Sakabe Masahide, Sakata Hirokazu, Matsui Hiroko, Ikeda Kazuo, Yamagishi Toshiyuki, Nakajima Yuji
Department of Anatomy and Cell Biology, Graduate School of Medicine, Osaka City University, Osaka, Japan.
Anat Rec (Hoboken). 2008 Jul;291(7):845-57. doi: 10.1002/ar.20708.
During early heart development at the looped heart stage, endothelial cells in the outflow tract and atrioventricular (AV) regions transform into mesenchyme to generate endocardial cushion tissue. This endocardial epithelial-mesenchymal transition (EMT) is regulated by several regulatory pathways, including the transforming growth factor-beta (TGFbeta), bone morphogenetic protein (BMP), and Rho-ROCK pathways. Here, we investigated the spatiotemporal expression pattern of ROCK1 mRNA during EMT in chick and examined whether TGFbeta or BMP could induce the expression of ROCK1. At the onset of EMT, ROCK1 expression was up-regulated in endothelial/mesenchymal cells. A three-dimensional collagen gel assay was used to examine the mechanisms regulating the expression of ROCK1. In AV endocardium co-cultured with associated myocardium, ROCK1 expression was inhibited by either anti-TGFbeta3 antibody, anti-ALK2 antibody or noggin, but not SB431542 (ALK5 inhibitor). In cultured preactivated AV endocardium, TGFbeta3 protein induced the expression of ROCK1, but BMP did not. AV endothelial cells that were cultured in medium supplemented with TGFbeta3 plus anti-ALK2 antibody failed to express ROCK1. These results suggest that the expression of ROCK1 is up-regulated at the onset of EMT and that signaling mediated by TGFbeta3/ALK2 together with BMP is involved in the expression of ROCK1.
在心脏发育早期的环状心脏阶段，流出道和房室（AV）区域的内皮细胞转变为间充质，以生成心内膜垫组织。这种心内膜上皮-间充质转化（EMT）受多种调节途径调控，包括转化生长因子-β（TGFβ）、骨形态发生蛋白（BMP）和Rho-ROCK途径。在此，我们研究了鸡EMT过程中ROCK1 mRNA的时空表达模式，并检测了TGFβ或BMP是否能诱导ROCK1的表达。在EMT开始时，ROCK1在内皮/间充质细胞中的表达上调。采用三维胶原凝胶试验来研究调节ROCK1表达的机制。在与相关心肌共同培养的AV心内膜中，抗TGFβ3抗体、抗ALK2抗体或头蛋白可抑制ROCK1的表达，但SB431542（ALK5抑制剂）则不能。在培养的预激活AV心内膜中，TGFβ3蛋白可诱导ROCK1的表达，但BMP则不能。在添加TGFβ3加抗ALK2抗体的培养基中培养的AV内皮细胞未能表达ROCK1。这些结果表明，ROCK1的表达在EMT开始时上调，并且TGFβ3/ALK2与BMP介导的信号传导参与了ROCK1的表达。
