NLRP3 inhibitors(Bengbu Medical College)

To investigate the mechanism of 2, 6-dimethoxy-1, 4-benzoquinone (DMQ), an active ingredients in fermented wheat germ extract, for inhibiting NLRP3 inflammasome activation and alleviating septic shock in mice.

Cultured murine bone marrow-derived macrophages (BMDM) stimulated with lipopolysaccharide (LPS) were treated with DMQ, followed by treatment with Nigericin, ATP, and MSU for activating the canonical NLRP3 inflammasome; the noncanonical NLRP3 inflammasome was activated by intracellular transfection of LPS, and AIM2 inflammasome was activated using Poly A: T.In human monocytic THP-1 cells, the effect of Nigericin on inflammasome activation products was examined using Western blotting and ELISA.Co-immunoprecipitation was performed to explore the mechanism of DMQ-induced blocking of NLRP3 inflammasome activation.In a male C57BL/6J mouse model of LPS-induced septic shock treated with 20 and 40 mg/kg DMQ, the levels of IL-1β and TNF-α in the serum and peritoneal lavage fluid were determined using ELISA, and the survival time of the mice within 36 h was observed.

Treatment with DMQ effectively inhibited LPS-induced activation of canonical NLRP3 inflammasome in mouse BMDM and human THP-1 cells and also inhibited non-canonical NLRP3 inflammasome activation in mouse BMDM, but produced no significant effect on AIM2 inflammasome activation.DMQ significantly blocked the binding between ASC and NLRP3.In the mouse models of septic shock, DMQ treatment significantly reduced the levels of IL-1β in the serum and peritoneal fluid and obviously prolonged survival time of the mice.

DMQ can effectively block ASC-NLRP3 interaction to inhibit NLRP3 inflammasome activation and alleviate LPSinduced septic shock in mice.

Coumarin was detected as one of the most abundant compounds by nontargeted analysis of natural product components in actual water samples prior to disinfection. More importantly, prechlorination of humic acid generated 3-hydroxycoumarin and monohydroxy-monomethyl-substituted coumarin with a total yield of ≤10.1%, which suggested the humic substance in raw water is an important source of coumarins. 7-Hydroxycoumarin, 6-hydroxy-4-methylcoumarin, 6,7-dihydroxycoumarin, and 7-methoxy-4-methylcoumarin were identified in raw water by high-performance liquid chromatography-tandem high-resolution mass spectrometry because only some coumarin standards were commercially available. Their chlorination generated monochlorinated and polychlorinated coumarins, and their structures were confirmed by the synthesized standards. These products could form at various dosages of chlorine and pH levels, and some with a concentration of 600 ng/L can be stable in tap water for days. 3,6,8-Trichloro-7-hydroxycoumarin, 3-chloro-7-methoxy-4-methylcoumarin, and 3,6-dichloro-7-methoxy-4-methylcoumarin were first identified in finished water with concentrations of 0.0670, 78.1, and 14.7 ng/L, respectively, but not in source water, suggesting that they are new DBPs formed during disinfection. The cytotoxicity of 3-chloro-7-methoxy-4-methylcoumarin in CHO-K1 cells was comparable to those of 2,6-dibromo-1,4-benzoquinone and 2,6-dichloro-1,4-benzoquinone in TIC-Tox analyses, suggesting that further investigation of their occurrence and control in drinking water systems is warranted.