PTEN在胚胎早期心脏发育过程中的作用

摘要

Objective Endogenous phosphatase and tensin homolog ( PTEN ) expresses in primitive streak during embryo gastrulation and mesoderm derived structure , included heart tube, later. We investigated the possibility that PTEN is involved in heart formation during early embryo development. Methods Un - incubated chicken embryos were cultured using the modified filter paper method in 37℃ incubator until HH4 stage. Subsequently PTENs was transfected into cardiogenesis area around anterior primitive streak. Half of transfected embryos were fostered at 37℃ for about 30hs continuously until the heart tube formation to study the role of PTEN in the processes of cell adhesion and migration; CSCs from the other half were isolated and cultured in 37 ℃ CO2 incubator in vitro until differentiating into myocardial tissue with beating pulse in order to study the role of PTEN in the processes of cardiac cell proliferation, differentiation and apoptosis.Results The result of whole chick embryos immunohistochemistry ( N - cadherin ) showed that chick heart tuhes transfected by PTEN - WT and PTEN - G129E appeared in reverse cyclization, polycystic malformation and so on, while the heart tubes from GFP group and PTEN - C124S group developed roughly normally. After transfection with PTEN, the beating frequency of myocardial tissue cultured in vitro decline faster than that in GFP group, with shorter maintaining time than control group. There was significantly difference between PTEN - WT group and the GFP group ( P ＜ 0. 05 ), while no significant difference among PTEN - WT group, PTEN - C1245 group and PTEN - G129E was revealed ( P ＞ 0. 05 ).Conclusion It suggests that PTEN increases the cell adhesion between the early embryo CSC and cell matrix, interferes with cell migration while promote the differentiation CSC , and accelerates the aging of myocardium.%目的 通过过表达PTEN,探讨胚胎早期原肠胚形成期PTEN表达及在早期心脏发生发育过程的作用和意义.方法 用改良滤纸鸡胚胎整体培养法取胚,37℃孵箱孵育待其发育到HH4期,对鸡胚心肌干细胞(Cardiac Stem Cell)进行PTEN电转染,转染成功后一部分鸡胚37℃孵箱继续培养30 h,待其发育形成心管,观察过表达PTEN对心管发生的影响;另一部分鸡胚在超净台内完成心肌干细胞的离体,体外培养使其分化发育成心肌组织,主要研究PTEN在细胞增殖、分化和凋亡过程中的作用.结果 30 h后鸡胚整体的免疫组织化学(N-cadherin)结果显示,PTEN-WT组和PTEN-G129E组鸡胚心管出现了反向环化、多囊化等畸形,GFP组和PTEN-C124S组心管发育受到的影响较小;转染后离体培养得到的心肌组织的跳动频率与GFP组下降的速度加快,且维持时间缩短,GFP组与PTEN-WT组差异有显著性(P<0.05),PTEN-WT组与PTEN-C124S组、PTEN-G129E组差异无显著性(P＞0.05).结论 PTEN蛋白酯酶可能参与早期胚胎心肌干细胞之间黏附分子的表达从而调节细胞迁移、分化发育,并加速了心肌组织的衰老.