),were treated with different concentrations of Luteolin(10,20,and 40 ?M),and the inhibitory effects of Luteolin on VSMC proliferation,migration,and TGFBR1 signaling pathway activation were investigated using MTS,EdU,Transwell,wound healing,and Western blot assays.In addition,the interaction between Luteolin and TGFBR1 was further verified using molecular docking and molecular dynamics simulation,the surface plasmon resonance(SPR),and the TGFBR1 enzyme activity assay.The role of TGFBR1 in the Luteolin-mediated inhibition of VSMC proliferation and migration was further verified using an overexpressing TGFBR1 cell model constructed by transfecting the A7r5 and HASMC cells with a TGFBR1 adenovirus(Ad-TGFBR1).In vivo experiments:HE staining and immunohistochemical methods were used to detect the inhibitory effects of Luteolin on intimal hyperplasia of the injured blood vessels and TGFBR1 signaling pathway activation using a left common carotid artery ligation mouse model.Subsequently,we verified the role of TGFBR1 in the Luteolin-mediated inhibition using a mouse model with TGFBR1 gene overexpression in the vascular tissue.Results:In vitro experiments:Luteolin could inhibit VSMC proliferation,migration,and the activation of the TGFBR1 signaling pathway in a dose-dependent manner,compared to the control group(P<0.05).The overexpression of the TGFBR1 gene could partially block the inhibitory effect of Luteolin on VSMC proliferation,migration,and TGFBR1 signaling activation(P<0.05).In vivo experiments:Luteolin could inhibit the intimal hyperplasia of injured blood vessels and the activation of the TGFBR1 signaling pathway,compared to the model group(P<0.05).The overexpression of TGFBR1 could partially block the inhibitory effects of Luteolin on the intimal hyperplasia of injured blood vessels and the activation of the TGFBR1 signaling pathway(P<0.05).In addition,molecular docking and molecular dynamics simulation,the SPR,and the TGFBR1 enzyme activity assay strongly indicated that Luteolin could directly dock onto TGFBR1,acting as a potential TGFBR1 inhibitor.Conclusions:Luteolin could inhibit VSMC proliferation,migration,and intimal hyperplasia by suppressing TGFBR1 activation and ultimately plays a therapeutic role in the treatment of restenosis and related diseases.