AM251, a cannabinoid receptor 1 antagonist, prevents human fibroblasts differentiation and collagen deposition induced by TGF-ß - An in vitro study

Autores da FMUP

• Maria Paula De Valadares Souto Pinto Serrão

  Autor

• Maria Augusta Vieira Coelho

  Autor

Participantes de fora da FMUP

• Correia-Sá, IB
• Carvalho, CM
• Machado, VA
• Carvalho, SO
• Marques, M

Unidades de investigação

• Biomedicina

Abstract

Previous studies showed that cannabinoid 1 receptor (CB1) is linked with skin fibrosis and scar tissue formation in mice. Therefore, the topical use of cannabinoids may have a role in the prevention or treatment of local fibrotic and wound healing diseases as hypertrophic scars or keloids. In this study, we asked whether CB1 activation or inactivation would change fibroblast differentiation into myofibroblast and collagen deposition in skin human fibroblast. Primary cultures of adult human fibroblasts were obtained from abdominal human skin. Cells were stimulated with transforming growth factor-beta (TGF-beta, 10ng/ml) and treated with a CB1 selective agonist (arachidonyl-2-chloroethylamide, ACEA 1 mu M) and an antagonist (AM251 1, 5 and 10 mu M). Alpha-smooth muscle actin (alpha-SMA) was quantified using Immunocytochemistry and Western Blot. Collagen was quantified with Sirius Red staining assay. Significance was assessed by One-way ANOVA. P < 0.05 was considered significant. TGF-beta significantly increases alpha-SMA expression. ACEA 1 mu M significantly increases collagen deposition but does not change alpha-SMA expression. AM251 10 mu M added in the absence and the presence of ACEA reduces alpha-SMA expression and collagen content in TGF-beta treated cells. AM251 shows a concentration-dependent effect over collagen deposition with a pIC50 of 5.5 (4.6-6.4). TGF-beta significantly increases CB1 receptor expression. CB1 inactivation with AM251 prevents fibroblasts differentiation and collagen deposition, induced by TGF-beta in human fibroblasts. The outcome supports that CB1 is a molecular target for wound healing disorders and in vivo and pre-clinical studies should be implemented to clarify this premise.

Keywords

• CB1 receptors; Fibroblasts; Skin; Fibrosis; TGF-beta; AM251