Periodontal bacteria in the brain-Implication for Alzheimer's disease: A systematic review.

Periodontitis is a chronic non-communicable disease caused by a dysbiotic microbiota. Pathogens can spread to the bloodstream, colonize other tissues or organs, and favor the onset of other pathologies, such as Alzheimer's disease (AD). Pathogens could permanently or transiently colonize the brain and induce an immune response. Thus, we analyzed the evidence combining oral bacteria's detection in the brain, both in animals and humans affected with AD. This systematic review was carried out following the PRISMA guideline. Studies that detected oral bacteria at the brain level were selected. The search was carried out in the Medline, Latindex, SciELO, and Cochrane Library databases. SYRCLE tool and Newcastle-Ottawa Scale were used for the risk of bias assessment. 23 studies were selected according to the eligibility criteria. Infection with oral pathogens in animals was related to developing neuropathological characteristics of AD and bacteria detection in the brain. In patients with AD, oral bacteria were detected in brain tissues, and increased levels of pro-inflammatory cytokines were also detected. There is evidence of a microbiological susceptibility to develop AD when the most dysbiosis-associated oral bacteria are present. The presence of bacteria in the brain is related to AD's pathological characteristics, suggesting an etiological oral-brain axis.

Neutrophil N1 and N2 Subsets and Their Possible Association with Periodontitis: A Scoping Review.

Periodontitis is a chronic non-communicable disease caused by dysbiotic changes that affect the subgingival microbiota. During periodontitis, neutrophils play a central role in the initial recognition of bacteria, and their number increases with the appearance of the first signs of periodontal inflammation. Recent evidence has led to the proposition that neutrophils can also functionally polarize, determining selective activity patterns related to different diseases. Two well-defined neutrophil phenotypes have been described, the pro-inflammatory N1 subset and the suppressor N2 subset. To date, it has not been established whether these different neutrophil subtypes play a role in the pathogenesis of periodontitis. Thus, this scoping review aimed to determine whether there was evidence to suggest that the neutrophils present in periodontal tissues can be associated with certain phenotypes. The research question, population, concept, and context sought to identify original articles, in humans, that detected the presence of neutrophils in the periodontal tissues of people affected by periodontitis. Based on the search strategy, we found 3658 studies. After removing the papers with abstracts not related to the outcome measures and eligibility criteria, 16 articles were included for qualitative analysis. Several studies identified the presence of different neutrophil subsets, specifically, the naive, pro- and para-inflammatory, hyper-reactive and hyper-active, and high- and low-responder phenotypes. The existing evidence demonstrates the presence of pro-inflammatory, hyper-reactive and high-responder neutrophils in periodontal tissues affected with periodontitis. There is no evidence demonstrating the presence of the N1 or N2 phenotypes in periodontal tissues during periodontitis. However, the existence of pro-inflammatory phenotypes, which increase NETosis and degranulation, and increase the production of pro-inflammatory cytokines, could be suggestive of the N1 phenotypes.

Gingival crevicular fluid as Biomarker's source for alzheimer's disease / Fluido crevicular gingival como biomarcador de enfermedad de Alzheimer

Abstract Periodontitis is a low-grade inflammatory disease caused by a subgingival dysbiotic microbiota. Multiple studies have determined the higher prevalence of tooth loss and poor oral hygiene in patients with Alzheimer's disease (AD). However, the periodontal diagnosis, periodontal bacteria or mediators has not been measured to date. Aim: To determine the periodontal status, the pro-inflammatory mediators, Porphyromonas gingivalis load, and Apoliporpotein E (ApoE) in patients with AD. A complete dental examination was performed on 30 patients, and cognitive status was determined by the Montreal Cognitive Assessment (MoCA). Subgingival microbiota and GCF samples were then taken from all patients from the deepest sites. Total DNA was isolated from the microbiota samples for the quantification of the 16S ribosomal subunit. Pro-inflammatory mediators and ApoE were quantified from the gingival crevicular fluid (GCF). Patients with AD had periodontitis stage III-IV in 80%, a higher concentration of pro-inflammatory and ApoE mediators, and a higher P. gingivalis load compared to healthy subjects. The pro-inflammatory mediators, P. gingivalis load had a negative correlation with the MoCA test scores. Finally, a ROC curve was performed to assess the specificity and sensitivity of ApoE levels, detecting an area of 84.9%. In AD patients, we found a more severe periodontitis, a higher levels of pro-inflammatory mediators, and higher bacterial load. In addition, there is an increase in ApoE that allows to clearly determine patients with health, periodontitis and periodontitis and AD.

