Macrophage-related gingival transcriptomic patterns and microbiome alterations in experimental periodontitis in nonhuman primates.

OBJECTIVE: This study examined the microbiome features specifically related to host macrophage polarization in health, initiation and progression of periodontitis, and in resolution samples using a nonhuman primate model of ligature-induced periodontitis. BACKGROUND: The oral microbiome is a complex of bacterial phyla, genera, and species acquired early in life into the individual autochthonous oral ecology. The microbiome changes overtime in response to both intrinsic and extrinsic stressors, and transitions to a dysbiotic ecology at sites of periodontal lesions. METHODS: Comparisons were made between the microbial and host features in young (≤7 years) and adult (≥12 years) cohorts of animals. Footprints of macrophage-related genes in the gingival tissues were evaluated using expression profiles including M0, M1, and M2 related genes. RESULTS: Within the gingival tissues, similar macrophage-related gene patterns were observed with significant increases with disease initiation and continued elevation throughout disease in both age groups. Approximately, 70% of the taxa were similar in relative abundance between the two groups; however, the adults showed a large number of OTUs that were significantly altered compared with the younger animals. Developing a correlation map identified three major node levels of interactions that comprised approximately ⅓ of the Operational Taxonomic Units (OTUs) that dominated the microbiomes across the samples. Also noted was a much greater frequency of significant correlations of individual OTUs with the macrophage phenotype markers, compared with disease and resolution samples in both age groups, with a greater frequency in the younger group. Moreover, these correlations were assigned to differentially expressed genes representing M0, M1, and M2-related phenotypes. A cluster analyses across the macrophage-related transcriptome and the OTUs demonstrated multiple somewhat distinct bacterial consortia, incorporating both commensal and putative pathogens, linked to the gene responses that differed in health, disease, and resolution samples. Finally, there were minimal alterations in the OTUs in individual clusters with specific macrophage-related responses in the younger group, while in the adult samples substantial variations were noted with genes from all macrophage phenotypes. CONCLUSIONS: The results confirmed important features that could reflect macrophage polarization in periodontal lesions, and provided some initial data supporting specific members of the oral microbiome feature prominently related to specific gene response patterns consistent with macrophages in the gingival tissues.

Biological Aging and Periodontal Disease: Analysis of NHANES (2001-2002).

INTRODUCTION: Periodontitis is a chronic inflammatory disease caused by multiple potential contributing factors such as bacterial biofilm infection of the tissues surrounding the teeth and environmental determinants and a dysregulated host response for modifying and resolving the inflammation. Because periodontal disease is a major public health concern with substantial increases in the prevalence and severity in aging populations, previous studies of periodontitis tended to approach the disease as an age-associated outcome across the life span. However, few investigations have considered that, as a chronic noncommunicable disease, periodontitis may not simply be a disease that increases with age but may contribute to more rapid biologic aging. OBJECTIVES: Increasing population data supports the potential disconnect between chronological aging and biologic aging, which would contribute to the heterogeneity of aging phenotypes within chronologic ages across populations. Thus, our aim was to test whether periodontal disease affects biological aging across the life span. METHODS: The prevalence of periodontitis in the adult US population is a portion of the assessment of the National Health and Nutrition Examination Survey (NHANES), which has been ongoing since 1971 through 2-y cycles sampling populations across the country. We used NHANES 2001-2002 to test the hypothesis that the presence/severity of periodontal disease as an exposure variable would negatively affect telomere length, a measure of biological aging, and that this relationship is modified by factors that also affect the progression of periodontitis, such as sex, race/ethnicity, and smoking. RESULTS: The data demonstrated a significant impact of periodontitis on decreasing telomere lengths across the life span. These differences were modulated by age, sex, race/ethnicity, and smoking within the population. CONCLUSION: The findings lay the groundwork for future studies documenting broader effects on biological aging parameters as well as potential intervention strategies for periodontitis in driving unhealthy aging processes. KNOWLEDGE TRANSFER STATEMENT: Periodontitis is a chronic inflammatory disease and dysregulated host response. Shortening of telomeres is a reflection of biologic aging. Decreased telomere lengths with periodontitis are seemingly related to chronic infection and persistent local and systemic inflammation. These findings suggest that periodontitis is not simply a disease of aging but may also transmit chronic systemic signals that could affect more rapid biological aging. Clinicians can use this outcome to recognize the role of periodontitis in driving unhealthy aging processes in patients.

