磷脂酰肌醇-3激酶在间充质干细胞成肌分化中的表达及肌损伤修复中的作用

Expression of phosphoinositide-3k inases in mesenchymal stem cells diffe rentiation of myoblasts and its effects on muscle injury repair

目的:以5-氮杂胞苷(5-azacytidine,5-Aza-CR)诱导间充质干细胞(mesenchymalstemcells,MSCs)成肌分化,探讨生肌调控因子Myf5的表达情况及磷脂酰肌醇-3激酶(phosphoinositide-3kinases,PI3K)在此分化全过程中的作用。方法:分离、纯化及扩增MSCs,以5-Aza诱导其成肌分化,RT-PCR测定Myf5的表达,免疫组化检测α-sarcomeric表达;Western-blot检测LY294002抑制前后磷酸化Akt蛋白水平的变化。结果:LY294002作用后,Myf5表达延迟至诱导后12h;诱导后10d,部分MSCs表达α-sarcomeric;磷酸化Akt在5-Aza诱导后蛋白水平逐渐增强,LY294002抑制明显下降。结论:MSCs成肌分化过程中,PI3K是重要的正向信号转导通路。

To investigate th e expression of Myf5and the effects of phosphoinosi-tide-3kinases(PI3K)in a ll the course of myoblast differe ntiation in mes-enchymal stem cells(MSCs )i nduced by 5-azacytidine(5-Aza-CR).METHODS:MSCs were sperated,purified ,and pr oliferated,and their my-obilast differentiated were induced by 5-Az a-CR.The gene expression of Myf5was assayed with reverse transc ription-ploy merase chain reaction(RT-PCR)method and the antigen expression ofα-sarcomer ic was detected with immunohistochemistry method.The changes of the activity of phosph o-rylation Akt before and after inhibi ted by LY294002were observ ed by Western-blot method.RESULTS:The time of Myf5expression was delay ed to 12h after inhibited by LY294002.Some of MSCs expressedα-sarcomeric a t 10day after induc-tion.The phosphorylation Akt activ ity of MSCs increased after induced b y5-Aza-CR but obviously decreased after inhibited by LY 29 4002.PI3K is a positive signal transduction path in the course of myoblast differentiation in MSCs.