Periplogenin inhibits pyroptosis of fibroblastic synoviocytes in rheumatoid arthritis through the NLRP3/Caspase-1/GSDMD signaling pathway.

Although the pathogenesis of rheumatoid arthritis (RA) remains unclear, an increasing number of studies have confirmed that pyroptosis of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) is an important factor affecting the progression of RA. Periplogenin (PPN) is a natural cardiac glycoside; reportedly, it exerts anti-inflammatory and analgesic effects in diseases by inhibiting cell growth and migration. This study aimed to determine the effect of PPN on the growth, migration, and invasion of RA-FLS and the potential mechanism of pyroptosis regulation. We discovered that PPN could inhibit the migration and invasion abilities of RA-FLS and block their growth cycle, down-regulate the secretion and activation of NLRP3, Caspase-1, GSDMD, IL-1β, and IL-18, and reduce the number of pyroptosis. In summary, PPN inhibited pyroptosis, reduced the release of inflammatory factors, and improved RA-FLS inflammation by regulating the NLRP3/Caspase-1/GSDMD signaling pathway.

Ma X ，Yang Y ，Li H ，Luo Z ，Wang Q ，Yao X ，Tang F ，Huang Y ，Ling Y ，Ma W ... -  《-》

SMAD2 inhibits pyroptosis of fibroblast-like synoviocytes and secretion of inflammatory factors via the TGF-β pathway in rheumatoid arthritis.

Mao X ，Wu W ，Nan Y ，Sun W ，Wang Y ... -  《-》

The ROS/GRK2/HIF-1α/NLRP3 Pathway Mediates Pyroptosis of Fibroblast-Like Synoviocytes and the Regulation of Monomer Derivatives of Paeoniflorin.

Hypoxia is an important factor in the development of synovitis in rheumatoid arthritis (RA). The previous study of the research group found that monomeric derivatives of paeoniflorin (MDP) can alleviate joint inflammation in adjuvant-induced arthritis (AA) rats by inhibiting macrophage pyroptosis. This study revealed increased levels of hypoxia-inducible factor- (HIF-) 1α and N-terminal p30 fragment of GSDMD (GSDMD-N) in fibroblast-like synoviocytes (FLS) of RA patients and AA rats, while MDP significantly inhibited their expression. Subsequently, FLS were exposed to a hypoxic environment or treated with cobalt ion in vitro. Western blot and immunofluorescence analysis showed increased expression of G protein-coupled receptor kinase 2 (GRK2), HIF-1α, nucleotide-binding oligomerization segment-like receptor family 3 (NLRP3), ASC, caspase-1, cleaved-caspase-1, and GSDMD-N. Electron microscopy revealed FLS pyroptosis after exposure in hypoxia. Next, corresponding shRNAs were transferred into FLS to knock down hypoxia-inducible factor- (HIF-) 1α, and in turn, NLRP3 and western blot results confirmed the same. The enhanced level of GSDMD was reversed under hypoxia by inhibiting NLRP3 expression. Knockdown and overexpression of GRK2 in FLS revealed GRK2 to be a positive regulator of HIF-1α. Levels of GRK2 and HIF-1α were inhibited by eliminating excess reactive oxygen species (ROS). Furthermore, MDP reduced FLS pyroptosis through targeted inhibition of GRK2 phosphorylation. According to these findings, hypoxia induces FLS pyroptosis through the ROS/GRK2/HIF-1α/NLRP3 pathway, while MDP regulates this pathway to reduce FLS pyroptosis.

Hong Z ，Zhang X ，Zhang T ，Hu L ，Liu R ，Wang P ，Wang H ，Yu Q ，Mei D ，Xue Z ，Zhang F ，Zhang L ... -  《-》

Natural compound library screening to identify berberine as a treatment for rheumatoid arthritis.

