Cannabinoid receptor-2 ligand beta-caryophyllene modulates inflammatory responses of bovine peripheral blood mononuclear cells in vitro

Beta-caryophyllene (BCP) is an agonist of cannabinoid receptor 2 (CNR2/CB2), exerting mostly anti-inflammatory effects in rodents, but not explored in cattle. Herein we conducted the first reductionist step to examine the effects of in-vitro supplementation of BCP in bovine peripheral blood mononuclear cells (PBMCs) on the inflammatory response under thermoneutral (TN) or heat stress (HS) conditions. We performed this work in two cohorts: in Study 1, PBMC were supplemented with BCP at 1 µM (n = 8), and in Study 2 PBMC from other cows were supplemented with BCP at 1 mM (n = 7). In each study, PBMC from each cow were assigned to controls (CTL) or BCP-supplemented and incubated in TN (38.5 °C) or HS (41.0 °C) conditions. Then, half of the cells from each cow were treated with lipopolysaccharide (LPS), and media were collected for analysis of cytokines/chemokines and cyclic adenosine monophosphate (cAMP) concentrations as indicator of CB2 activation. In addition, RNA and proteins were extracted from plate-adhered monocyte-enriched-PBMCs (MEPs), for gene-expression and proteomics analyses. In each study, the effects of BCP, LPS, HS, and interactions were analyzed. We found in both studies that LPS and BCP (at 1 µM or at 1 mM) lowered CNR2 gene expression in MEPs. At 1 mM, BCP inhibited gene expression of LPS-induced NFKB1, amplified TNFA, and suppressed IL10, and tended to reduce cAMP in medium, possibly indicating CB2 activation. Post-LPS, BCP (1 mM) increased monocyte-chemoattractant-protein-1 and tended to enhance interleukin-1-alpha, beta, and macrophage-inflammatory-protein-1-alpha in medium; while HS increased macrophage-inflammatory-protein-1-beta. Proteomics of LPS-activated MEPs revealed immune-related proteins and canonical pathways enriched by BCP. Overall, BCP supplementation in vitro modulated immune function in PBMC from dairy cows, with similar effects in TN and HS conditions; thus, BCP could be studied as a potential feed supplement in cattle that may induce moderate immune-modulating effects. Further studies should investigate the in vivo effects of BCP on immune function in cattle.