Bushen Huoxue decotion-containing serum prevents chondrocyte pyroptosis in a m6A-dependent manner in facet joint osteoarthritis

Background: Facet joint osteoarthritis (FJOA) is a common lumbar osteoarthritis characterized by degeneration of small joint cartilage. Bushen Huoxue decotion (BSHXD) has good therapeutic effects on OA. Our work aimed to further probe the pharmacological effects of BSHXD-containing serum (BSHXD-CS) on FJOA and define underlying the mechanisms invovled.

Methods: To establish a FJOA cell model, primary rat chondrocytes were treated with LPS. The mRNA and protein expressions were assessed using qRT-PCR and western blot, respectively. The secretion levels of pro-inflammatory cytokines were measured by ELISA. Cell viability was determined by CCK8 assay. The global m⁶A level was detected by the kit, and NLRP3 mRNA m⁶A level was determined by Me-RIP assay. The molecular interactions were analyzed by RIP and RNA pull-down assays.

Results: BSHXD-CS treatment relieved LPS-induced cell injury, inflammation, NLRP3 inflammasome and Pyroptosis in chondrocytes (all p < 0.05). LPS-induced NLRP3 upregulation in chondrocytes was related to its high m⁶A modification level (p < 0.05). It was also observed that BSHXD-CS reduced LPS-induced m⁶A modification in chondrocytes via repressing STAT3 (all p < 0.05), suggesting BSHXD-CS could repress NLRP3 expression via m⁶A-dependent manner. Moreover, DAA, a m⁶A specific inhibitor, was proved to strengthen the protectively roles of BSHXD-CS on LPS-challenged pytoptosis (all p < 0.05).

Conclusion: BSHXD-CS inhibited NLRP3 inflammasome activation and Pyroptosis in chondrocytes to repress OA progression by reducing RNA m⁶A modification.

Bushen Huoxue decotion; Facet joint osteoarthritis; NLRP3; Pyroptosis; STAT3; m(6)A.