CB2 receptor-mediated effects of pro-inflammatory macrophages influence survival of cardiomyocytes

Aims: The endocannabinoid system and cannabinoid receptor 2 (CB2 receptor) have been associated with modulation of inflammatory response and myocardial adaptation after ischemic injury. In order to elucidate CB2 receptor-related effects during cellular interactions, we investigated cardiomyocyte survival and macrophage function in vitro. Main methods: Murine embryonic (eCM) and adult (CM) cardiomyocytes, murine macrophages (MO), and their subtypes M1 (M1-MO) and M2 (M2-MO) were derived from wildtype- (WT) and C82 receptor-deficient (Cnr2(-/-)) mice. Cells were cultured separately or in co-culture under normoxia or hypoxia (2% O-2) and pro-inflammatory stimulation using interferon (IFN)-gamma. Besides immunohistochemistry, we also measured mRNA expression (Taqman (R)) and performed FACS-analysis of cardiomyocytes. Macrophage migration was assessed using Boyden chamber assay. Key findings: We found a significant induction of CB2 receptor mRNA and protein in murine eCM as well as M1- and M2-MO in vitro following cultivation under hypoxia or stimulation with IFN-gamma. A significantly higher amount of apoptotic Cnr2(-/-)-CMs was found after incubation under hypoxia when compared to WT-CMs. We observed a significantly stronger migration potential in Cnr2(-/-)-M1-MOs towards the supernatant of apoptotic CM, than in corresponding WT-cells. Co-culture revealed a significantly higher loss of eCMs and induction of their apoptosis after cultivation with Cnr2(-/-)-Ml-MOs. Production of TNF-alpha in M1-MOs was dependent on CB2 receptor stimulation by anandamide. Significance: Our data provide novel insights into CB2 receptor-mediated protection of cardiomyocytes during hypoxia and pro-inflammatory stimulation. We show CB2 receptor-dependent effects on migration and function of M1-MOs in interaction with cardiomyocytes, thereby influencing their survival. (C) 2014 Elsevier Inc. All rights reserved.