C-反应蛋白对体外培养乳鼠心肌细胞的影响  

Effect of C-reactive protein on myocardial cells in suckling mice cultured in vitro

作　　者：刘剑南[1] 陈相健[1] 戴健[1] 马继政[1] 杨笛[1] 张寄南[1] 

机构地区：[1]南京医科大学第一附属医院心内科暨心血管病研究所,江苏省南京市210029

出　　处：《中国临床康复》2006年第21期76-78,i0004,共4页Chinese Journal of Clinical Rehabilitation

基　　金：江苏省高新技术研究(BG2003033)~~

摘　　要：目的:应用C-反应蛋白干预体外培养的乳鼠心肌细胞,观察C-反应蛋白对心肌细胞的影响。方法:实验于2005-05/10在南京医科大学第一附属医院心内科暨心血管病研究所完成。取新生1～3d的SD乳鼠,无菌操作取出心脏,去除大血管和心房后将心脏剪碎,加入适量0.125%胰蛋白酶反复消化至完全。所得消化液中加入含血清的DMEM培养基,离心后弃上清,沉淀用含0.1%Brdu、体积分数为0.15新生牛血清的DMEM培养基混悬。细胞密度稀释至6×108L-1,接种至六孔板中,放入37℃,体积分数为0.05的CO2孵育箱内培养。24h后换液除去Brdu,取第3天细胞进行研究。采用充氮气孵育4h模拟缺氧造成心肌细胞损伤(缺氧组),以未缺氧组做对照,缺氧组与未缺氧组均加用不同浓度的C-反应蛋白(0,10,55,100mg/L)进行干预,测定细胞培养上清心肌肌钙蛋白Ⅰ的含量及脂质过氧化产物丙二醛与超氧化物歧化酶的含量以观察C-反应蛋白对心肌细胞的影响,并通过免疫组织化学观察C-反应蛋白在细胞的分布情况。结果:①细胞形态及搏动率的改变:未缺氧组细胞搏动率总体随C-反应蛋白浓度的增加而增加,但在缺氧4h组,细胞搏动率在C-反应蛋白55mg/L时达到最高峰犤(132±9)次/min,P<0.05犦。②培养细胞上清肌钙蛋白Ⅰ含量的变化:在未缺氧组和缺氧4h组,肌钙蛋白Ⅰ均随C-反应蛋白浓度的增加而增高,且C-反应蛋白浓度为100mg/L时的增幅最高犤C-反应蛋白为0,10,55,100mg/L时的肌钙蛋白Ⅰ浓度:缺氧组分别为(0.789±0.066),(1.198±0.101),(1.979±0.151),(3.714±0.288)μg/L;未缺氧组分别为(0.332±0.034),(0.377±0.054),(0.527±0.057),(0.867±0.077)μg/L,P<0.05犦。③细胞上清超氧化物歧化酶、丙二醛含量变化:各组丙二醛含量的变化趋势与肌钙蛋白Ⅰ相似,随C-反应蛋白浓度的增加而增加,而超氧化物歧化酶活力则随C-反应蛋白浓度的增加而降低。④免疫组织化学结果显AIM： To observe the effect of C-reactive protein （CRP） on myocardial cells by interfering myocardial cells cultured in vitro with CRP. METHODS： The experiment was conducted in the Institute of Cardiovascular Disease,First Affiliated Hospital of Nanjing Medical University from May to October 2005. Newly born SD suckling mice （1-3 days） were selected to take out the hearts, cut off great vessels and atrium under sterile condition. The hearts were cut into little slices and repeatedly digested by 0.125 % of trypase. The digestive juice was mixed with DMEM, which containing serums and was removed off supernatant after centrifuging. The remaining was suspended with DMEM nutritive medium containing 0.1% of new bern bull serum with the volume fraction of 0.15. Cell density was diluted to 6×10^2L^-1 and inoculated into the 6-pore board culturing in the 5% of CO2 incubation at 37℃. At 24 hours after that, fluid was exchanged to exclude Brdu, and cells were studied on the 3^rd day. Cells in the hypoxia group were cultured in nitrogen gas without oxygen for 4 hours to establish myocardial cell injury and cells in the non-hypoxia group were taken as control. Both of the two groups were exposed to CRP at different concentration （0, 10, 55, 100μmg/L）. The content of cardiac troponin Ⅰ （CTnⅠ）, lipid peroxidative product malondialdehyde （MDA） and superoxide dismutase （SOD） activity in supernatant of cells were measured to observe the effects of CRP on myocardial cells, and the distribution of CRP in cells was observed by immunohistochemiscal method. RESULTS： ①Changes of cellular shape and beating rate： The beating rate of cells in non-hypoxia group totally increased with the increased concentration of CRP, whereas in the 4-hour hypoxia group, the beating rate reached to the peak when the CRP was 55 mg/L [（132±9） times per minute,P 〈 0.05].②Changes of content of CTN1 in supematant of cultured cells： In non-hypexia group and 4-hour hypoxia group, content of CTN1 increased wi

关 键 词：C-反应蛋白质 心肌 细胞 缺氧 肌钙蛋白Ⅰ 

分 类 号：R318[医药卫生—生物医学工程]