LNK对人巨细胞病毒感染平滑肌细胞血管新生微环境的影响

Effect of LNK on angiogenesis microenvironment of smooth muscle cells infected with human cytomegalovirus

目的研究淋巴细胞连接蛋白(LNK)对人巨细胞病毒(HCMV)感染平滑肌细胞(VSMC)血管新生微环境的影响及作用机制.方法经过细胞培养及细胞转染将细胞分为正常组、对照组、空载组和LNK组.进行细胞增殖实验,酶联免疫吸附法检测白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、干扰素γ(IFN-γ)水平,化学发光法检测促血管生成素-1(Ang-1)、促血管生成素-2(Ang-2)及血管内皮生长因子-A(VEGF-A)水平,Western blot法检测血清转化生长因子β1(TGFβ1)、转化生长因子β1受体(TGFβ1R1)相对表达量.结果与正常组比较,对照组、空载组、LNK组OD值均升高(P<0.05).与对照组比较,空载组、LNK组OD值较低(P<0.05).与空载组相比,LNK组OD值较低(P<0.05).与正常组细胞比较,对照组、空载组、LNK组细胞IL-6、TNF-α、IFN-γ水平均升高(P<0.05),与对照组、空载组细胞比较,LNK组细胞IL-6、TNF-α、IFN-γ水平均降低(P<0.05).与正常组比较,对照组、空载组、LNK组Ang-1降低,Ang-2、VEGF-A升高(P<0.05),与对照组比较,空载组、LNK组Ang-1升高,Ang-2、VEGF-A降低(P<0.05),与空载组比较,LNK组Ang-1升高,Ang-2、VEGF-A降低(P<0.05).与正常组比较,对照组、空载组、LNK组TGFβ1及TGFβR1表达升高(P<0.05),与对照组比较,空载组TGFβ1、TGFβR1表达升高,LNK组TGFβ1、TGFβR1表达降低(P<0.05),与空载组比较,LNK组TGFβ1、TGFβR1表达降低(P<0.05).结论LNK转染对HCMV感染VSMC有一定干预作用,能够降低炎症反应,通过调控Ang-2、VEGF-A、TGFβ1、TGFβ1R1蛋白水平,改善血管新生微环境.

OBJECTIVE To observe the effect of lymphocyte connexin(LNK)on angiogenic microenvironment of vascular smooth muscle cells(VSMC)infected with human cytomegalovirus(HCMV)and analyze the action mechanisms.METHODS The cells were divided into the normal group,the control group,the no-load group and the LNK group through cell culture and cell transfection.Cell proliferation assays were performed,the levels of interleukin-6(IL-6),tumor necrosis factor-α(TNF-α)and interferon-γ(IFN-γ)were detected by using enzyme-linked immunosorbent assay,the levels of angiopoietin-1(Ang-1),angiopoietin-2(Ang-2)and vascular endothelial growth factor-A(VEGF-A)were detected by means of chemiluminescence,and the relative expression levels of serum transforming growth factorβ1(TGFβ1)and transforming growth factorβ1 receptor(TGFβ1 R1)were detected by Western blot.RESULTS The OD values of the control group,the no-load group and the LNK group were significantly higher than those of the normal group(P<0.05).The OD values of the no-load group and the LNK group were significantly lower than those of the control group(P<0.05).The OD value of the LNK group was significantly lower than that of the no-load group(P<0.05).The levels of IL-6,TNF-αand IFN-γof the control group,the no-load group and the LNK group were significantly higher than those of the normal group(P<0.05),the levels of IL-6,TNF-αand IFN-γof the LNK group were significantly lower than those of the control group and the no-load group(P<0.05).The Ang-1 level of the control group,the no-load group and the LNK group was significantly lower than that of the normal group,the levels of Ang-2 and VEGF-A were significantly elevated(P<0.05);the Ang-1 level of the no-load group and the LNK group was significantly higher than that of the control group,and the levels of Ang-2 and VEGF-A of the no-load group and the LNK group were significantly lower than those of the control group(P<0.05);the Ang-1 level of the LNK group was significantly higher than that of the no-load group,and the levels of Ang-2 and VEGF-A of the LNK group were significantly lower than those of the no-load group(P<0.05).The expression levels of TGFβ1 and TGFβR1 of the control group,the no-load group and the LNK group were significantly higher than those of the normal group(P<0.05);the levels of TGFβ1 and TGFβR1 of the no-load group were significantly higher than those of the control group,and the levels of TGFβ1 and TGFβR1 of the LNK group were significantly lower than those of the control group(P<0.05);the levels of TGFβ1 and TGFβR1 of the LNK group were significantly lower than those of the no-load group(P<0.05).CONCLUSION LNK transfection has certain intervention effect on VSMC infected with HCMV,and it can reduce the inflammatory response and improve the angiogenic microenvironment by regulating the levels of Ang-2,VEGF-A,TGFβ1 and TGFβ1 R1 proteins.