Apoptosis Transcriptional Profile Induced by Porphyromonas gingivalis HmuY.

This study aimed at evaluating the transcriptional profile of apoptosis-related genes after in vitro stimulation of peripheral blood mononuclear cells (PBMCs) derived from individuals with periodontitis (P) and healthy nonperiodontitis (NP) control subjects with P. gingivalis HmuY protein. PBMCs from the P and NP groups were stimulated with HmuY P. gingivalis protein, and the expression of genes related to apoptosis was assessed by custom real-time polymerase chain reaction array (Custom RT2 PCR Array). Compared with the NP group, the P group showed low relative levels of apoptosis-related gene expression, downregulated for FAS, FAS ligand, TNFSF10 (TRAIL), BAK1, CASP9, and APAF1 after P. gingivalis HmuY protein stimulation. Furthermore, the P group exhibited low levels of relative gene expression, downregulated for CASP7 when the cells were not stimulated. Our data suggest that P. gingivalis HmuY protein might participate differently in the modulation of the intrinsic and extrinsic apoptosis pathways.

Porphyromonas gingivalis HmuY stimulates expression of Bcl-2 and Fas by human CD3+ T cells.

BACKGROUND: Apoptosis is a highly controlled process of cell death that can be induced by periodontopathogens. The present study aimed to investigate the expression of Fas and Bcl-2 proteins by CD3+ T cells in vitro under stimulation by total Porphyromonas gingivalis antigens and purified recombinant P. gingivalis HmuY protein. RESULTS: CD3+ T cells derived from CP patients and stimulated with HmuY expressed higher levels of Bcl-2 compared to identical cells stimulated with P. gingivalis crude extract or cells derived from NP control subjects (p = 0.043). CONCLUSION: The authors hypothesize that P. gingivalis HmuY plays a role in the pathogenesis of chronic periodontitis, possibly by reducing or delaying apoptosis in T cells through a pathway involving the Bcl-2 protein.

Prevotella intermedia and periodontitis are associated with severe asthma.

BACKGROUND: Periodontitis, an inflammatory disease of multibacterial etiology that affects the protective and supporting tissues surrounding teeth, can influence the course of respiratory diseases, such as asthma, due to epithelial alterations arising from inflammatory and immunological processes, bronchial remodeling, or by the aspiration of pathogenic colonizers found in periodontal pockets. This study evaluated the levels of periodontal pathogens Prevotella intermedia, Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Aggregatibacter actinomycetemcomitans in the subgingival biofilm of individuals with and without severe asthma. METHODS: A case-control study enrolling 457 individuals (220 with asthma and 237 without asthma) was conducted at the Program for Control of Asthma in Bahia (ProAR) Clinic located in Salvador, Bahia, Brazil. A structured questionnaire was used to obtain data on sociodemographic, health status, and lifestyle habits. A clinical periodontal assessment was performed, including bleeding on probing, probing depth, and clinical attachment level. Subgingival biofilm was collected at the deepest site of each sextant, and bacterial DNA was extracted. Quantitative real-time PCR analysis was performed to detect and relatively quantify periodontopathogens in the biofilm. RESULTS: Statistically significant positive associations were found between periodontitis and severe asthma, (odds ratio [OR]adjusted] : 4.00; 95% confidence interval [CI]: 2.26 to 7.10). High levels of P. intermedia were found in association with the presence of severe asthma (ORadjusted : 2.64; 95% CI: 1.62 to 4.39; P < 0.01). CONCLUSIONS: The present results suggest that periodontitis and P. intermedia are associated with severe asthma. However, the functional consequences of this dysbiosis upon asthma susceptibility and its phenotypes remain unclear.

Production of interferon-gamma, interleukin-6, and interleukin-1ß by human peripheral blood mononuclear cells stimulated with novel lys-gingipain synthetic peptides.

