Discovery and characterisation of novel myokines using innovative proteomic analysis

肥胖与代谢紊乱的发展有关，包括胰岛素抵抗和II型糖尿病。尽管运动在预防和治疗这些疾病方面具有已知的功效，但这种治疗方法和对所涉及机制的研究却令人惊讶地被忽视了。近年来，在宿主实验室和其他实验室的工作中，产生了肌因子的概念，肌因子是一种由收缩骨骼肌产生和释放的蛋白质，以协调运动对代谢紊乱的有益影响。自从原型肌因子IL-6的鉴定以来，已经描述了选择数量的肌因子，但通常是偶然发现的。因此，本研究旨在鉴定骨骼肌在运动时分泌的新型肌因子，重要的是，确定它们的生物学相关性。为此，在第一步中，将运动小鼠的蛋白质组与休息小鼠的蛋白质组进行比较。为了便于体内检测释放的肌因子，将采用创新的蛋白质组稳定同位素标记细胞培养物中的氨基酸（SILAC）加标方法，使用在宿主实验室进行的体外和体内SILAC标记实验制备的肌肉裂解物。然后，通过生物信息学分析来询问表现出运动反应的蛋白质的分泌序列。然后通过代谢筛选实验在体外和体内评估推定的肌因子对代谢功能的意义。该研究项目确定的推定新型肌因子将提供治疗靶点，并有助于了解运动对疾病状况的有益影响，如胰岛素抵抗和II型糖尿病。

Obesity is associated with the development of metabolic disorders, including insulin resistance and type II diabetes. Despite the known efficacy of exercise in preventing and treating these conditions, this therapeutic approach and research into the mechanisms involved, is surprisingly overlooked. In recent years, work in the host laboratory and others, has given rise to the concept of the myokine, a protein produced and released from contracting skeletal muscle in order to orchestrate the beneficial effects of exercise on metabolic disorders. Since the identification of the proto-typical myokine IL-6, a select number of myokines have been described, but often discovered serendipitously. Hence, this study aims to identify novel myokines which are secreted by the skeletal muscle in response to exercise and importantly, determine their biological relevance. To this end, in a first step the proteome of an exercising mouse will be compared with the proteome of a rested mouse. To facilitate the in vivo detection of released myokines, the innovative proteomic stable isotope labelling with amino acids in cell culture (SILAC) spike-in approach will be adopted, using prepared muscle lysates from in vitro and in vivo SILAC labelling experiments carried out at the host laboratory. Proteins exhibiting an exercise response will then be interrogated for secretory sequences via bioinformatical analyses. The significance of putative myokines on metabolic function will then be assessed in vitro and in vivo by metabolic screening experiments. The putative novel myokines identified by this research project will provide therapeutic targets and help to understand the beneficial effects of exercise on disease conditions, such as insulin resistance and type II diabetes.