Comparative Analysis of Gene Expression Patterns for Oral Epithelium-Related Functions with Aging.

Epithelial cells and functions of the epithelium are critical to the health of the oral cavity. We used a nonhuman primate model to profile the transcriptome of gingival tissues in health across the lifespan and hypothesized that in older animals, epithelial-related transcriptome patterns would reflect epithelial cells that are aggressively responsive to the surrounding environment and less able to modulate and resolve the noxious challenge from the bacteria. Rhesus monkeys (n = 34) with a healthy periodontium were distributed into four groups: ≤3 years (young), 3-7 years (adolescent), 12-16 years (adult), and 18-23 years (aged), and a buccal gingival sample from the premolar/molar region of each animal was obtained. RNA was subjected to a microarray analysis (GeneChip® Rhesus Macaque Genome Array, Affymetrix), and 336 genes examined that are linked to epithelium and epithelial cell functions categorized into 9 broad functional groups: extracellular matrix and cell structure; extracellular matrix remodeling enzymes; cell adhesion molecules, cytoskeleton regulation; inflammatory response; growth factors; kinases/cell signaling; cell surface receptors; junction associated molecules; autophagy/apoptosis; antimicrobial peptides; and transcription factors. Total of 255 genes displayed a normalized signal >100, and differences across the age groups were observed primarily in extracellular matrix and cell structure, cell adhesion molecules, and cell surface receptor gene categories with elevations in the aged tissues. Keratins 2, 5, 6B, 13, 16, 17 were all significantly increased in healthy-aged tissues versus adults, and keratins 1 and 2 were significantly decreased in young animals. Approximately 15 integrins are highly expressed in the gingival tissues across the age groups with only ITGA8, ITGAM (CD11b), and ITGB2 significantly increased in the aged tissues. Little impact of aging on desmosomal/hemidesmosomal genes was noted. These results suggest that healthy gingival aging has a relatively limited impact on the broader functions of the epithelium and epithelial cells, with some effects on genes for extracellular matrix and cell adhesion molecules (e.g., integrins). Thus, while there is a substantial impact of aging on immune system targets even in healthy gingiva, it appears that the epithelial barrier remains reasonably molecularly intact in this model system.

Epidemiologic evaluation of Nhanes for environmental Factors and periodontal disease.

Periodontitis is a chronic inflammation that destroys periodontal tissues caused by the accumulation of bacterial biofilms that can be affected by environmental factors. This report describes an association study to evaluate the relationship of environmental factors to the expression of periodontitis using the National Health and Nutrition Examination Study (NHANES) from 1999-2004. A wide range of environmental variables (156) were assessed in patients categorized for periodontitis (n = 8884). Multiple statistical approaches were used to explore this dataset and identify environmental variable patterns that enhanced or lowered the prevalence of periodontitis. Our findings indicate an array of environmental variables were different in periodontitis in smokers, former smokers, or non-smokers, with a subset of specific environmental variables identified in each population subset. Discriminating environmental factors included blood levels of lead, phthalates, selected nutrients, and PCBs. Importantly, these factors were found to be coupled with more classical risk factors (i.e. age, gender, race/ethnicity) to create a model that indicated an increased disease prevalence of 2-4 fold across the sample population. Targeted environmental factors are statistically associated with the prevalence of periodontitis. Existing evidence suggests that these may contribute to altered gene expression and biologic processes that enhance inflammatory tissue destruction.

