Loss of Mrgprd+ sensory nerves delays wound healing by suppressing keratinocyte migration and exacerbating inflammatory states in fibroblasts and immune cells

Summary: Abstract Body: The role of sensory neuronal subsets in the wound healing impairment of type 2 diabetes (T2D) is poorly understood. We have noted reduced Mrgprd+ nerve afferents in skin of diet-induced T2D mice, leading to our hypothesis that this nerve subset participates in normal wound healing. Mice with diphtheria toxin-ablated Mrgprd+ nerves (Mrgprd-) had a 25% reduction in histological wound closure (p<0.01) at 5 days after wounding, correlating with reduced keratinocyte (KC) migration from the proliferating wound edge (0.8 vs. 1.6 mm, p<0.05) (r=0.68, p<0.01). KC proliferation was unaffected. Wound healing was similarly reduced when the Mrgprd receptor was genetically deleted from the intact nerve (p<0.01). Single-cell RNAseq (scRNAseq) of unwounded skin from Mrgprd-ablated mice had fewer follicular stem cells (SC) (7.1% of total cells vs. 10.8%) and more immune-responsive fibroblasts (5.7% vs. 9.4%) than wildtype littermates (p<0.001). Analysis of wounded skin by scRNAseq with a focus on immune cells showed persistence of the M1-like macrophage population at day 5 post-wounding, with these cells having upregulation of genes (>log 2-fold change, p<0.05) associated with inflammatory responses and delayed wound healing, among them Hif1a, Ldha, Mct4, Nos2, Arg2, Tnf, Il1b, and Ccl2 (all p<0.001). There were also increases in neutrophils (p<0.01), as well as reductions in M2-like and monocyte-derived dendritic cells (MoDCs) (both p<0.001) when compared to control littermate wounds. Our findings suggest a key role for Mrgprd+ afferents in wound healing, with Mrgprd loss leading to reduced KC stem cells, impaired fibroblast activation, and increased recruitment of inflammatory immune populations. Lisa Maccio Maretto¹, Natalie Bourand¹, Jacqueline Wang¹, Robin Rong¹, Ziyou Ren¹, Daniela Menichella^{2, 3}, Nihal Kaplan¹, Amy Paller¹ 1. Dermatology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States. 2. Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States. 3. Neurology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States. Stem Cell Biology, Tissue Regeneration and Wound Healing