Efeitos do tratamento com endocanabinóides na resposta inflamatória associada à doença do enxerto-contra-hospedeiro em camundongos

Abstract

Graft-versus-host disease (GVHD) —a secondary complication of bone marrow transplantation—leads the host to develop a systemic inflammatory illness with high mortality and morbidity. Current therapies lack prophylactic effectiveness and commonly fail to provide an immunological balance between inflammation and immunosupression. The endocannabinoid, Anandamide (AEA) and its saturated analogue Palmitoylethanolamide (PEA) are immunomodulatory endogenous molecules produced on demand and rapidly hydrolyzed. In this present work, we investigate the effects of these N-Acetylethanolamines (NAEs) in the complex pathology of GVHD. First, we evaluated the effects of fatty acid amine hydrolase (FAAH) irreversible inhibitor (MAFP) that upon inhibition of FAAH leads to an increase of endogenous NAEs. This treatment led to an increase in survival of 80%. Next, mice were treated with exogenous AEA and it increased survival to 60% and reduce GVHD’s clinical signs. Treatment with AEA also led to a reduction of the number of CD3, CD4, and CD8 cells while reducing the activation of CD4 and CD8 cells but it does not reduce inflammatory cytokines; however, it increases IL-10 in the intestines of mice. Interestingly, AEA reduces Mac-1α, thus lowering adhesion of transplanted cells in mesenteric veins. Furthermore, the effects caused by anandamide treatment are due to CB2 receptor, whereas AEA effects were abolished by CB2 antagonist. These effects are mimicked by JWH133 - a CB2 selective agonist – an, again, abolished by treatment with CB2 antagonist. Therefore, we tested the level of similarity between AEA and PEA, and capability of docking in CB2 receptor of PEA to understand if this would be an interesting target. We found a 63-85% similarity between these molecules in following energy refinement and 82-93% after docking refinement, and a found a predominance of the electrostatic force over the steric regarding their contributions in the overlay of the molecules. Subsequently, we investigate the effect of PEA and the treatment led to an increase of 80% in survival. Furthermore, PEA protected the intestine against damage, reduced the number of CD8 cells and reduced the activation of both CD4 and CD8. It also did not alter pro-inflammatory cytokine release, but increased IL10 and it did interfere with both rolling and adhesion of cells. Interestingly, however, the survival effect appears to be related to CB1 interaction and not CB2 and PEA led to a reduction in mice diarrhea and this effect was abolished by CB1 antagonist. Thus, this study shows that molecular similarity is not the only factor determinant to its interaction in vivo, and other factors could interplay in this response. Furthermore, both CB1 and CB2 receptors can play different roles in the protection against GVHD by endocannabinoids.