Fibroblast-like synoviocytes (FLS) play a critical role on the exacerbation and deterioration of rheumatoid arthritis (RA). Aberrant activation of FLS pyroptosis signaling is responsible for the hyperplasia of synovium and destruction of cartilage of RA. This study investigated the screened traditional Chinese medicine berberine (BBR), an active alkaloid extracted from the Coptis chinensis plant, that regulates the pyroptosis of FLS and secretion of inflammatory factors in rheumatoid arthritis. First, BBR was screened using a high-throughput drug screening strategy, and its inhibitory effect on RA-FLS was verified by in vivo and in vitro experiments. Second, BBR was intraperitoneally administrated into the collagen-induced arthritis rat model, and the clinical scores, arthritis index, and joint HE staining were evaluated. Third, synovial tissues of CIA mice were collected, and the expression of NLRP3, cleaved-caspase-1, GSDMD-N, Mst1, and YAP was detected by Western blot. The administration of BBR dramatically alleviated the severity of collagen-induced arthritis rat model with a decreased clinical score and inflammation reduction. In addition, BBR intervention significantly attenuates several pro-inflammatory cytokines (interleukin-1β, interleukin-6, interleukin-17, and interleukin-18). Moreover, BBR can reduce the pyroptosis response (caspase-1, NLR family pyrin domain containing 3, and gasdermin D) of the RA-FLS in vitro, activating the Hippo signaling pathway (Mammalian sterile 20-like kinase 1, yes-associated protein, and transcriptional enhanced associate domains) so as to inhibit the pro-inflammatory effect of RA-FLS. These results support the role of BBR in RA and may have therapeutic implications by directly repressing the activation, migration of RA-FLS, which contributing to the attenuation of the progress of CIA. Therefore, targeting PU.1 might be a potential therapeutic approach for RA. Besides, BBR inhibited RA-FLS pyroptosis by downregulating of NLRP3 inflammasomes (NLRP3, caspase-1) and eased the pro-inflammatory activities via activating the Hippo signaling pathway, thereby improving the symptom of CIA.

Zhang L ，Tan M ，Mao J ，Zhang J ，Wang XY ，Zhang Y ，Duo RX ，Hao JY ，Shen HL ... -  《-》

The monomer derivative of paeoniflorin inhibits macrophage pyroptosis via regulating TLR4/ NLRP3/ GSDMD signaling pathway in adjuvant arthritis rats.

This study was aimed to investigate the effect of monomer derivative of paeoniflorin (MDP) on macrophage pyroptosis mediated by TLR4/NLRP3/GSDMD signaling pathway in adjuvant arthritis (AA) rats. Wistar rats were divided into normal group, AA model group, MDP (50 mg/kg) group and MTX (0.5 mg/kg) group. The expression of TLR4, NLRP3 and GSDMD in macrophage were detected by immunofluorescence assay. The expression of TLR4 and the ratio of macrophage pyroptosis were analyzed by flow cytometry. Cell morphology was observed by scanning electron microscopy. The cytokine levels of IL-18 and IL-1β were detected by ELISA. The expressions of proteins related to macrophage pyroptosis were detected by western blot. MDP has a therapeutic effect on rats AA by reducing the secondary inflammation and improving pathological changes. The results of X-ray imaging and ultrasound images showed that MDP could inhibit bone erosion, soft tissue swelling, and joint space narrowing. Macrophage pyroptosis was found in secondary inflammation of AA rats. The expressions of TLR4, MyD88, NLRP3, Caspase-1, ASC, and GSDMD-N in macrophage were increased, the levels of IL-18 and IL-1β were enhanced, and the morphology of pyroptosis could be observed. MDP could inhibit macrophage polarization and macrophage pyroptosis, and down-regulated the cytokine levels of IL-18 and IL-1β in AA rats. MDP could regulate the M1/M2 ratio, decreased the ratio of macrophage pyroptosis and down-regulated the expressions of TLR4, MyD88, NLRP3, Caspase-1, ASC, and GSDMD-N in vivo and in vitro. Abnormal activation of TLR4/NLRP3/GSDMD signaling pathway may be involved in macrophage pyroptosis in AA rats. The therapeutic effect of MDP on AA rats is related to the inhibition of macrophage pyroptosis by regulating the TLR4/NLRP3/GSDMD signaling pathway.

Xu L ，Wang H ，Yu QQ ，Ge JR ，Zhang XZ ，Mei D ，Liang FQ ，Cai XY ，Zhu Y ，Shu JL ，Tai Y ，Wei W ，Zhang LL ... -  《-》