Identification of Novel Circulating Non-hematopoietic Cells Orchestrating Tissue Fibrosis after Injury

PURPOSE: The majority of circulating cells that are recruited to sites of injury are derived from the hematopoietic system, mainly consisting of myeloid and lymphoid cells. Whether non-hematopoietic mesenchymal cells are present in the circulation and play a role in tissue remodeling after injury has remained a topic of debate. METHOD: We used lineage tracing with transgenic Col1-GFP as well as VavCre-mTmG mouse models, parabiosis, flow cytometry, confocal microscopy, and single-cell RNA sequencing (scRNA-seq) to analyze circulating cellular subpopulations recruited to excisional wounds and ischemic skin flaps at acute (3 and 7 days) and chronic (1, 3, and 6 months) stages. Next, GFP-expressing circulating collagen producing cells were systemically depleted using GFP-specific CD8+ T killer cells (Just eGFP death inducing, or JEDI) and the effect on tissue repair was determined using scRNA-seq and analysis of collagen ultrastructure. RESULTS: We identified non-hematopoietic circulating fibroblast-like cells, termed ‘fibrocirculators’, that exhibit stem cell characteristics and produce collagen. These cells were most abundant during the chronic remodeling stage of tissue repair (1 month after injury) and persisted within the tissue up to 6 months after injury. Fibrocirculators exhibited multiple interactions with myeloid cells and their recruitment was triggered by the tumor necrosis factor (TNF) pathway, specifically by TNFSF12/TWEAK. Targeted systemic depletion of fibrocirculators using JEDI T cells accelerated wound healing, decreased fibrotic collagen architecture, and reduced pro-fibrotic gene expression of wound fibroblasts. CONCLUSION: We have identified a novel population of circulating non-hematopoietic mesenchymal cells that are recruited to sites of injury and exhibit pro-fibrotic characteristics. Targeted depletion of these cells may lead to novel therapeutic opportunities for chronic fibrotic conditions such as hypertrophic scars and pulmonary fibrosis.