Low Shear Stress Induced Endothelial Cell Autophagy And Related Gene Expression

Background and PurposeWith the improvement of Chinese people’s living standard,the diet structure has also changed accordingly,leading to the incidence of coronary heart disease year by year.Therefore,it is particularly important to further explore the pathogenesis of CHD and provide new targets for clinical diagnosis and treatment.Atherosclerosis caused by blood sugar,blood lipids,blood pressure and other factors is the root cause of coronary heart disease.Under the action of these systemic factors,atherosclerosis should be evenly distributed in the coronary artery trees.However,in the clinical diagnosis and treatment process,we found that atherosclerosis mostly occurs in the vascular curvature and bifurcation sites,which are closely related to the shear force generated by blood flow.Studies have shown that the shear force generated by blood flow can not only affect the migration,proliferation and apoptosis of endothelial cells,but also regulate the transcription of many genes and the expression of proteins,thus regulating the occurrence and development of atherosclerosis.Autophagy is a process in which eukaryotic cells engulf their own organelles or cytoplasmic proteins,encapsulate into vesicles,and then fuse with lysosomes to form autophagosomes so as to degrade their inclusions.Previous studies have shown that autophagy occurs in endothelial cells,smooth muscle cells and macrophages in the process of atherosclerosis.At present,there is still no definite conclusion on how shear force induces autophagy in endothelial cells.This project aims to study the changes of autophagy protein and gene expression in endothelial cells under the effect of low shear force.MethodsHuman umbilical Vein Cells(HUVECs)were selected as the research object.The state of low shear force was simulated in vitro under the effect of parallel plate fluid cavity system to study the changes of autophagy and m RNA of endothelial cells under the effect of low shear force.The specific process is as follows:1.Changes in the expression of autophagy related protein LC3 under low shear forceThe morphological changes of HUVECs under low shear stress at 0h,1h,2h,4h and 6h were observed by microscope.The protein levels of LC3 I and LC3 II were also verified by Western blot.2.Transcriptomic sequencing was performed to screen the m RNA changes of endothelial cells under the effect of low shear forceAt 0 and 4 hours after HUVECs was loaded with 1.5dyne/cm2 low shear force,Trizol extracted the total RNA of the cells for transcriptomic sequencing,m RNA differential expression analysis,and GO and KEGG enrichment analysis.Result1.Low shear force can induce autophagy of endothelial cellsWestern Blot results showed that compared with HUVECs without shear force,autophagy-related protein LC3 II levels increased after 1h,2h,4h and 6h low shear forces.LC3 I levels decreased,and protein expression levels showed the greatest difference at 4 h.2.Transcriptome sequencing resultsCompared with the control group,109 genes were differentially expressed after 4hours of low shear force,among which 30 genes were up-regulated and 79 genes were down-regulated.The enrichment analysis showed that the major pathways of differential expression genes were: the oxidative phosporylation pathway and Parkinson’s disease Pathway.Conclusion:1.Low shear force can regulate autophagy of endothelial cells.2.Under the effect of low shear force,differential genes were mainly concentrated in the Parkinson’s disease pathway and the oxidative phosporylation pathway,both of which are important pathways of Oxidative stress and mitophagy.It is speculated that low shear force may be involved in the development of atherosclerosis through mitophagy.