摘要
目的探讨不同严重程度内质网应激对过氧化物酶体增殖物激活受体α(peroxisome proliferator-activated receptor alpha,PPARα)表达的影响。方法以小鼠肝细胞AML-12为研究对象,内质网应激诱导剂衣霉素(Tunicamycin,TM)以时间梯度、浓度梯度细胞诱导不同严重程度的内质网应激。采用实时荧光定量PCR方法和免疫印迹方法检测葡萄糖调节蛋白78(glucose regulated protein 78,GRP78)、CCAAT/增强子结合蛋白同源蛋白(C/EBP-homologous protein,CHOP)、PPARα在内质网应激过程中的基因和蛋白动态表达;用免疫荧光染色方法检测PPARα蛋白在内质网应激过程中的表达情况和位置变化。结果在TM分别以不同时间和不同浓度诱导AML-12细胞内质网应激的过程中,随TM刺激时间延长与浓度增加,CHOP、GRP78 mRNA和蛋白水平也逐渐升高;而PPARαmRNA和蛋白呈现先升高后下降的表达趋势。在免疫荧光检测中,TM刺激早期PPARα主要在细胞质中表达,晚期集中在细胞核内;低浓度TM刺激时PPARα的位置主要表达在细胞质,而高浓度TM刺激后,PPARα聚集在细胞核中。结论细胞轻度内质网应激时,PPARα表达升高,而在细胞严重内质网应激时PPARα表达下降。
Objective To investigate the effects of endoplasmic reticulum stress on the expression of peroxisome proliferator-activated receptor alpha(PPARα) in different severity. Methods AML-12 cells were used as the research object. Endoplasmic reticulum stress inducer-Tunicamycin(TM), stimulated AML-12 cells with time gradient and concentration gradient to induce different severity of endoplasmic reticulum stress. Real-time quantitative PCR and Western blotting were used to detect the expressions of GRP78, CHOP, PPARα mRNAs and proteins in endoplasmic reticulum stress induced by TM at different times and concentrations. Immunofluorescence staining was used to detect the expression and positional changes of PPARα protein in the endoplasmic reticulum stress induced by TM at different times and concentrations respectively. Results During the induction of endoplasmic reticulum stress in AML-12 cells at different time and different concentrations respectively, the expressions of CHOP and GRP78 mRNAs and proteins were increased gradually. However, PPARα mRNA and protein were increased first and then decreased. In the immunofluorescence assay, TM stimulated early expression of PPARα mainly in the cytoplasm, and lately concentrated in the nucleus;PPARα was mainly expressed in the cytoplasm when stimulated by low concentration of TM and accumulated in the nucleus after stimulation with high concentration of TM. Conclusion In the process of TM induced endoplasmic reticulum stress in AML-12 cells, the expression of PPARα is increased in the mild endoplasmic reticulum stress, but in the severe endoplasmic reticulum stress, the expression of PPARα is decreased.
作者
徐玲
田原
张向颖
王慧娟
任锋
XU Ling;TIAN Yuan;ZHANG Xiangying;WANG Huijuan;REN Feng(Beijing Institute of Hepatology,Beijing YouAn Hospital,Capital Medical University,Beijing 100069;Second Clinical Hospital,Shanxi Medical University,China)
出处
《胃肠病学和肝病学杂志》
CAS
2020年第7期799-804,共6页
Chinese Journal of Gastroenterology and Hepatology
基金
国家自然科学基金项目(81770611)
北京市科技计划重大专项(D161100002716004)
天晴肝病基金项目(TQGB20180213)
科技部传染病重大专项项目(2018ZX10301407-005-002,2018ZX10302205-004-004)。
