Roles of GDF3 in the differentiation and function of the adipocyte

描述（申请人提供）：TGF-（超家族信号）调控体外前脂肪细胞的确定和分化。这些分泌的配体在刺激和抑制脂肪形成中起作用。骨形态发生蛋白（BMPs） 2、4和7刺激了前体细胞系向脂肪细胞的确定和随后的分化。TGF-超家族成员GDF8/肌生长抑制素可抑制bmp7诱导的脂肪细胞分化。TGF-(在体内和体外均对脂肪生成具有拮抗作用。我们假设TGF-超家族成员GDF3也在调节脂肪形成和/或影响成熟脂肪细胞的功能中发挥重要作用。首先，GDF3在脂肪组织中表达，并在白色脂肪库中显著上调，以应对高脂肪饮食，脂肪细胞和间质血管部分不成比例地增加。其次，体内通过腺病毒转导GDF3的过度表达导致对高脂肪饮食的成脂作用更敏感，这反映在体重增加、脂肪组织质量、瘦素水平升高和脂肪细胞大小增大上。相反，我们对Gdf3-/-小鼠的研究表明，尽管摄入了更多的热量，但Gdf3-/-小鼠对饮食诱导的肥胖具有保护作用，包括正常的身体和脂肪组织质量和脂肪细胞大小。因此，GDF3可能参与调节脂肪形成和/或成熟脂肪细胞的功能。该应用程序的目的如下：具体目的1-检查GDF3对细胞增殖，存活和分化的影响。我们将研究外源性GDF3蛋白对两种脂肪形成细胞系的增殖、有丝分裂克隆扩增、凋亡和分化的影响。特异性目的2-评估GDF3对成熟脂肪细胞功能的影响。将分析完全分化的3T3-L1细胞，以确定外源性GDF3蛋白给药是否对脂肪生成、脂肪分解和其他成熟脂肪细胞功能有影响。具体目标3-定义脂肪细胞中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。我们将进一步确定GDF3与其受体的相互作用以及与影响脂肪形成的其他超家族成员的相互作用。具体目的4-研究体内脂肪组织中GDF3过表达的后果。我们将在强力霉素的可逆控制下，产生在脂肪组织中选择性表达GDF3的转基因小鼠。

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.