Discussion
Brucea javanica  (Linn.) Merr. (Simaroubaceae) is a traditional Chinese medicine commonly used for the treatment of inflammatory diseases and malaria (Huang et al., 2017; Yang et al., 2013). YDZG, a quassinoid glucoside, is firstly purified and identified by Zhang et al in 1983 (Zhang et al., 1983). However, as it stands now, quite few papers were found to report its bioactivity. Our previous study indicated that YDZG exhibited anti-complementary activity(Zhan et al., 2018). In the present study, evidence for the direct and covalent inhibition of NLRP3 expression by YDZG is provided. Also, the remarkable anti-inflammatory activity of YDZG was demonstrated through the attenuation of NLRP3 inflammasome activation both in vitro andin vivo . Taken together, these results suggest that YDZG can be used as a lead compound to design new therapeutics against NLRP3-driven diseases.
Previous studies have identified compounds that can directly target NLRP3 to suppress NLRP3 activation (Jiang et al., 2017; Marchetti et al., 2018). In contrast, our results showed that YDZG did not directly bind to NLRP3. However, YDZG could enhance TRIM31 expression, an NLRP3 protein inhibitor, to block NLRP3 inflammasome activation. In addition, activation of the NLRP3 inflammasome is thought be regulated at transcriptional and post-translational levels (Jo et al., 2015). The first signal in inflammasome activation is a priming signal, induced by the toll-like receptor (TLR)/nuclear factor (NF)-kB pathway, to upregulate NLRP3 expression (Swanson et al., 2019). NF-κB plays an important role in regulating immune responses (Liang et al., 2004; Zhang & Ghosh, 2001). However, aberrant activation of NF-κB is linked to an up-regulation of pro-inflammatory mediators in ALI (Everhart et al., 2006). NF-κB binding sequences have also been identified in pro-inflammatory genes such as inducible IL-6 and TNF-α (Cho et al., 2007; Xie et al., 2012). In our study, results suggested that YDZG inhibited LPS-induced NF-κB activation to block the expression of NLRP3.
ROS production can stimulate NLRP3 inflammasome activation and inflammatory cascade reactions (Heid et al., 2013; Shirasuna et al.; Zhou et al.). When stimulated by oxidative stress, Nrf2 is dissociated from Keap1, transferred to the nucleus, and combined with antioxidant response elements (ARE), where it activates the transcription of the HO-1 gene to achieve antioxidant effects (Bao et al.; Sawle et al.; Yang et al., 2015). A recent study found that Nrf2 negatively regulates NLRP3 inflammasome activity by inhibiting ROS (Hu et al.). Our results indicated that YDZG promoted Nrf2 translocation from the cytoplasm into the nucleus and inhibited ROS generation.
As a primary organ of respiration, the lung encounters constant exposure to foreign particles and infectious agents. Chronic respiratory diseases and acute lung injuries are common and widespread problems affecting millions of people worldwide, especially in developing countries (Bellani et al., 2016; Guerin et al., 2015; Lorente et al., 2015). In spite of considerable efforts, there are still no Food and Drug Administration-approved treatments for ALI (Standiford & Ward, 2016). In this study, the protective effect of YDZG on an LPS-induced ALI model was evaluated. Results revealed that YDZG could significantly protect mice from lung damage and respiratory dysfunction. Recent data has shown that NLRP3 inflammasome is involved in the development of ALI (Dolinay et al., 2012). IL-1β is a central pro-inflammatory cytokine in the initiation of inflammation and transient expression of IL-1β alone can induce acute lung injury (Kolb et al., 2001). Additionally, application of the neutralizing antibodies for IL-18, IL-1β, and IL-1R antagonists has reduced lung injury on ALI mouse and rat models (Kuipers et al., 2012; Mulligan & Ward, 1992; Wu et al., 2013). In this study, YDZG decreased IL-1β in the BALF, serum, and lung tissue of mice and reduced NLRP3 and caspase-1 p20 in the lung.
In summary, our results indicated that YDZG effectively inhibited NLRP3 inflammasome activation in macrophages and mouse model of ALI. We discovered that YDZG suppressed NLRP3 inflammasome via the reduction of NF-κB and Nrf2 activation and the promotion of TRIM31 expression, suggesting that YDZG could be a potential agent of treatment with NLRP3-driven ALI.