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Relevance of Caspase-1 and Nlrp3 Inflammasome on Inflammatory Bone Resorption in A Murine Model of Periodontitis.

ABSTRACT: This study investigates the role of NLRP3 inflammasome and its main effector Caspase-1 in inflammation and alveolar bone resorption associated with periodontitis. Heat-killed Aggregatibacter actinomycetemcomitans (Aa) was injected 3x/week (4 weeks) into gingival tissues of wild-type (WT), Nlrp3-KO and Caspase1-KO mice. Bone resorption was measured by µCT and osteoclast number was determined by tartrate-resistant acid phosphatase (TRAP) staining. Inflammation was assessed histologically (H/E staining and immunofluorescence of CD45 and Ly6G). In vitro studies determined the influence of Nlrp3 and Caspase-1 in Rankl-induced osteoclast differentiation and activity and on LPS-induced expression of inflammation-associated genes. Bone resorption was significantly reduced in Casp1-KO but not in Nlrp3-KO mice. Casp1-KO mice had increased in osteoclast numbers, whereas the inflammatory infiltrate or on gene expression were similar to those of WT and Nlrp3-KO mice. Strikingly, osteoclasts differentiated from Nlrp3-deficient macrophages had increased resorbing activity in vitro. LPS-induced expression of Il-10, Il-12 and Tnf-? was significantly reduced in Nlrp3- and Casp1-deficient macrophages. As an inceptive study, these results suggest that Nlrp3 inflammasome does not play a significant role in inflammation and bone resorption in vivo and that Caspase-1 has a pro-resorptive role in experimental periodontal disease.

SUBMITTER: Rocha FRG

PROVIDER: S-EPMC7210885 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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Relevance of Caspase-1 and Nlrp3 Inflammasome on Inflammatory Bone Resorption in A Murine Model of Periodontitis.

Rocha Fernanda R G FRG Delitto Andrea E AE de Souza Joao A Chaves JAC González-Maldonado Laura A LA Wallet Shannon M SM Rossa Junior Carlos C

Scientific reports 20200508 1

This study investigates the role of NLRP3 inflammasome and its main effector Caspase-1 in inflammation and alveolar bone resorption associated with periodontitis. Heat-killed Aggregatibacter actinomycetemcomitans (Aa) was injected 3x/week (4 weeks) into gingival tissues of wild-type (WT), Nlrp3-KO and Caspase1-KO mice. Bone resorption was measured by µCT and osteoclast number was determined by tartrate-resistant acid phosphatase (TRAP) staining. Inflammation was assessed histologically (H/E stai ...[more]

PMID: 32385413

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Project description:High mobility group box 1 (HMGB1) is a non-histone DNA-binding protein that is secreted into the extracellular milieu in response to inflammatory stimuli. The secreted HMGB1 mediates various inflammatory diseases, including periodontitis; however, the underlying mechanisms of HMGB1-induced periodontal inflammation are not completely understood. Here, we examined whether anti-HMGB1 neutralizing antibody inhibits periodontal progression and investigated the molecular pathology of HMGB1 in vitro and in vivo. In vitro analysis indicated that HMGB1, granulocyte-macrophage colony-stimulating factor (GM-CSF), and interleukin-1β (IL-1β) were secreted in response to tumor necrosis factor-α (TNF-α) stimuli in human gingival epithelial cells (HGECs) and human monocytic leukemia cells (THP-1) treated with phorbol myristate acetate. Increased levels of GM-CSF and IL-1β were observed in the conditioned media from TNF-α-stimulated HGECs and THP-1 in vitro Simultaneous stimulation with TNF-α and anti-HMGB1 antibody significantly decreased TNF-α-induced inflammatory cytokine secretion. Experimental periodontitis was induced in mice using Porphyromonas gingivalis-soaked ligatures. The extracellular translocation was confirmed in gingival epithelia in the periodontitis model mice by immunofluorescence analysis. Systemic administration of anti-HMGB1 neutralizing antibody significantly inhibited translocation of HMGB1. The anti-HMGB1 antibody inhibited periodontal inflammation, expression of IL-1β and C-X-C motif chemokine ligand 1 (CXCL1), migration of neutrophils, and bone resorption, shown by bioluminescence imaging of myeloperoxidase activity, quantitative reverse transcription-PCR (RT-PCR), and micro-computed tomography analysis. These findings indicate that HMGB1 is secreted in response to inflammatory stimuli caused by periodontal infection, which is crucial for the initiation of periodontitis, and the anti-HMGB1 antibody attenuates the secretion of a series of inflammatory cytokines, consequently suppressing the progression of periodontitis.

Project description:BackgroundPhosphoinositide 3-kinase (PI3K) is located within cells, and is involved in regulating cell survival, proliferation, apoptosis and angiogenesis. The purpose of this study was to investigate the role of PI3K in the process of bone destruction in apical periodontitis, and provide reference data for the treatment of this disease.MethodsThe relative mRNA expression of PI3K, Acp5 and NFATc1 in the normal human periodontal ligament and in chronic apical periodontitis were analyzed by real-time quantitative polymerase chain reaction (RT-qPCR). A mouse model of apical periodontitis was established by root canal exposure to the oral cavity, and HE staining was used to observe the progress of apical periodontitis. Immunohistochemical staining was used to detect the expression of PI3K and AKT in different stages of apical periodontitis, while enzymatic histochemical staining was used for detection of osteoclasts. An Escherichia coli lipopolysaccharide (LPS)-mediated inflammatory environment was also established at the osteoclast and osteoblast level, and osteoclasts or osteoblasts were treated with the PI3K inhibitor LY294002 to examine the role of PI3K in bone resorption.ResultsThe expression of PI3K, Acp5 and NFATc1 genes in chronic apical periodontitis sample groups was significantly increased relative to healthy periodontal ligament tissue (P < 0.05). Mouse apical periodontitis was successfully established and bone resorption peaked between 2 and 3 weeks (P < 0.05). The expression of PI3K and Akt increased with the progression of inflammation, and reached a peak at 14 days (P < 0.05). The gene and protein expression of PI3K, TRAP and NFATc1 in osteoclasts were significantly increased (P < 0.05) in the E. coli LPS-mediated inflammatory microenvironment compared to the normal control group. Meanwhile in osteoblasts, the gene and protein expression of PI3K, BMP-2 and Runx2 were significantly reduced (P < 0.05) in the inflammatory microenvironment. With the addition of LY294002, expressions of bone resorption-related factors (TRAP, NFATc1) and bone formation-related factors (BMP-2, Runx2) significantly decreased (P < 0.05).ConclusionsUnder the inflammatory environment induced by LPS, PI3K participates in the occurrence and development of chronic apical periodontitis by regulating the proliferation and differentiation of osteoclasts and osteoblasts.

Dataset Information

Relevance of Caspase-1 and Nlrp3 Inflammasome on Inflammatory Bone Resorption in A Murine Model of Periodontitis.

Publications

Relevance of Caspase-1 and Nlrp3 Inflammasome on Inflammatory Bone Resorption in A Murine Model of Periodontitis.

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