Cannabinoid receptor type 2 is time-dependently expressed during skin wound healing in mice

Abstract

Dynamic localization of CB2R and quantitative analysis of CB2R mRNA during skin wound healing in mice were performed. Co-localization of CB2R with F4/80 or α-SMA was detected by double-color immunofluorescence microscopy. A total of 110 male mice were divided into control, injury, and postmortem groups. Sixty-five mice were sacrificed, followed by sampling at 0.5 h-21 days post-injury. Five mice without incision were used as control. The other 40 mice that received incised wound were sacrificed at 5 days after injury. The samples were collected at 0 h-3 days postmortem. In the uninjured controls, CB2R immunoreactivity was detected in the epidermis, hair follicles, sebaceous glands, dermomuscular layer, and vascular smooth muscle. In the incision groups, polymorphonulcear cells, macrophages, and myofibroblasts showed positive staining for CB2R. Morphometrically, the average ratios of CB2R-positive cells were more than 50 % at 5 days post-wounding, whereas it was <50 % at the other posttraumatic intervals. The average ratios of CB2R-positive macrophages maximized at 3 days post-wounding, and the average ratios of CB2R-positive myofibroblasts peaked at 5 days post-wounding. The relative quantity of CB2R mRNA expression maximized at posttraumatic 5 days in comparison with control as detected by real-time PCR, with an average ratio of >4.10, which was also confirmed by Western blotting. There was no significant change for CB2R protein within 6 h postmortem and for mRNA within 3 h postmortem as compared with the control group. In conclusion, dynamic distribution and expression of CB2R suggest that CB2R is involved in modulating macrophages and myofibroblasts in response to inflammatory event and repair process in mouse skin wound healing, and CB2R is available as a marker for wound age determination.

Pharmacological activation of cannabinoid 2 receptor attenuates inflammation, fibrogenesis, and promotes re-epithelialization during skin wound healing

“…Taking into account our previous finding that expression of CB 2 receptor was up-regulated during skin wound healing in mice (Zheng et al, 2012), we presumed CB 2 receptor might regulate skin wound healing, and pharmacomodulation of its activity might be a novel perspective for wound therapy. In the present study, we verified this hypothesis using Gp1a (highly selective CB 2 receptor agonist) and AM630 (highly selective CB 2 receptor antagonist) in mice skin excisional wounds in vivo and HaCaT keratinocytes in vitro.…”

Mustard vesicants alter expression of the endocannabinoid system in mouse skin

“…Endocannabinoids function by binding to cannabinoid receptors and lipid signaling molecules such as the nuclear receptor peroxisome proliferator-activated receptor alpha (PPARα) (Di Marzo et al, 2001; Dubrac et al, 2011; Kendall et al, 2013; O'Sullivan et al, 2010). Two major G protein-coupled endocannabinoid receptors have been identified in the skin, CB1 and CB2 (Galiegue et al, 1995; Kupczyk et al, 2009; Pertwee, 2014; Stander et al, 2005; Zheng et al, 2012). Whereas binding of AEA to CB1 is involved in controlling keratinocyte growth, differentiation and apoptosis (Maccarrone et al, 2003; Paradisi et al, 2008), endocannabinoid signaling through CB2 regulates genes mediating lipid biosynthesis, immune cell signaling, cell migration and inflammation (Dobrosi et al, 2008; Kishimoto et al, 2005; Oka et al, 2006; Zheng et al, 2012).…”

“…Two major G protein-coupled endocannabinoid receptors have been identified in the skin, CB1 and CB2 (Galiegue et al, 1995; Kupczyk et al, 2009; Pertwee, 2014; Stander et al, 2005; Zheng et al, 2012). Whereas binding of AEA to CB1 is involved in controlling keratinocyte growth, differentiation and apoptosis (Maccarrone et al, 2003; Paradisi et al, 2008), endocannabinoid signaling through CB2 regulates genes mediating lipid biosynthesis, immune cell signaling, cell migration and inflammation (Dobrosi et al, 2008; Kishimoto et al, 2005; Oka et al, 2006; Zheng et al, 2012). PPARα has also been identified in the skin, specifically in keratinocytes, sebaceous glands and T-cells, and it is thought to play a role in wound re-epithelialization, sebocyte differentiation, and the resolution of inflammation (Di-Poi et al, 2004; Dubrac et al, 2011; Michalik et al, 2001).…”

Vitality and wound-age estimation in forensic pathology: review and future prospects

“…Several studies showed that the level of argininosuccinate lyase mRNA is stable for 18 h postmortem, that of the sodium-coupled neutral amino acid transporter (SNAT2) mRNA is stable for 48 h postmortem, and those of microtubule-associated protein 1A/1B-light chain 3 (LC3)-II and sequestosome 1 (p62) protein are stable for 4 days postmortem [28,46,47]. In contrast, cannabinoid receptor type-2 mRNA was degraded significantly at 3 h postmortem, and matrix metalloproteinase-2 and the tissue inhibitors of metalloproteinase-2 mRNA were significantly degraded at 12 h postmortem [27,48]. The degradation of RNA and protein caused by postmortem effects -especially putrefaction, decomposition, and desiccation -is inevitable after death.…”

“…After wounding, however, the mRNA levels of cytokines and enzymes typically change sooner than the protein levels and the histomorphology [25][26][27][28]. Hence, assays based on mRNA are suitable for estimating the age of early-stage wounds.…”