Associations of serum concentrations of IgG,IgA, IgM and interleukin-1beta with early-onset periodontitis classification and race

BACKGROUND: The significance of serum concentrations of various antibodies and cytokines in the pathogenesis of early-onset periodontitis (EOP) is not well understood. Recent reports suggest differences between young blacks and whites in certain humoral responses, regardless of periodontal status. This study was undertaken to compare the serum concentrations of IgG, IgA, IgM, and IL-1beta in EOP subjects with that of healthy controls, and to study the effect of race on these levels. MATERIAL AND METHODS: This case-control study included 228 individuals, 19-25 years old who were selected from a larger population examined in the National Survey of Oral Health of United States Children in 1986/1987. The subjects were classified by their EOP status and they included 166 subjects with EOP and 62 healthy controls. Blood samples were used to assess the serum concentrations of IgG, IgM, IgA, IgG subclass, and IL-1beta. RESULTS: The serum concentrations of IgG, IgG subclasses, IgA, and IgM in blacks were not significantly different in the generalized, localized and incidental EOP groups as compared to the healthy controls. The serum IL-1beta concentration was slightly and uniformly lower in the EOP groups than in the control group, although not statistically significant. Blacks had significantly higher serum concentrations of total IgG, and of IgG1, IgG2 and IgG3 than whites and Hispanics. Hispanics had significantly higher serum concentrations of IgM and IgG4 than whites and blacks. Hispanics also had a significantly higher serum concentration of IL-1beta than blacks. CONCLUSIONS: Total antibody response in blacks is not associated with EOP classification. Race has a significant effect on serum antibody concentrations irrespective of disease classification, with blacks having significantly higher serum concentrations of IgG1, IgG2 and IgG3 than whites and Hispanics.     

Effects of induced periapical abscesses on rat pregnancy outcomes

Objective

The objective of this study was to determine the effects of induced periapical abscesses on pregnant rats.

Design

In 1/2 of the animals (n = 16), the pulps of the maxillary right molars were exposed to the oral environment, which resulted in a periapical abscess. The other 1/2 (n = 16) were sham-operated. 1/2 of the animals of both groups became pregnant 2 weeks later. The pregnancy duration, and weight and number of pups were assessed at delivery. Serum, liver and uterine horn samples were taken from all animals at euthanasia and serum IL-6, endothelin-1, TNF-α, IL-10, cortisol and insulin were determined by ELISA. Liver concentrations of IL-6, CRP and IL-6 and uterine horn concentrations of IL-6, vascular endothelial growth factor (VEGF), TNF-α, IL-10 and IL-1-β were assessed by ELISA. Blood glucose concentrations were determined using a glucometer. Outcome variables were compared by factorial ANOVA, a post hoc Tukey test, and Pearson’s correlation test.

Results

Pregnant rats with periapical abscesses had a significantly longer pregnancy and delivered pups with a significantly higher birthweight (p < 0.05). They had significantly higher concentrations of IL-6, VEGF, IL-1-β, and IL-10 within the uterine horn and IL-6, CRP and TNF-α within the liver (p < 0.01). Blood glucose and serum TNF-α, IL-6, endothelin-1, IL-10, and insulin concentrations were significantly higher in the pregnant animals with pulpal abscesses (p < 0.01).

Conclusion

The significant increase in serum TNF-α, taken together with significant increases in blood glucose and serum insulin concentrations, suggest that animals with induced periapical abscesses developed insulin resistance, which significantly affected their pregnancy outcomes.     

Genetic variation in the recognition of Porphyromonas gingivalis antigens in mice

T-cell cytokine profiles, anti-Porphyromonas gingivalis antibodies and Western blot analysis of antibody responses were examined in BALB/c, CBA/CaH, C57BL6 and DBA/2J mice immunized intraperitoneally with different doses of P. gingivalis outer membrane antigens. Splenic CD4 and CD8 cells were examined for intracytoplasmic interleukin (IL)-4, interferon (IFN)-gamma and IL-10 by FACS analysis and levels of anti-P. gingivalis antibodies in the serum samples determined by enzyme-linked immunosorbent assay. Western blot analysis was performed on the sera from mice immunized with 100 microg of P. gingivalis antigens. The four strains of mice demonstrated varying degrees of T-cell immunity, although the T-cell cytokine profiles exhibited by each strain were not affected by different immunizing doses. While BALB/c and DBA/2J mice exhibited responses that peaked at immunizing doses of 100-200 microg of P. gingivalis antigens, CBA/CaH and C57BL6 demonstrated weak T-cell responsiveness compared with control mice. Like the T-cell responses, serum antibody levels were not dose dependent. DBA/2J exhibited the lowest levels of anti-P. gingivalis antibodies followed by BALB/c with CBA/CaH and C57BL6 mice demonstrating the highest levels. Western blot analysis showed that there were differences in reactivity between the strains to a group of 13 antigens ranging in molecular weight from 15 to 43 kDa. Antibody responses to a number of these bands in BALB/c mice were of low density, whereas CBA/CaH and C57BL6 mice demonstrated high-density bands and DBA/2J mice showed medium to high responses. In conclusion, different immunizing doses of P. gingivalis outer membrane antigens had little effect on the T-cell cytokine responses and serum anti-P. gingivalis antibody levels. Western blot analysis, however, indicated that the four strains of mice exhibited different reactivity to some lower-molecular-weight antigens. Future studies are required to determine the significance of these differences, which may affect the outcome of P. gingivalis infection.     