Resumen La periodontitis es una enfermedad crónica no transmisible que se caracteriza por generar una inflamación sistémica de bajo grado causada por una microbiota disbiótica subgingival. Múltiples estudios han determinado la mayor prevalencia de pérdida de dientes y mala higiene bucal en pacientes con enfermedad de Alzheimer (EA). Sin embargo, el diagnóstico periodontal, bacterias periodontales o mediadores pro-inflamatorio no se ha medido hasta la fecha. Determinar el estado periodontal, los mediadores pro-inflamatorios, la carga de Porphyromonas gingivalis y la apoliporpoteína E (ApoE) en pacientes con EA. Se realizó un examen odontológico completo en 30 pacientes y el estado cognitivo se determinó mediante la Evaluación Cognitiva de Montreal (MoCA). Luego, se tomaron muestras de microbiota subgingival y FCG de todos los pacientes de los sitios más profundos. Se aisló el DNA total de las muestras de microbiota para la cuantificación de la subunidad ribosómica 16S. Los mediadores pro-inflamatorios y la ApoE se cuantificaron a partir del líquido crevicular gingival (GCF). Los pacientes con EA tenían periodontitis en estadio III-IV en 80%, una mayor concentración de mediadores pro-inflamatorios y ApoE, y una mayor carga de P. gingivalis en comparación con los sujetos sanos. Los mediadores pro-inflamatorios y la carga de P. gingivalis tuvieron una correlación negativa con las puntuaciones de la prueba MoCA. Finalmente, se realizó una curva ROC para evaluar la especificidad y sensibilidad de los niveles de ApoE, detectando un área del 84,9%. En los pacientes con EA encontramos una periodontitis más severa, mayores niveles de mediadores pro-inflamatorios y mayor carga bacteriana. Además, un aumento de ApoE que permite determinar claramente a los pacientes con salud, periodontitis y periodontitis y EA.

Natural Killer T (NKT) Cells and Periodontitis: Potential Regulatory Role of NKT10 Cells.

Natural killer T (NKT) cells constitute a unique subset of T lymphocytes characterized by specifically interacting with antigenic glycolipids conjugated to the CD1d receptor on antigen-presenting cells. Functionally, NKT cells are capable of performing either effector or suppressor immune responses, depending on their production of proinflammatory or anti-inflammatory cytokines, respectively. Effector NKT cells are subdivided into three subsets, termed NKT1, NKT2, and NKT17, based on the cytokines they produce and their similarity to the cytokine profile produced by Th1, Th2, and Th17 lymphocytes, respectively. Recently, a new subgroup of NKT cells termed NKT10 has been described, which cooperates and interacts with other immune cells to promote immunoregulatory responses. Although the tissue-specific functions of NKT cells have not been fully elucidated, their activity has been associated with the pathogenesis of different inflammatory diseases with immunopathogenic similarities to periodontitis, including osteolytic pathologies such as rheumatoid arthritis and osteoporosis. In the present review, we revise and discuss the pathogenic characteristics of NKT cells in these diseases and their role in the pathogenesis of periodontitis; particularly, we analyze the potential regulatory role of the IL-10-producing NKT10 cells.

Periodontitis and alzheimer's disease

BACKGROUND: Alzheimer's disease (AD), the main cause of dementia in the adult population, is characterized by a progressive loss of cognitive function. It is considered that neuroinflammation plays a fundamental role in its onset and progression. The bacteria present in the disbiotic microbiome generated during the course of periodontitis (PE) are capable of inducing a systemic inflammatory response, exacerbating the production of proinflammatory mediators that have the potential to spread to the systemic circulation. MATERIAL AND METHODS: A literature review was made using the databases Scielo, PubMed, EBSCO and key words "Alzheimer disease", "Periodontitis", "Neurodegeneration", "Inflammation mediators", "Elderly". RESULTS: Several hypotheses point to similar pathophysiological pathways in the establishment of AD and PE, sharing cellular and molecular proinflammatory characteristics. In periodontitis, locally produced cytokines and pro-inflammatory products spread from the ulcerated periodontal pocket into the systemic circulation, or around the trigeminal nerve terminals, which allows the passage of bacteria or their products to the brain. This fact leads to the formation of plaques of amyloid peptide and intraneuronal neurofibrillar tangles (NFTs) that activate the glial cells producing a significant increase in proinflammatory cytokines in the affected regions that lead to a loss of neuronal synapses and neurodegeneration, contributing to the progression of AD. CONCLUSIONS: This review of the literature contributes to the understanding of the pathological pathways shared by both diseases such as oxidative damage and inflammation. There is not enough evidence to determine an association between this two pathologies, so it is considered necessary to conduct studies for determine if periodontitis is capable of inducing or exacerbating the neuroinflammation that will trigger AD

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Oral-Gut-Brain Axis in Experimental Models of Periodontitis: Associating Gut Dysbiosis With Neurodegenerative Diseases.

