去血清饥饿条件下肌卫星细胞增殖及自噬蛋白LC3、Beclin1的表达  

Effects of serum-free starvation on proliferative capacity of muscle satellite cells and expression of autophagy-related proteins LC3 and Beclin1

作　　者：刘通[1] 于佳妮[2] 刘悦[1] 陈欢 邝伟川[1] 王小寅[1] 张淑静 江烨[1] 邱晓佳[1] 文希[1] Liu Tong;Yu Jiani;Liu Yue;Chen Huan;Kuang Weichuan;Wang Xiaoyin;Zhang Shujing;Jiang Ye;Qiu Xiaojia;Wen Xi(Department of Acupuncture and Rehabilitation,Guangdong Second Hospital of Traditional Chinese Medicine,Guangzhou 510095,Guangdong Province,China;Department of Rehabilitation,Guangdong Hospital of Traditional Chinese Medicine,Guangzhou 510120,Guangdong Province,China;Department of Acupuncture and Moxibustion,First Affiliated Hospital of Nanjing Medical University,Nanjing 210000,Jiangsu Province,China;School of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]广东省第二中医院针灸康复科,广东省广州市510095 [2]广东省中医院康复科,广东省广州市510120 [3]南京医科大学第一附属医院针灸科,江苏省南京市210000 [4]北京中医药大学中医学院,北京市100029

出　　处：《中国组织工程研究》2019年第11期1657-1661,共5页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金青年基金(81704179);项目负责人:刘通;国家自然科学基金青年基金(81503641);项目负责人:陈欢~~

摘　　要：背景:自噬可促进低氧环境下肌卫星细胞的存活,然而基于自噬有明显的时效性,不同时间的自噬水平与肌卫星细胞增殖能力间的关系如何未见探讨。目的:探讨不同时间去血清饥饿对大鼠肌卫星细胞增殖能力及自噬微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3,LC3)及Beclin1表达的变化。方法:原代分离培养雄性SD大鼠(广州中医药大学动物实验中心提供,体质量约120 g)多裂肌卫星细胞,取第3代细胞,培养至对数生长期后,分别设置正常血清组(体积分数10%胎牛血清)、空白血清(仅含培养基,无胎牛血清)6 h组、空白血清12 h组、空白血清24 h组,CCK-8检测各组细胞的增殖情况,并采用Western-blot法和RT-PCR法检测LC3及Beclin1蛋白及基因的表达。结果与结论:(1)细胞增殖能力:空白血清诱导12 h后细胞增殖能力较正常血清组及诱导6 h时下降(P <0.01),诱导24h后细胞增殖能力较诱导12h下降(P<0.05);(2)LC3及Beclin1蛋白及基因的表达:空白血清诱导6 h后,LC3Ⅱ/LC3Ⅰ及Beclin1蛋白及基因的表达较正常血清组明显升高(P <0.05),诱导12 h及诱导24 h后LC3Ⅱ/LC3Ⅰ蛋白表达较诱导6 h时明显升高(P <0.01),Beclin1的表达较诱导6 h时升高(P <0.05),LC3及Beclin1基因的表达较6 h均升高(P <0.05);(3)结果证实,去血清饥饿可以诱导肌卫星细胞发生自噬,自噬在去血清早期(12 h内)可保护细胞的增殖能力,12 h后细胞可出现过度自噬,不利于细胞的增殖。BACKGROUND:Autophagy can promote the survival of muscle satellite cells in hypoxic environment.However,based on the obvious time-dependent nature of autophagy,the relationship between the level of autophagy and the proliferation ability of muscle satellite cells at different time has not been explored.OBJECTIVE:To observe the effect of different time of serum-free starvation on the proliferative capacity and expression of autophagy-related proteins,microtubule-associated protein 1 light chain 3(LC3)and Beclin1,in rat muscle satellite cells.METHODS:Primary multifidus muscle satellite cells were isolated and cultured from male Sprague-Dawley rats(provided by the Animal Experimental Center,Guangdong University of Chinese Medicine in China,about 120 g of weight).Passage 3 cells cultured in logarithmic phase were assigned into normal serum group(10%fetal bovine serum),6-hour blank serum group,12-hour blank serum,and 24-hour blank serum group.Proliferative capacity of cells was detected by cell counting kit-8,while the expression of LC3 and Beclin1 at protein and mRNA levels was determined by western blot and RT-PCR.RESULTS AND CONCLUSION:(1)No difference was found between the 6-hour blank serum group and the normal serum group in term of the proliferative capacity(P>0.05),while the proliferative capacity of cells in the 12-hour blank serum group was lower than that in the normal serum group and 6-hour blank serum group(P<0.01).The proliferative capacity of cells in the 24-hour blank serum group was lower than that in the 12-hour blank serum group(P<0.05).(2)Compared with the normal serum group,the protein and mRNA expressions of LC3II/LC3I and Beclin1 in the 6-hour blank serum group increased significantly(P<0.05),while compared with the 6-hour blank serum group,the protein and mRNA expressions of LC3II/LC3I and Beclin were increased significantly in the 12-hour and 24-hour blank serum groups(P<0.01 or P<0.05).To conclude,serum-free starvation can induce autophagy in muscle satellite cells,and the autophagy can protec

关 键 词：去血清 饥饿 肌卫星细胞 增殖 自噬 LC3 BECLIN1 大鼠 去血清饥饿条件 过度自噬 肌卫星细胞自噬 肌 骨骼 组织工程 

分 类 号：R456[医药卫生—治疗学] R318[医药卫生—临床医学]