骨骼肌间质生肌内皮细胞（MEC）的来源、命运决定机制及在维持肌稳态中的作用

Intermyofiber progenitors within skeletal muscle are heterogeneous, which include fibro-adipogenic progenitors (FAPs), endothelial and myo-endothelial cells and other multipotent mesenchymal progenitors. These cells play important roles in maintaining skeletal muscle homeostasis and muscle regeneration, but their precise origin and function are still poorly understood. We ablated bone morphogenetic protein receptor type 1a (BMPR1a) in Myf5 progenitors and found that mutant mice displayed increased intramuscular fat and reduced myofiber size in selected muscles or following muscle injury. In skeletal muscle, a cell fraction distinct from satellite cells with bipotent differentiation potential (muscle and fat) is found responsible for the phenotype switch (muscle into fat) when BMPR1a is ablated. This cell fraction is called interstitial myo-endothelial cell (MEC). A major function of MEC in vivo is to inhibit intramuscular adipogenesis, both through a cell-autonomous mechanism and a cell–cell interaction mechanism by inhibiting FAP differentiation. In this grant proposal, we will focus on mechanistic study of MEC fate specification and the signaling pathways through which MECs inhibit intramuscular adipogenesis. Furthermore, our preliminary data have revealed a marker expressed by Myf5+BMPR1a+ interstitial MECs but not satellite cells or FAPs, and we will use the CreER mouse driven by this gene to trace the developmental origin of these cells, their contribution to developing and adult skeletal muscle and to investigate the biological significance of these cells. These studies will help to unravel many unanswered questions in the field and to develop new therapeutic strategies for treating neuromuscular diseases.

骨骼肌间质前体细胞具有异质性, 包括纤维-脂肪祖细胞(FAP)，内皮细胞和其他多功能间质干细胞，它们对维持骨骼肌稳态和损伤后修复起重要作用，但是这些前体细胞的来源和功能不完全清楚。 我们发现在小鼠Myf5表达的细胞内敲除骨形成蛋白受体1a (BMPR1a) 导致部分骨骼肌纤维变细，肌肉间质脂肪组织增生。受影响的Myf5阳性细胞不是肌卫星细胞，而是间质的生肌内皮细胞(MEC)。MEC有成肌分化和脂肪分化潜能，敲除 BMPR1a 能促使其向脂肪细胞分化，MEC还具有阻止骨骼肌内FAP向脂肪分化的功能。本课题拟运用人骨骼肌MEC特异标志物进行谱系示踪，结合基因干预、ChIP-Seq等方法，深入研究MEC的来源、命运决定机制及对维持肌稳态的作用，解析MEC抑制骨骼肌FAP脂肪分化的信号通路。本研究将为预防及治疗疾病和老龄化带来的骨骼肌内脂肪浸润提供新的理论依据，为治疗肌营养不良症等疾病提供新思路。

在哺乳动物骨骼肌发育和损伤修复过程中,有许多不同来源的成肌干细胞如骨骼肌实质中的肌卫星细胞（SC）和间质中的生肌内皮细胞（MEC）、mesoangioblast及微细血管旁周细胞等发挥重要作用。由于缺乏特异标志物，小鼠MEC无法和血管内皮细胞分开，影响了对它的进一步研究和功能开发。我们通过筛查人源MEC（可有效分离）并对其转录组进行测序分析，筛检出MEC特异表达标志物 Apelin，该蛋白也同时表达于小鼠MEC。利用AplnCreER/+：ZsGreen/tdTomato 小鼠进行谱系示踪显示Apln表达的细胞仅在Sca-1+CD31+组分中检出，而在肌卫星细胞SC和纤维脂肪祖细胞FAP中没有表达，提示Apelin是小鼠生肌内皮细胞的特异标志物，研究显示MEC参与肌纤维形成。我们发现细胞黏附因子KirrelA 在MEC中高表达，其表达量是SC中的5倍。KirrelA 在成肌细胞分化融合和骨骼肌损伤修复过程中表达升高，敲低KirrelA同时抑制新生肌管和成熟肌管的生成，说明它在调控成肌细胞融合中起重要作用。