A Histopathological and Immunohistochemical Study of Wound Healing Mechanism and Biological Effect of Low-Energy Laser Irradiation in Type 2 Diabetic Model Mice

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Diabetes mellitus (DM) is a typical life style related disease, in which hyperglycemia is caused by an absolute defect (type 1 DM) or relative defect (type 2 DM) of insulin. Specifically, type 2 DM is considered a life style related disease. DM is known to cause a capillary vessel disorder due to the hyperglycemia and often brings about delayed wound healing ; however, the mechanism behind this delayed wound healing is uncertain and the effect of laser irradiation using a mouse model is insufficient. Thus, this paper studies histopathological and immunohistochemical changes using the db/db mouse model in order to investigate the wound healing mechanism in type 2 DM as well as the biological effects of wound healing by low-energy laser irradiation. This DM model mice (db/db) were compared with a normal mice (m+/m+). The different levels of inflammatory cell infiltration in the wound site, capillary dilatation and regeneration, and proliferation of fibroblasts and collagen fibers were observed over time, and delayed wound healing was studied histopathologically. Moreover, db/db of laser irradiation was compared with db/db of non laser irradiation, finding capillary dilatation and regeneration, the proliferation of fibroblasts and collagen fibers prominent. Mast cells seen by metachromasia in toluidine blue pH 2.5 staining were observed in the dermis around the margin on the wound site of m+/m+ and db/db of non laser irradiation, capillary dilatation and regeneration, and proliferation of the fibroblasts and collagen fibers surroundings. The number of mast cells over time was lower in db/db of non laser irradiation than in m+/m+, however, more degranulation was seen in db/db of laser irradiation than in db/db of non laser irradiation. According to immunohistochemical findings, HSP70 was positively immunoreactive in neutrophils, macrophages, vascular endothelial cells, fibroblasts and epidermis. Moreover, the appearance of HSP70 was lower in db/db of non laser irradiation than in m+/m+. The appearance level of HSP70 in vascular endothelial cells was higher in db/db of laser irradiation than in db/db of non laser irradiation. NOS2 kept positively high in vascular endothelial cells continuously until 7 days post-operatively in db/db of the non laser irradiation compared with m+/m+. On the other hand, the level of NOS2 was low in macrophages and vascular endothelial cells in db/db of laser irradiation. Moreover, while the appearance of FGF2 was identified on 2 days in neutrophils in m+/m+, it was seen by 5 days in db/db of non laser irradiation, suggesting a delayed appearance time. The positive appearance of FGF2 was high in vascular endothelial cells and fibroblasts in db/db of laser irradiation. These findings suggest a delayed wound healing process in db/db of non laser irradiation by the decrease in the mast cell number and mast cell degranulation, decreased expression of HSP70 and FGF2, and increased expression of NOS2. It is speculated that low-energy laser irradiation improves the above findings and delayed wound healing process in db/db of non laser irradiation.