肉仔鸡卫星细胞氧化应激时MAPK信号通路  

Study on Oxidative Stress-Activated MAPK Signaling Pathway in Broilers Breast Muscle Satellite Cells

作　　者：王成[1] 武书庚[1] 张海军[1] 岳洪源[1] 翟永功[2] 齐广海[1] 

机构地区：[1]中国农业科学院饲料研究所/农业部饲料生物技术重点开放实验室,北京100081 [2]北京师范大学生命科学学院,北京100875

出　　处：《中国农业科学》2010年第20期4286-4294,共9页Scientia Agricultura Sinica

基　　金：国家科技支撑计划(2006BAD12B06);云南省科技创新强省计划(2007AD003);国家自然科学基金(31072048)

摘　　要：【目的】在地塞米松(DEX)诱导骨骼肌卫星细胞氧化应激的体外条件下,筛选丝裂原活化蛋白激酶(MAPKs)信号系统中起关键作用的信号通路。【方法】将体外培养的肉仔鸡胸肌骨骼肌卫星细胞(SCs)分别进行处理:Ⅰ、对照组;Ⅱ、DEX;Ⅲ、p38MAPK抑制剂;Ⅳ、DEX+p38MAPK抑制剂;Ⅴ、JNK抑制剂;Ⅵ、DEX+JNK抑制剂;Ⅶ、ERK5抑制剂;Ⅷ、DEX+ERK5抑制剂;Ⅸ、ERK1/2抑制剂;Ⅹ、DEX+ERK1/2抑制剂。除处理Ⅰ和处理Ⅱ外,其它处理首先用抑制剂预处理30min,弃去培养基后,再按处理设置分别加入含DEX的培养基或者正常培养基继续处理24h。处理结束后,分别测定细胞培养液中丙二醛(MDA)、活性氧自由基(ROS)、超氧化物歧化酶(SOD)和谷胱甘肽硫转酶(GST)含量或活性;同时采用RT-PCR方法检测通路蛋白及SOD与GST基因表达量。【结果】抑制剂单独处理组MDA和ROS的产生量显著低于DEX处理(P<0.05);DEX+ERK5抑制剂处理与DEX+ERK1/2抑制剂处理的MDA和ROS产生量显著高于ERK5抑制剂处理和ERK1/2抑制剂处理(P<0.05);DEX+p38MAPK抑制剂处理与DEX+JNK抑制剂处理组的MDA和ROS产生量与p38MAPK处理和JNK处理的差异不显著(P>0.05)。抑制剂单独处理组的SOD和GST活性比DEX处理高(P<0.01);DEX+ERK5抑制剂处理与DEX+ERK1/2抑制剂处理的SOD和GST活性极显著低于ERK5抑制剂处理组和ERK1/2抑制剂处理组(P<0.01),DEX+p38MAPK抑制剂处理与DEX+JNK抑制剂处理的SOD和GST活性与p38MAPK抑制剂处理和JNK抑制剂处理的差异不显著(P>0.05)。基因表达结果表明,DEX能够抑制GST和SOD基因的表达,抑制率达37%以上。与抑制ERK5和ERK1/2通路不同,抑制p38MAPK和JNK通路后,DEX对GST和SOD基因的表达无明显抑制作用。【结论】在DEX诱导的肉仔鸡胸肌骨骼肌SCs产生的氧化应激过程中,p38MAPK和JNK通路起着关键作用。【Objective】Mitogen-activated protein kinases（MAPKs） signaling pathway was investigated to screen which one plays a key role in broiler skeletal muscle satellite cells（SCs） under oxidative stress induced by dexamethasone（DEX） in vitro. 【Method】The broilers skeletal muscle SCs were assigned into one of the ten treatments：control（I） ,DEX（Ⅱ） ,inhibitor p38 MAPK（Ⅲ） ,DEX plus inhibitor p38 MAPK（Ⅳ） ,inhibitor JNK（Ⅴ） ,DEX plus inhibitor JNK（Ⅵ） ,inhibitor ERK5（Ⅶ） ,DEX plus inhibitor ERK5（Ⅷ） ,inhibitor ERK1/2（Ⅸ） ,DEX plus inhibitor ERK1/2（Ⅹ） .The treatments exclude treatments I andⅡwere treated with inhibitors for 30 min before treating with DEX and normal medium for 24 h.The relative production of malonaldehyde（MDA） ,reactive oxidative species（ROS） ,the activities of superoxide dismutase（SOD） and glutathione S-transferase（GST） were measured,respectively.Polymerase chain reaction（PCR） was employed to assess SOD and GST gene expression.【Result】 Compared with DEX,the MDA and ROS concentrations were decreased（P0.05） in all the inhibitors-treated groups.Theconcentration of MDA and ROS in treatment with inhibitors were higher than that in treatment with DEX（P0.05） The MDA and ROS concentrations of treatment with DEX plus inhibitor ERK5 and of treatment with DEX plus inhibitor ERK 1/2 were significantly higher than that of treatment with inhibitor ERK5 and treatment with inhibitor ERK1/2（P〈0.05） .However,there were no significant differences in MDA and ROS concentrations among treatment with DEX plus inhibitor p38 MAPK,treatment with DEX plus inhibitor JNK and treatment with inhibitor p38 MAPK,treatment with inhibitor JNK（P〈0.05） .The activities of SOD and GST of all the inhibitors-treated groups were significantly higher than those of DEX group（P0.01） .The activities of SOD and GST of treatment with DEX plus inhibitor ERK5 and treatment with DEX plus inhibitor ERK 1/2 were significantly lower than th

关 键 词：氧化应激 骨骼肌卫星细胞 MAPK P38 JNK 

分 类 号：S831[农业科学—畜牧学]