Role of Gs-alpha in regulation of adipose tissue growth and function

Gs-α 在调节脂肪组织生长和功能中的作用

• 批准号: 8349770
• 负责人: Lee Weinstein
• 金额: $ 16.57万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8349770
• 关键词: Acute; Adipocytes; Adipose tissue; Adrenergic Agents; Alleles; Body Temperature; Brown Fat; CCAAT-Enhancer-Binding Protein-beta; Catecholamines; Cells; Cyclic AMP; Diet; Embryo; Energy Metabolism; Environment; Exons; Fatty acid glycerol esters; Fibroblasts; GTP-Binding Protein alpha Subunits, Gs; Genes; Genetic Recombination; Growth; Gs alpha mutations; Heart; Human; Hypoglycemia; In Vitro; Insulin Resistance; Insulin-Like Growth Factor I; Knock-out; Lead; Lipids; Lipolysis; Liver; Mediating; Metabolic Activation; Modeling; Morbid Obesity; Mus; Muscle; Normal Cell; Obesity; Pathway interactions; Pattern; Phenotype; Production; Regulation; Relative (related person); Role; Signal Pathway; Site; Skeletal Muscle; Sympathetic Nervous System; Thermogenesis; Tissues; Transgenic Mice; Urine; Weight Gain; adiponectin; adrenergic; base; beta-adrenergic receptor; gene induction; glucose tolerance; improved; in vivo; insulin sensitivity; lipid biosynthesis; lipid metabolism; programs; recombinase; response; uncoupling protein 1

We generated mice with Gs-alpha deficiency in adipose tissue (FGsKO mice) by repeated matings of aP2-cre recombinase transgenic mice with floxed Gs-alpha mice which have loxP recombination sites surrounding Gs-alpha exon 1. Results to date show that FGsKO mice had poor survival and decreased linear growth, particularly those mice in which Gs-alpha expression in adipose tissue was severely reduced. The cause of these effects are unclear, although insulin-like growth factor 1 (IGF1) levels were reduced by 50% in FGsKO mice. In this subset of mice, white adipose tissue (WAT) pads were almost absent. Fibroblasts from FGsKO embryos had significantly reduced adipogenic conversion in the presence of agents known to induce adipogenesis. These in vivo and in vitro results confirm that Gs-alpha is critical for normal adipogenesis. FGsKO mice which survived had also had reduced relative fat mass with smaller WAT pads and smaller adipocytes with less lipid content per cell. In contrast, the interscapular brown adipose tissue (BAT) pads were larger than normal and the cells had increased lipid stores with a more unilocular distribution. This histological pattern in BAT is consistent with reduced metabolic activation presumably due to an inability of the sympathetic nervous system to stimulate lipolysis via beta-adrenergic/Gs-alpha pathways. Consistent with this, the expression of PGC-1, uncoupling protein 1 (UCP1) and other genes associated with lipid metabolism were markedly reduced in BAT from FGsKO mice. FGsKO mice were hypoglycemic and hypoinsulinemic relative to controls and had improved glucose tolerance and insulin sensitivity on both regular and high-fat diets, consistent with their lean phenotype. Two forms of adaptive thermogenesis, cold- and diet-induced thermogenesis, are mediated by increased sympathetic nervous system activity. FGsKO mice are cold intolerant and cold-induced thermogenesis is proabably markedly impaired as FGsKO mice placed in a cold environment do not maintain their body temperature or raise their expression of UCP1 in BAT. This is consistent with the known role for BAT in cold-induced thermogenesis and the results in FGsKO mice that BAT fails to be activated by sympathetic stimulation despite the fact that their sympathetic activity, as determined by urine catecholamine levels, was markedly increased. In contrast, diet-induced thermogenesis is maintained and in fact greater than normal in FGsKO mice based upon the observations that FGsKO mice fail to gain weight on a high-fat diet and have markedly increase their energy expenditure on a high-fat diet. These results suggest that cold- and diet-induced thermogenesis can occur in separate tissues and we propose that muscle is the main site for diet-induced thermogenesis in these mice. To further examine this we looked at sympathetic activity and PGC-1alpha induction after either acute cold or high fat diet in control mice. In response to high fact diet sympathetic activity only increased in skeletal muscle, with no change in heart, liver, or brown fat. In contrast sympathetic activity increased in all tissues after acute cold exposure. PGC-1alpha was markedly induced in BAT after cold exposure and much less so after high fat diet. Finally, these results as well as results in heterozygous FGsKO mice suggest that adipose tissue is not the site whereby germline Gs-alpha mutations on the maternal allele lead to severe obesity and insulin resistance. We are presently generating a Ucp1-cre transgenic mice which if successful will allow us to generated BAT-specific Gs-alpha mice. In addition we are trying to generate more WAT-specific Gs-alpha knockout models using other cre lines such as adiponectin-cre transgenic mice.

