Pyroptosis is triggered by inflammasomes after its activation by various inflammatory stimulations, including lipopolysaccharide (LPS) and improper pH. This may result in programmed death of the affected cell. It is well known that NLRP1 and NLRP3 inflammasomes mediate the production of various cytokines in inflammatory disorders; however, it is still unknown whether NLRP1 and NLRP3 inflammasomes can influence the LPS‑induced pyroptosis in the progression of knee osteoarthritis (KOA). In the present study, the correlation between the NLRP inflammasomes and fibroblast‑like synoviocytes (FLSs) pyroptosis was investigated in vivo and in vitro. Human synovial samples were collected from KOA patients and the expression of NLRP1 and NLRP3 inflammasomes was analyzed. Human FLS were isolated in vitro and stimulated with LPS. To determine whether NLRP1 and NLRP3 inflammasomes are involved in FLS pyroptosis, NLRP1 and NLRP3 small interfering RNAs (siRNAs) were used. The results showed that the expression of NLRPs and inflammasome‑related proteins were upregulated and FLS stimulated with LPS+ATP resulted in cell pyroptosis. However, LPS+ATP‑induced pyroptosis was attenuated by NLRP1 and NLRP3 siRNAs. The results of the present study indicate that LPS‑induced FLS pyroptosis may be mediated by either NLRP1 or NLRP3 inflammsomes. Overall, based on the data obtained from patients and in vitro cells, the present finsings showed that NLRP1 and NLRP3 inflammasomes are highly involved in the FLS inflammation and pyroptosis. Furthermore, inhibition of NLRP1 and NLRP3 led to a remarkable reduction of pyroptosis‑related cytokines. Thus, NLRP1 and NLRP3 inflammasomes may be important in the pathogenesis of OA and may represent a novel therapeutic target.

Fibroblast-like synoviocytes (FLSs) are the main effector cells of knee osteoarthritis (KOA) synovial fibrosis. Our last report showed that NLRP1 and NLRP3 inflammasomes may mediate LPS/ATP-induced FLSs pyroptosis in KOA. In the present study, we found an elevated hypoxia-inducible factor-1α (HIF-1α) level in the synovial tissue of KOA model rats, and inhibiting the increase of HIF-1α could improve synovial fibrosis in rats. Subsequently, we established LPS/ATP-induced model in FLSs mimicking the inflammatory environment of KOA. FLSs transfected with siRNA HIF-1α showed a reduced cell death; meanwhile, the relative expression of pyroptosis-related proteins was also downregulated. Additionally, FLSs transfected with or without siRNA GSDMD were exposed to hypoxia. GSDMD silencing can significantly reduce both gene and protein levels of fibrogenic markers transforming growth factor-β (TGF-β), procollagen-lysine, 2-oxoglutarate 5-dioxygenase2 (PLOD2), collagen type I α1 chain (COL1A1), and tissue inhibitor of metalloproteinases 1 (TIMP1). Taken together, our findings indicate that increased HIF-1α is highly involved in the KOA synovial fibrosis. Moreover, elevated HIF-1α may aggravate synovial fibrosis via FLS pyroptosis.

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Transient receptor potential ankyrin 1 (TRPA1) is a membrane-associated cation channel, widely expressed in neuronal and non-neuronal cells. Recently, emerging evidences suggested the crucial role of TRPA1 in the disease progression of osteoarthritis (OA). Therefore, we aimed to investigate whether TRPA1 mediate lipopolysaccharide (LPS)-induced inflammatory responses in primary human OA fibroblast-like synoviocytes (OA-FLS). The expression of TRPA1 in LPS-treated OA-FLS was assessed by polymerase chain reaction (PCR) and western blot (WB), and the functionality of TRPA1 channel by Ca influx measurements. Meanwhile, production of interleukin (IL)-1β, tumor necrosis factor (TNF)-α, IL-6, matrix metalloproteinase (MMP)-1, and MMP-3 in LPS-treated cells was measured by immunoassay. Histological observation after inhibition of TRPA1 was also performed in rats with LPS-induced inflammatory arthritis. After being induced by LPS, the gene and protein expression of TRPA1 was increased in the time-dependent or dose-dependent manner. Meanwhile, Ca influx mediated by TRPA1 in human OA-FLS was also enhanced. In addition, pharmacological inhibition and gene silencing of TRPA1 downregulated the production of IL-1β, TNF-α, IL-6, MMP-1, and MMP-3 in LPS-treated FLS. Finally, synovial inflammation and cartilage degeneration were also reduced by the TRPA1 antagonist. We found the LPS caused the increased functional expression of TRPA1, the activation of which involved in LPS-reduced inflammatory responses in primary human OA-FLS, and the inhibition of TRPA1 produces protective effect in LPS-induced arthritis.