IL-1β induces changes in expression of core circadian clock components PER2 and BMAL1 in primary human chondrocytes through the NMDA receptor/CREB and NF-κB signalling pathways

The circadian clock is a specialised cell signalling circuit present in almost all cells. It controls the timing of key cell activities such as proliferation and differentiation. In osteoarthritis, expression of two components of the circadian clock, BMAL1 and PER2 is altered in chondrocytes and this change has been causally linked with the increase in proliferation and altered chondrocyte differentiation in disease. IL-1β, an inflammatory cytokine abundant in OA joints, has previously been shown to induce changes in BMAL1 and PER2 expression in chondrocytes. The purpose of this study is to identify the mechanism involved. We found IL-1β treatment of primary human chondrocytes led to activation of NMDA receptors as evidenced by an increase in phosphorylation of GluN1 and an increase in intracellular calcium which was blocked by the NMDAR antagonist MK801. Levels of phosphorylated CREB were also elevated in IL-1β treated cells and this effect was blocked by co-treatment of cells with IL-1β and the NMDAR antagonist MK-801. Knockdown of CREB or inhibition of CREB activity prevented the IL-1β induced increase in PER2 expression in chondrocytes but had no effect on BMAL1. Phosphorylated p65 levels were elevated in IL-1β treated chondrocytes indicating increased NF-κB activation. Inhibition of NF-κB activity prevented the IL-1β induced reduction in BMAL1 expression and partially mitigated the IL-1β induced increase in PER2 expression in chondrocytes. These data indicate that the NMDAR/CREB and NF-κB signalling pathways regulate the core circadian clock components PER2 and BMAL1 in chondrocytes. Given that changes in expression of these clock components have been observed in a wide range of diseases, these findings may be broadly relevant for understanding the mechanism leading to circadian clock changes in pathology.