MME+ fibro-adipogenic progenitors are the dominant adipogenic population during fatty infiltration in human skeletal muscle

Fatty infiltration, the ectopic deposition of adipose tissue within skeletal muscle, is mediated via the adipogenic differentiation of fibro-adipogenic progenitors (FAPs). We used single-nuclei and single-cell RNA sequencing to characterize FAP heterogeneity in patients with fatty infiltration. We identified an MME+ FAP subpopulation which, based on ex vivo characterization as well as transplantation experiments, exhibits high adipogenic potential. MME+ FAPs are characterized by low activity of WNT, known to control adipogenic commitment, and are refractory to the inhibitory role of WNT activators. Using preclinical models for muscle damage versus fatty infiltration, we show that many MME+ FAPs undergo apoptosis during muscle regeneration and differentiate into adipocytes under pathological conditions, leading to their depletion. Finally, we utilized the varying fat infiltration levels in human hip muscles to show the depletion of MME+ FAPs in fatty infiltrated human muscle. Altogether, we have identified the dominant adipogenic FAP subpopulation in skeletal muscle. For bulk RNAseq, mRNA from sorted skeletal muscle FAPs (PDGFRA/eGFP+ MME+/-). 5 biological replicates per group. For human sn- and scRNA-seq, we harvested muscles from fatty (GM) and non-fatty infiltrated (RF) muscles. One sample per condition (scRNAseq), and pooled muscles (n=5) in 2 technical replicates (snRNAseq). For mouse scRNAseq, we harvested muscles from WT mouse. One biological sample.

脂肪浸润（fatty infiltration）指骨骼肌内异位沉积脂肪组织的病理过程，其发生由成纤维脂肪祖细胞（fibro-adipogenic progenitors, FAPs）的成脂分化介导。本研究采用单细胞核RNA测序（single-nuclei RNA sequencing）与单细胞RNA测序（single-cell RNA sequencing）技术，对脂肪浸润患者体内的FAP异质性进行了表征。我们鉴定出一类MME+ FAP亚群，经体外功能表征与移植实验证实，该亚群具备极高的成脂分化潜能。MME+ FAP的特征为WNT信号通路活性较低——已知WNT信号可调控成脂定向分化，且该亚群对WNT激活剂的抑制作用具有抗性。本研究利用肌肉损伤与脂肪浸润的临床前模型，证实大量MME+ FAP在肌肉再生过程中发生凋亡，并在病理状态下分化为脂肪细胞，最终导致该亚群耗竭。最后，我们借助人类髋部肌肉中不同程度的脂肪浸润样本，验证了脂肪浸润的人类骨骼肌内MME+ FAP确实发生了耗竭。综上，本研究成功鉴定出骨骼肌内占主导地位的成脂性FAP亚群。
针对批量RNA测序（bulk RNAseq），我们从分选得到的骨骼肌FAP（PDGFRA/eGFP+ MME+/-）中提取mRNA，每组设置5个生物学重复。
针对人类单细胞核RNA测序与单细胞RNA测序，我们分别采集脂肪浸润（GM）与非脂肪浸润（RF）的肌肉组织；单细胞测序每组设置1个样本，单细胞核测序则将5份肌肉组织混合后设置2个技术重复。
针对小鼠单细胞RNA测序，我们采集野生型（wild type, WT）小鼠的肌肉组织，设置1个生物学重复。