Mouse Models to Delineate a Unique Metabolic and Skeletal Network

描述(由申请人提供)：这项建议的主要目标是利用我们实验室创建的小鼠模型来表征小鼠染色体(Chr)6中间区两个相邻基因的多态对骨骼和代谢的影响。我们最初在两个近交系(C3H/HeJ[C3H]和C57BL/6[B6])的杂交中发现了一个对峰值骨获取和IGF-I有强烈影响的数量性状基因座(QTL)。随后，我们创造了一个同源小鼠B6.C3H-6T(即6T)，其中该基因座被放置在B6背景上。6T雌性小鼠具有显著的骨骼和代谢表型，包括极低的峰值骨量，骨形成减少，血清IGF-I水平降低，瘦素水平受到抑制，骨髓和肝脏肥胖症显著，以及对高脂饮食的胰岛素敏感性。此外，我们能够通过增加膳食脂肪摄入量挽救6T的骨小梁表型。我们发现Chr6QTL在一个很小的基因组区域内携带至少6个基因，参与脂肪细胞和成骨细胞的分化，其中两个基因Alox5和PPARG不仅在6T骨髓基质细胞(MSCs)中受到抑制，而且携带有影响其转录的多态。此外，我们还发现了一个染色体倒位，其断裂点正好位于Alox5的远端，位于基质分化因子-1(SDF-1)基因5‘端的高度保守区域附近。因此，我们推测6T同源基因有一个功能调节子，即在另一个调控基因的控制下，由一组不连续的基因组成的遗传单位，它参与了多效性功能。为了验证这一假设，我们提出了两个特定的目标：1-在高脂饮食和PPARG激动剂治疗后，6T，ALOX5-/-和B6小鼠的全面代谢和骨骼表型；2-利用6T，ALOX5-/-和B6小鼠在高脂饮食和PARG激动剂治疗下的MSCs、脂肪细胞和破骨细胞的体外研究，描绘该调节基因如何影响Wnt/B-catenin和IGF-I网络。在第二个目的中，我们还将使用TOPGAL 6T和B6报告小鼠来确定Wnt/B-catenin系统的体内活性。 与公共健康的相关性这项提案的成功完成将为了解一个独特的新陈代谢和骨骼网络提供新的线索，并将为了解饮食脂肪和骨髓脂肪与骨骼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身上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.