需肌醇酶-1-c-Jun氨基末端激酶通路在周期性应力介导的成肌细胞凋亡中的作用  

The role of inositol requiring enzyme-1-c-Jun N-terminal kinase signaling pathways in stress-induced apoptosis of myoblasts

作　　者：夏晨蕾 丁弦[1] 孙文娜[1] 贺苗[1] 王芳[1] 姜文心[1] 张彩霞[1] 张强[1] 刘文[1] 张月[1] 阎潇[1] 姚如永[1] 袁晓[1] 

机构地区：[1]青岛大学医学院附属青岛市立医院口腔医学中心,266071

出　　处：《中华实验外科杂志》2015年第5期1127-1129,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（31170891）;山东省卫生厅重点资助项目（2011HD001）;青岛市市立医院总院长科研专项基金青年基金资助项目（Ⅷ院2013VYW12）

摘　　要：目的 观察在周期性应力介导成肌细胞凋亡中内质网应激需肌醇酶-1（IRE-1）-c-jun氨基端激酶（JNK）信号通路的作用.方法 以大鼠L6成肌细胞为实验对象,构建体外培养-力学刺激模型.应用应力加载装置分别加1、2、6、12、24 h的周期性张应力（拉伸变形率为15％,频率为10次循环/分）,对照组不加力（0 h）.Hochest 33258染色、膜联蛋白V（Annexin V）-异硫氰酸荧光素（FITC）/碘化丙锭（PI）流式细胞术分别检测凋亡细胞的数目及凋亡率;反转录-聚合酶链反应（RT-PCR）检测内质网应激相关因子C/EBP同源蛋白（CHOP）、凋亡信号调节激酶-1（ASK-1）、肿瘤坏死因子受体相关因子2（TRAF2）和JNK mRNA的表达变化;加入IRE-1特异性抑制剂STF-083010,检测ASK-1、TRAF2和JNK mRNA的表达变化.结果 随应力加载时间的延长,凋亡细胞的数目、凋亡率呈一致上升趋势,且在24h达峰值[（14.34±0.77）个];CHOP、ASK-1、TRAF2mRNA表达量随着应力加载时间的延长逐渐上升,并在24 h达最高值,分别为4.19±1.76、3.46±0.84、3.78±1.38（P ＜0.05）,JNK mRNA的表达量在0～2h逐渐上升后逐渐下降,随后又逐渐上升在24 h达最高值（3.04±0.79,P＜0.05）;加入大鼠IRE-1特异性抑制剂STF-083010后,TRAF2、ASK-1表达量均明显减少（P＜0.05）.结论 周期性张应力可诱导成肌细胞的凋亡,凋亡率随着应力加载时间的延长而升高.CHOP、TRAF2、ASK-1、JNK mRNA的表达量升高,提示内质网相关的凋亡途径参与了应力介导的成肌细胞的凋亡.加入IRE-1抑制剂后,TRAF2、ASK-1、JNK mRNA表达量均明显降低,提示IRE-1-JNK通路参与了应力介导的成肌细胞凋亡.Objective To explore the role of inositol requiring enzyme-1 （IRE-1）-c-Jun N-terminal kinase （JNK） signaling pathways in stretch-induced apoptosis of myoblasts.Methods As experimental objects,L6 rat myoblasts were used to construct the culture-mechanical stimulation model in vitro.Then cells were subjected to 15% surface elongation with a frequency of 10 cycles/min cyclic stretch using multi-channel stress loading system.Cells were harvested after different durations （1,2,6,12,24 h） of cyclic exposure.After cyclic stretch,Hochest 33258 staining and Annexin V-fluoresceine isothiocyanate （FITC）/propidium iodide （PI） staining was performed to observe the morphology of the apoptotic cells.Reverse transcriptase-polymerase chain reaction （RT-PCR） was used to detect the mRNA expression of endoplasmic reticulum stress related factors [C/EBP homologous protein （CHOP）,tumor necrosis factor receptor associated factor2 （TRAF2）,apoptosis signal regulating kinase-1 （ASK-1） and JNK].IRE-1 specific inhibitor （STF-083010） was used to detect the TRAF2,ASK-1 and JNK mRNA expression to investigate the role of IRE-1-JNK signaling pathway in stretch-induced apoptosis.Results The stretch-induced apoptosis rate of myoblasts was increased in a time-dependent manner,peaked at 24 h （14.34 ± 0.77）.CHOP,TRAF2 and ASK-1 mRNA expression levels were increased with the extension of stretch time,and reached the maximum at 24 h [（4.19 ± 1.76）,（3.46 ±0.84）,and （3.78 ± 1.38）,P 〈0.05];but JNK mRNA expression was decreased followed by an increase range from 0 h to 2 h,and then increased again within 24 h and peaked at 24 h （3.04 ± 0.79,P 〈 0.05）.After treatment with IRE-1 specific inhibitor （STF-083010）,the real-time PCR results showed that TRAF2 and ASK-1 mRNA expression was significantly decreased （P 〈 0.05）.Conclusion Cyclic stretch can induce myoblast apoptosis and within 24 h the apoptosis rate of myoblasts was increased in a time-dependent manner.The increase of CHOP,

关 键 词：需肌醇酶-1 C-JUN氨基端激酶 内质网应激 周期性张应力 脱噬作用 

分 类 号：R966[医药卫生—药理学]