Clinical benefits of oral irrigation for periodontitis are related to reduction of pro-inflammatory cytokine levels and plaque

BACKGROUND: Although a growing body of evidence indicates that oral irrigation with water has therapeutic benefits in periodontitis, the mechanisms of action have not been elucidated. The aims of this study were: (1) to analyze the effects of oral irrigation (Water Pik Oral Irrigator) on the clinical signs of adult periodontitis (AP) and on the levels of interleukin-1 beta (IL-beta), prostaglandin-E2 (PGE2), interleukin-10 (IL-10) and interferon-gamma (IFN-gamma) in GCF, and (2) to analyze the influence of the periodontitis-related IL-1 genotype (IL-1GT) on these variables. METHOD: A single-center, blinded study in otherwise healthy humans (n= 52) with localized mild to moderate AP was carried out, using the following groups: group A (n= 12), no oral hygiene for 14 days; group B (n=20), routine oral hygiene (ROH) for 14 days; group C (n=20), supra-gingival oral irrigation plus ROH for 14 days. Group A patients were crossed-over to group C for 14 days (=day 28) after a professional prophylaxis. Group assignment was randomized by a coin toss, with the exception of group A subjects, who were self-selected as per recommendations of the internal review board for human subjects. GCF was sampled from 3 study teeth per patient and analyzed for IL-1 beta, PGE2, IL-10 and IFN gamma by ELISA on days 0, 7, 14 and 28. Probing pocket depths (PPD), clinical attachment levels (CAL), bleeding on probing (BOP), gingival index (GI) and plaque index (PI) were measured by a calibrated examiner (TWS) on days 0, 14 and 28. Analysis of covariance was performed using SAS 6.12 and Proc Mixed with group and IL-1GT as the factors and the baseline levels as the covariate, with output being least squares means and least significant difference (LSD). Significant differences were declared if the p-value for the F-statistic was < or =0.05. RESULTS: Oral irrigation plus ROH resulted in a significant reduction in PPD, BOP, GI and PI, as well as IL-beta levels by 7 days and PGE2 levels by 14 days, relative to ROH or no oral hygiene. Interestingly, decreased IL-1 beta levels in patients using oral irrigation plus ROH was accompanied by a trend for increased levels of the "anti-inflammatory" cytokine IL-10. ROH reduced GI, BOP and PI, and PGE2 levels by 14 days, but had no effect on IL-1 beta or IL-10 levels relative to no oral hygiene. The effects of no oral hygiene were reversed by a prophy followed by oral irrigation plus ROH for 14 days. No clinical differences were evident between IL-1 GT (+) patients (n= 1) and GT (-) patients (n=40), but the former had significantly elevated levels of GCF IL-10 and borderline increases in IL-1 beta (p=0.07). CONCLUSIONS: Oral irrigation with water for 14 days had an improved therapeutic benefit for AP over that of routine oral hygiene alone and this improvement was accompanied by a down-modulation of the pro-inflammatory cytokine profile in GCF.     

