NLRP6 suppresses the inflammatory response of human periodontal ligament cells by inhibiting NF-κB and ERK signal pathways.

AIM: To explore the function and mechanisms of NLRP6 (NOD-, LRR- and pyrin domain-containing 6) in the inflammatory response of human periodontal ligament cells (HPDLCs). METHODOLOGY: Tissues associated with apical periodontitis were obtained from three patients who underwent endodontic microsurgery. The expression of NLRP6 in 3 human apical periodontitis tissues and HPDLCs was examined by immunohistochemistry and immunofluorescence, respectively. The expressions of NLRP6, Phospho(p)- p65, p65, IκB-α, p- IκB-α, ERK, p- ERK, NLRP3, Pro interleukin (IL)-1ß, Pro caspase-1 and apoptosis-associated speck-like protein containing a CARD (ASC) were examined by western blot. The gene expression and secretion of proinflammatory cytokines were detected using quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Data were analysed statistically with independent sample t-tests. RESULTS: NLRP6 was expressed in inflammatory periapical tissues and HPDLCs. Lipopolysaccharide (LPS) from Escherichia coli induced NLRP6 in HPDLCs (P < 0.05). After silencing NLRP6, E. coli LPS-induced activation of NF-κB and ERK signalling was enhanced, which was also accompanied by elevated levels of IL-6 and tumour necrosis factor-α (TNF-α) (P < 0.05). Moreover, knockdown of NLRP6 led to up-regulation of NLRP3, Pro IL-1ß and Pro caspase-1 (P < 0.05), whereas down-regulation of ASC (P < 0.05), which may contribute to unchanged levels of IL-1ß in HPDLCs inflammation. CONCLUSION: NLRP6 was functionally expressed in inflamed periapical tissues and HPDLCs. NLRP6 negatively regulated the production of IL-6 and TNF-α in HPDLCs inflammation by inhibiting NF-κB and ERK signal pathways.

NLRP3 inflammasome may regulate inflammatory response of human periodontal ligament fibroblasts in an apoptosis-associated speck-like protein containing a CARD (ASC)-dependent manner.

AIM: To explore the role of NLRP3 (NACHT [nucleotide-binding oligomerization], LRR [leucine-rich repeat] and PYD [pyrin domain] domains-containing protein 3) inflammasome in the inflammatory response of human periodontal ligament fibroblasts (HPDLFs). METHODOLOGY: The expression of NLRP3 and apoptosis-associated speck-like protein containing a CARD (ASC) in inflammatory periapical tissues and HPDLFs was examined by immunohistochemical and immunofluorescent staining. HPDLFs were stimulated with muramyl dipeptide (MDP) and lipopolysaccharide (LPS) from E. coli with or without the silencing of ASC. The expression of NLRP3, ASC and caspase-1 was examined using quantitative real-time polymerase chain reaction. The secretion of proinflammatory cytokines, including interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumour necrosis factor-α (TNF-α) was measured in the cell supernatant with an enzyme-linked immunosorbent assay. Data were statistically analysed using independent sample t-tests. RESULTS: Immunohistochemistry and immunocytochemistry staining revealed that NLRP3 and ASC were expressed in HPDLFs and inflammatory periapical tissues. MDP and LPS promoted the expression of NLRP3, ASC and caspase-1 in HPDLFs (P < 0.05). The secretion of proinflammatory cytokines was also increased with MDP and LPS stimulation (P < 0.05). After silencing ASC, the secretion of IL-1ß induced by MDP and LPS was significantly attenuated (P < 0.05). CONCLUSION: In HPDLFs, MDP and LPS activated NLRP3 inflammasome and induced IL-1ß secretion. ASC plays an important role in this inflammatory response.