Periodontitis is considered a non-communicable chronic disease caused by a dysbiotic microbiota, which generates a low-grade systemic inflammation that chronically damages the organism. Several studies have associated periodontitis with other chronic non-communicable diseases, such as cardiovascular or neurodegenerative diseases. Besides, the oral bacteria considered a keystone pathogen, Porphyromonas gingivalis, has been detected in the hippocampus and brain cortex. Likewise, gut microbiota dysbiosis triggers a low-grade systemic inflammation, which also favors the risk for both cardiovascular and neurodegenerative diseases. Recently, the existence of an axis of Oral-Gut communication has been proposed, whose possible involvement in the development of neurodegenerative diseases has not been uncovered yet. The present review aims to compile evidence that the dysbiosis of the oral microbiota triggers changes in the gut microbiota, which creates a higher predisposition for the development of neuroinflammatory or neurodegenerative diseases.The Oral-Gut-Brain axis could be defined based on anatomical communications, where the mouth and the intestine are in constant communication. The oral-brain axis is mainly established from the trigeminal nerve and the gut-brain axis from the vagus nerve. The oral-gut communication is defined from an anatomical relation and the constant swallowing of oral bacteria. The gut-brain communication is more complex and due to bacteria-cells, immune and nervous system interactions. Thus, the gut-brain and oral-brain axis are in a bi-directional relationship. Through the qualitative analysis of the selected papers, we conclude that experimental periodontitis could produce both neurodegenerative pathologies and intestinal dysbiosis, and that periodontitis is likely to induce both conditions simultaneously. The severity of the neurodegenerative disease could depend, at least in part, on the effects of periodontitis in the gut microbiota, which could strengthen the immune response and create an injurious inflammatory and dysbiotic cycle. Thus, dementias would have their onset in dysbiotic phenomena that affect the oral cavity or the intestine. The selected studies allow us to speculate that oral-gut-brain communication exists, and bacteria probably get to the brain via trigeminal and vagus nerves.

Alzheimer's Disease-Like Pathology Triggered by Porphyromonas gingivalis in Wild Type Rats Is Serotype Dependent.

Periodontal disease is a disease of tooth-supporting tissues. It is a chronic disease with inflammatory nature and infectious etiology produced by a dysbiotic subgingival microbiota that colonizes the gingivodental sulcus. Among several periodontal bacteria, Porphyromonas gingivalis (P. gingivalis) highlights as a keystone pathogen. Previous reports have implied that chronic inflammatory response and measurable bone resorption are observed in young mice, even after a short period of periodontal infection with P. gingivalis, which has been considered as a suitable model of experimental periodontitis. Also, encapsulated P. gingivalis strains are more virulent than capsular-defective mutants, causing an increased immune response, augmented osteoclastic activity, and accrued alveolar bone resorption in these rodent experimental models of periodontitis. Recently, P. gingivalis has been associated with Alzheimer's disease (AD) pathogenesis, either by worsening brain pathology in AD-transgenic mice or by inducing memory impairment and age-dependent neuroinflammation middle-aged wild type animals. We hypothesized here that the more virulent encapsulated P. gingivalis strains could trigger the appearance of brain AD-markers, neuroinflammation, and cognitive decline even in young rats subjected to a short periodontal infection exposure, due to their higher capacity of activating brain inflammatory responses. To test this hypothesis, we periodontally inoculated 4-week-old male Sprague-Dawley rats with K1, K2, or K4 P. gingivalis serotypes and the K1-isogenic non-encapsulated mutant (GPA), used as a control. 45-days after periodontal inoculations with P. gingivalis serotypes, rat´s spatial memory was evaluated for six consecutive days in the Oasis maze task. Following functional testing, the animals were sacrificed, and various tissues were removed to analyze alveolar bone resorption, cytokine production, and detect AD-specific biomarkers. Strikingly, only K1 or K2 P. gingivalis-infected rats displayed memory deficits, increased alveolar bone resorption, pro-inflammatory cytokine production, changes in astrocytic morphology, increased Aß1-42 levels, and Tau hyperphosphorylation in the hippocampus. None of these effects were observed in rats infected with the non-encapsulated bacterial strains. Based on these results, we propose that the bacterial virulence factors constituted by capsular polysaccharides play a central role in activating innate immunity and inflammation in the AD-like pathology triggered by P. gingivalis in young rats subjected to an acute experimental infection episode.

