摘要
目的观察心肌微血管内皮细胞条件培养液对心肌细胞的影响及通心络的干预作用。方法制备3种心肌微血管内皮细胞条件培养液:正常内皮细胞条件培养液(N-CM),同型半胱氨酸诱导损伤的心肌微血管内皮细胞条件培养液(H-CM)和通心络干预的心肌微血管内皮细胞条件培养液(T-CM)。将心肌细胞随机分为4组:正常对照组、N-CM组、T-CM组及H-CM组,采用相应培养液培养。倒置显微镜观察心肌细胞形态,酶联免疫吸附法检测细胞上清液心肌肌钙蛋白T(cTnT)含量,JC-1试剂盒测定细胞中线粒体膜电位(MMP),Western blot法检测细胞色素C(Cyt C)和钙网蛋白(CRT)蛋白表达。结果与正常对照组比较,N-CM组细胞形态无明显变化,细胞上清液cTnT含量、MMP水平、CRT及Cyt C蛋白表达无明显变化(P>0.05);与N-CM组比较,H-CM组细胞皱缩,细胞边缘模糊,有大量坏死细胞漂浮,细胞上清液cTnT含量、CRT及Cyt C蛋白表达升高,MMP水平降低(P<0.01);与H-CM组比较,T-CM组细胞无明显皱缩,细胞边缘较清楚,坏死漂浮细胞减少,细胞上清液cTnT含量、CRT及Cyt C蛋白表达降低,MMP水平升高(P<0.05,P<0.01)。结论受损心肌微血管内皮细胞的条件培养液可损伤心肌细胞,其损伤机制可能与CRT介导的线粒体功能受损有关,通心络可抑制该损伤过程。
Objective To investigate the influence of conditioned medium of cardiac microvascu- lar endothelial cells on cardiomyocytes and the protective effect of Tongxinluo. Methods Three kinds of conditioned cardiac microvascular endothelial cell conditioned medium were prepared： the normal cardi- ac microvascular endothelial cell-conditioned medium （normal-conditioned medium, N-CM）, the homo- cysteine-induced microvascular endothelial cell-conditioned medium （homocysteine-conditioned medi- um ,H-CM） and the cardiac microvascular endothelial cell-conditioned medium with Tongxinluo （~Tongxin- luo-conditioned medium ,T-CM）. The cardiomyocytes were randomly divided into four groups ： the normal group, the N-CM group, the T-CM group and the H-CM group, corresponding conditioned medium were used to culture cardiomyocytes. The morphology of cardiomyocytes were observed with an inverted mi- croscope, the contents of cardiac troponin T （cTnT） in cell supernatant were detected by enzyme-linked immunosorbent assay, the mitochondrial membrane potential （MMP） of the cells were detected by JC-1reagent, and the protein expression levels of Cytochrome C （Cyt C） and calreticulin （CRT） were detec- ted by Western blot. Results Compared with the normal group, the cells morphology, the contents of cTnT in cell supernatant, the MMP, and the protein expression levels of Cyt C and CRT did not significant- ly changed in the N-CM group （P 〉0.05） . Compared with N-CM group, the cells were shrinkaged, the cell edge was fuzzy, and there were a large number of dead cells floated in conditioned medium, the con- tents of cTnT in cell supernatant was increased, and the protein expression levels of Cyt C as well as CRT were up-regulated, while the MMP was decreased in the H-CM group（P 〈0.01 ）. Compared with H- CM group, there was no obvious change in morphology, the cell edge was clear, and the dead cells floa- ted in culture medium were decreased, the contents of cTnT in cell supernatant was decreased, and th
出处
《中国中西医结合杂志》
CAS
CSCD
北大核心
2017年第11期1334-1338,共5页
Chinese Journal of Integrated Traditional and Western Medicine
基金
国家重点基础研究发展计划(No.2012CB518600)
石家庄市科学技术研究与发展计划项目(No.161462343)
关键词
通心络
心肌微血管内皮细胞
心肌细胞
共培养
线粒体
钙网蛋白
Tongxinluo
cardiac microvascular endothelial cells
cardiomyocytes
co-culture
mi- tochondrial membrane potential
calreticulin
