Effect Of Fluid Shear Stress-induced Expression Of CX3CR1 In Human Vascular Endothelial Cells

Objective:Shear stress(the frictional force generated by the blood flow,and wall strain)have long been known to play a role in vascular development and in maintaining vascularhomeostasis.CX3C R1isauniquereceptorforchemokine Fractalkine(FKN,CX3CL1),also is a GPCR,can induced the adhesion and chemota xis characteristics of Fractalkine.The aim of this study is to investigate the expression of CX3CR1 in human vascular endothelial cells through loading different intensity and time of flow shear stress.Methods:1.EA.hy926 cell lines as experimental object.Using M199complete medium cultured cells.Observed cell morphology under optical and electron microscope.Through immunohistochemical and cell growth curve to identify cell lines;2.Loaded different intensity(0,2.37,4.14,7.11,9.47,14.21,17.76 dyn/cm~2)shear stress.By real-time fluorescent quantitative PCR method to detect cells C X3CR1 mRN A expression,Western blotting to detect CX3C R1 protein expression in EA.hy926 cells;cell immunofluorescence techniqu positioning the protein expression of CX3CR1,determine the optimum expression intensity.3.Loaded different time(0,1,2,4,6,8,10 h)shear stress(4.14 dyn/cm~2),detected the mRN A and protein expression of CX3C R1 in cells,respectively.Cell immunofluorescence to locate CX3CR1 expression.Results:1.Cell lines growth can be covered with single layer in 2~3 d,a spindle cells,vortex pattern growth,extend the visible lumen after sample structure,can be grown into stratified by observed with optical microscopes.Found that the transverse and longitudinal cutting Webel-Palade corpuscle by the transmission electron microscopy.Observed cytoplasm of tan particles under immunohistochemical.2.Different shear stress intensity with endothelial cells,qRT-PCR,Western-blotting test are shown in 4.14 dyn/cm~2,CX3CR1expression quantity highest,and the rest of the group compared between group statistica lly difference(P<0.05).3.Different shear stress time with endothelial cells,qRT-PCR tests revealed in 2 h,C X3CR1 mRNA highest,Western blotting-tests revealed in 4 h protein expression level the highest.Conclusion:Different intensity and time of fluid shear stress can regulate the expression of CX3C R1.Therefore,CX3C R1 for further s tudy on endothelial cell mechanics,chemical coupling effect of important signal molecule mechanism provides the basis.