Inhibitory effect of serotype a of Aggregatibacter actinomycetemcomitans on the increased destructive potential of serotype b.

OBJECTIVE: The serotype b of Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) induces higher cytokine production in dendritic cells (DCs) compared with the other serotypes. However, this increased immunostimulatory potential was modified when DCs were co-infected with the other A. actinomycetemcomitans serotypes. This study aimed to analyze whether the production of interferon gamma (IFN-γ), C-reactive protein (CRP), matrix metalloproteinase (MMP)-2, and MMP-9, as well as the activity of osteoclasts, also varies when DCs are co-infected with the A. actinomycetemcomitans serotypes. MATERIALS AND METHODS: Human DCs were stimulated with the A. actinomycetemcomitans serotypes using the following stimulatory conditions: serotype a/b/c/a+b/a+c/b+c/a+b+c. The IFN-γ, CRP, and MMP-2 levels were quantified by ELISA. The active form of MMP-9 was quantified using fluorescent functional assays. The MMP-2 gelatinolytic activity was identified by zymogram. The osteoclast activity was determined by quantifying the TRAP expression and resorption-pit formation using cytochemistry and osteoassays. RESULTS: Higher levels of IFN-γ, CRP, MMP-2, MMP-9, and osteoclast activity were detected when DCs were stimulated with the serotype b of A. actinomycetemcomitans compared with the others. This increased immunostimulatory potential attributed to serotype b diminished when DCs were co-infected with the serotype a. CONCLUSIONS: This study provides new insights into the virulence of A. actinomycetemcomitans and reveals important differences in the immunostimulatory and pro-destructive potential among its serotypes.

Immunostimulatory activity of low-molecular-weight hyaluronan on dendritic cells stimulated with Aggregatibacter actinomycetemcomitans or Porphyromonas gingivalis.

OBJECTIVES: Periodontitis is a chronic inflammatory disease characterized by tooth-supporting tissue destruction, which is elicited by the host's immune response triggered against periodonto-pathogen bacteria. During periodontal tissue destruction, extracellular matrix components are metabolized and fragmented. Some extracellular matrix component-derived fragments, such as low-molecular-weight hyaluronan (LMW-HA), have potent immunogenic potential, playing a role as damage-associated molecular patterns (DAMPs) during activation of immune cells. Dendritic cells (DCs) play a central role in the host's immune response displayed during periodontitis; thus, this study aimed to analyze whether LMW-HA has an immunostimulatory activity on DCs when stimulated with periodonto-pathogen bacteria. MATERIALS AND METHODS: LMW-HA-treated and non-treated DCs were stimulated with Aggregatibacter actinomycetemcomitans or Porphyromonas gingivalis and the mRNA expression for cytokines tumor necrosis factor-α (TNF-alpha), interleukin-1ß (IL-1B), interleukin-6 (IL-6), and interleukin-23 (IL-23A) was quantified by RT-qPCR. In addition, transcription factors interferon regulatory factor 4 (IRF4), interferon regulatory factor 8 (IRF8), neurogenic locus notch homolog protein 2 (NOTCH2), and basic leucine zipper ATF-like transcription factor 3 (BATF3), involved in DC activation, were analyzed. RESULTS: Higher expression levels of TNF-alpha, IL-1B, IL-6, and IL-23A were detected in LMW-HA-treated DCs after bacterial infection, as compared with non-treated DCs. When LMW-HA-treated DCs were infected with A. actinomycetemcomitans, higher levels of IRF4, NOTCH2, and BATF3 were detected compared with non-treated cells; whereas against P. gingivalis infection, increased levels of IRF4 and NOTCH2 were detected. CONCLUSION: LMW-HA plays an immunostimulatory role on the immune response triggered by DCs during infection with A. actinomycetemcomitans or P. gingivalis. CLINICAL RELEVANCE: Detection of extracellular matrix component-derived fragments produced during periodontal tissue destruction, such as LMW-HA, could explain at least partly unsuccessful periodontal treatment and the chronicity of the disease.

Serotype a of Aggregatibacter actinomycetemcomitans down-regulates the increased serotype b-induced cytokine and chemokine production in dendritic cells.

