GSE176360
Single-cell chromatin landscapes supporting fibroblast polarization drives skin regeneration versus fibrosis in adult reindeer
In adult mammals, skin wound healing has evolved to favor rapid repair through the formation of fibrotic scar. These dermal scars are dysfunctional and may lead to chronic disfigurement and disability, yet the biologic mechanisms that drive fibrosis and prevent tissue regeneration remain unknown. Here, we report that reindeer (Rangifer tarandus) antler velvet exhibits regenerative wound healing, whereas identical full-thickness injury in dorsal back skin of the same animal forms fibrotic scar. This regenerative capacity is retained even following ectopic transplantation of velvet to a scar-forming site, demonstrating that this latent regenerative capacity is innate to velvet cells and independent of local factors derived from the growing antler. Single cell RNA-sequencing of uninjured skin revealed a marked divergence in resting fibroblast transcriptional states and immunomodulatory function. Uninjured velvet fibroblast shared a striking resemblance with human fetal fibroblasts whereas uninjured back skin fibroblasts exhibited an overrepresentation of pro-inflammatory genes resembling adult human fibroblasts. Identical skin injury resulted in site-specific fibroblast polarization; back fibroblasts exacerbated the inflammatory response, whereas velvet fibroblasts adopted an immunosuppressive state and reverted back to a regeneration-competent ground state. Consequently, velvet wounds exhibited an accelerated adoption of anti-inflammatory immune states and an expedited resolution of immune response. This study demonstrates reindeer as a novel comparative mammalian model to study both adult skin regeneration (velvet) and scar formation (back skin) within the same animal. Our study underscores the importance of fibroblast heterogeneity in shaping local immune cell functions that ultimately polarize wound healing outcomes. Purposeful, acute modulation of fibroblast-mediated immune signaling represents an important therapeutic avenue to mitigate scar and improve wound healing. Overall design: Back and velvet skin were wounded by making 12mm circular excisions to remove a full thickness piece of tissue. Healing back and velvet skin (along with uninjured D0 skin) were harvested at 7 days following initial injury. To investigate regenerative (velvet) and non-regenerative (back) wound healing mechanisms, we isolated single-cells from all  samples and processed according to 10X Genomics Chromium Next GEM Single Cell ATAC Library & Gel Bead Kit v1 Chemistry Guidelines as per the manufacturer's protocol. In brief, single cells were sorted based on forward versus side scatter gating into 0.1% BSA–PBS and partitioned into Gel Bead-In-EMulsions (GEMs) using 10x GemCodeTM Technology. Next-Generation Sequencing was performed using the Illumina NovaSeq SP and S1 Flow cells using paired-end dual indexing. All raw FASTQs were aligned to the bovine (Bos taurus) reference genome generated using cellranger-atac mkref pipeline. The resulting filtered peak barcode matrix in hdf5 format of the paired back and antler samples for each time point were processed using Signac v.1.2.1.