The mammalian muscularized diaphragm is essential for respiration and defects in the developing diaphragm cause a common and frequently lethal birth defect, congenital diaphragmatic hernia (CDH). Human genetic studies have implicated more than 150 genes and multiple molecular pathways in CDH, but few of these have been validated because of the expense and time to generate mouse mutants. The pleuroperitoneal folds (PPFs) are transient embryonic structures in diaphragm development and defects in PPFs lead to CDH. We have developed a system to culture PPF fibroblasts from E12.5 mouse embryos and show that these fibroblasts, in contrast to the commonly used NIH 3T3 fibroblasts, maintain expression of key genes in normal diaphragm development. Using pharmacological and genetic manipulations that result in CDH in vivo, we also demonstrate that differences in proliferation provide a rapid means of distinguishing healthy and impaired PPF fibroblasts. Thus, the PPF fibroblast cell culture system is an efficient tool for assaying the functional significance of CDH candidate genes and molecular pathways and will be an important resource for elucidating the complex etiology of CDH.

哺乳动物肌肉化的横膈膜对于呼吸至关重要，而发育中的横膈膜缺陷会导致常见且经常致命的出生缺陷，即先天性横膈疝 (CDH)。人类基因研究已涉及 CDH 中的 150 多个基因和多种分子途径，但由于产生小鼠突变体的费用和时间，其中很少有得到验证。胸膜襞 (PPF) 是膈肌发育过程中的短暂胚胎结构，PPF 的缺陷会导致 CDH。我们开发了一种从 E12.5 小鼠胚胎中培养 PPF 成纤维细胞的系统，并表明这些成纤维细胞与常用的 NIH 3T3 成纤维细胞相比，在正常膈肌发育中保持关键基因的表达。使用导致体内CDH 的药理学和基因操作，我们还证明了增殖的差异提供了一种快速区分健康和受损 PPF 成纤维细胞的方法。因此，PPF成纤维细胞培养系统是检测CDH候选基因和分子通路功能意义的有效工具，将成为阐明CDH复杂病因的重要资源。