Age and Periodontal Health - Immunological View.

PURPOSE OF THE REVIEW: Aging clearly impacts a wide array of systems, in particular the breadth of the immune system leading to immunosenescence, altered immunoactivation, and coincident inflammaging processes. The net result of these changes leads to increased susceptibility to infections, increased neoplastic occurrences, and elevated frequency of autoimmune diseases with aging. However, as the bacteria in the oral microbiome that contribute to the chronic infection of periodontitis is acquired earlier in life, the characteristics of the innate and adaptive immune systems to regulate these members of the autochthonous microbiota across the lifespan remains ill defined. RECENT FINDINGS: Clear data demonstrate that both cells and molecules of the innate and adaptive immune response are adversely impacted by aging, including in the oral cavity, yielding a reasonable tenet that the increased periodontitis noted in aging populations is reflective of the age-associated immune dysregulation. Additionally, this facet of host-microbe interactions and disease needs to accommodate the population variation in disease onset and progression, which may also reflect an accumulation of environmental stressors and/or decreased protective nutrients that could function at the gene level (ie. epigenetic) or translational level for production and secretion of immune system molecules. SUMMARY: Finally, the majority of studies of aging and periodontitis have emphasized the increased prevalence/severity of disease with aging, all based upon chronological age. However, evolving areas of study focusing on "biological aging" to help account for population variation in disease expression, may suggest that chronic periodontitis represents a co-morbidity that contributes to "gerovulnerability" within the population.

Cross-talk between clinical and host-response parameters of periodontitis in smokers.

BACKGROUND AND OBJECTIVE: Periodontal diseases are a major public health concern leading to tooth loss and have also been shown to be associated with several chronic systemic diseases. Smoking is a major risk factor for the development of numerous systemic diseases, as well as periodontitis. While it is clear that smokers have a significantly enhanced risk for developing periodontitis leading to tooth loss, the population varies regarding susceptibility to disease associated with smoking. This investigation focused on identifying differences in four broad sets of variables, consisting of: (i) host-response molecules; (ii) periodontal clinical parameters; (iii) antibody responses to periodontal pathogens and oral commensal bacteria; and (iv) other variables of interest, in a population of smokers with (n = 171) and without (n = 117) periodontitis. MATERIAL AND METHODS: Bayesian network structured learning (BNSL) techniques were used to investigate potential associations and cross-talk between the four broad sets of variables. RESULTS: BNSL revealed two broad communities with markedly different topology between the populations of smokers, with and without periodontitis. Confidence of the edges in the resulting network also showed marked variations within and between the periodontitis and nonperiodontitis groups. CONCLUSION: The results presented validated known associations and discovered new ones with minimal precedence that may warrant further investigation and novel hypothesis generation. Cross-talk between the clinical variables and antibody profiles of bacteria were especially pronounced in the case of periodontitis and were mediated by the antibody response profile to Porphyromonas gingivalis.

Differential Gene Expression Profiles Reflecting Macrophage Polarization in Aging and Periodontitis Gingival Tissues.

Recent evidence has determined a phenotypic and functional heterogeneity for macrophage populations. This plasticity of macrophage function has been related to specific properties of subsets (M1 and M2) of these cells in inflammation, adaptive immune responses and resolution of tissue destructive processes. This investigation hypothesized that targeted alterations in the distribution of macrophage phenotypes in aged individuals, and with periodontitis would be skewed towards M1 inflammatory macrophages in gingival tissues. The study used a non-human primate model to evaluate gene expression profiles as footprints of macrophage variation in healthy and periodontitis gingival tissues from animals 3-23 years of age and in periodontitis tissues in adult and aged animals. Significant increases in multiple genes reflecting overall increases in macrophage activities were observed in healthy aged tissues, and were significantly increased in periodontitis tissues from both adults and aged animals. Generally, gene expression patterns for M2 macrophages were similar in healthy young, adolescent and adult tissues. However, modest increases were noted in healthy aged tissues, similar to those seen in periodontitis tissues from both age groups. M1 macrophage gene transcription patterns increased significantly over the age range in healthy tissues, with multiple genes (e.g. CCL13, CCL19, CCR7 and TLR4) significantly increased in aged animals. Additionally, gene expression patterns for M1 macrophages were significantly increased in adult health versus periodontitis and aged healthy versus periodontitis. The findings supported a significant increase in macrophages with aging and in periodontitis. The primary increases in both healthy aged tissues and, particularly periodontitis tissues appeared in the M1 phenotype.

