MOLECULAR PATHOGENESIS OF CRANIOFACIAL MALFORMATIONS

在已知的致畸因素中，乙醇是美国最大的公共卫生问题，约5%的先天性心脏、中枢神经系统和颅面畸形都是由乙醇引起的。尽管它具有巨大的意义，但对乙醇胚胎病的分子发病机制知之甚少。颅神经脊源性细胞(CNCC)是所有发育中的颅面结构的重要组成部分。小鸡或小鼠胚胎暴露在乙醇中会导致颅面部过度发育。MSX2和信号分子BMP4的表达增加与后脑CNCC的发育性程序性死亡有关。MSX2的过度表达会导致颅面畸形，与酒精暴露的结果惊人地相似。该项目的目标是确定Msx1、MSX2和/或BMP4是否与乙醇诱导的颅面畸形相关的过度细胞死亡有关。具体目的如下：(1)确定MSX2转基因小鼠颅面畸形是否是由于MSX2过表达细胞过度死亡所致。(2)确定Msx1过表达是否影响CNCC的存活和随后的小鼠胚胎颅面发育。(3)研究乙醇暴露对小鼠胚胎Msx1、MSX2和/或Bmp4基因表达的影响。(4)确定乙醇是否影响斑马鱼胚胎CNCC的存活和随后的颅面发育，以及这些影响是否由BMP4和/或MSX蛋白介导。(5)确定Msx1或Msx2基因功能缺失突变是否对乙醇所致小鼠颅面畸形易感性。(6)鉴定子宫内高表达MSX2或宫内暴露于乙醇的小鼠胚胎CNCC中受MSX2转录调控的基因。除了增加我们对疾病发病机制和正常发育的了解外，识别参与这一过程的基因最终可能提供一种诊断方法，用于识别乙醇胚胎病风险增加的妊娠。

Among known teratogens, ethanol represents the greatest public health problem in the U.S., playing an etiological role in approximately 5% of all congenital cardiac, central nervous system, and craniofacial malformations. Despite its enormous significance, little is known regarding the molecular pathogenesis of ethanol embryopathy. Cranial neural-crest derived cells (CNCC) are a major constituent of all developing craniofacial structures. Exposure of chick or mouse embryos to ethanol results in excessive craniofacial development. Increased expression of MSX2 and the signaling molecule BMP4 is associated with developmentally-programmed death of CNCC in the hindbrain. Over-expression of MSX2 results in craniofacial malformations strikingly similar to those associated with ethanol exposure. The goal of this project is to determine whether MSX1, MSX2, and/or BMP4 are involved in the excessive cell death associated with ethanol-induced craniofacial malformations. The Specific Aims are as follows: (1) To determine whether craniofacial malformations in MSX2 transgenic mice result from excessive death of MSX2- over-expressing cells. (2) To determine whether MSX1 over-expression affects survival of CNCC and subsequent craniofacial development in mouse embryos. (3) To determine whether ethanol exposure alters Msx1, Msx2, and/or Bmp4 gene expression in mouse embryos. (4) To determine whether ethanol affects survival of CNCC and subsequent craniofacial development in zebrafish embryos and whether these effects are mediated by BMP4 and/or MSX proteins. (5) To determine whether loss-of-function mutations at the Msx1 or Msx2 locus susceptibility to ethanol- induced craniofacial malformations in mice. (6) To identify genes that are transcriptionally regulated by MSX2 in CNCC of mouse embryos that over-express MSX2 or have been exposed to ethanol in utero. In addition to increasing our understanding of disease pathogenesis and of normal development, the identification of genes involved in this process may ultimately provide a diagnostic method for identifying pregnancies at increased risk for ethanol embryopathy.