BACKGROUND: Periodontitis is a progressive inflammatory process, and its pathogenesis is related to the presence of a dysbiotic subgingival biofilm that elicits the immune response. Porphyromonas gingivalis is a keystone pathogen, and its Lys-gingipain (Kgp) virulence factor is involved in the pathogen-host interaction through the production of cytokines by host cells, but the specific mechanisms of this interaction have not been elucidated. The present study evaluated the in vitro production of interferon-gamma (IFN-γ), interleukin (IL)-6, and IL-1ß cytokines in response to antigenic stimulation of peripheral blood mononuclear cells (PBMCs) with novel Kgp synthetic peptides. METHODS: Our previous in silico study predicted 16 immunogenic peptides from Kgp protein. Nine peptides derived from different regions of the protein were chemically synthesized. The synthetic peptides Kgp12, 17, and 18 were selected based on the immunoglobulin G immunoreactivity in the serum of patients with periodontitis (P) and individuals without periodontitis (WP), and they were used in in vitro stimulation of PBMC derived from groups P and WP. Enzyme-linked immunosorbent assay and microsphere-based flow cytometric assay were used to verify the levels of the cytokines produced in PBMC cultures after 48 hours. RESULTS: Kgp12, 17, and 18 peptides induced lower production of IFN-γ. Kgp12 induced higher levels of IFN-γ in WP than in P individuals. Kgp12 induced higher production of IL-6 and IL-1ß compared with the other stimuli. CONCLUSION: The novel Kgp synthetic peptides tested herein are immunogenic peptides (epitopes) since they induced the production of cytokines by PBMC and therefore may be useful tools in evaluating the pathogen-host interaction.

Porphyromonas gingivalis antigens differently participate in the proliferation and cell death of human PBMC.

OBJECTIVE: Modulation of cell-mediated immunity by microorganisms in periodontal diseases has been widely studied; however, the proliferative activity and/or programmed death of mononuclear cells under periodontopathogenic stimuli are not yet well understood. The aim of this study was to investigate in vitro proliferation and death of peripheral blood mononuclear cells (PBMC) upon stimulation with Porphyromonas gingivalis (Pg) antigens. DESIGN: In 19 patients with chronic periodontitis (CP) and 16 controls without periodontitis (NP) the following clinical parameters were evaluated: bleeding on probing, probing depth, and clinical attachment level. PBMC were cultured under Pg stimuli and apoptosis/necrosis and proliferation assays were carried out for 18 and 48 h, respectively. Fluorescence of labelled cells was determined using flow cytometry. RESULTS: PBMC of CP and NP subjects exhibited a lower proliferative response to Pg LPS (p<0.05) and HmuY protein (p<0.001) compared with non-stimulated cells. Early apoptosis was induced by Pg LPS (p<0.01) and Pg extract (p<0.05), whilst all antigens induced late apoptosis (Pg LPS: p<0.001; Pg extract: p<0.001; HmuY: p<0.01) and necrosis (Pg LPS: p<0.01; Pg extract: p<0.001; HmuY: p<0.001). Pg LPS induced higher late apoptosis than HmuY (p<0.05). Only Pg LPS-induced necrosis tended to be higher in CP compared with NP. CONCLUSIONS: The inhibitory effect of cell proliferation caused by Pg LPS and HmuY protein is not observed when these antigens comprise Pg extract. Despite induced apoptosis, some still unknown mechanism determines the inflammatory outcome in cell death stimulated by HmuY.

Evidence for the presence of ß-D-galactofuranosyl residues (1->3)-linked to a-D-mannopyranose on the cell membrane in all developmental stages of Trypanosoma cruzi

The lipopeptidophosphoglycan (LPPG) component isolated from epimastigote forms of Trypanosoma cruzi has antigenic properties. Using a rabbit antiserum aginst LPPG, specific for ß-D-galactofuranosyl residues (1-3)-linked to alfa-Dmannopyranose, we identified, by indirect immunofluorescence, the presence of epitopes containing ß-D-galactofuranosyl structures on the cell membrane of epimastigote, amastigote and trypomastigote forms of the parasite. The results demonstrade that all developmental stages of T. cruzi contain similar antigenic determinants on their cell body and flagellar membrane