Utility of salivary biomarkers for demonstrating acute myocardial infarction.

The comparative utility of serum and saliva as diagnostic fluids for identifying biomarkers of acute myocardial infarction (AMI) was investigated. The goal was to determine if salivary biomarkers could facilitate a screening diagnosis of AMI, especially in cases of non-ST elevation MI (NSTEMI), since these cases are not readily identified by electrocardiogram (ECG). Serum and unstimulated whole saliva (UWS) collected from 92 AMI patients within 48 hours of chest pain onset and 105 asymptomatic healthy control individuals were assayed for 13 proteins relevant to cardiovascular disease, by Beadlyte technology (Luminex(®)) and enzyme immunoassays. Data were analyzed with concentration cut-points, ECG findings, logistic regression (LR) (adjusted for matching for age, gender, race, smoking, number of teeth, and oral health status), and classification and regression tree (CART) analysis. A sensitivity analysis was conducted by repetition of the CART analysis in 58 cases and 58 controls, each matched by age and gender. Serum biomarkers demonstrated AMI sensitivity and specificity superior to that of saliva, as determined by LR and CART. The predominant discriminators in serum by LR were troponin I (TnI), B-type natriuretic peptide (BNP), and creatine kinase-MB (CK-MB), and TnI and BNP by CART. In saliva, LR identified C-reactive protein (CRP) as the biomarker most predictive of AMI. A combination of smoking tobacco, UWS CRP, CK-MB, sCD40 ligand, gender, and number of teeth identified AMI in the CART decision trees. When ECG findings, salivary biomarkers, and confounders were included, AMI was predicted with 80.0% sensitivity and 100% specificity. These analyses support the potential utility of salivary biomarker measurements used with ECG for the identification of AMI. Thus, saliva-based tests may provide additional diagnostic screening information in the clinical course for patients suspected of having an AMI.

A bidirectional relationship of oral-systemic responses: observations of systemic host responses in patients after full-mouth extractions.

OBJECTIVE: This investigation tested the hypothesis that systemic inflammatory responses would be attenuated by minimizing the oral microbial burden in patients with moderate to severe periodontitis. STUDY DESIGN: Patients (n = 73) scheduled for full-mouth extractions were categorized as case type I/II (gingivitis/mild periodontitis) or case type III/IV (moderate/severe periodontitis). Serum levels of acute phase proteins (APPs) and immunoglobulin G (IgG) antibody were assessed at baseline and through 1 year after extraction. RESULTS: At baseline, the levels of multiple APPs (e.g., fibrinogen, C-reactive protein) and antibodies to periodontal pathogens were significantly higher with case type III/IV vs I/II. These differences were sustained 12 months after extractions for most APPs. CONCLUSIONS: The results demonstrated that removal of disease by full-mouth extraction of teeth altered the overall burden of challenge to the host. Continued elevation in various APPs in the III/IV group suggested a potential underlying constitutive difference in systemic response characteristics of this population.

Smoking-related cotinine levels and host responses in chronic periodontitis.

