Myostatin功能抑制时机体通过mTOR和AMPK对话调控能量再分配的机制研究

Myostatin is an important member of the energy metabolism network, its functional signals in close contact with the body's two energy sensors mTOR and AMPK. But the regulatory mechanism is unclear among them, especially, little is known about the signaling networks when myostatin function is inhibitted.How to jointly coordinate the body's energy redistribution through interaction has not yet commenced research.Myostatin is a negative regulator of skeletal muscle growth because myostatin deficiency causes increased muscle mass in several species, myostatin-null mice also have reduced adiposity, with decreased adipocyte cell size and number. However, myostatin overexpression in mouse adipose tissue not only decreases adipocyte size, but also confers resistance to diet-induced obesity, consistent with its antiadipogenic roles in vitro. Systemic overexpression, however, achieved by im injection of myostatin-producing CHO cells, results in reduced skeletal muscle mass and a nearly complete loss of WAT. Similarly, sc injection of myostatin significantly reduces adiposity, but does not affect muscle mass or total body weight, possibly due to differences in the levels of bioactive protein in the two models.whereas inhibition of myostatin signaling in only adipose tissue did not affect body composition. This indicates that the regulation of adiposity in myostatin null models is likely to be secondary to the metabolic changes associated with increased skeletal muscle mass. So we presume they may jointly coordinate the body's energy balance through Cross-talk among myostatin and AMPK and mTOR.The project intends to prove how mTOR and AMPK signaling pathway to transmit signal in the energy metabolism regulation of skeletal muscle and adipose tissue of lack of myostatin function on two levels from the cells and individual, through the body's immune mice and in vitro C2C12 and 3T3L-1 co-cultured in Transwell method, on the level of signaling networks myostatin regulation of body's skeletal muscle and adipose tissue systemic energy metabolism. We want to verify how to interact between the two pathways mTOR and AMPK, simultaneously to find the differences of time course and effect in each signal. so we clarify the mechanism of myostatin regulation of energy metabolism through expliciting network interaction of these two signaling pathways while the lack of myostatin function. Further we could improve the content of the body skeletal muscle and adipose tissue of the myostatin regulation of energy metabolism network system.It can provide a reliable theoretical basis, For we appropriately use of myostatin inhibitors and other drugs, or by means of transformation of myostatin and other. It may also improve the economic benefits of animal husbandry, and treat muscle atrophy, obesity and type 2 diabetes.

Myostatin作为能量代谢网络的重要成员，与机体两大能量感受器mTOR和AMPK联系密切，但在整体信号网络水平三者之间如何协调仍不清楚，特别是myostatin功能抑制时它们如何通过交互作用共同协调机体能量再分配尚未见文献报道。本项目拟从细胞和个体两个层面，通过建立成肌细胞与前体脂肪细胞Transwell共培养体系和体内注射Myostatin无活性蛋白获得免疫小鼠，探索myostatin功能抑制时mTOR和AMPK通路如何相互传递信号、各信号发生的时间历程及效应强弱的差异，从而勾勒出myostatin介导的这两个信号通路可能的交互网络关系与引发的级联反应线路，以便深入了解myostatin对机体能量代谢的调控机制，丰富其调控机体能量代谢网络体系内容。为正确利用myostatin抑制剂或通过改造myostatin等手段，提高畜牧业经济效益和治疗肌萎缩、肥胖症以及2型糖尿病等提供理论基础。

Myostatin能够负调控骨骼肌生长，但其在脂肪发育和机体代谢中的作用，至今未取得明确的结果。1、本课题组在用Transwell 培养法建立C2C12细胞与3T3-L1 细胞最佳的共培养模式的过程中，探索了不同的共培养方式对两类细胞生长增值的影响，总结出一种体外细胞接触式共培养装置及其培养操作方法，已提出申请专利1项。2、通过构建载体，诱导表达了Myostatin重组蛋白，利用乳化抗原主动免疫新西兰雄兔制抗血清，通过ELISA测定抗体应答水平；用纯化的Myostatin无活性蛋白免疫小鼠，可明显增加处理组及其后代的体重；同时探索了微波辅助SDS-PAGE胶检测蛋白的染色方法，发明了一种具有广泛用途的小鼠心脏采血装置（发表SCI论文2篇、获授权发明专利1项）。 3、发现Myostatin基因外显子1的267碱基发生A→G的同义突变，在所检测牛群中存在3种基因型(AA、GG、AG)，A等位基因为优势基因。该位点在4个牛群体中均处于中度多态。卡方检验表明皖东牛、皖南牛和和牛处于哈代-温伯格平衡状态(P>0.05)，西门塔尔牛处于哈代-温伯格不平衡状态(P>0.05)。（发表论文1篇）。 4、我们发现，在冻存C2C12细胞与3T3-L1细胞时，抑制微丝蛋白G-actin组装可提高细胞的冷冻复苏率（发表SCI论文2篇，提交会议论文1篇）。5、Rho/Rho激酶信号转导通路是体内普遍存在的一条信号转导通路，参与多种细胞功能。本课题组总结了Rho激酶与脂肪细胞分化的研究进展；并研究了不同浓度羊栖菜多糖对秀丽隐杆线虫脂肪代谢的影响（发表论文2篇）。 总之，通过课题组的共同努力，初步勾勒出了Myostatin介导的这两个信号通路可能的交互网络关系与引发的级联反应线路，深入地了解了Myostatin对机体能量代谢的调控机制，丰富了其调控机体能量代谢网络体系内容。为进一步正确利用Myostatin抑制剂或通过改造Myostatin等手段，提高畜牧业经济效益和治疗肌萎缩、肥胖症以及2型糖尿病等提供了理论基础。