摘要
成肌分化是动物肌肉发育的关键环节,与其健康和生产性能密切相关。为研究成肌细胞抗氧化应激的机制,试验分离了绵羊胎儿成肌细胞,利用地塞米松处理分化中的成肌细胞建立氧化应激模型;试验分为4组,包括正常分化48 h组(N 48 h)、地塞米松处理分化48 h组(Dex 48 h)、正常分化72 h组(N 72 h)、地塞米松处理分化72 h组(Dex 72 h),通过倒置显微镜观察各组细胞的形态变化,利用ROS检测试剂盒检测ROS含量,用RNA-seq检测NF-κB和Nrf2信号通路相关基因mRNA的表达情况,并采用皮尔森相关系数法分析相关性,用STRING数据库分析蛋白的互作关系。结果表明,地塞米松诱导的氧化应激对成肌分化具有明显的抑制作用,且抑制作用随处理时间的延长而增加;ROS含量较各自对照组均显著增加,且随着处理时间的延长而增加。由RNA-seq分析结果可知,与对照组相比,地塞米松处理组NF-κB和Nrf2信号通路被激活,上游基因P65、IKK2、IκBα、IκBβ、Nrf2、Keap1以及相应下游靶基因SOD1、SOD2、FHC、HO-1、GSH-Px的mRNA相对表达量均显著升高,各基因之间存在不同程度的相关性以及错综复杂的蛋白互作关系,其中Keap1基因是联系2条通路的重要枢纽。综上所述,在成肌细胞分化过程中,NF-κB和Nrf2信号通路交互联系,共同在地塞米松诱导的成肌细胞氧化应激中发挥抗氧化作用。
Myogenic differentiation is a key link in animal muscle development and is closely related to its health and production performance.To study the mechanism of anti-oxidative stress of myoblasts,in this study,sheep fetal myoblasts were isolated,the oxidative stress model were established by treating differentiating myoblasts with dexamethasone.The test was divided into 4 groups,including normal differentiation 48 h(N 48 h),dexamethasone treatment differentiation 48 h(Dex 48 h),normal differentiation 72 h(N 72 h),dexamethasone treatment differentiation 72 h(Dex 72 h).The cell morphological changes of each group were observed by inverted microscopy,the ROS content was detected by ROS detection kit,and the mRNA expression of genes associated with NF-κB and Nrf2 signaling pathways was detected by RNA-seq.The correlation of genes was analyzed by Pearson correlation coefficient method,and the protein interaction relationship was analyzed in the STRING database.The results showed that the myogenic differentiation was significantly inhabited under the treatment of oxidative stress induced by dexamethasone,and the inhibitory effect was enhanced with prolongation of the treatment time.The ROS content significantly increased compared to the respective control groups,and increased with prolongation of the treatment time.RNA-seq analysis showed that the NF-κB and Nrf2 signaling pathways were activated in dexamethasone treatment group,the mRNA expression levels of upstream genes P65,IKK2,IκBα,IκBβ,Nrf2,Keap1 and corresponding downstream target genes SOD1,SOD2,FHC,HO-1,GSH-Px were significantly higher than those in the control.There were different degrees of correlation and intricate protein interactions among these genes,in which Keap1 was an important hub connecting the two pathways.In summary,during the myoblast differentiation,NF-κB and Nrf2 signaling pathways interacted and jointly played an antioxidant role in the dexamethasone-induced oxidative stress of myoblasts.
作者
李君
王鹤洁
安洁
李俊玲
窦敏敏
翟竹汇
何浪
李振月
秦健
杜荣
LI Jun;WANG Hejie;AN Jie;LI Junling;DOU Minmin;ZHAI Zhuhui;HE Lang;LI Zhenyue;QIN Jian;DU Rong(College of Veterinary Medicine,Shanxi Agricultural University,Taigu 030801,China;College of Life Sciences,Shanxi Agricultural University,Taigu 030801,China;Center of Experiment Teaching,Shanxi Agricultural University,Taigu 030801,China)
出处
《山西农业科学》
2023年第11期1252-1259,共8页
Journal of Shanxi Agricultural Sciences
基金
国家自然科学基金项目(32272964,31872438)
山西省基础研究计划(20210302123394)
山西省回国留学人员科研资助项目(2022-102)
山西省留学回国人员科技活动择优资助项目
山西农业大学横向科技项目(2015HX12)
山西农业大学中青年拔尖创新人才支持计划(BJRC201204)。
