MOUSE MODELS TO DELINEATE A UNIQUE METABOLIC AND SKELETAL NETWORK

描述（由申请人提供）：本提案的主要目标是使用我们实验室创建的小鼠模型，表征小鼠染色体（Chr）6中部区域两个相邻基因多态性对骨骼和代谢的影响。我们最初发现了一个数量性状位点（QTL），强烈影响峰值骨收购和IGF-I在两个近交系（C3 H/HeJ[C3 H]和C57 BL/6[B6]）之间的交叉。我们随后创建了同源小鼠B6.C3H-6 T（即6 T），其中该基因座被置于B6背景上。6 T雌性小鼠具有显著的骨骼和代谢表型，包括非常低的峰值骨量、骨形成减少、低血清IGF-I、抑制的瘦素浓度、显著的骨髓和肝脏肥胖以及对高脂肪喂养的胰岛素敏感性。此外，我们能够通过增加膳食脂肪摄入来挽救6 T的小梁表型。我们发现Chr 6 QTL在一个非常小的基因组区域内携带至少6个基因，这些基因参与脂肪细胞和成骨细胞的分化，我们发现其中的两个，Alox 5和Pparg，不仅在6 T骨髓基质细胞（MSC）中受到抑制，而且还携带影响其转录的多态性。此外，我们还发现了一个染色体倒位，其断裂点正好位于Alox 5的远端，位于基质分化因子-1（SDF-1）基因的高度保守区域5'附近。因此，我们假设6 T同源基因具有功能调节子，即由另一个调节基因控制下的非连续基因组组成的遗传单位，并且其参与多效性功能。为了验证这一假设，我们提出了两个具体的目标：1-对高脂饮食和用Pparg激动剂治疗后的6 T、Alox 5-/-和B6小鼠进行综合代谢和骨骼表型分析; 2-利用来自6 T的MSC、脂肪细胞和破骨细胞的体外研究描绘该调节子如何影响Wnt/B-连环蛋白和IGF-I网络，高脂饮食和Pparg激动剂的Alox 5-/-和B6小鼠。在第二个目标中，我们还将使用TOPGAL 6 T和B6报告小鼠确定Wnt/B-连环蛋白系统的体内活性。 与公共卫生的相关性成功完成这一提案将为独特的代谢和骨骼网络提供新的线索，也将为膳食脂肪和骨髓肥胖与骨获得的关系提供重要的见解。更重要的是，这项工作可能会导致我们在小鼠和人类中识别骨质疏松症基因的一般方法发生巨大变化。

DESCRIPTION (provided by applicant): The primary goal of this proposal is to characterize the skeletal and metabolic effects of polymorphisms in two adjacent genes in the mid-region of mouse chromosome (Chr) 6, using mouse models created in our laboratory. We initially found a quantitative trait locus (QTL) that strongly influenced peak bone acquisition and IGF-I in a cross between two inbred strains (C3H/HeJ[C3H], and C57BL/6[B6]). We subsequently created a congenic mouse B6.C3H-6T (i.e. 6T) in which this locus was placed on a B6 background. 6T female mice had a remarkable skeletal and metabolic phenotype including very low peak bone mass, reduced bone formation, low serum IGF-I, suppressed leptin concentrations, significant marrow and hepatic adiposity, and insulin sensitivity in response to high fat feeding. In addition, we were able to rescue the trabecular phenotype of 6T by increasing dietary fat intake. We found that the Chr 6 QTL carried at least 6 genes within a very small genomic region that were involved in both adipocyte and osteoblast differentiation, and we showed two of these, Alox5 and Pparg, not only were suppressed in 6T marrow stromal cells (MSCs) but also carried polymorphisms which affected their transcription. Furthermore, we discovered a chromosomal inversion with a break point just distal to Alox5, in the vicinity of a highly conserved region 5' to the stromal differentiation factor-1 (SDF-1) gene. Hence we postulate that the 6T congenic has a functioning regulon, i.e. a genetic unit composed of a non-contiguous group of genes under the control of another regulatory gene, and which is involved in pleiotropic functions. To test that hypothesis, we propose two specific aims: 1-comprehensive metabolic and skeletal phenotyping of 6T, Alox5-/- and B6 mice on high fat diets and after treatment with Pparg agonists; 2-delineation of how this regulon influences the Wnt/B-catenin and IGF-I networks utilizing in vitro studies of MSCs, adipocytes and osteoclasts from 6T, Alox5-/- and B6 mice on high fat diets and Pparg agonists. In this 2nd aim we will also determine the in vivo activity of the Wnt/B-catenin system using TOPGAL 6T and B6 reporter mice. Relevance to Public Health Successful completion of this proposal will shed new light on a unique metabolic and skeletal network, and will also provide important insight into the relationship of dietary fat and marrow adiposity to bone acquisition. More importantly, this work could lead to a sea change in our general approach to identifying osteoporosis genes in mice and humans.