Roles of Gs Alpha and interaction with Wnt signaling in regulation of adipocyte d

描述（由申请人提供）：脂肪细胞和成骨细胞可能共享驻留在骨髓中的共同前体。这是由临床观察和动物研究模型表明，骨髓肥胖和骨密度之间的反比关系。阐明骨髓基质细胞如何分化成脂肪细胞和成骨细胞的机制可能对代谢性疾病如肥胖症和骨疾病如骨质疏松症产生影响。我们的研究集中在Gsa，一个异源三聚体G蛋白亚基，介导的环AMP依赖性信号的G蛋白偶联受体，包括甲状旁腺激素（PTH）/PTH相关肽（PTHrP）受体（PPR）在这一机制中的潜在作用。PPR信号传导被认为促进成骨细胞分化，并且还可以抑制脂肪细胞分化。这些作用可能部分由经典Wnt信号传导介导。该实验室的研究显示，在成骨细胞谱系早期靶向缺失Gsa的小鼠（Osterix-Cre：Gsa（fl/fl））在出生时表现出骨质疏松症和多发性骨折，沿着骨髓肥胖增加。该实验室先前报告称，Osterix-Cre：Gsa（fl/fl）小鼠骨细胞分泌的典型Wnt信号传导抑制剂sclerostin的表达增加，Wnt靶基因的基因表达减少。基于这些观察结果，我们假设Gsa在早期成骨细胞谱系的细胞中表达，在成骨细胞和脂肪细胞分化的调节中起作用，并且这种作用部分由经典Wnt信号传导介导。使用体外集落形成单位测定的初步研究表明，在成骨细胞谱系早期靶向缺失Gsa导致总祖细胞和骨祖细胞数量减少，但不改变脂肪祖细胞的数量。然而，问题仍然存在，来自Osterix-Cre：Gsa fl/fl小鼠的细胞的成脂潜力增加是否是由于脂肪细胞祖细胞数量增加或Gsa缺失成骨细胞转化为脂肪细胞。特定目的I的实验通过使用双荧光报告基因膜靶向串联二聚体番茄/绿色荧光蛋白来鉴定表达Osterix：Cre的脂肪细胞，从而推测其最初为成骨细胞谱系，从而解决了这一问题。Specific Aim II的实验解决了Wnt信号在早期成骨细胞中Gsa信号如何调节脂肪细胞和成骨细胞分化中的潜在作用。首先，这些实验将研究Osterix-Cre：Gsa小鼠中Wnt活性是否降低。其次，我们将研究早期成骨细胞中的Gsa信号传导是否与β-连环蛋白稳定化上游的Wnt信号传导相互作用。该问题将通过测试子宫内锂暴露对Osterix-Cre：Gsa fl/fl小鼠骨量、骨髓肥胖以及脂肪祖细胞和骨祖细胞相对数量的影响来解决。 公共卫生相关性：这项研究的重点是脂肪细胞和骨细胞如何从骨髓中发育的机制。这可能有助于改善对肥胖和骨质疏松症等疾病的理解和治疗。

DESCRIPTION (provided by applicant): Adipocytes and osteoblasts may share a common precursor residing in the bone marrow. This is suggested by clinical observations and animal research models which demonstrate an inverse relationship between bone marrow adiposity and bone density. Elucidation of the mechanisms governing how bone marrow stromal cells differentiate into adipocytes and osteoblasts may carry implications for metabolic diseases such as obesity and of bone diseases such as osteoporosis. Our studies focus on the potential role in this mechanism of Gsa, a heterotrimeric G protein subunit that mediates cyclic AMP-dependent signaling of G protein-coupled receptors including the parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptor (PPR). PPR signaling is believed to promote osteoblast differentiation and may also suppress adipocyte differentiation. These actions may be mediated in part by canonical Wnt signaling. Studies from this laboratory revealed that mice with targeted deletion of Gsa early in the osteoblast lineage (Osterix-Cre:Gsa(fl/fl)) exhibit osteoporosis and multiple fractures at birth along with increased bone marrow adiposity. This laboratory previously reported that Osterix-Cre:Gsa(fl/fl) mice have increased expression of sclerostin, an inhibitor of canonical Wnt signaling secreted by osteocytes, and decreased gene expression of Wnt target genes. Based on these observations, we hypothesized that Gsa expressed in cells of the early osteoblast lineage has a role in regulation of osteoblast and adipocyte differentiation, and that this role was mediated in part by canonical Wnt signaling. Preliminary studies using in vitro colony forming unit assays suggested that targeted deletion of Gsa early in the osteoblast lineage leads to fewer numbers of total progenitor cells and osteoprogenitor cells, but does not change the number of adipocyte progenitors. The question remains, however, whether the increased adipogenic potential of cells derived from Osterix-Cre:Gsa fl/fl mice is accounted for by increased adipocyte progenitor number or conversion of Gsa null osteoblasts into adipocytes. The experiments of Specific Aim I address this question by using the the double fluorescent reporter gene membrane-targeted tandem dimer tomato/green fluorescent protein to identify adipocytes that expressed Osterix:Cre and thus presumably were originally of the osteoblast lineage. The experiments of Specific Aim II address the potential role of Wnt signaling in how Gsa signaling in early osteoblasts regulates adipocyte and osteoblast differentiation. Firstly, these experiments will study whether there is decreased Wnt activity in Osterix-Cre:Gsa mice. Secondly, we will study whether Gsa signaling in early osteoblasts interact Wnt signaling upstream of b-catenin stabilization. This question will address by testing the effects of in utero lithium exposure on Osterix-Cre:Gsa fl/fl mice upon bone mass, bone marrow adiposity, and relative numbers of adipocyte progenitors and osteoprogenitors. PUBLIC HEALTH RELEVANCE: This study focuses on mechanisms underlying how fat cells and bone cells develop from the bone marrow. This may help improve understanding and treatment of diseases such as obesity and osteoporosis.