BACKGROUND AND OBJECTIVE: Smoking has been reported to increase the risk of periodontal disease by disrupting the balance of immune responses and tissue repair processes; however, this risk varies among smokers. Cotinine levels in saliva are routinely used to measure the level of smoking, and reflect the quantity of nicotine, and other smoking-related xenobiotics that challenge host systems. This study delineated characteristics of inflammatory mediators in saliva and serum antibody responses to both periodontal pathogens and commensal bacteria in smokers as they related to cotinine levels. MATERIALS AND METHODS: This case-control study (n = 279) examined salivary inflammatory mediator responses [interleukin (IL)-1ß, IL-10, prostaglandin E2, myeloperoxidase and plasminogen activator inhibitor-1], and serum IgG antibody responses to three periodontal pathogens (Aggregatibacter actinomyce-temcomitans, Porphyromonas gingivalis, Treponema denticola) and five commensal oral microorganisms (Veillonella parvula, Streptococcus sanguis, Prevotella loescheii, Actinomyces naeslundii, Capnocytophaga ochracea). RESULTS: The patients were stratified into health (n = 30), gingivitis (n = 55) and periodontitis (n = 184); cotinine levels correlated with reported smoking habits in health, less so with gingivitis, and were not correlated in periodontitis. Of the inflammatory mediators/acute phase proteins, only IL-1ß levels were positively associated (p < 0.001) with the pack years and cotinine levels. As might be predicted, patients with periodontitis smoked more (p < 0.001) and had higher levels of cotinine. IL-1ß and antibody to A. actinomycetemcomitans, P. gingivalis and T. denticola were significantly higher in the patients with periodontitis than either patients with gingivitis or who were healthy. CONCLUSIONS: Generally, antibody to the pathogens and commensals was lower with decreased cotinine levels. Smoking exacerbated differences in both inflammatory mediators and three antibody in periodontal disease compared to healthy subjects.

Oral epithelial cell responses to multispecies microbial biofilms.

This report describes the use of a novel model of multispecies biofilms to stimulate profiles of cytokines/chemokines from oral epithelial cells that contribute to local inflammation in the periodontium. Streptococcus gordonii (Sg)/S. oralis (So)/S. sanguinis (Ss) and Sg/Fusobacterium nucleatum (Fn)/Porphyromonas gingivalis (Pg) biofilms elicited significantly elevated levels of IL-1α and showed synergistic stimulatory activity compared with an additive effect of the 3 individual bacteria. Only the Sg/Actinomyces naeslundii (An)/Fn multispecies biofilms elicited IL-6 levels above those of control. IL-8 was a primary response to the Sg/An/Fn biofilms, albeit the level was not enhanced compared with a predicted composite level from the monospecies challenges. These results represent some of the first data documenting alterations in profiles of oral epithelial cell responses to multispecies biofilms.

Antibacterial effects of blackberry extract target periodontopathogens.

BACKGROUND AND OBJECTIVE: Antimicrobial agents provide valuable adjunctive therapy for the prevention and the control of oral diseases. Limitations in their prolonged use have stimulated the search for new, naturally occurring agents with more specific activity and fewer adverse effects. Here we sought to determine the antibacterial properties of blackberry extract (BBE) in vitro against oral bacterial commensals and periodontopathogens. MATERIAL AND METHODS: The effects of whole and fractionated BBE on the metabolism of 10 different oral bacteria were evaluated using the colorimetric water-soluble tetrazolium-1 assay. The bactericidal effects of whole BBE against Fusobacterium nucleatum were determined by quantitating the numbers of colony-forming units (CFUs). Cytotoxicity was determined in oral epithelial (OKF6) cells. RESULTS: BBE at 350-1400 µg/mL reduced the metabolic activity of Porphyromonas gingivalis, F. nucleatum and Streptococcus mutans. The reduced metabolic activity observed for F. nucleatum corresponded to a reduction in the numbers of CFUs following exposure to BBE for as little as 1 h, indicative of its bactericidal properties. An anthocyanin-enriched fraction of BBE reduced the metabolic activity of F. nucleatum, but not of P. gingivalis or S. mutans, suggesting the contribution of species-specific agents in the whole BBE. Oral epithelial cell viability was not reduced following exposure to whole BBE (2.24-1400 µg/mL) for ≤ 6 h. CONCLUSION: BBE alters the metabolic activity of oral periodontopathogens while demonstrating a minimal effect on commensals. The specific antibacterial properties of BBE shown in this study, along with its previously demonstrated anti-inflammatory and antiviral properties, make this natural extract a promising target as an adjunct for prevention and/or complementary therapy of periodontal infections.

