Vibrio alginolyticus Triggers Inflammatory Response in Mouse Peritoneal Macrophages via Activation of NLRP3 Inflammasome

Abstract

Vibrio alginolyticus is a food-borne marine Vibrio that causes gastroenteritis, otitis media, otitis externa, and septicemia in humans. The pathogenic mechanisms of V. alginolyticus have previously been studied in aquaculture animals; however, the underlying mechanisms in mammals remain unknown. In this study, an in vitro model of mouse peritoneal macrophages infected with V. alginolyticus was established. qPCR results revealed that V. alginolyticus induced the transcription levels of various cytokines, including IL-1β, IL-12, IL-18, TNF-α, IL-17, IL-6, IFN-γ, and IL-10, and the secretion level of IL-1β is the most significant. Inhibition assays with Ac-YVAD-CHO (a caspase-1 inhibitor) and Z-VAD-FMK (a pan-caspase inhibitor) were conducted to determine whether caspase-1 or caspase-11 is involved in V. alginolyticus-triggered IL-1β secretion. Results showed that IL-1β secretion was partly inhibited by Ac-YVAD-CHO and absolutely blocked by Z-VAD-FMK. To explore the sensed pattern recognition receptors, several NLR family members and the AIM2 receptor were detected and many receptors were upregulated especially NLRP3. Moreover, the NLRP3 protein displayed a puncta-like surrounding cell nucleus, which signified that the NLRP3 inflammasome was activated in response to V. alginolyticus infection. Inhibition assays with glyburide and CA-074 methyl ester (K+ outflow inhibitor and cathepsin B inhibitor) blocked IL-1β secretion, which demonstrated the essential role of the NLRP3 inflammasome in inflammatory response. To better understand how V. alginolyticus affects IL-1β release, the NLRP3 inflammasome was detected with doses ranging from 0.1 to 10 MOIs and time periods ranging from 3 to 12 h. Results showed that V. alginolyticus-mediated NLRP3 inflammasome activation was in a time- and dose-dependent manner and IL-1β release peaked at MOI of 1 for 12 h. Most importantly, blocking the NLRP3 inflammasome with inhibitors and the use of NLRP3-/- and caspase-1/11-/- mice could attenuate pro-inflammatory cytokine secretion, such as IL-1β, IL-6, IL-12, and TNF-α. Taken together, our study first found that the NLRP3 inflammasome plays vital roles in V. alginolyticus triggered inflammatory response in mouse peritoneal macrophages. This may provide reference information for the development of potential anti-inflammatory treatments against V. alginolyticus infection.

Inhibition of circular RNA circ_0138959 alleviates pyroptosis of human gingival fibroblasts via the microRNA-527/caspase-5 axis

“…The HGFs in each group were scraped off and washed with PBS (C0221A; Beyotime, Shanghai, China) twice, before resuspending in caspase-1 staining solution (1:30). After 1 h of incubation in a 5% CO 2 incubator at 37°C, PI staining solution (1:100) was added to resuspend the cells [ 26 ].…”

“…Protein expressions of caspase-1, Capase-4, and cleaved N-terminal gasdermin D (GSDMD-N) were measured by Western blotting [ 26 ]. Proteins were extracted from cells and tissues by RIPA (Sigma-Aldrich, Missouri, USA) reagents and were detected using the BCA method (P0012; Beyotime, Shanghai, China).…”

Gut Microbiota-Related Inflammation Factors as a Potential Biomarker for Diagnosing Major Depressive Disorder

“…Inflammatory response is a very complex biological process, which has an important role in many diseases ( Costa et al., 2021 ; Li et al., 2021 ; Lu et al., 2021 ; Wang J. et al., 2021 ). Previous studies showed that inflammatory response played a critical role in the onset of depression, and the disturbance of inflammation levels could increase the incidence of depression ( Leonard, 2018 ; Colasanto et al., 2020 ).…”

A. caviae infection triggers IL-1β secretion through activating NLRP3 inflammasome mediated by NF-κB signaling pathway partly in a TLR2 dependent manner

“…The female wild type (WT) C57BL/6 mice, TLR2-deficient (TLR2 −/− ) mice, NLRP3-deficient (NLRP3 −/− ) mice, caspase-1/11-deicient (caspase-1/11 −/− ) mice were kept in our lab, housed in the pathogen-free equipment and fed with sterile diet. The mouse peritoneal macrophages (PMs) were prepared referring to previous methods [ 17 , 18 ]. The cell purity was analysed by flow cytometry using CD11b (BioLegend).…”

“…A. hydrophila induced IL-1β release and disturbed head kidney macrophages mitochondrial dysfunction and apoptosis [ 16 ]. Moreover, IL-1β would promote TNF-α, IL-6 and IL-12 secretion to aggravate inflammation and cell death in mice macrophages infected with Vibrio alginolyticus and Vibrio harveyi [ 17 , 18 ]. However, whether A. caviae infection induces IL-1β release and the underlying molecular mechanism is needed to be further explored.…”