Multiomics profiling of young and old quiescent skeletal muscle stem cells [aging WGBS]

To uncover new pathways that are important for skeletal muscle stem cell aging, we performed multiomics profiling, including transcriptomics, DNA methylomics, proteomics, and metabolomics on quiescent muscle stem cells from young and old mice. Our goals were to discover pathways that have been overlooked by isolated profiling approaches and to gain insight into which changes are causal, compensatory, correlational, and consequential. In our work, we found that glutathione metabolism is a key pathway of muscle stem cell aging that involves a compensatory feedback loop. Follow-up experiments showed that old muscle stem cells actually form a dichotomy between glutathione-high muscle stem cells and glutathione-low muscle stem cells. RNA-Seq showed that glutathione-high old muscle stem cells are able to synthesize adequate glutathione and thus compensate adequately for oxidative stress with increased glutathione turnover, while glutathione-low old muscle stem cells have failed to compensate for oxidative stress metabolically and instead show increased inflammatory signaling. Overall design: C57BL/6 male mice were used at two ages: young (4 months) and old (22 months). Quiescent muscle stem cells were isolated from limb muscles by FACS (CD45- CD31- Sca1- VCAM+) and immediately used for analysis. Each replicate represents the muscle stem cells from a single animal. There are 4 replicates in each group.

为探明调控骨骼肌干细胞衰老的关键新通路，我们对年轻与老年小鼠的静息态肌肉干细胞开展了多组学谱分析（multiomics profiling），涵盖转录组学、DNA甲基化组学、蛋白质组学与代谢组学。本研究旨在发掘单一组学分析手段所遗漏的通路，并阐明哪些变化属于因果性、代偿性、相关性与后果性改变。
研究发现，谷胱甘肽代谢是肌肉干细胞衰老的关键通路，其涉及代偿性反馈环路。后续实验表明，老年肌肉干细胞实际上呈现二分特征，可分为高谷胱甘肽肌肉干细胞与低谷胱甘肽肌肉干细胞两类。RNA测序（RNA-Seq）结果显示，高谷胱甘肽老年肌肉干细胞可合成足量谷胱甘肽，通过增强谷胱甘肽周转充分代偿氧化应激；而低谷胱甘肽老年肌肉干细胞无法在代谢层面代偿氧化应激，反而呈现炎症信号通路活化增强的表型。
整体实验设计：选用C57BL/6品系雄性小鼠，设置两个年龄组：年轻组（4月龄）与老年组（22月龄）。通过荧光激活细胞分选术（FACS）从四肢肌肉中分离静息态肌肉干细胞，分选标记为CD45⁻ CD31⁻ Sca1⁻ VCAM⁺，并立即用于后续分析。每个生物学重复取自单只小鼠的肌肉干细胞，每组设置4个生物学重复。