Bone remodeling-associated salivary biomarker MIP-1α distinguishes periodontal disease from health.

BACKGROUND AND OBJECTIVE: The field of salivary diagnostics lacks an accepted and validated biomarker of alveolar bone remodeling. To address this, we examined levels of salivary biomolecules specifically associated with biological aspects of bone remodeling in subjects with chronic periodontitis in a case-control study. MATERIAL AND METHODS: Levels of macrophage inflammatory protein-1α (MIP-1α), osteoprotegerin, C-telopeptide pyridinoline cross-links of type I collagen and ß-C-terminal type I collagen telopeptide in unstimulated whole saliva of 80 subjects (40 subjects with moderate to severe chronic periodontitis and 40 sex- and age-matched healthy control subjects) were measured using enzyme immunosorbent assays. Saliva was collected before clinical examination, which included probing depth, clinical attachment loss and bleeding on probing. RESULTS: The mean level of MIP-1α in subjects with periodontitis was 18-fold higher than in healthy subjects (p < 0.0001). Clinical periodontal indices correlated significantly with MIP-1α levels (p < 0.0001). Of the biomolecules examined, MIP-1α demonstrated the greatest ability to discriminate between periodontal disease and health as determined by the area under the curve (0.94) and classification and regression tree analysis (sensitivity 94% and specificity 92.7%). Osteoprotegerin levels were elevated 1.6-fold (p = 0.055), whereas C-telopeptide pyridinoline cross-links of type I collagen and ß-C-terminal type I collagen telopeptide levels were below the level of detection in the majority of subjects. CONCLUSION: These findings suggest that the chemokine MIP-1α may aid in identifying periodontitis. Future longitudinal studies are warranted to determine whether this biomarker can help in ascertaining the progression of bone loss in subjects with periodontal disease.

Epithelial interleukin-8 responses to oral bacterial biofilms.

An in vitro model of bacterial biofilms on rigid gas-permeable contact lenses (RGPLs) was developed to challenge oral epithelial cells. This novel model provided seminal data on oral biofilm-host cell interactions, and with selected bacteria, the biofilms were more effective than their planktonic counterparts at stimulating host cell responses.

Smoking and periodontal disease: discrimination of antibody responses to pathogenic and commensal oral bacteria.

Smoking is an independent risk factor for the initiation, extent and severity of periodontal disease. This study examined the ability of the host immune system to discriminate commensal oral bacteria from pathogens at mucosal surfaces, i.e. oral cavity. Serum immunoglobulin (Ig)G antibody reactive with three pathogenic and five commensal oral bacteria in 301 current smokers (age range 21-66 years) were examined by enzyme-linked immunosorbent assay. Clinical features of periodontal health were used as measures of periodontitis. Antibody to the pathogens and salivary cotinine levels were related positively to disease severity; however, the antibody levels were best described by the clinical disease unrelated to the amount of smoking. The data showed a greater immune response to pathogens than commensals that was related specifically to disease extent, and most noted in black males. Significant correlations in individual patient responses to the pathogens and commensals were lost with an increasing extent of periodontitis and serum antibody to the pathogens. Antibody to Porphyromonas gingivalis was particularly distinct with respect to the discriminatory nature of the immune responses in recognizing the pathogens. Antibody responses to selected pathogenic and commensal oral microorganisms differed among racial groups and genders. The antibody response to the pathogens was related to disease severity. The level of antibody to the pathogens, and in particular P. gingivalis, was correlated with disease severity in black and male subsets of patients. The amount of smoking did not appear to impact directly serum antibody levels to these oral bacteria.