OBJECTIVE: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Mixed infection with the different A. actinomycetemcomitans serotypes is frequent in periodontitis patients; accordingly, the role of this bacterial species in the pathogenesis of periodontitis may differ depending whether patients or periodontal lesions harbour one or more of the A. actinomycetemcomitans serotypes. We hypothesized that different combinations of these serotypes could be associated with distinct host responses and hence different inflammatory patterns. This investigation was aimed to assess whether the increased immuno-stimulatory potential attributed to the serotype b of A. actinomycetemcomitans on immune cells is able to be modified during co-infection with other A. actinomycetemcomitans serotypes. METHODS: Dendritic cells (DCs) were obtained from healthy subjects and stimulated with the different A. actinomycetemcomitans serotypes or their purified LPS using the following stimulatory conditions: serotype a, b, or c, and the combinations a+b, a+c, b+c, or a+b+c. The cytokine, CCR, and CCL levels were quantified by qPCR and ELISA. RESULTS: Higher levels of cytokines, CCRs, and CCLs were induced when DCs were stimulated with the serotype b of A. actinomycetemcomitans compared with the same cells stimulated with the other serotypes. When DCs were co-infected, these levels decreased in comparison with the serotype b-stimulation alone, in particular when the serotype a was present in the mixed infection. CONCLUSIONS: The increased immuno-stimulatory potential attributed to the serotype b was modified when DCs were co-infected with other A. actinomycetemcomitans serotypes, in particular, when the serotype a was present, the DC response diminished.

Differential human Th22-lymphocyte response triggered by Aggregatibacter actinomycetemcomitans serotypes.

OBJECTIVE: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on lipopolysaccharide (LPS) antigenicity. When T lymphocytes were stimulated with these serotypes, different patterns of T-helper (Th)1 and Th17-type of immune responses were reported. Recently, two new Th phenotypes have been described and named Th9 and Th22 lymphocytes; however, their role in the pathogenesis of periodontitis remains unclear. This study aimed to investigate the potential Th9 and/or Th22 lymphocyte responses when stimulated with autologous dendritic cells infected with different A. actinomycetemcomitans serotypes. METHODS: Monocyte-derived dendritic cells and naïve CD4+ T lymphocytes were obtained from healthy donors and stimulated with different serotypes of A. actinomycetemcomitans at a multiplicity of infection MOI=102 or their purified LPS (10-50ng/ml). The levels for the Th9 and Th22-associated cytokines, as well as the transcription factor master-switch genes implied in their differentiation Spi-B and AhR, were quantified by qPCR and ELISA. RESULTS: When stimulated with the serotype b of A. actinomycetemcomitans, higher levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-α were detected in dendritic cells, as well as higher levels of IL-22 and AhR were detected in T lymphocytes, when compared with stimulation with the other serotypes. CONCLUSIONS: The serotype b of A. actinomycetemcomitans has a higher capacity of trigger Th22-type of immune response in both dendritic cells and T lymphocytes. These data allow us to suggest that, when the serotype b of A. actinomycetemcomitans is a significant part of the subgingival biofilm, the Th22 polarization might be triggered within the periodontal lesion.

Differential expression of CC chemokines (CCLs) and receptors (CCRs) by human T lymphocytes in response to different Aggregatibacter actinomycetemcomitans serotypes

ABSTRACT In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Recently, our research group has reported a differential immunogenicity when T lymphocytes were stimulated with these different serotypes. In particular, it was demonstrated that the serotype b of A. actinomycetemcomitans has a stronger capacity to trigger Th1- and Th17-type cytokine production. Objective This study aimed to quantify the expression of different CC chemokines (CCLs) and receptors (CCRs) in T lymphocytes stimulated with the different A. actinomycetemcomitans serotypes. In addition, the expression of the transcription factors T-bet, GATA-3, RORC2, and Foxp3, master-switch genes implied in the Th1, Th2, Th17, and T-regulatory differentiation, respectively, was analyzed in order to determine T-cell phenotype-specific patterns of CCL and CCR expression upon A. actinomycetemcomitans stimulation. Material and Methods Human naïve CD4+ T lymphocytes were obtained from healthy subjects and stimulated with autologous dendritic cells primed with the different A. actinomycetemcomitans serotypes. The expression levels for the chemokines CCL1, CCL2, CCL3, CCL5, CCL11, CCL17, CCL20, CCL21, CCL25, and CCL28, as well as the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 were quantified by qPCR. Similarly, the expression levels for the transcription factors T-bet, GATA-3, RORC2, and Foxp3 were quantified and correlated with the CCL and CCR expression levels. Results Higher expression levels of CCL2, CCL3, CCL5, CCL20, CCL21, CCL28, CCR1, CCR2, CCR5, CCR6, CCR7, and CCR9 were detected in T lymphocytes stimulated with the serotype b of A. actinomycetemcomitans compared with the other serotypes. In addition, these higher expression levels of CCLs and CCRs positively correlated with the increased levels of T-bet and RORC2 when T lymphocytes were stimulated with the serotype b. Conclusion A T-lymphocyte response biased towards a Th1- and Th17-pattern of CCL and CCR expression was detected under stimulation with the serotype b of A. actinomycetemcomitans.

