Gordon Sloan ¹ Philippe D. Donatien ² Rosario Privitera ² Pallai Shillo ¹ Sharon Caunt ¹ Dinesh Selvarajah ¹ Praveen Anand ^2* Solomon Tesfaye ¹

• ¹ The University of Sheffield, Sheffield, England, United Kingdom
• ² Imperial College London, London, United Kingdom

Background: Identifying distinct mechanisms and biomarkers for painful diabetic peripheral neuropathy (DPN) are required for advancing the treatment of this major global unmet clinical need. We previously provided evidence in calf skin biopsies that disproportion between reduced sensory small nerve fibers and increased blood vessels may distinguish painful from non-painful DPN. We proposed that overexposure of the reduced nerve fibers in DPN to increased hypoxemia-induced vasculature and related algogenic factors, e.g. Nerve Growth Factor (NGF), leads to neuropathic pain.To further investigate this proposed mechanism, we have now studied more proximal thigh skin biopsies, to see if the same disproportion between increased vasculature and decreased nerve fibers generally differentiates painful-DPN from painless-DPN.Methods: 28 subjects with type 2 diabetes (T2DM) and 13 healthy volunteers (HV) underwent detailed clinical and neurophysiological assessments; based on the neuropathy composite score of the lower limbs (NIS(LL)) plus 7 tests. T2DM subjects were subsequently divided into three groups: painful-DPN (n=15), painless-DPN (n=7) and No-DPN (n=6). All subjects underwent skin punch biopsy from the upper lateral thigh 20cm below the anterior iliac spine.Results: Skin biopsies showed decreased PGP 9.5-positive intra-epidermal nerve fiber (IENF) density in both painful-DPN (p< 0.0001) and painless-DPN (p=0.001). Vascular marker von Willebrand Factor (vWF) density was markedly increased in painful-DPN vs. other groups, including painless-DPN (p=0.01). There was a resulting significant decrease of the ratio of intra-epidermal nerve fiber density to vasculature, PGP9.5:vWF, in painful-DPN vs. painless DPN (p=0.05). These results were similar in pattern to those observed in these HV and T2DM groups previously in distal calf biopsies; however, the increase in vWF was much higher and nerve fiber density much lower in the calf than thigh for painful-DPN. Thigh skin vWF density was significantly correlated with several metabolic (waist/hip ratio, HbA1c), clinical (e.g. pain score), and neurophysiological measures.This study supports our proposal that increased dermal vasculature, and its disproportionate ratio to reduced nociceptors, may help differentiate painful-DPN from painless-DPN. This disproportion is greater in distal calf than proximal thigh skin, hence neuropathic pain in DPN is length-dependent, and first localized to the distal lower limbs, mainly feet.