| ObjectiveSkin wound healing is continuous and complex process, which has been simply divided into three phases:inflammation, proliferation, and remodeling. Within each phase, a myriad of orchestrated reactions and interactions between cells, cytokine and inflammatory mediators are put into action. cannabinoid receptor type 2 (CB2R), is a G-protein-coupled receptor that activates in a multitude of signal transductional pathways. CB2R is predominantly expressed in peripherally, such as immune system and hematopoietic cells. More recent studies have found that CB2R is involved in a wide range of disparate diseases and tissue injuries, and plays an important role of anti-inflammatory and anti-fibrosis. In this study, the level of CB2R in skin incised wound was detected by immunohistochemical technique and western blotting. The aim of this investigation is to explore the possible function of CB2R in skin wound healing and its applicability to the wound age estimation.Materials and MethodsA total of 50 mice, each weighting 35-40g, were divided into ten groups randomly, one control group and nine experiment groups, A 1.5cm-long full-thickness incision deep to the fascia was made with a scalpel in the skin layer on the central dorsum of each mouse under sterile technique after the mouse was anesthetized by inhalating diethyl ether. After injury, each mouse was housed individually and fed with sterilized food and distilled water to prevent from infection. The 1.5cm×1.0cm specimens were taken from the wounded site after the mice were executed by cervical dislocation at Oh,6h,12h, 1d,3d,5d,7d, 10d and 14d postinjury to prepare for the subsequent procedures. The level of CB2R in mouse skin incised wound was detected by immunohistochemical staining and Western blot, and the non-incised mouse skin was used as control. The number and ratio of CB2R positive cells were counted and analyzed by microscope. The band AverGray was analyzed by software of Fluochem V2.0. The data were analyzed comparatively by the software of SPSS13.0 for Windows, and there was statistical significance comparing with the neighboring group when P<0.05.ResultsCB2R immuroreactivity was detected in epidemical layer, hair follicle, sebaceous gland, dermomuscular layer, vascular smooth muscle, nerve fiber adventitia and perimysium in the control skin; weak CB2R immunostaining was detected in polymorphonuclear cells (PMNs) in the wound and peripheral region in aged 6h, a large number of CB2R-positive mononuclear cells (MNCs) were identified from 12h to 24h, Afterwards, the CB2R-positive cells were mostly MNCs and spindle-shaped fibroblastic cells (FBCs) at 3d postinjury, and mainly in spindle-shaped fibroblastic cells from 5d to 14d postinjury. The ratio of the CB2R-positive cells increased gradually in the wound specimens from Oh to 3d, and maximized at 5d, and decreased from 7d to 14d postinjury. The bands of CB2R expression were observed throughout the wound healing stages by Western blotting, and maximized at 5d.ConclusionThe results suggested that CB2R was expressed in PMNs, MNCs, FBCs during healing process of skin wound in mice, and may be used as a newmarker for the wound age determination.
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