Differential expression of CC chemokines (CCLs) and receptors (CCRs) by human T lymphocytes in response to different Aggregatibacter actinomycetemcomitans serotypes.

UNLABELLED: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Recently, our research group has reported a differential immunogenicity when T lymphocytes were stimulated with these different serotypes. In particular, it was demonstrated that the serotype b of A. actinomycetemcomitans has a stronger capacity to trigger Th1- and Th17-type cytokine production. OBJECTIVE: This study aimed to quantify the expression of different CC chemokines (CCLs) and receptors (CCRs) in T lymphocytes stimulated with the differentA. actinomycetemcomitans serotypes. In addition, the expression of the transcription factors T-bet, GATA-3, RORC2, and Foxp3, master-switch genes implied in the Th1, Th2, Th17, and T-regulatory differentiation, respectively, was analysed in order to determine T-cell phenotype-specific patterns of CCL and CCR expression upon A. actinomycetemcomitans stimulation. MATERIAL AND METHODS: Human naïve CD4+ T lymphocytes were obtained from healthy subjects and stimulated with autologous dendritic cells primed with the differentA. actinomycetemcomitans serotypes. The expression levels for the chemokines CCL1, CCL2, CCL3, CCL5, CCL11, CCL17, CCL20, CCL21, CCL25, and CCL28, as well as the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 were quantified by qPCR. Similarly, the expression levels for the transcription factors T-bet, GATA-3, RORC2, and Foxp3 were quantified and correlated with the CCL and CCR expression levels. RESULTS: Higher expression levels of CCL2, CCL3, CCL5, CCL20, CCL21, CCL28, CCR1, CCR2, CCR5, CCR6, CCR7, and CCR9 were detected in T lymphocytes stimulated with the serotype b of A. actinomycetemcomitans compared with the other serotypes. In addition, these higher expression levels of CCLs and CCRs positively correlated with the increased levels of T-bet and RORC2 when T lymphocytes were stimulated with the serotype b. CONCLUSION: A T-lymphocyte response biased towards a Th1- and Th17-pattern of CCL and CCR expression was detected under stimulation with the serotype b ofA. actinomycetemcomitans.

Differential expression of CC chemokines (CCLs) and receptors (CCRs) by human T lymphocytes in response to different Aggregatibacter actinomycetemcomitans serotypes

In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Recently, our research group has reported a differential immunogenicity when T lymphocytes were stimulated with these different serotypes. In particular, it was demonstrated that the serotype b of A. actinomycetemcomitans has a stronger capacity to trigger Th1- and Th17-type cytokine production.Objective This study aimed to quantify the expression of different CC chemokines (CCLs) and receptors (CCRs) in T lymphocytes stimulated with the differentA. actinomycetemcomitans serotypes. In addition, the expression of the transcription factors T-bet, GATA-3, RORC2, and Foxp3, master-switch genes implied in the Th1, Th2, Th17, and T-regulatory differentiation, respectively, was analysed in order to determine T-cell phenotype-specific patterns of CCL and CCR expression upon A. actinomycetemcomitans stimulation.Material and Methods Human naïve CD4+ T lymphocytes were obtained from healthy subjects and stimulated with autologous dendritic cells primed with the differentA. actinomycetemcomitans serotypes. The expression levels for the chemokines CCL1, CCL2, CCL3, CCL5, CCL11, CCL17, CCL20, CCL21, CCL25, and CCL28, as well as the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 were quantified by qPCR. Similarly, the expression levels for the transcription factors T-bet, GATA-3, RORC2, and Foxp3 were quantified and correlated with the CCL and CCR expression levels.Results Higher expression levels of CCL2, CCL3, CCL5, CCL20, CCL21, CCL28, CCR1, CCR2, CCR5, CCR6, CCR7, and CCR9 were detected in T lymphocytes stimulated with the serotype b of A. actinomycetemcomitans compared with the other serotypes. In addition, these higher expression levels of CCLs and CCRs positively correlated with the increased levels of T-bet and RORC2 when T lymphocytes were stimulated with the serotype b.Conclusion A T-lymphocyte response biased towards a Th1- and Th17-pattern of CCL and CCR expression was detected under stimulation with the serotype b ofA. actinomycetemcomitans.