Effect of compressive force on the expression of inflammatory cytokines and their receptors in osteoblastic Saos-2 cells

OBJECTIVE: In orthodontic tooth movement, some cytokines released from periodontal ligament fibroblasts and alveolar bone osteoblasts on the pressure side can alter the normal processes of bone remodelling, resulting in physiological bone resorption. We examined the effect of compressive force and interleukin (IL)-1 type I receptor antagonist (IL-1ra) on the expression of inflammatory cytokines that promote osteoclast formation, as well as on their receptors, in osteoblastic Saos-2 cells. DESIGN: The cells were cultured in Dulbecco's modified Eagle medium containing 10% fetal bovine serum with or without continuous compressive force (0.5-3.0 g/cm(2)) and/or IL-1ra for up to 24h. The gene expression levels of the cytokines and their receptors were estimated by determining mRNA levels using real-time PCR; the protein levels were determined using ELISA or immunohistochemical staining. RESULTS: The expression of IL-1beta, IL-1 receptor, IL-6, IL-6 receptor, IL-8 receptor, IL-11 and tumor necrosis factor-alpha (TNFalpha) increased depending on the strength and duration of the compressive force, whereas the expression of IL-8, IL-11 receptor and TNFalpha receptor did not change with the application of compressive force. The expression of cytokines and their receptors produced by 3.0 g/cm(2) of compressive force decreased with the simultaneous addition of IL-1ra and the decrease was remarkable in IL-8 receptor, IL-11 and TNFalpha. CONCLUSION: These results indicate that mechanical stress induces the production of inflammatory cytokines and their receptors in osteoblasts and the phenomenon is enhanced by the autocrine action of IL-1beta, which is increased in amount by mechanical stress.     

Infection-mediated early-onset periodontal disease in P/E-selectin-deficient mice

BACKGROUND: Retrospective and correlation studies suggest that early-onset periodontal disease may be due to a deficiency in phagocyte function, a pathogenic oral biofilm, and/or dysregulated gingival cytokine expression. Increased susceptibility to periodontal disease is therefore thought to result from multiple risk factors. METHODS: We tested this hypothesis prospectively using P/E-selectin adhesion molecule deficient mice that mimic the human syndrome leukocyte adhesion deficiency II. RESULTS: Our studies demonstrate that, in comparison to wild type animals, P/E-/- mice exhibit: spontaneous, early onset alveolar bone loss which is significant by 6 weeks of age; a 10-fold elevation in bacterial colonization of their oral cavities; and elevated gingival tissue levels of the bone resorptive cytokine IL-1alpha. Alveolar bone loss is completely prevented by prophylactic antibiotic therapy. CONCLUSIONS: These experiments provide the first prospective evidence for the multiple risk factor hypothesis of periodontal disease, and validate the first animal model for early onset periodontitis in which both the microbiota and host response can be systematically manipulated. P/E-/- animals should be useful in testing the virulence of putative periodontal pathogens, in determining the role of host resistance factors in periodontitis, in exploring the proposed relationship(s) between infection mediated alveolar bone loss and systemic health disorders, and exploring their genetic relationships.     

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目的探讨中性多形核白细胞(PMN)在重度慢性牙周炎发病机制中的作用。方法2005年11月至2006年3月,于四川大学华西口腔医学院收集牙周健康者和重度慢性牙周炎患者各4人作为研究对象,以梯度密度离心法分离外周血PMN,并与趋化肽fMLP作用,酶联免疫吸附试验(ELISA)法测定其分泌白细胞介素(IL)-1β和IL-8的质量浓度。结果重度慢性牙周炎患者PMN分泌IL-1β和IL-8的质量浓度高于牙周健康者(P<0.05),且fMLP对PMN的刺激作用具有时间依赖性,随作用时间延长,PMN分泌IL-1β和IL-8的质量浓度升高(P<0.01)。结论重度慢性牙周炎患者PMN具有"过度炎症反应性"特征,可分泌高质量浓度的IL-1β和IL-8,在牙周炎发病机制中具有一定作用。     

Human T‐cell responses to oral streptococci in human PBMC‐NOD/SCID mice

We investigated cellular and humoral immune responses to oral biofilm bacteria, including Streptococcus mutans, Streptococcus anginosus, Streptococcus sobrinus, and Streptococcus sanguinis, in NOD/SCID mice immunized with human peripheral blood mononuclear cells (hu‐PBMC‐NOD/SCID mice) to explore the pathogenicity of each of those organisms in dental and oral inflammatory diseases. hu‐PBMC‐NOD/SCID mice were immunized by intraperitoneal injections with the whole cells of the streptococci once a week for 3 weeks. FACS analyses were used to determine the percentages of various hu‐T cell types, as well as intracellular cytokine production of interleukin‐4 and interferon‐γ. Serum IgG and IgM antibody levels in response to the streptococci were also determined by enzyme‐linked immunosorbent assay. S. anginosus induced a significant amount of the proinflammatory cytokine interferon‐γ in CD4⁺ and CD8⁺ T cells in comparison with the other streptococci. However, there was no significant differences between the streptococci in interleukin‐4 production by CD4⁺ and CD8⁺ T cells after inoculation. Further, S. mutans significantly induced human anti‐S. mutans IgG, IgG₁, IgG₂, and IgM antibodies in comparison with the other organisms. In conclusion, S. anginosus up‐regulated Th1 and Tc1 cells, and S. mutans led to increasing levels of their antibodies, which was associated with the induction of Th2 cells. These results may contribute to a better understanding of human lymphocyte interactions to biofilm bacteria, along with their impact on dental and mucosal inflammatory diseases, as well as endocarditis.     

