癌胚抗原相关黏附分子1对CVB3感染后CAR表达及心肌损伤的影响

Effect of CC1 on expression of CAR and myocardial injury after infection of CVB3

目的探讨癌胚抗原相关黏附分子1(CEACAM1或CC1)对柯萨奇病毒(CVB3)感染后柯萨奇病毒-腺病毒受体(CAR)表达和继发心肌损伤的影响。方法构建过表达小鼠CC1重组病毒,包装重组慢病毒pLVX-CEACAM 1-ZsGreen-Puro(rLV-CEACAM 1)并测定慢病毒生物学滴度。分为CC1正常组、CC1过表达组、CC1正常+CVB3组和CC1过表达+CVB3组;各组使用AnnxinV-PE/7-AAD双染法检测心肌细胞凋亡率,CCK8检测细胞活性;qPCR检测CAR基因表达;Western blot检测CAR蛋白表达,ELISA检测肿瘤坏死因子α(TNF-α)及白细胞介素1β(IL-1β)水平。结果(1)CEACAM1重组载体测序提示目的基因序列连接,证明小鼠CEACAM1重组病毒载体构建成功,测定重组慢病毒滴度为1.5×10^11 TU/L。(2)CC1过表达+CVB3组较其他组心肌细胞凋亡率明显升高,心肌细胞增殖最低(P<0.05)。(3)CAR基因相对表达量在CC1过表达+CVB3组最高,而在CC1正常组最低,CAR蛋白表达也有类似结果;CC1过表达组较CC1正常组胞内CVB3相对表达量显著升高(P<0.05)。(4)CC1过表达组TNF-α、IL-1β水平较高,CVB3感染后较前明显升高。结论 CC1可能可以促进CVB3感染心肌细胞后心肌组织或细胞上CAR的表达,CAR可能是CC1调控CVB3感染心肌致心肌损伤过程的潜在作用靶点。

Aim To investigate the effect of carcinoembryonic antigen-related cell adhesion molecule 1 ( CC1 ) on the expression of coxsackie and adenovirus receptor ( CAR) and secondary myocardial injury after coxsackievirus B3 (CVB3 ) infection. Methods The overexpressed mouse CC1 recombinant virus was constructed, and the recombinant lentivirus pLVX-CEACAM 1 -ZsGreen-Puro (rLV-CEACAM 1 ) was packaged and the biological titer of lentivirus was determined. It was divided into CC1 normal cell group, CC1 overexpression group, CC1 normal +CVB3 group and CC1 overexpression+ CVB3 group. The apoptosis rate of cardiomyocytes was detected by AnnxinV-PE/ 7-AAD double staining in each group, and cell activity was detected by CCK8. The expression of CAR gene was detected by qPCR. The expression of CAR protein was detected by Western blot, tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL-1β) were measured by ELISA. Results ( 1) Recombinant vector sequencing CEACAM1 showed a gene sequence connection, which proved mice CEACAM1 recombinant virus vector was built, determination of recombinant lentivirus was 1.5× 10^11 TU/L.(2 ) Apoptosis rate of cardiac myocytes in CC1 overexpression+CVB3 group was significantly higher than that in other groups, and the proliferation rate of cardiac myocytes was the lowest ( P<0. 05 ).( 3 ) Relative expression of CAR gene was the highest in CC1 overexpression+CVB3 group, and the lowest in CC1 normal group. Relative expression of CVB3 was significantly higher in CC1 overexpression group than in CC1 normal group (P<0. 05 ).(4) Level of TNF-α,IL-1β were higher in CC1 overexpression group, which increased significantly after CVB3 infection,Conclusion CC1 may promote the expression of CAR in cardiac tissue or cell after CVB3 infected cardiac myocytes,CAR might be a potential target for CC1 to regulate the process of cardiac injury caused by CVB3 infection.