Variability of the dendritic cell response triggered by different serotypes of Aggregatibacter actinomycetemcomitans or Porphyromonas gingivalis is toll-like receptor 2 (TLR2) or TLR4 dependent.

BACKGROUND: Different serotypes of Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis have been shown to induce differential dendritic cell (DC) responses. This study investigates whether cytokine and CC-chemokine receptor (CCR) production by DCs stimulated with different serotypes of A. actinomycetemcomitans or P. gingivalis is Toll-like receptor 2 (TLR2) and/or TLR4 dependent. METHODS: DCs were obtained from healthy individuals and primed at a multiplicity of infection (MOI) of 10(2) with different A. actinomycetemcomitans or P. gingivalis serotypes in the presence or absence of anti-TLR2 or anti-TLR4 blocking antibodies. TLR2 and TLR4 expression, CCR5 and CCR6 expression, and interleukin (IL)-1ß, IL-10, IL-12, and IL-23 expression and secretion were quantified by flow cytometry, real-time reverse-transcription polymerase chain reaction, and enzyme-linked immunosorbent assay. RESULTS: When DCs were stimulated with serotype b of A. actinomycetemcomitans or serotype K1 of P. gingivalis, higher levels of TLR2 or TLR4, respectively, were detected compared to DCs stimulated with the other serotypes. Similarly, higher levels of cytokines and CCRs were detected in serotype b- or serotype K1-primed DCs compared to the others, and these increased levels positively correlated with levels of TLR2 or TLR4. When TLR2 signaling was blocked using a specific anti-TLR2 monoclonal antibody, serotype b-induced cytokine and CCR expression was inhibited; when TLR4 signaling was blocked, serotype K1-induced response was inhibited. CONCLUSIONS: These results demonstrate that the variability of secretion of cytokines and expression of CCRs detected in DCs stimulated with different serotypes of A. actinomycetemcomitans or P. gingivalis is TLR2 or TLR4 dependent, respectively.

Differential expression of CC chemokines (CCLs) and receptors (CCRs) by human T lymphocytes in response to different Aggregatibacter actinomycetemcomitans serotypes.

In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Recently, our research group has reported a differential immunogenicity when T lymphocytes were stimulated with these different serotypes. In particular, it was demonstrated that the serotype b of A. actinomycetemcomitans has a stronger capacity to trigger Th1- and Th17-type cytokine production. Objective This study aimed to quantify the expression of different CC chemokines (CCLs) and receptors (CCRs) in T lymphocytes stimulated with the different A. actinomycetemcomitans serotypes. In addition, the expression of the transcription factors T-bet, GATA-3, RORC2, and Foxp3, master-switch genes implied in the Th1, Th2, Th17, and T-regulatory differentiation, respectively, was analyzed in order to determine T-cell phenotype-specific patterns of CCL and CCR expression upon A. actinomycetemcomitans stimulation. Material and Methods Human naïve CD4+ T lymphocytes were obtained from healthy subjects and stimulated with autologous dendritic cells primed with the different A. actinomycetemcomitans serotypes. The expression levels for the chemokines CCL1, CCL2, CCL3, CCL5, CCL11, CCL17, CCL20, CCL21, CCL25, and CCL28, as well as the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 were quantified by qPCR. Similarly, the expression levels for the transcription factors T-bet, GATA-3, RORC2, and Foxp3 were quantified and correlated with the CCL and CCR expression levels. Results Higher expression levels of CCL2, CCL3, CCL5, CCL20, CCL21, CCL28, CCR1, CCR2, CCR5, CCR6, CCR7, and CCR9 were detected in T lymphocytes stimulated with the serotype b of A. actinomycetemcomitans compared with the other serotypes. In addition, these higher expression levels of CCLs and CCRs positively correlated with the increased levels of T-bet and RORC2 when T lymphocytes were stimulated with the serotype b. Conclusion A T-lymphocyte response biased towards a Th1- and Th17-pattern of CCL and CCR expression was detected under stimulation with the serotype b of A. actinomycetemcomitans.

Activation of RANKL-induced osteoclasts and memory T lymphocytes by Porphyromonas gingivalis is serotype dependant.

