MOLECULAR PATHOGENESIS OF CRANIOFACIAL MALFORMATIONS

在已知的致畸物质中，乙醇是美国最大的公共卫生问题，在大约5%的先天性心脏、中枢神经系统和颅面畸形中起病因作用。尽管其巨大的意义，很少有人知道关于乙醇胚胎病的分子发病机制。颅神经嵴衍生细胞（CNCC）是所有发育中颅面结构的主要组成部分。鸡或小鼠胚胎暴露于乙醇导致颅面过度发育。MSX 2和信号分子BMP 4的表达增加与后脑中CNCC的发育程序性死亡相关。MSX 2的过度表达导致颅面畸形，与乙醇暴露相关的颅面畸形惊人相似。该项目的目标是确定MSX 1，MSX 2和/或BMP 4是否参与与乙醇诱导的颅面畸形相关的过度细胞死亡。具体目的如下：（1）确定MSX 2转基因小鼠的颅面畸形是否由MSX 2过表达细胞的过度死亡引起。(2)确定MSX 1过表达是否影响小鼠胚胎CNCC的存活和随后的颅面发育。(3)确定乙醇暴露是否改变小鼠胚胎中Msx 1、Msx 2和/或Bmp 4基因的表达。(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BMP 4和/或MSX蛋白介导。(5)确定Msx 1或Msx 2基因座的功能缺失突变是否对乙醇诱导的小鼠颅面畸形易感。(6)鉴定过表达MSX 2或在子宫内暴露于乙醇的小鼠胚胎CNCC中受MSX 2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理解外，识别参与这一过程的基因可能最终提供一种诊断方法，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.