Polybacterial challenge effects on cytokine/chemokine production by macrophages and dendritic cells.

OBJECTIVE: To investigate the polymicrobial infection of periodontal disease, which elicits inflammatory mediators/cytokines/chemokines in the local gingival tissues, and a polybacterial challenge of antigen-presenting cells, e.g. macrophages and dendritic cells (DCs), at the mucosal surface. MATERIALS AND METHODS: The cytokine/chemokine profiles of human macrophages and DCs in response to polybacterial challenges were investigated. RESULTS: Oral Gram-negative bacteria elicited significantly greater IL-8 levels from macrophages, compared to Gram-positive bacteria. Gram-positive bacteria did not show synergism in inducing this chemokine from macrophages. In contrast, pairs of oral Gram-negative bacteria elicited synergistic production of IL-8 by macrophages. Similar results were not observed with TNFα, which only appeared additive with the polybacterial challenge. Selected Gram-negative bacterial pairs synergized in IL-6 production by immature DCs. In mature DCs (mDCs), a Porphyromonas gingivalis/Fusobacterium nucleatum and Porphyromonas intermedia/F. nucleatum polybacterial challenge resulted in significant synergism for IL-6 and TNFα levels. However, only the Pi/Fn combination synergized for IL-12 production and there appeared to be no polybacterial effect on IL-10 production by the mDCs. CONCLUSIONS: These results indicate that a polybacterial challenge of cells linking innate and adaptive immune responses results in varied response profiles that are dependent upon the characteristics of the microorganisms that are components of the polybacterial complex.

Molecular characterization of Treponema denticola infection-induced bone and soft tissue transcriptional profiles.

Treponema denticola is associated with subgingival biofilms in adult periodontitis and with acute necrotizing ulcerative gingivitis. However, the molecular mechanisms by which T. denticola impacts periodontal inflammation and alveolar bone resorption remain unclear. Here, we examined changes in the host transcriptional profiles during a T. denticola infection using a murine calvarial model of inflammation and bone resorption. T. denticola was injected into the subcutaneous soft tissue over the calvaria of BALB/c mice for 3 days, after which the soft tissues and the calvarial bones were excised. RNA was isolated and analysed for transcript profiling using Murine GeneChip arrays. Following T. denticola infection, 2905 and 1234 genes in the infected calvarial bones and soft tissues, respectively, were differentially expressed (P

Supernatants from oral epithelial cells and gingival fibroblasts modulate human immunodeficiency virus type 1 promoter activation induced by periodontopathogens in monocytes/macrophages.

Bacterial and host cell products during coinfections of Human Immunodeficiency Virus type 1-positive (HIV-1(+)) patients regulate HIV-1 recrudescence in latently infected cells (e.g. T cells, monocytes/macrophages), impacting highly active antiretroviral therapy (HAART) failure and progression of acquired immunodeficiency syndrome. A high frequency of oral opportunistic infections (e.g. periodontitis) in HIV-1(+) patients has been demonstrated; however, their potential to impact HIV-1 exacerbation is unclear. We sought to determine the ability of supernatants derived from oral epithelial cells (OKF4) and human gingival fibroblasts (Gin-4) challenged with periodontal pathogens, to modulate the HIV-1 promoter activation in monocytes/macrophages. BF24 monocytes/macrophages transfected with the HIV-1 promoter driving the expression of chloramphenicol acetyltransferase (CAT) were stimulated with Porphyromonas gingivalis, Fusobacterium nucleatum, or Treponema denticola in the presence of supernatants from OKF4 or Gin4 cells either unstimulated or previously pulsed with bacteria. CAT levels were determined by enzyme-linked immunosorbent assay and cytokine production was evaluated by Luminex beadlyte assays. OKF4 and Gin4 supernatants enhanced HIV-1 promoter activation particularly related to F. nucleatum challenge. An additive effect was observed in HIV-1 promoter activation when monocytes/macrophages were simultaneously stimulated with gingival cell supernatants and bacterial extracts. OKF4 cells produced higher levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukins -6 and -8 in response to F. nucleatum and P. gingivalis. Preincubation of OKF4 supernatants with anti-GM-CSF reduced the additive effect in periodontopathogen-induced HIV-1 promoter activation. These results suggest that soluble mediators produced by gingival resident cells in response to periodontopathogens could contribute to HIV-1 promoter activation in monocytes/macrophages, albeit this effect is most notable following direct stimulation of the cells with oral gram-negative bacteria.

