基于“脾主肌肉”理论研究益气健脾活血法调控mTOR信号通路干预衰老糖尿病大鼠肌量减损的分子机制

Sarcopenia is a geriatric syndrome with high morbidity, which induces tumble, bone fracture as well as loss of life independence, decreases quality of life and expected lifespan, increases hospital admission rate and mortality. Diabetes accelerates the process of sarcopenia. Nowadays, pharmaceutical intervention for diabetic sarcopenia is inadequate, and some anti-diabetic medicine may be detrimental to the health of skeletal muscles. The “spleen governing muscle” theory in Traditional Chinese Medicine suggests the possibility of treating this disease from the “spleen”. Chinese herbal medicine may become the new therapy for the treatment of diabetic sarcopenia. In the previous National Science Found Research (No.8150151388), it is found that ShenQi compound with invigorating Qi and spleen with activating blood circulation may be helpful in maintaining muscle mass, which induces differential gene expression in mTOR signaling network. It is well known that the balance between anabolism and catabolism is determinant for muscle mass, however, mTOR signal pathway gathers and integrates multiple cell stimulate factors to regulate protein metabolism. Therefore, it is presumed that ShenQi compound has effects on protein metabolism via modulating mTOR signaling network, thus prevent muscle loss in diabetic rats. However, the mechanism is unclear. This research will focus on protein metabolism to study the effect of invigorating Qi and spleen with activating blood circulation therapy on prevention of muscle mass loss in geriatric diabetic rats. Futhermore, The mTOR signaling phospho antibody array and western blot technology will be used to explore the potential molecular mechanism.

肌少症是一种患病率高且危害巨大的老年综合症，导致跌扑、骨折、失能等，降低生活质量及预期寿命，增加再入院率及死亡率。糖尿病加速肌少症的进程，目前缺乏有效的药物治疗手段，且部分降糖药有损于骨骼肌健康。中医“脾主肌肉”理论提示从脾论治该病的可能性，中医药有望成为糖尿病性肌少症治疗的新方向。申请人在既往自然科学基金项目（No.8150151388）中发现益气健脾活血之参芪复方有助于保持肌量，且引起mTOR信号通路差异基因表达；肌量取决于蛋白质合成与分解代谢之间的平衡，而mTOR信号通路汇聚和整合多种细胞刺激信号调控蛋白质代谢。因此，推测益气健脾活血法保持肌量的作用与调控mTOR信号通路、影响蛋白质代谢有关，但具体机制不明。本研究拟以蛋白质代谢为入口，运用mTOR信号通路磷酸化抗体芯片、蛋白免疫印迹等技术，阐释益气健脾活血法调控mTOR信号通路、影响蛋白质代谢、干预衰老糖尿病大鼠肌量减损的分子机制。

肌少症是一种患病率高且危害大的老年综合症，增加跌扑、骨折、失能等风险，降低生活质量及预期寿命，增加再入院率及死亡率。目前缺乏有效的药物治疗手段，糖尿病加速肌少症的进程且部分降糖药有损于骨骼肌健康。中医治疗“痿症”理论及经验丰富，有望成为肌少症治疗的新选择。既往研究证明参芪复方具有良好的糖调节作用，同时该方改善老年虚劳、痿症、疲乏等也疗效确切。因而本课题深入探讨参芪复方改善衰老/糖尿病相关性肌少症的效应及机制。肌量是肌少症的核心诊断指标，取决于蛋白质合成与分解代谢之间的动态平衡，而mTOR信号通路汇聚和整合多种细胞刺激信号调控蛋白质代谢。本研究以蛋白质代谢为切入点，运用mTOR信号通路磷酸化抗体芯片、蛋白免疫印迹、免疫荧光/组化、RT-qPCR、透射电镜等技术，研究中药复方对衰老/糖尿病大鼠骨骼肌组织、细胞、分子层次的效应及机制。研究内容包括肌量、肌力及肌功能；血糖、胰岛素抵抗指数、氧化应激指标；蛋白质合成/分解代谢信号通路关键靶点表达；mTOR信号网络磷酸化位点表达；myostatin、PAX 7及PDGFR-a表达等。研究发现，中药复方改善血糖稳态、胰岛素抵抗以及氧化应激指标，提升衰老/糖尿病大鼠运动能力，有利于维持骨骼肌纤维横截面积、减少骨骼肌组织炎性浸润等；上调肌浆蛋白浓度及肌卫星细胞PAX 7表达、抑制Myostatin表达；mTOR磷酸化抗体芯片分析提示AKT1 (Phospho-Thr450)、RSK1/2/3/4 (Phospho-Ser221/227/218/232)、P70S6K (Phospho-Thr229)、PTEN (Phospho-Ser370)是参芪复方潜在作用靶点；抑制肌间纤维及脂质沉积，下调纤维/脂肪祖细胞PDGFR-a和纤维化相关靶点mRNA表达，提示其参与调控细胞外基质重塑、为后续研究奠定了基础。本课题确证参芪复方干预衰老/糖尿病骨骼肌减损的多靶点调控作用，尤其是通过抑制myostatin表达、调控mTOR信号网络下游蛋白质合成信号通路，延缓衰老及糖尿病病理状态下肌量丢失、阻遏肌少症发生发展。本课题阐释经典中医理论、明确中医药干预肌少症的科学依据。