Aged skeletal muscle is characterized by poor muscle recovery following disuse coinciding with an impaired muscle pro-inflammatory macrophage response. Macrophage inflammatory status is regulated by its metabolic state, but little is understood of macrophage metabolism and its relation to macrophage inflammation in the context of muscle recovery and aging. Therefore, the purpose of this study was to thoroughly characterize macrophage metabolism and inflammation in aged muscle during early recovery following disuse atrophy using single cell transcriptomics and functional assays. Young (4–5 months) and old (20–22 months) male C57BL/6 mice underwent 14 days of hindlimb unloading followed by 4 days of ambulatory recovery. CD45+ cells were isolated from solei muscles and analyzed using 10x Genomics single cell RNA sequencing. We found that aged pro-inflammatory macrophage clusters were characterized with an impaired inflammatory and glycolytic transcriptome, and this dysregulation was accompanied by a suppression of HIF-1α and its immediate downstream target, Glut1. As a follow-up, bone marrow-derived macrophages were isolated from a separate cohort of young and old mice at 4-d recovery and were polarized to a pro-inflammatory phenotype and used for glycolysis stress test, phagocytosis activity assay, and targeted GC-MS metabolomics. Aged bone marrow-derived pro-inflammatory macrophages were characterized with impaired glycolysis and phagocytosis function, decreased succinate and an accumulation of glycolytic metabolic intermediates overall supporting reduced glycolytic flux and macrophage function. Our results indicate that the metabolic reprograming and function of aged skeletal muscle pro-inflammatory macrophages are dysfunctional during early recovery from disuse atrophy possibly attributing to attenuated regrowth.

中文翻译：

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废用性萎缩后早期恢复期间老年比目鱼肌中巨噬细胞代谢重编程被破坏

衰老的骨骼肌的特点是废用后肌肉恢复不良，同时肌肉促炎巨噬细胞反应受损。巨噬细胞炎症状态受其代谢状态调节，但人们对巨噬细胞代谢及其与肌肉恢复和衰老背景下巨噬细胞炎症的关系知之甚少。因此，本研究的目的是使用单细胞转录组学和功能测定来彻底表征废用性萎缩后早期恢复期间老化肌肉中的巨噬细胞代谢和炎症。年轻（4-5 个月）和年老（20-22 个月）雄性 C57BL/6 小鼠接受 14 天的后肢卸载，然后进行 4 天的动态恢复。从比目肌中分离出 CD45+ 细胞，并使用 10x Genomics 单细胞 RNA 测序进行分析。我们发现，衰老的促炎巨噬细胞簇的特征是炎症和糖酵解转录组受损，并且这种失调伴随着 HIF-1α 及其直接下游靶标 Glut1 的抑制。作为后续研究，在 4 天恢复时从一组年轻和年老小鼠中分离出骨髓来源的巨噬细胞，并将其极化为促炎表型，用于糖酵解应激测试、吞噬活性测定和靶向 GC -MS代谢组学。衰老的骨髓源性促炎巨噬细胞的特点是糖酵解和吞噬功能受损、琥珀酸盐减少以及糖酵解代谢中间体的积累，总体上支持糖酵解通量和巨噬细胞功能的减少。我们的结果表明，衰老骨骼肌促炎巨噬细胞的代谢重编程和功能在废用性萎缩的早期恢复过程中出现功能障碍，可能归因于再生减弱。