A novel murine model for chronic inflammatory alveolar bone loss.

BACKGROUND AND OBJECTIVE: Chronic inflammatory bowel disease (IBD) demonstrates some similarities to the dysregulated chronic immunoinflammatory lesion of periodontitis. Trinitrobenzene sulphonic acid (TNBS) and dextran sodium sulphate (DSS) administered to rodents have been shown to elicit inflammatory responses that undermine the integrity of the gut epithelium in a similar manner to IBD in humans. The objective of this study was to evaluate the ability of these chemicals to elicit periodontal inflammation as a novel model for alveolar bone loss. MATERIAL AND METHODS: Mice were treated by oral application of TNBS twice a week, or with DSS in the diet over a period of 18 weeks. Alveolar bone loss was assessed on the defleshed skull using morphometric measures for area of bone resorption. RESULTS: The TNBS-treated animals tolerated oral administration with no clinical symptoms and gained weight at a similar rate to normal control animals. In contrast, DSS exerted a systemic response, including shortening of colonic tissue and liver enzyme changes. Both TNBS and DSS caused a localized action on periodontal tissues, with alveolar bone loss observed in both maxilla and mandibles, with progression in a time-dependent manner. Bone loss was detected as early as week 7, with more severe periodontitis increasing over the 18 weeks (p < 0.001). Young (7-month-old) and old (12-month-old) mice with severe combined immunodeficiency were treated with TNBS for a period of 7 weeks and did not develop significant bone loss. CONCLUSION: These data show that oral administration of TNBS or DSS provokes alveolar bone loss in concert with the autochthonous oral microbiota.

Oral bacteria induce a differential activation of human immunodeficiency virus-1 promoter in T cells, macrophages and dendritic cells.

INTRODUCTION: The human immunodeficiency virus (HIV) can integrate into T cells, macrophages and dendritic cells resulting in a latent infection. Reports have also demonstrated that various microbial and host cell factors can trigger HIV reactivation leading to HIV recrudescence, potentially undermining highly active antiretroviral therapies. METHODS: This study evaluated the capacity of oral bacteria associated with chronic periodontal infections to stimulate HIV promoter activation in various cell models of HIV latency. RESULTS: T cells (1G5) challenged with oral bacteria demonstrated a dose-response of HIV promoter activation with a subset of the bacteria, as well as kinetics that were generally similar irrespective of the stimuli. Direct bacterial challenge of the T cells resulted in increased activation of approximately 1.5- to 7-fold over controls. Challenge of macrophages (BF24) indicated different kinetics for individual bacteria and resulted in consistent increases in promoter activation of five fold to six fold over basal levels for all bacteria except Streptococcus mutans. Dendritic cells showed increases in HIV reactivation of 7- to 34-fold specific for individual species of bacteria. CONCLUSION: These results suggested that oral bacteria have the capability to reactivate HIV from latently infected cells, showing a relationship of mature dendritic cells > immature dendritic cells > macrophages > or = T cells. Expression of various pattern recognition receptors on these various cell types may provide insight into the primary receptors/signaling pathways used for reactivation by the bacteria.