TMEM182基因调控猪骨骼肌卫星细胞成肌分化的研究  

Study on the Regulation of Myogenic Differentiation of Porcine Skeletal Muscle Satellite Cells by Gene TMEM182

作　　者：龚宇轩 黑伟 鲍武 陈佳仪 李萌[1] 郭晓红[1] 李步高[1] GONG Yuxuan;HEI Wei;BAO Wu;CHEN Jiayi;LI Meng;GUO Xiaohong;LI Bugao(College of Animal Science,Shanxi Agricultural University,Taigu 030801,China)

机构地区：[1]山西农业大学动物科学学院,太谷030801

出　　处：《畜牧兽医学报》2025年第4期1676-1688,共13页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金面上项目(32272846);山西省重点研发计划项目(202102140601005)。

摘　　要：旨在探究TMEM182对猪骨骼肌卫星细胞增殖和成肌分化的调控作用。本研究选取饲养在相同条件下马身猪和大白猪各3头(180日龄,健康去势公猪),采集各组织;选取1周龄仔猪分离猪骨骼肌卫星细胞;通过qRT-PCR检测TMEM182在猪不同组织的表达谱、在不同猪种肌肉组织中的表达差异以及在骨骼肌卫星细胞不同分化天数的时序表达模式;通过在卫星细胞过表达TMEM182(OE-TMEM182)和干扰TMEM182 (si-TMEM182),采用qRT-PCR检测增殖标志基因的表达变化,采用EdU试验检测EdU阳性细胞数量变化;诱导猪骨骼肌卫星细胞成肌分化后,通过qRT-PCR、 Wstern blot、免疫荧光染色等技术检测成肌分化关键基因mRNA和蛋白的表达水平以及对肌管融合的影响。结果显示,TMEM182在猪肌肉组织中均高表达,在其他组织中几乎不表达;与马身猪相比,TMEM182在大白猪肌肉组织中表达量更高(P<0.05);随着卫星细胞分化天数的增加,TMEM182的表达量在分化第4天和5天时上升,分化第6天时下降,在分化第0天时表达量最低,第7天时表达量最高(P<0.05);互作蛋白结果显示,OE-TMEM182组CDH15、TMEM25、HS3ST1、SEC11A的表达量极显著降低(P<0.01),VWC2L的表达量显著降低(P<0.05);在细胞增殖过程中,过表达TMEM182后极显著下调CDK1、PCNA、Ki67的表达(P<0.01),且EdU阳性细胞数极显著减少(P<0.01)。干扰TMEM182后极显著上调CDK4、CDK1(P<0.01),显著上调Ki67的表达(P<0.05),且EdU阳性细胞数极显著增加(P<0.01)。在成肌分化过程中,过表达TMEM182后,MyoG、MyHC表达量极显著降低(P<0.01),MyoD表达量显著降低(P<0.05);干扰TMEM182后,MyoD、MyoG、MyHC的表达量极显著上升(P<0.01),Myf5的表达量显著上升(P<0.05);同时,TMEM182可能通过抑制PI3K-Akt信号通路调控猪卫星细胞的增殖和分化。本研究结果表明,TMEM182为猪骨骼肌卫星细胞增殖及分化过程中的一个负调控因子,推测其为猪骨骼肌生长发育的一个关键�This study aimed to investigate the regulatory impact of TMEM182 on the proliferation and differentiation of porcine skeletal muscle satellite cells.This study utilized 3 each of healthy castrated boars,Mashen pigs and Large White pigs,housed under the same conditions for 180 days of age,were selected for collection of each tissue;one-week-old piglets were selected for isolation of porcine skeletal muscle satellite cells;qRT-PCR was used to investigate the expression profile of TMEM182 in different pig tissues,compare expression patterns in muscle tissues across various pig species,and analyze the temporal expression changes in skeletal muscle satellite cells during different stages of differentiation.By overexpressing TMEM182(OETMEM182)and interfering with TMEM182(si-TMEM182)in satellite cells,we utilized qRTPCR to analyze the expression changes of proliferation marker genes and conducted EdU experiments to observe changes in the number of EdU-positive cells.Following the induction of myogenic differentiation in porcine skeletal muscle satellite cells,various techniques including qRTPCR,Western blot,and immunofluorescence staining techniques were utilized to examine alterations in the mRNA and protein expression levels of genes associated with myogenic differentiation,as well as their impact on myotube fusion.The study results showed that TMEM182 had high expression levels in muscle tissues but was nearly absent in other tissues.In comparison to Mashen pigs,TMEM182 was notably more expressed in Large White pigs tissues(P<0.05).As satellite cells differentiation progressed,at the 4th and 5th days of differentiation,there was an increase in TMEM182 expression.The expression level was the lowest on the 0th day of differentiation.However,at the 6th day of differentiation,there was a decrease in TMEM182 expression.Conversely,at the 7th day of differentiation,TMEM182 expression was at its peak(P<0.05).The results of the intercalated proteins showed that the expression of CDH15,TMEM25,HS3ST1,and SEC11A was highly signi

关 键 词：TMEM182 肌肉特异性 骨骼肌卫星细胞 增殖 成肌分化 

分 类 号：S828.2[农业科学—畜牧学]