Role of fatty acyl-CoA synthetases in the development of lipid induced insulin resistance in skeletal muscle cells

2型糖尿病的特点是血液中葡萄糖的慢性升高，其在全球的流行率正在上升。肥胖和循环中与之相关的高水平的脂质和脂肪酸是2型糖尿病发病的关键风险因素。在2型糖尿病的发展过程中，脂质不仅储存在脂肪组织中，还储存在其他组织中，如骨骼肌。胰岛素刺激骨骼肌摄取葡萄糖，但当细胞内脂质代谢产物浓度增加时，这一功能受到损害。这种对胰岛素作用的抵抗力已被证明与活性脂肪酸和下游脂肪衍生物的浓度有关。活性脂肪酸，即酰基-辅酶A分子是由脂肪酰基-辅酶A合成酶家族产生的。我们的实验室证明，这些蛋白质在脂肪酸摄取中也起着重要作用，它们的酶活性受胰岛素信号的调节。然而，关于酰辅酶A合成酶在胰岛素抵抗和2型糖尿病发生中的作用，人们知之甚少。这项研究建议的主要假设是，酰基辅酶A合成酶的减少将防止或减轻胰岛素抵抗的发展，因为在这种条件下会产生更少的酰基辅酶A分子和其他反应性脂质代谢产物。详细分析这一过程的模型系统是将肌肉细胞分化为多核肌管。这些肌管是在体外培养的，这使得可以精确地控制条件，并在时间进程实验中监控胰岛素抵抗的发展。这些肌管是经过基因操作的，因此它们的酰辅酶A合成酶活性降低。然后通过生化分析来分析脂肪酸诱导的胰岛素抵抗的进展。哺乳动物的酰基辅酶A合成酶家族有13种不同的同工酶，先前的研究表明，它们既有重叠的功能，也有独特的功能。因此，对酰辅酶A合成酶家族蛋白的mRNA水平、蛋白质水平和酶活性的分析将被用来确定与胰岛素抵抗发生发展相关的特定酶。这种方法首先在体外系统中使用，但将通过对肥胖症和糖尿病受试者的人类肌肉活检进行分析来补充。这些研究的结果将确定酰辅酶A合成酶家族蛋白与骨骼肌中胰岛素抵抗的发展之间的相关性，并确定与这一过程相关的特定同工酶。据设想，对这些酶的药物抑制可能是未来治疗或预防2型糖尿病的一种治疗方法。

Type 2 diabetes is characterised by the chronic elevation of glucose in the blood, and its worldwide prevalence is increasing. Obesity and the associated high levels of lipids and fatty acids in the circulation are key risk factors in the pathogenesis of type 2 diabetes. During the development of type 2 diabetes, lipids are not only stored in adipose tissue but also in other tissues like skeletal muscle. Insulin stimulates the uptake of glucose into skeletal muscle, but this function is impaired when the intracellular concentration of lipid metabolites is increased. This resistance towards the action of insulin has been shown to correlate with the concentration of activated fatty acids and downstream lipid derivatives. Activated fatty acids, i.e. acyl-CoA molecules are generated by the fatty acyl-CoA synthetase family of enzymes. Our lab demonstrated that these proteins also have an important role in fatty acid uptake, and that their enzyme activity is regulated by insulin signaling. However, little is known about the function of acyl-CoA synthetases in the development of insulin resistance and type 2 diabetes. The main hypothesis of this research proposal is that a reduction of acyl-CoA synthetases will prevent or diminish the development of insulin resistance, because less acyl-CoA molecules and other reactive lipid metabolites will be produced under these conditions. The model system for the detailed analysis of this processes are muscle cells differentiated into multinucleated myotubes. These myotubes are cultivated in vitro, which allows to control conditions precisely and to monitor the development of insulin resistance in time course experiments. The myotubes are genetically manipulated so that they have a reduced acyl-CoA synthetase enzyme activity. The progression of fatty acid-induced insulin resistance is then analysed by biochemical assays. There are thirteen different mammalian isoenzymes of the acyl-CoA synthetase family, and previous research has shown that they have overlapping but also unique functions. Therefore, analysis of the acyl-CoA synthetase family proteins on the level of mRNA, protein and enzyme activity will be used to identify the specific enzymes relevant for the development of insulin resistance. This approach is first used on the in vitro system, but will be complemented by the analysis of human muscle biopsies from obese and diabetic subjects. The results from these studies will define the correlation between the acyl-CoA synthetase family proteins and the development of insulin resistance in skeletal muscle, and identify the specific isoenzymes relevant for this process. It is envisaged that pharmacological inhibition of these enzymes could be a future therapeutic approach in the treatment or prevention of type 2 diabetes.