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BackgroundThe anti-inflammatory properties of the cannabinoid 2 receptor (CB2R) in injury and inflammatory diseases have been widely substantiated. Specifically, the anti-inflammatory effect of CB2R may be achieved by regulating macrophage polarisation. Several research findings suggested that the activation of CB2R could attenuate inflammation by reducing pro-inflammatory M1 macrophage polarisation and promoting anti-inflammatory M2 polarisation. However, considering CB2R inhibits fibrosis and M2 promotes fibrosis, that the activation of CB2R may lead to an increase in M2 macrophages seems contradictory. Therefore, we hypothesised that the activation of CB2R to attenuate inflammation is not achieved by up-regulating M2 macrophages.MethodsWe established an incised wound model using mouse skin and used this to evaluate the effect of CB2R agonists (JWH133 or GP1a) and an antagonist (AM630) on wound healing. At various post-injury intervals, we used western blot analysis, immunofluorescence staining, enzyme-linked immunosorbent assay and quantitative reverse transcription polymerase chain reaction assays to determine CB2R protein expression, M1/M2 macrophage infiltration, and the protein and gene expression of M1/M2-associated markers and cytokines in skin lesions.ResultsActivation of CB2R significantly reduced M1 macrophage infiltration and slightly increased M2 macrophage infiltration. Similarly, gene expression and protein levels of M1-associated markers and cytokines (interleukin [IL]-6, IL-12, CD86 and inducible nitric oxide synthase) were significantly down-regulated after CB2R agonist administration; in contrast, markers and cytokines were increased in the CB2R antagonist–treated group. Conversely, the administration of agonists slightly increased gene expression and protein levels of M2-associated markers and cytokines (IL-4, IL-10, CD206 and arginase-1 [Arg-1]); however, a statistical significance at most time points post-injury was not noted.ConclusionIn summary, our findings suggested that during incised skin wound healing in mice, increased levels of CB2R may affect inflammation by regulating M1 rather than M2 macrophage subtype polarisation. These results offer a novel understanding of the molecular mechanisms involved in the inhibition of inflammation by CBR2 that may lead to new treatments for cutaneous inflammation.

Studies have shown that cannabinoid CB2 receptors are involved in wound repair, however, its physiological roles in fibrogenesis remain to be elucidated. In the present study, the capacity of cannabinoid CB2 receptors in the regulation of skin fibrogenesis during skin wound healing was investigated. To assess the function of cannabinoid CB2 receptors, skin excisional BALB/c mice were treated either the cannabinoid CB2 receptor selective agonist, GP1a, or antagonist, AM630. Skin fibrosis was assessed by histological analysis and profibrotic cytokines were determined by immunohistochemistry, immunofluorescence staining, reverse transcription-quantitative polymerase chain reaction and immunoblotting in these animals. GP1a decreased collagen deposition, reduced the levels of transforming growth factor (TGF)-β1, TGF-β receptor I (TβRI) and phosphorylated small mothers against decapentaplegic homolog 3 (P-Smad3), but elevated the expression of its inhibitor, Smad7. By contrast, AM630 increased collagen deposition and the expression levels of TGF-β1, TβRI and P-Smad3. These results indicated that cannabinoid CB2 receptors modulate fibrogenesis and the TGF-β/Smad profibrotic signaling pathway during skin wound repair in the mouse.

The study was focused on time-dependent expressions of paired-box transcription factor 7 (Pax7) and myoblast determination protein (MyoD) during skeletal muscle wound healing. An animal model of skeletal muscle contusion was established in 40 Sprague-Dawley male rats. Samples were taken at 1, 3, 5, 7, 9, 13, 17, and 21 days after injury, respectively (five rats in each posttraumatic interval). Five rats were employed as control. By morphometric analysis, the data based on the number of Pax7(+)/MyoD(-), Pax7(+)/MyoD(+), and Pax7(-)/MyoD(+) cells were highly correlated with the wound age. Pax7 and MyoD expressions were upregulated after injury by Western blot and quantitative real-time PCR assays. The relative quantity of Pax7 protein peaked at 5 days after injury, which was >1.13, and decreased thereafter. Similarly, the relative quantity of MyoD mRNA expression peaked at 3 days after injury, which was >2.59. The relative quantity of Pax7 protein >0.73 or mRNA expression >2.38 or the relative quantity of MyoD protein >1.33 suggested a wound age of 3 to 7 days. The relative quantity of MyoD mRNA expression >2.02 suggested a wound age of 1 to 7 days post-injury. In conclusion, the expressions of Pax7 and MyoD are upregulated in a time-dependent manner during skeletal muscle wound healing, suggesting that Pax7 and MyoD may be potential markers for wound age estimation in skeletal muscle.

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