CD34+ Cell-derived Fibroblast–Macrophage Crosstalk Drives Limb Ischemia Recovery through the OSM–ANGPTL Signaling Axis

CD34+ cells improve the perfusion and function of ischemic limbs in humans and mice. However, there is no direct evidence of the differentiation potential and functional role of these cells in the ischemic muscle microenvironment. Here, we combined the single-cell RNA sequencing and genetic lineage tracing technology and then provided exact single-cell atlases of normal and ischemic limb tissues in human and mouse, consequently found that bone marrow-derived macrophages with antigen-presenting function migrated to the ischemic site, while resident macrophages underwent apoptosis. The macrophage oncostatin M (OSM) regulatory pathway was specifically turned on by ischemia. Simultaneously, bone marrow CD34+-derived pro-regenerative fibroblasts were recruited to the ischemia niche, where they received macrophage-released OSM, and promoted angiopoietin-like protein (ANGPTL)-associated angiogenesis. These findings provided novel mechanisms on the cellular events and cell–cell communications during tissue ischemia and regeneration, and provided new evidence that CD34+ cells serve as fibroblast progenitors promoting tissue regeneration. Overall design: The muscle was obtained from four groups of hind limb ischemia model mice (Sham, HLI, tdTomato+ cells sorted from Cd34-CreERT2;R26-tdTomato mice, and tdTomato+ cells sorted from Bone marrow transplanted Cd34-CreERT2;R26-tdTomato mice) and two amputees (Ctrl and PAD), the muscle cells were discarded. Mononuclear single-cell suspensions were used to perform Single-cell RNA sequencing (scRNA-seq).