Activation of Keratinocyte Gq-linked G-Protein Coupled Receptors Regulates Degeneration of Cutaneous Nerves

Painful diabetic neuropathy (PDN) is one of the most common complications of diabetes, affecting 25% of diabetic patients. DPN is characterized by small-fiber degeneration, which can progress to complete loss of cutaneous innervation and is accompanied by neuropathic pain. Uncovering the mechanisms underlying degeneration of cutaneous nerves in PDN remains a major challenge to finding effective and disease-modifying therapy. Keratinocytes are closely juxtaposed to cutaneous nerve terminals potentially enabling communication between keratinocytes and cutaneous afferents. Our aim is to identify mechanisms by which keratinocytes communicate with cutaneous afferents and how this signaling regulates axonal degeneration underlying neuropathic pain in PDN. In this study, we utilized a chemogenetic approach using DREADD receptors, synthetic receptors based on the structure of human muscarinic receptors that can be activated by the synthetic ligands clozapine and clozapine N-oxide (CNO). We genetically expressed stimulatory DREADD (hM3Dq) into K14 basal keratinocytes (K-14) in mice as a tool for mimicking the activation of Gq-linked G protein-coupled receptors (GPCRs) in K14-expressing basal keratinocytes. Histological characterization of the skin from mice expressing hM3Dq in K-14 positive cells revealed a clear thickening of the epidermis (as shown by H&E staining) due to K-14 expressing cell hyperproliferation (as shown by BrdU incorporation). Additionally, we observed reduced innervation of the epidermis (as shown by PGP9.5 staining), indicating that activation of K-14 Gq-linked GPCRs drives nerve fibers degeneration in the epidermis. Furthermore, transcriptional profiling of activated K-14 from the skin of K14-hM3Dq mice revealed downregulation of genes involved in neuronal survival and neurite outgrowth, including Nerve Growth Factor (NGF), artemin, a member of the Glial Cell-Derived Neurotrophic Factor (GDNF) ligand family, and Semaphorin 3D. Our results suggest that basal keratinocyte Gq-linked GPCRs could represent highly “druggable” and easily accessible targets for the development of therapeutics that by reversing axonal degeneration of cutaneous nerves in a pathological conditions such as PDN and could ameliorate the associated neuropathic pain. RO1-Supplement 610-5210100-60054666, RO1-610-5210100-60057550, P&F award-610-5202000-60056119.