AUTHOR=Li Weijie , Wang Kexin , Liu Yudong , Wu Hao , He Yan , Li Congchong , Wang Qian , Su Xiaohui , Yan Shikai , Su Weiwei , Zhang Yanqiong , Lin Na 

TITLE=A Novel Drug Combination of Mangiferin and Cinnamic Acid Alleviates Rheumatoid Arthritis by Inhibiting TLR4/NFκB/NLRP3 Activation-Induced Pyroptosis

JOURNAL=Frontiers in Immunology

VOLUME=Volume 13 - 2022

YEAR=2022

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2022.912933

DOI=10.3389/fimmu.2022.912933

ISSN=1664-3224

ABSTRACT=As a traditional Chinese medicine (TCM)-originated disease-modifying anti-rheumatic prescription, Baihu-Guizhi decoction (BHGZD) exerts a satisfying clinical efficacy for rheumatoid arthritis (RA) therapy. Although we previously verified its immunomodulatory and anti-inflammatory activities, bioactive compounds (BACs) of BHGZD and the underlying mechanisms remain unclear. Herein, an integrative research strategy combining UFLC-Q-TOF-MS/MS, gene expression profiling, network calculation, pharmacokinetic profiling, surface plasmon resonance, microscale thermophoresis, and pharmacological experiments was carried out to identified the putative targets of BHGZD and underlying bioactive compounds. Further, in vitro and in vivo experiments were established to determine the drug effect and pharmacological mechanism. As a result, the calculation and functional modularization based on the interaction network of "RA-related gene-BHGZD effective gene" screened the TLR4/PI3K/AKT/NFκB/NLRP3 signaling-mediated pyroptosis to be one of the candidate effective targets of BHGZD for reversing the imbalance network of "immune-inflammation" during RA progression. In addition, both mangiferin (MG) and cinnamic acid (CA) were identified as representative BACs acting on that target, for the strong binding affinities between compounds and target proteins, well pharmacokinetic features, and similar pharmacological effects to BHGZD. Notably, both BHGZD and the two-BAC-combination of MG and CA effectively improved disease severity of adjuvant-induced arthritis modified rat model, including elevating pain thresholds, relieving joint inflammation and bone erosion via inhibiting NF-κB via TLR4/PI3K/AKT signaling to suppress the activation of the NLRP3 inflammasome, leading to the down-regulation of downstream caspase-1, the reduced release of IL-1β and IL-18 and the modulation of GSDMD-mediated pyroptosis. The consistent data were obtained based on the in vitro pyroptosis cellular models of Raw264.7 and MH7A cells induced by LPS/ATP. In conclusion, our data offer an evidence that MG and CA combination identified from BHGZD may interact with TLR4/PI3K/AKT/NFκB signaling to inhibit NLRP3 inflammasome activation and modulate pyroptosis, which provides the novel representative BACs and pharmacological mechanisms of BHGZD against active RA. The findings may shed new light on the mechanisms of the TCM formula, and promote the modernization development of TCM and drug discovery.