lncRNA TCONS00815878对猪骨骼肌卫星细胞分化的影响

The effect of lncRNA TCONS00815878 on differentiation of porcine skeletal muscle satellite cells

猪骨骼肌发育是一个复杂的生物学过程,其中骨骼肌卫星细胞分化是影响骨骼肌发育的重要环节。近年来发现长链非编码RNA(long non-coding RNA,lncRNA)在骨骼肌卫星细胞分化中具有重要作用。为探究lncRNA TCONS00815878对猪骨骼肌卫星细胞分化的影响,本研究利用qRT-PCR技术检测出生7 d内大白仔猪6种组织(心脏、脾脏、肺脏、肾脏、背肌和腿肌)及从胚胎期到出生后5个不同时间点(35 d、45 d、55 d胚胎及产后第7 d和第200 d后腿肌肉组织)TCONS00815878的表达情况;利用反义核苷酸(antisense oligonucleotides,ASO)在猪骨骼肌卫星细胞中敲低TCONS00815878,检验分化标记基因MyoD、MyoG和MyHC表达情况;通过生物信息学分析预测TCONS00815878靶基因,并利用DAVID软件在线预测其靶基因的功能与通路。结果表明:TCONS00815878在猪心肌和腿肌中高表达;仔猪出生后7 d内,TCONS00815878在猪肌肉组织中表达量不断升高,第7 d达到高峰;在猪骨骼肌卫星细胞增殖和分化过程中,TCONS00815878在分化期表达量不断上升,且在分化30 h表达量达到峰值;敲低TCONS00815878后,MyoD、MyoG和MyHC基因表达量降低,其中MyoD表达量显著下降(P<0.05)。此外,功能预测结果发现,其靶基因富集到糖酵解和丙酮酸代谢等与骨骼肌卫星细胞分化相关的多个生物学过程。本研究推测,lncRNA TCONS00815878可能对猪骨骼肌卫星细胞的分化起促进作用。

Porcine skeletal muscle development is a complex biological process,and differentiation of skeletal muscle satellite cells is an important part of skeletal muscle development.In recent years,it has been found that lncRNA plays an important role in the differentiation of skeletal muscle satellite cells.Here we investigate the effect of lncRNA TCONS00815878 on the differentiation of porcine skeletal muscle satellite cells.We first used q RT-PCR to detect the expression levels of TCONS00815878 in six tissues(heart,spleen,lung,kidney,back muscles and leg muscles)of Yorkshire piglets within seven days of birth.At the same time,the expression levels of TCONS00815878 at five different time points from the embryonic stage to the postnatal stage(35 d,45 d,55 d of embryos,and 7 d,200 d of postpartum leg muscles)were examined.The expression of the differentiation marker genes Myo D,MyoG and My HC was examined by knocking down TCONS00815878 in porcine skeletal muscle satellite cells using antisense oligonucleotides(ASO).The target gene of TCONS00815878 was predicted by bioinformatics analysis,and the function and pathway of its target gene were predicted online using DAVID software.The results showed that TCONS00815878 had the highest expression level in pig myocardium and leg muscles.Within seven days after birth,TCONS00815878 increased in the muscle tissue of pigs,and reached the peak of expression level on the 7 th day.During the process of proliferation and differentiation of porcine skeletal muscle satellite cells,the expression level of TCONS00815878 increased during the differentiation stage and peaked at 30 h of differentiation.After knocking down TCONS00815878,the expression levels of MyoD,MyoG and My HC were decreased,but the expression level of MyoD was significantly decreased(P<0.05).In addition,functional predictions revealed that the target gene of TCONS00815878 is enriched in multiple biological processes,such as glycolysis and pyruvate metabolism,related to skeletal muscle satellite cell differentiation.This study speculates that lncRNA TCONS00815878 may promote the differentiation of porcine skeletal muscle satellite cells.