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Analysis of the mechanisms of mammalian palate and limb morphogenesis

哺乳动物上颚和肢体形态发生机制分析

基本信息

批准号：
12670016
负责人：
TAKIGAWA Toshiya
金额：
$ 2.5万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

Cleft palate, the most frequent congenital craniofacial birth defect in humans, is caused by genetic and/or environmental perturbations in the multi-step process of palate development. Mutations in the TGF-β3 gene are associated with non-syndromic cleft palate in humans. However, little is known about why cleft palate is variable in its phenotype and penetrance and why mutation analysis using specific gene-deficient mice does not necessarily predict phenotypic consequences.In this study, we developed a novel suspension culture method of fetal mouse palatal shelves. By using the 'single palatal shelf in suspension culture' , we evaluated the differentiation of palatal medial edge epithelial (MEE) cells and its roles in palatal fusion. In addition, we established TGF-β3-defficient mice in various strains (C57BL/6J, 129/Sv, FVB/N, and ICR) as model systems for analyzing phenotypic variability of genetic cleft palate and the synergism of the master and modifier genes that regulate mammalian palatogenesis. In vivo analysis indicated that TGF-β3-null mutation causes severe cleft palate in inbred strains but partial cleft palate in outbred strains. In vitro analysis suggested that phenotypic variation of cleft palate in TGFβ3-defficient mice is closely associated with the difference in MEE cell differentiation between inbred and outbred mouse strains. However, such severe cleft palate in TGF-β3-null mice in inbred strains was partially rescued with a DNA methyltransferase inhibitor and produced milder phenotypes similar to those observed in outbred strains. Our results strongly suggest that TGFβ3-associated or -interacting imprinting genes may act as modifier genes and bring about phenotypic variation of mammalian non-syndromic cleft palate.

腭裂是人类最常见的先天性颅面畸形，是由腭裂发育过程中的遗传和/或环境干扰引起的。TGF-β3基因突变与人类非综合征性腭裂相关然而，很少有人知道为什么腭裂是可变的，在其表型和突变率，为什么使用特定的基因缺陷小鼠的突变分析不一定预测表型consequents.In这项研究中，我们开发了一种新的悬浮培养方法胎鼠腭架。本实验采用悬浮培养的单腭架，研究腭内侧缘上皮细胞的分化及其在腭融合中的作用。此外，我们建立了不同品系的TGF-β3缺陷小鼠（C57 BL/6 J，129/Sv，FVB/N和ICR）作为模型系统，用于分析遗传性腭裂的表型变异性以及调节哺乳动物腭裂发生的主基因和修饰基因的协同作用。体内分析表明，TGF-β3无效突变导致近交系中的严重腭裂，而远交系中的部分腭裂。体外分析表明，TGFβ3基因缺陷小鼠腭裂的表型变异与近交系和远交系小鼠间腭上皮细胞分化的差异密切相关。然而，在近交系中TGF-β3缺失小鼠中的这种严重腭裂被DNA甲基转移酶抑制剂部分挽救，并产生与远交系中观察到的表型相似的较温和的表型。我们的研究结果强烈提示，TGFβ3相关或相互作用的印记基因可能作为修饰基因，引起哺乳动物非综合征性腭裂的表型变异。