IL-17A stimulates the expression of inflammatory cytokines via celecoxib-blocked prostaglandin in MC3T3-E1 cells

Objective

The prostaglandins (PGs) released from osteoblasts can alter the process of bone remodelling. Recently, we showed that compressive force induced the expression of pro-inflammatory cytokine interleukin (IL)-17s and their receptors in osteoblastic MC3T3-E1 cells and that IL-17A was expressed most highly. Consequently, in the current study we examined the effect of IL-17A and/or celecoxib on PGE₂ production and the expression of cyclooxygenases (COXs) and inflammatory cytokines in MC3T3-E1 cells. We also examined the effects of PGE₂ and cyclohexamide on the expression of inflammatory cytokines.

Methods

Cells were cultured with or without IL-17A (0.1, 1.0, or 10 ng/ml) in the presence or absence of 10 μM celecoxib, a specific inhibitor of COX-2, for up to 72 h. Cells were pretreated with or without 10 μg/ml cycloheximide, protein synthesis inhibitor, for 30 min, and then cultured with 10 ng/ml IL-17A for 24 h. Cells were also cultured with or without 1.5 ng/ml PGE₂ for 24 h. PGE₂ production was determined by ELISA. The expression of COX-1, COX-2, IL-1α, IL-6, IL-8, IL-11, and TNF-α mRNAs and proteins was determined by real-time PCR and ELISA, respectively.

Results

The expression of COX-2, IL-1α, IL-6, IL-8, IL-11, and TNF-α, as well as PGE₂ production increased in the presence of IL-17A, whereas COX-1 expression did not change. Celecoxib blocked the stimulatory effect of IL-17A on the expression of COX-2, IL-1α, IL-6, IL-8, and IL-11 as well as PGE₂ production, whereas it did not block TNF-α expression. Cycloheximide pretreatment suppressed the expression of IL-17-induced inflammatory cytokines. The expression of IL-1α, IL-6, IL-8, and IL-11 increased by the addition of PGE₂, whereas TNF-α expression was not affected.

Conclusion

These results suggest that IL-17A stimulates the expression of bone resorption-related inflammatory cytokines through an autocrine mechanism involving celecoxib-blocked PGs, mainly PGE₂, in osteoblasts.     

Effect of smoking on gingival crevicular fluid cytokine profile during experimental gingivitis

BACKGROUND: Cigarette smoking is a significant risk factor in the pathogenesis of periodontal disease, able to influence both the subgingival microbiota and host responses. AIM: The aim of the present study was to determine the influence of smoking on the amount of IL-1beta, IL-4 and IL-8 in gingival crevicular fluid (GCF) during experimental gingivitis in humans. MATERIAL AND METHODS: Twenty-two healthy subjects, 10 smokers and 12 non-smokers, participated in the study. After professional cleaning, they performed optimal hygiene to reach perfect clinical gingival health. Oral hygiene measures were ceased for a period of 10 days. Clinical indices, including plaque index (PI), gingival index (GI), probing pocket depth (PPD) and bleeding on probing (BOP), were assessed 2 days before (day -2), at the beginning (day 0) and at the end of the experimental gingivitis period (day 10). At the same time, GCF was collected from 12 sites in each patient, by means of durapore filter membranes. Total amounts of IL-1beta, IL-4 and IL-8 were determined by enzyme-linked immunoadsorbent assay. RESULTS: Clinical data revealed that both smokers and non-smokers showed an increase in PI, GI and BOP scores during the experiment. Although no differences were noted with regard to PI at day 10, the GI and BOP were significantly less pronounced in smokers than non-smokers (p < 0.005). Non-smokers showed higher total amounts of IL-4 but lower amounts of IL-8 than smokers, throughout the experiment. Total amounts of IL-1beta and IL-8 increased significantly during plaque accumulation in both groups. IL-4 remained stable for the smoker group and decreased for the non-smoker group. CONCLUSIONS: The present results indicate that smoking interferes with cytokine production. When performing studies regarding the pathogenesis of periodontitis, the smoking status of the participants needs to be taken into consideration.