Mast Cells in Diabetes and Diabetic Wound Healing

Abstract

Mast cells (MCs) are granulated, immune cells of the myeloid lineage that are present in connective tissues. Apart from their classical role in allergies, MCs also mediate various inflammatory responses due to the nature of their secretory products. They are involved in important physiological and pathophysiological responses related to inflammation, chronic wounds, and autoimmune diseases. There are also indications that MCs are associated with diabetes and its complications. MCs and MC-derived mediators participate in all wound healing stages and are involved in the pathogenesis of non-healing, chronic diabetic foot ulcers (DFUs). More specifically, recent work has shown increased degranulation of skin MCs in human diabetes and diabetic mice, which is associated with impaired wound healing. Furthermore, MC stabilization, either systemic or local at the skin level, improves wound healing in diabetic mice. Understanding the precise role of MCs in wound progression and healing processes can be of critical importance as it can lead to the development of new targeted therapies for diabetic foot ulceration, one of the most devastating complications of diabetes.

Keywords: Diabetes mellitus; Diabetic foot ulcer; Mast cells; Wound healing.

Fig. 1

Main mast cell functions in diabetes

Fig. 2

Skin MC degranulation, assessed by toluidine blue and/or tryptase immunostaining, is increased in patients with DM and is associated with inflammation. a Representative images of toluidine blue-stained MCs in forearm skin (top panel) and of tryptase-immunostained MCs in foot skin specimens (bottom panel) from subjects with and without DM (scale bar 10 µm). Black arrows show non-degranulated MCs and red arrows show degranulated MCs. Degranulated MCs were in proximity to inflammatory cells (blue arrows). b The total number and c percentage of degranulated MCs stained with toluidine blue were increased in forearm skin specimens from subjects with DM. d MC degranulation was also increased in foot skin specimens from subjects with DM stained with tryptase, while e the total number of MC was not different. *p < 0.05. f–h A positive correlation was observed between degranulated MCs and the f dermis inflammatory cells, the serum levels of g IL-6 and h TNFα. (Ref. [77])

Fig. 3

Functional MCs are required for proper wound healing. Wound healing progress was evaluated over a 10-day period in non-DM and DM mice DSCG treated or not treated and MC-deficient mice. a, b Wound healing was delayed in C57BL6 DM mice compared to non-DM mice and DSCG pre-treatment accelerated it from days 6 to 10 post wounding. DSCG had no effect on non-DM mice. c–f Wound healing was delayed in KitW/KitW-v mice with or without DM, when compared to their respective non-DM or DM WBB6F1 controls. Topical SP improved healing at day 10 post wounding in both non-DM and DM WBB6F1 mice, but failed to have an effect in either non-DM KitW/KitW-v or DM KitW/KitW-v mice. g, h Wound healing was delayed in MC-deficient KitW-sh/KitW-sh mice when compared to their respective B6.Cg controls. *p < 0.05. (Ref. [77])

Fig. 4

Synthesis and validation of mast cell stabilization by MCS-01. a Chemical synthesis of MCS-01. In rat RBL-2H cells MCS-01 reduced b Ca²⁺ influx measured as % relative fluorescence of Fluo-4, c β-hex release measured as % of total β-hex in cell lyses at 30 min, and d TNFα release measured at 3 h of MCS-01 stimulation (0.033–30 µM) after thapsigargin (Tg, 1 µM) activation (n = 3). MCS-01 (100 µM) treatment inhibited SP (2 μM)-mediated e β-hex release (after 30 min pre-treatment), f TNFα or g IL-8 release (after 24 h pre-treatment) from human mast cells. h Development of MCS-0-releasing bandage for topical delivery: the schematic of alginate bandage fabrication. Mixture of alginate polymer, laponite, and MCS-01 drug is added to acrylic molds and is frozen at − 20° C. The molds are then placed in a lyophilizer to remove ice crystals, leaving polymer meshes with large voids. The dried polymer meshes are then placed in a large 100 mM CaCl₂ bath for 15 min to form ionic cross-linking for gelation. Alginate bandages are formed. i In vitro release of MCS-01 from alginate bandages. Data shown as mean ± SD (n ≥ 3, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001) (Ref. [89])

Fig. 5

Topical MCS-01 applied either pre or post wounding and DSCG (i.p. injected pre-wounding) accelerated wound healing in diabetic mice. a Experimental design of in vivo wound healing in STZ-induced diabetic mice. 6-mm full-thickness wounds were created on the dorsum. Effects after 10 days of topical treatment with MCS-01-loaded bandages (800 µg/bandage), applied either before or after wounding (MCS-pre or MCS-post, respectively), were compared to that of vehicle-only bandages (blank) or intraperitonially administered mast cell stabilizer disodium cromoglycate (DSCG). Wounds before and after bandage application are shown to the right. b DSCG or pre-MCS-01 treatment reduced skin mast cell degranulation assessed by toluidine blue staining compared to blank controls. c Wound size measurement, d wound re-epithelialization analysis, and e representative images on day 10 (D10) post wounding showed improvement in treatment groups compared to blank controls. Topical post-MCS-01 treatment and DSCG also f rescued the elevation in MMP-9 protein expression at D10 vs D0 observed in diabetic mice. g D10 counts of M1 macrophages were elevated upon post-MCS-01 treatment, but reduced with pre-MCS-01 and DSCG treatments. h M2 macrophages showed similar trends as M1. Data shown as mean ± SD (n ≥ 3, *p ≤ 0.05, **p ≤ 0.01. Results in Fig. 2d are based on two-sample t test comparisons between blank and pre-MCS-01 and DSCG) (Ref. [89])