Heparan Sulfate in Skeletal Development and Diseases

描述(由申请人提供)：此竞争性修订申请是对NOT-OD-09-058的响应：NIH宣布可为竞争性修订申请提供恢复法案资金。硫酸乙酰肝素(HS)在多种组织的发育中起着重要作用。了解HS在骨骼发育过程中的作用具有重要的临床意义。遗传性多发性骨软骨病(HME)是一种遗传性疾病，以多发性骨软骨瘤的形成为特征，在美国影响着数千人。HME患者携带EXT1或EXT2杂合性突变，它们共同编码HS生物合成所必需的糖基转移酶。尽管HME的致病基因已被鉴定，但其发病机制仍有许多谜团和未解之谜。最令人费解的问题之一是人类HME的表现与携带相同基因的小鼠的表型之间的明显差异。最重要的是，应该忠实地模仿人类HME的ext1和ext2杂合子小鼠都对外生骨发育具有抵抗力，尤其是在长骨中。我们最近做了一个令人惊讶的观察，我们的新的Ext1条件性基因敲除小鼠(Col2a1-CreERT；Ext1flx/flx)，它基于一小部分软骨细胞中随机纯合的Ext1缺失，在长骨中以100%的外显率发展成外骨瘤，并表现出人类HME的其他骨骼缺陷，其程度在任何以前的突变小鼠中都没有见过。这一竞争性修订建议增加第四个目标(使用新的HME小鼠模型来研究HME的遗传和细胞机制)来表征这些突变的小鼠。通过这些研究，我们希望建立Col2a1-CreERT；Ext1flx/Flox小鼠作为第一个与HME直接相关的小鼠模型，确定杂合性缺失在HME发病机制中的意义，并为进一步研究骨软骨瘤发生发展的信号缺陷提供基础信息。 公共卫生相关性：硫酸乙酰肝素对正常的骨骼发育是必不可少的，人类遗传性骨病遗传性多发性外生症(HME)的存在就证明了这一点，这种疾病是由硫酸乙酰肝素生物合成所必需的基因突变引起的。在这次竞争性修订中，我们将描述我们的新型条件性基因敲除小鼠模型的特征，该模型表现出与人类HME前所未有的表型相似性。拟议的研究将建立一种新的HME小鼠模型，并对这种人类疾病的致病机制产生新的见解。

DESCRIPTION (provided by applicant): This competitive revision application is in response to NOT-OD-09-058: NIH Announces the Availability of Recovery Act Funds for Competitive Revision Applications. Heparan sulfate (HS) plays essential roles in development of various tissues. To understand how HS functions during skeletal development is of a great clinical importance. Hereditary multiple exostoses (HME) is a genetic disorder characterized by the formation of multiple osteochondromas ("exostoses"), which affects several thousands of people in the US. Individuals with HME carry heterozygous mutations of EXT1 or EXT2, which jointly encode a glycosyltransferase essential for the biosynthesis of HS. Despite the identification of its causative genes, there are a number of enigmas and unanswered questions on the pathogenic mechanism of HME. One of the most puzzling questions is the stark discrepancy between human HME manifestations and the phenotype of mice carrying the same genotype. Most importantly, both Ext1 and Ext2 heterozygous mice, which are supposed to mimic human HME faithfully, are resistant to the development of exostoses, especially in long bones. We have recently made a surprising observation that our new Ext1 conditional knockout mice (Col2a1- CreERT;Ext1flox/flox), which is based on stochastic homozygous Ext1 deletion in a small fraction of chondrocytes, develop exostoses in long bones at a 100% penetrance and phenocopy other skeletal defects of human HME to a degree not seen in any of the previous mutant mice. This competitive revision proposes to add the fourth aim (Investigate the genetic and cellular mechanisms of HME using novel HME mouse models) to characterize these mutant mice. By these studies, we wish to establish the Col2a1-CreERT;Ext1flox/flox mouse as the first mouse model directly relevant to HME, determine the significance of loss-of-heterozygocity in the pathogenesis of HME, and obtain baseline information for the future study to elucidate the signaling defects underlying osteochondroma development. PUBLIC HEALTH RELEVANCE: Heparan sulfate is essential for normal bone development, as illustrated by the existence of the human genetic bone disorder hereditary multiple exostosis (HME), which is caused by mutations of genes essential for heparan sulfate biosynthesis. In this competitive revision, we will characterize our novel conditional knockout mouse model that exhibits unprecedented levels of phenotypic similarities with human HME. The proposed studies will establish a novel mouse model for HME and generate new insights into the pathogenic mechanism of this human disorder.