Molecular Anatomy of Functional Differentination Affected by Combination of Gap Junctions, Connexins

Cx45缺陷的ES细胞在体外被诱导分化为收缩的心肌细胞。由于细胞间通讯的缺陷，它们的收缩不协调。在体内，我们创建了在所有体细胞（Cx45-KO）、心肌细胞（Cx45-CA）和血管内皮细胞（Cx45-Tie 2）中缺乏Cx45的小鼠。我们发现在每个突变体的表型有趣的差异。Cx45-KO小鼠在E10期死亡，显示房室传导阻滞和早期心脏内血管垫缺陷。Ca 2+依赖性转录因子Nfatcl在Cx45-KO内皮细胞中处于无活性的胞质形式。Cx45-CA小鼠于E10期死亡，表现为房室传导阻滞，但无内膜垫缺损。Nfatcl在Cx45-CA内皮细胞中以活性核形式存在。致死阶段表明传导阻滞是Cx45-KO小鼠死亡的主要原因。相反，Cx45-Tie 2小鼠没有表现出 关于我们 任何异常。此外，我们在缺乏Cx45的情况下创建了缺乏另一种心脏连接蛋白Cx43的小鼠。它们与Cx45-KO小鼠相同，表明Cx45构成了早期心脏中最重要的间隙连接蛋白。接下来，我们创建了心肌肌钙蛋白T（cTnT）缺陷小鼠。因为它是收缩器官的重要组成部分，cTnT缺陷小鼠没有显示任何收缩。其致死期与上述Cx45突变体相同。他们表现出由于心脏发育迟缓导致的内膜垫缺陷。只有当心肌层和内膜层Cx45均缺失时，内膜垫缺损才明显。我们的模型中，Cx45调节内皮细胞的上皮间质转化，需要进一步的研究，但该缺陷不是由生长迟缓引起的。与海外合作者的合作正在进行中，揭示了Cx45在神经元/神经胶质/神经嵴细胞中的组织特异性功能。Cx45在这些细胞类型中的作用将在不久的将来变得明显。少

Cx45-deficient ES cells were induced to differentiate into contracting cardiac myocytes in vitro. Their contractions were uncoordinated due to defective intercellular communication. In vivo, we created mice lacking Cx45 in all the somatic cells (Cx45-KO), in cardiac myocytes (Cx45-CA), and in vascular endothelial cells (Cx45-Tie2). We found intriguing differences of phenotypes in each mutant. The Cx45-KO mice died at the E10 stage, showing atrioventricular conduction block and an endocardial cushion defect in the early heart. The Ca2+-dependent transcription factor Nfatcl was in inactive, cytoplasmic form in the Cx45-KO endocardial endothelium. The Cx45-CA mice died at the E10 stage, showing atrioventricular conduction block, but without the endocardial cushion defect. Nfatcl was in active nuclear form in the Cx45-CA endocardial endothelium. The lethal stage shows that the conduction block is the primary cause of death in the Cx45-KO mice. The Cx45-Tie2 mice, in contrast, did not show … More any abnormality. Moreover, we created mice lacking another cardiac connexin Cx43, in the absence of Cx45. They were identical to the Cx45-KO mice, indicating that Cx45 constitutes the most significant gap junction protein in the early heart.Next, we created cardiac troponin T (cTnT)-deficient mice. Because it is an essential component of the contracting apparatus, the cTnT-deficient mice did not show any contractions. Their lethal stage was the same as the above Cx45-mutants. They showed the endocardial cushion defect due to retarded cardiac development. Nfatcl localization, however, indicated active nuclear form.The endocardial cushion defect was only apparent when Cx45 was absent in both of the myocardial and the endocardial layer. Our model, in which Cx45 regulates the epithelial-mesenchymal transformation in the endocardial endothelium, needs further revise by the future study, though the defect is not caused by growth retardation. Collaborations with the overseas coworkers are in progress, revealing the tissue specific functions of Cx45 in neuronal/glial/neural crest cells. Roles of Cx45 in these cell types will be evident in the near future. Less