AIM: Destructive periodontitis is associated with a Th1-Th17 immune response and activation of RANKL-induced osteoclasts. In addition, Porphyromonas gingivalis K1 and K2 serotypes induce a strong Th1-Th17 response. This study aimed to investigate whether these P. gingivalis serotypes induce higher osteoclasts activation, by increased Th17-associated RANKL production, and an antigen-specific memory T-lymphocyte response. MATERIAL AND METHODS: The RANKL production and TRAP(+) osteoclast induction were quantified on naïve T lymphocytes stimulated with dendritic cells primed with the P. gingivalis serotypes. The T-bet, GATA-3, RORC2 and Foxp3 expression was correlated with RANKL production. The frequency of proliferating memory T lymphocytes in response to P. gingivalis serotypes was determined in both periodontitis and healthy subjects. RESULTS: T lymphocytes stimulated by K1 or K2-primed dendritic cells elicited higher levels of RANKL and TRAP(+) osteoclasts than cells stimulated with the other serotypes. RANKL positively correlated with RORC2. Whereas periodontitis patients had a higher frequency of memory T lymphocytes responding to K1 or K2, healthy subjects had a higher frequency of memory T lymphocytes responding to K4 or K(-) . CONCLUSIONS: P. gingivalis serotypes K1 and K2, but not others, are associated with an increased production of the osteoclastogenesis-related factor RANKL. This important information suggests that these serotypes could elicit a greater bone resorption in vivo and have a role in the periodontitis pathogenesis.

Serotype-dependent response of human dendritic cells stimulated with Aggregatibacter actinomycetemcomitans.

AIM: Different serotypes of Aggregatibacter actinomycetemcomitans have been described based on the lipopolysaccharide (LPS)-O-polysaccharide antigenicity. In turn, a distinct effect of A. actinomycetemcomitans serotypes has been described on cell proliferation and pro-inflammatory cytokine production in different human cells. This study was aimed to investigate the differential dendritic cell (DC) response when stimulated with different bacterial strains belonging to the most prevalent serotypes of A. actinomycetemcomitans (a-c). MATERIALS AND METHODS: Dendritic cells were obtained from healthy subjects and stimulated with increasing multiplicity of infection (MOI = 10(-1) -10(2)) of A. actinomycetemcomitans, serotypes a-c, or their lipopolysaccharide (10-50 ng/ml). The levels for interferon (IFN)-γ, tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-5, IL-6, IL-10, IL-12 and IL-23 were quantified by real-time RT-PCR and ELISA. RESULTS: Variable DC responses were detected when stimulated with the different strains of A. actinomycetemcomitans. DCs stimulated with A. actinomycetemcomitans strains belonging to the serotype b or their purified LPS expressed higher levels of IL-1ß, IL-6, IL-12, IL-23, IFN-γ and TNF-α than DCs stimulated with the other serotypes. CONCLUSIONS: Aggregatibacter actinomycetemcomitans strains belonging to the serotype b demonstrated a higher capacity to trigger Th1 and Th17-type cytokine production on DCs. These increased potential is likely explained by a higher immunogenicity of their LPS.

Morphological effects of mesenchymal stem cells and pulsed ultrasound on condylar growth in rats: a pilot study.

AIM: The aim of this study was to assess and describe the morphological effects of an intra-articular iniection of Mesenchymal Stem Cells (MSCs) and/or Low Intensity Pulsed Ultrasound (LIPUS) stimulation on the mandibular condyles of growing rats, using cone beam computed tomography (CBCT) and histology. METHODS: Twenty-six young (23-day-old) rats were divided into 5 groups identified as LIPUS-stimulated (20 minutes daily using 50 mW/cm2, 1MHz, 0.2 millisecond pulses), MSCs injected (1 x 10(5) cells/kg), LIPUS + MSCs, medium inlected, and untreated controls. All treatments were performed in the left temporomandibular joint of each rat (TMJs). At day 21, CBCTs were obtained for cephalometric analysis and 3D reconstructions. After animal sacrifice, left and right TMJ sections were histologically prepared and examined. The Wilcoxon sign rank test and the Kruskal-Wallis 2 test were applied for statistical comparison. RESULTS: Imaging results showed that left condyles were wider in all LIPUS-treated groups (p < 0.05), while the LIPUS-only group had a greater left sagittal condylar length. LIPUS-treated groups displayed a lower midline shift to the right (p < 0.02). No significant differences were observed in the MSC group. Bone marrow morphology and vascularity differed between the groups as LIPUS-treated groups exhibited increased vascularity in the erosive cartilage zone. CONCLUSION: It was established that LIPUS and MSC application to the TMJ region of growing rats favoured transverse condylar growth, while LIPUS application alone may enhance sagittal condylar development.The MSC injection model had little effect on sagittal condylar growth.