Roles of GDF3 in the differentiation and function of the adipocyte

描述(申请人提供)：转化生长因子-(超家族信号调节前体脂肪细胞的测定和分化的体外。这些分泌的配体在促进和抑制脂肪生成方面发挥作用。骨形态发生蛋白(BMPs)2、4和7可促进前体细胞向脂肪细胞的分化。BMP7诱导的脂肪细胞分化可被另一个超家族成员GDF8/myostatin抑制。在体外和体内，转化生长因子-β对脂肪生成也有拮抗作用。我们推测，GDF3，另一个转化生长因子超家族成员，也是本方案的主题，在调节脂肪形成和/或影响成熟脂肪细胞的功能方面也发挥着重要作用。首先，GDF3在脂肪组织中表达，并在白色脂肪堆积中显著上调，以响应高脂肪饮食，脂肪细胞和间质血管部分不成比例地增加。其次，通过腺病毒转导的GDF3在体内的过度表达导致对高脂肪饮食的成脂效应更敏感，表现为更大的体重增加、脂肪组织质量、更高的瘦素水平和更大的脂肪细胞尺寸。相反，我们对GDF3-/-小鼠的研究揭示了对饮食诱导的肥胖的保护作用，包括正常的身体和脂肪组织质量以及脂肪细胞大小，尽管热量摄入更多。因此，GDF3可能具有调节脂肪生成和/或成熟脂肪细胞功能的作用。应用的目的如下：特定目的1-检测GDF3对细胞增殖、存活和分化的影响。我们将检测外源GDF3蛋白对两种成脂细胞系的增殖、有丝分裂克隆扩增、凋亡和分化的影响。特异性目的2-评价甘草酸甘油三酯对成熟脂肪细胞功能的影响。将对完全分化的3T3-L1细胞进行分析，以确定外源性GDF3蛋白注射是否对脂肪生成、脂肪分解和其他成熟脂肪细胞功能有影响。具体目标3-确定脂肪细胞中GDF3信号的基本成分，并确定是否与棕色/白色脂肪细胞的命运决定有关。我们将进一步定义GDF3与其受体以及与其他影响脂肪形成的超家族成员之间的相互作用。特定目标4-检查GDF3在体内脂肪组织中过度表达的后果。我们将产生转基因小鼠，在药物多西环素的可逆控制下，在脂肪组织中选择性表达GDF3。

DESCRIPTION (provided by applicant): TGF-( superfamily signaling regulates pre-adipocyte determination and differentiation in vitro. These secreted ligands play roles in the stimulation and inhibition of adipogenesis. The determination and subsequent differentiation of precursor cell lines into adipocytes is stimulated by bone morphogenetic proteins (BMPs) 2, 4 and 7. BMP7-induced adipocyte differentiation can be inhibited by GDF8/myostatin, another TGF-( superfamily member. TGF-( is also antagonistic to adipogenesis in vitro and in vivo. We hypothesize that GDF3, another TGF-( superfamily member and the subject of this proposal, also plays an important role in regulating adipogenesis and/or in affecting the function of mature adipocytes. First, GDF3 is expressed in adipose tissues and is significantly up regulated in white adipose depots in response to high fat diets, with disproportionate increases in the adipocyte and stromal vascular fractions. Secondly, over expression of GDF3, in vivo, by adenoviral transduction results in a greater sensitivity to the adipogenic effects of high fat diets, reflected by greater weight gain, adipose tissue mass, higher leptin levels, and greater adipocyte cell size. Conversely, our studies of Gdf3-/- mice have revealed a protection from diet- induced obesity, including normal body and adipose tissue mass and adipocyte cell sizes, despite a greater caloric intake. Therefore, GDF3 may function in to regulate adipogenesis and/or mature adipocyte function. The aims of the application are as follows: Specific aim 1- Examine the effects of GDF3 on cell proliferation, survival, and differentiation. We will examine the effects of exogenous GDF3 protein on the proliferation, mitotic clonal expansion, apoptosis, and differentiation of two adipogenic cell lines. Specific aim 2- Assess GDF3's effects on mature adipocyte function. Fully differentiated 3T3-L1 cells will be analyzed to determine if exogenous GDF3 protein administration has effects on lipogenesis, lipolysis, and other mature adipocyte functions. Specific aim 3- Define the essential components of GDF3 signaling in adipocytes and determine if there is a link to brown/white adipocyte fate decisions. We will further define GDF3's interactions with its receptors and with other superfamily members that affect adipogenesis. Specific aim 4- Examine the consequences of GDF3 over expression in adipose tissues in vivo. We will generate transgenic mice which express GDF3 selectively in adipose tissues under the reversible control of the drug, doxycycline.