Background/Purpose: Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by activated synovial fibroblasts in which pro-inflammatory cytokine interleukin-1β (IL-1β) mediates inflammation. The endocannabinoid system (ECS) is comprised of two evolutionarily conserved cannabinoid receptors 1 and 2 (CB1 and CB2) which elicit their effects through Gα_i/o mediated signaling. However, the mechanism through which ECS activation can modulate inflammatory signaling in RA is poorly understood.

Methods: Human RASFs were obtained from patients diagnosed with RA according to the ACR guidelines (7 female, 2 male, average age 50 ± 43 years). RASFs were pre-treated with 10 or 20 µM of JWH-015 for 10 minutes prior to the addition of IL-1β (10 ng/mL). Stimulation duration was for 30 minutes for signaling studies or 24 hours to evaluate the production of IL-6, IL-8, PGE₂, and cyclooxygenase (Cox) enzymes. Conditioned media was used for the quantification of IL-6, IL-8, and PGE₂ by ELISA and cell lysates were prepared for the analysis of IL-1β signaling proteins like p-P38, p-JNK, p-ERK, and p-TAK-1^Thr184/187 using Western immunoblotting. To understand the role of CB2, knockdown studies were performed using CB2 siRNA for 48 hours prior to the addition of JWH-015 and IL-1β.

Results: Pretreatment of JWH-015 inhibited IL-1β-induced IL-6 (39 and 50% at 10 and 20 µM, respectively) and IL-8 (20 and 47% at 10 and 20 µM, respectively) production in human RASFs (p<0.05; n=3). Evaluation of the signaling pathways using Western immunoblotting showed JWH-015 reduced the expression of phosphorylated TAK-1^Thr184/187 by 12% and 27% at 10 and 20 µM concentration, respectively. Phosphorylated JNK1 was also inhibited by 11% and 22% at 10 and 20 µM and JNK2 by 18% and 64% at 10 and 20 µM, respectively (p<0.05; n=3). Knockdown of CB2 expression using siRNA approach showed almost 40% reduction in the production of IL-β-induced IL-6 and IL-8 and also abrogated the protective effect of JWH-015 (p< 0.05; n=3), suggesting CB2 as a receptor central to JWH-015’s anti-inflammatory action. We also observed CB2 knockdown resulted in a significant decrease of IL-1β-induced Cox-2 expression by ~58%, which resulted in a marked inhibition of PGE₂ production by ~80% (p<0.05; n=3).

Conclusion: Our study provides novel evidence that CB2 activation is anti-inflammatory in response to IL-1β stimulation in RASFs, and JWH-015 elicits anti-inflammatory effects through CB2, not CB1, in human RASFs. These findings suggest that JWH-015 may be tested as an adjunct therapy option to reduce inflammation and the perception of local pain through the ECS.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/cannabinoid-receptor-2-agonist-jwh-015-inhibits-interleukin-1%ce%b2-induced-inflammation-in-rheumatoid-arthritis-synovial-fibroblasts/