蛋白激酶C在脂多糖致心肌细胞骨架结蛋白损伤中的作用  

Role of protein kinase C in process of influence of lipopolysaccharide on myocardial desmin

作　　者：薛翔[1] 肖利民[1] 张青[1] 徐小元[1] 金春华[1] 

机构地区：[1]南方医科大学基础医学院医学基础实验教学中心,广州510515

出　　处：《重庆医学》2007年第24期2519-2521,I0025,共4页Chongqing medicine

基　　金：国家自然科学基金(30570707);广东省自然科学基金(05004721)

摘　　要：目的观察脂多糖(lipopolysaccharide,LPS)对心肌细胞结蛋白(desmin)结构形态和数量的影响,并探讨蛋白激酶C (protein kinase C,PKC)信号通路在LPS作用过程中起的作用。方法体外培养Wistar乳鼠心肌细胞,随机分为4组,分别是正常对照组、LPS组(100ng/ml LPS)、Calphostin C处理组(40mmol/L Calphostin C+100ng/ml LPS)和12-o-十四烷酰佛波醋酸酯- 13(12-o-tetradecanoylphorbol-13-acetate,TPA)处理组(100mmol/L TPA),每组分别处理6h和8h。使用免疫荧光技术标记心肌结蛋白,在共聚焦显微镜下观察细胞内结蛋白的结构、分布并计算细胞内结蛋白荧光强度。结果正常对照组结蛋白均匀分布于除细胞核的胞质内,并在局部有明显的聚集增加,荧光强度为(45.57±2.96);LPS 6h组结蛋白分布和荧光强度与正常组相比差异无统计学意义,LPS 8h组结蛋白在细胞内分布与正常相比差异有统计学意义,且荧光强度显著减弱(31.33±2.23,P<0.01);Calphostin C 6h和8h处理组的结蛋白分布以及荧光强度与正常组接近;TPA处理6h即可造成结蛋白荧光显著减弱(33.30±1.15,P<0.01),且结蛋白有异常的纤维样聚集。结论LPS可以影响心肌细胞内结蛋白的结构、分布和数量,并存在一定的时效关系,而Calphostin C可抑制LPS造成的心肌细胞骨架蛋白结蛋白的改变,而TPA则可在较短的时间内造成与LPS相似的结果,提示PKC可能参与了LPS对结蛋白破坏的过程。Objective To observe the influence of lipopolysaccharide （LPS） on desmin of myocardial cells in rats and the role of protein kinase C in this process. Methods Wistar rat cardiac myocytes （2d old） were isolated and cultured in vitro, cells were divided into control group （with DMEM for 6h and 8h）, LPS group （with 100ng/ml LPS stimulation for 6h and 8h）, Calphostin C group （with 40mmol/L Calphostin C＋ 100ng/ml LPS for 6h and 8h）, and 12-o-tetradecanoylphorbol-lS-acetate （TPA） group （with 100mmol/L TPA for 6h and 8h）. Myocardial desmin were observed by immunoflurescent staining with confocal microscope and then the density of desmin was analyed. Results In control group, desmin was localized in the cytoplasm and accumulated in somewhere. After LPS simulation for 6h, desmin in myocytes did not show much difference compared with control group. After LPS simulation for 8h, desmin was localized disorderly and the fluorescent density of desmin decreased significantly （31.33±2.23, P〈0.01） compared with control group （45.57±2.96）. In Caiphostin C group, the localization and fluorescent density of desmin in myocytes were similar as control group. In TPA 6h group, the fluorescent density of desmin in myocytes decreased significantly （33.30± 1.15, P〈 0. 01） compared with control. Conclusion LPS stimulation can affect the distribution and concentration of desmin in myocytes, Calphostin C can inhibit these effects, TPA show similar effects as LPS. These results indicate that PKC may participate in the LPS signaling process of influence of LPS on myocardial desmin.

关 键 词：结蛋白 心肌 细胞骨架 蛋白激酶C 脂多糖 

分 类 号：R363[医药卫生—病理学]