Increased number of Langerhans cells in the epidermis of diabetic foot ulcers correlates with healing outcome

Abstract

Langerhans cells (LCs) are a specialized subset of epidermal dendritic cells. They represent one of the first cells of immunologic barrier and play an important role during the inflammatory phase of acute wound healing. Despite considerable progress in our understanding of the immunopathology of diabetes mellitus and its associated comorbidities such as diabetic foot ulcers (DFUs), considerable gaps in our knowledge exist. In this study, we utilized the human ex vivo wound model and confirmed the increased epidermal LCs at wound edges during early phases of wound healing. Next, we aimed to determine differences in quantity of LCs between normal human and diabetic foot skin and to learn if the presence of LCs correlates with the healing outcome in DFUs. We utilized immunofluorescence to detect CD207+ LCs in specimens from normal and diabetic foot skin and DFU wound edges. Specimens from DFUs were collected at the initial visit and 4 weeks later at the time when the healing outcome was determined. DFUs that decreased in size by >50 % were considered to be healing, while DFUs with a size reduction of <50 % were considered non-healing. Quantitative assessment of LCs showed a higher number of LCs in healing when compared to non-healing DFU's. Our findings provide evidence that LCs are present in higher number in diabetic feet than normal foot skin. Healing DFUs show a higher number of LCs compared to non-healing DFUs. These findings indicate that the epidermal immune barrier plays an important role in the DFU healing outcome and may offer new therapeutic avenues targeting LC in non-healing DFUs.

Fig. 1. Number of intra-epidermal LCs changes during a course of acute wound healing

Hematoxylin and eosin staining of ex vivo human skin wounds at a time of wounding (0hrs) and 3 and 7 days post wounding. Full arrows demarcate a wound edge while empty ones point at migrating epithelial tongue at day 3 and newly formed multilayered epidermis on day 7 post wounding (n=3) (a). Same specimens were stained using CD207/Langerin antibody, a Langerhans cell marker. White arrows point at the positive LCs staining. Dotted line demarcates basement membrane (b). Enlargements of the CD207 positive staining are shown (c). Scale bar 50μm.

Fig. 2. Human diabetic plantar foot skin shows increased presence of LCs

Histology of diabetic and normal plantar foot skin showing thick epidermis and cornified layer (a) Number of 207/Langerin positive cells is increased in diabetic (n=5) when compared to normal foot skin (n=3). (b) A bar graph depicts number of Langerin positive cells quantified per mm of tissue. Data are presented as means ± S.D (c). Scale bar 50μm.

Fig. 3. Number of Langerhans cells is increased in the epidermis of healing DFUs

Hematoxylin and eosin staining of healing and non-healing diabetic foot ulcers showing distinctive chronic wound morphology characterized by hyperproliferative epidermis (HE) and thick cornified layer (CL) with a presence of nuclei. Dotted line represents epidermal- dermal junction (a) The most representative images of immunofluorescent staining of CD207/Langerin in non-healing (n=6) and healing (n=6) DFUs at week 0 and 4 weeks after shows increase in number of LCs in healing DFUs at both weeks (b) Quantification of CD207/Langerin positive cells in healing and non-healing DFUs per mm of tissue is shown (c). Scale bar 50μm.