我们通过将 aP2-cre 重组酶转基因小鼠与 floxed Gs-alpha 小鼠重复交配，产生了脂肪组织中 Gs-alpha 缺乏的小鼠（FGsKO 小鼠），floxed Gs-alpha 小鼠在 Gs-alpha 外显子 1 周围具有 loxP 重组位点。迄今为止的结果表明，FGsKO 小鼠的存活率较差且线性生长下降，特别是那些 Gs-alpha 表达的小鼠 脂肪组织中的含量严重减少。尽管 FGsKO 小鼠中胰岛素样生长因子 1 (IGF1) 水平降低了 50%，但造成这些影响的原因尚不清楚。在这组小鼠中，白色脂肪组织（WAT）垫几乎不存在。在已知诱导脂肪生成的试剂存在下，来自 FGsKO 胚胎的成纤维细胞显着降低了脂肪生成转化。这些体内和体外结果证实，Gs-α 对于正常脂肪生成至关重要。存活的 FGsKO 小鼠的相对脂肪量也减少了，WAT 垫更小，脂肪细胞也更小，每个细胞的脂质含量也更少。相比之下，肩胛间棕色脂肪组织（BAT）垫比正常细胞大，并且细胞的脂质储存增加，且分布更加单房。 BAT 中的这种组织学模式与代谢激活减少相一致，这可能是由于交感神经系统无法通过 β-肾上腺素能/Gs-α 途径刺激脂肪分解所致。与此相一致的是，FGsKO 小鼠的 BAT 中 PGC-1、解偶联蛋白 1 (UCP1) 和其他与脂质代谢相关的基因的表达显着降低。与对照组相比，FGsKO 小鼠出现低血糖和低胰岛素血症，并且在常规和高脂肪饮食中葡萄糖耐量和胰岛素敏感性均得到改善，这与它们的瘦表型一致。适应性产热的两种形式，寒冷和饮食诱导的产热，是由交感神经系统活动增加介导的。 FGsKO 小鼠不耐受寒冷，并且寒冷诱导的生热作用可能明显受损，因为置于寒冷环境中的 FGsKO 小鼠无法维持体温或提高 BAT 中 UCP1 的表达。这与 BAT 在冷诱导产热中的已知作用一致，并且 FGsKO 小鼠的结果表明，尽管根据尿液儿茶酚胺水平确定，BAT 的交感活性显着增加，但 BAT 未能被交感神经刺激激活。相比之下，基于FGsKO小鼠在高脂肪饮食中体重未增加并且在高脂肪饮食中能量消耗显着增加的观察结果，饮食诱导的生热作用在FGsKO小鼠中得以维持，并且实际上高于正常水平。这些结果表明，寒冷和饮食诱导的生热作用可以发生在不同的组织中，我们认为肌肉是这些小鼠中饮食诱导的生热作用的主要部位。为了进一步研究这一点，我们观察了对照小鼠在急性寒冷或高脂肪饮食后的交感神经活动和 PGC-1α 诱导。为了应对高事实饮食，交感神经活动仅在骨骼肌中增加，而心脏、肝脏或棕色脂肪没有变化。相反，在急性寒冷暴露后，所有组织的交感神经活动均增加。 BAT 中的 PGC-1α 在寒冷暴露后显着被诱导，而在高脂肪饮食后则少得多。最后，这些结果以及杂合 FGsKO 小鼠的结果表明，脂肪组织并不是母体等位基因上种系 Gs-α 突变导致严重肥胖和胰岛素抵抗的部位。我们目前正在培育 Ucp1-cre 转基因小鼠，如果成功，我们将能够培育出 BAT 特异性 Gs-alpha 小鼠。 此外，我们正在尝试使用其他 cre 系（例如脂联素-cre 转基因小鼠）生成更多 WAT 特异性 Gs-alpha 敲除模型。