Abstract
The NLRP3 (NACHT, LRR and PYD domains-containing protein 3) inflammasome has been implicated in a variety of diseases, including atherosclerosis, neurodegenerative diseases, and infectious diseases. Thus, inhibitors of NLRP3 inflammasome have emerged as promising approaches to treat inflammation-related diseases. The aim of this study was to explore the effects of juglone (5-hydroxyl-1,4-naphthoquinone) on NLRP3 inflammasome activation. The inhibitory effects of juglone on nitric oxide (NO) production were assessed in lipopolysaccharide (LPS)-stimulated J774.1 cells by Griess assay, while its effects on reactive oxygen species (ROS) and NLRP3 ATPase activity were assessed. The expression levels of NLRP3, caspase-1, and pro-inflammatory cytokines (IL-1β, IL-18) and cytotoxicity of juglone in J774.1 cells were also determined. Juglone was non-toxic in J774.1 cells when used at 10 μM (p < 0.01). Juglone treatment inhibited the production of ROS and NO. The levels of NLRP3 and cleaved caspase-1, as well as the secretion of IL-1β and IL-18, were decreased by treatment with juglone in a concentration-dependent manner. Juglone also inhibited the ATPase activities of NLRP3 in LPS/ATP-stimulated J774.1 macrophages. Our results suggested that juglone could inhibit inflammatory cytokine production and NLRP3 inflammasome activation in macrophages, and should be considered as a therapeutic strategy for inflammation-related diseases.
Highlights
Innate immune cells, including macrophages and dendritic cells, play crucial roles in the initiation of inflammatory immune responses upon activation of inflammasome complexes that induce the maturation of inflammatory cytokines [1]
To determine the juglone concentration range that is non-toxic to J774.1 cells, cells were treated with increasing concentrations of juglone (3.1–50 μM), followed by 3-(4,5-dimethylthiazol2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay
Juglone reduced the levels of cleaved caspase-1 in a concentration-dependent manner (Figure 4b). These results suggested that juglone could inhibit the interaction of NLRP3 and apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC) and subsequent NLRP3 inflammasome formation by downregulating the expression of NLRP3
Summary
Innate immune cells, including macrophages and dendritic cells, play crucial roles in the initiation of inflammatory immune responses upon activation of inflammasome complexes that induce the maturation of inflammatory cytokines [1]. Pattern-recognition receptors (PRRs) recognize pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs), initiating signaling cascades that lead to inflammation and release of inflammatory cytokines [2,3]. Toll-like receptors (TLRs) and NOD-like receptors (NLRs) called PRR are membrane-bound and cytoplasmic receptors, respectively, which recognize PAMPs and DAMPs [4]. The immune responses initiated by the interaction between TLRs and PAMPs are mediated through MyD88-dependent and TRIF-dependent pathways [2,3,5]. When TLRs recognize and bind lipopolysaccharides (LPS) or other extracellular PAMPs, the transcription factor nuclear factor (NF)-κB is activated and induces the expression of several pro-inflammatory cytokines, including interleukin (IL)-1β and IL-18 [6].
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