Levels of interleukin 1β in tissue from sites of active periodontal disease

Interleukin 1 beta is a potent bone resorptive cytokine which also mediates soft tissue destruction through the stimulation of prostaglandin production, and the induction of collagenase and other proteases. This constellation of activities suggests a role for IL-1 beta in the pathogenesis of human periodontitis. Levels of IL-1 beta were therefore determined in tissue obtained from (1) diseased, active (2) diseased, inactive and (3) healthy sites from 12 patients with destructive adult periodontitis. Disease activity was defined as attachment loss of greater than or equal to 2.5 mm, as determined by sequential probing and the tolerance method. IL-1 beta was extracted from homogenates of tissue biopsies taken at surgery, and levels were quantified by ELISA. IL-1 beta was found to be present in most patient tissue samples, with levels ranging from 0-82 ng/ml. Disease active sites had higher IL-1 beta levels (p less than 0.05) than inactive and healthy sites. Diseased inactive sites were divided into 2 groups, those losing small amounts of attachment (0.5-2.0 mm, worsening sites) and those which showed no change or attachment gain (stable sites). Stable diseased sites had IL-1 beta levels which were comparable to those found in healthy sites, and which were significantly different from active sites (p less than 0.02). Worsening sites had IL-1 beta levels intermediate between the levels in stable and active sites. Detection of disease activity occurred more frequently at sites with IL-1 beta levels greater than 25 ng/ml (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Crevicular fluid level of β-glucuronidase in relation to clinical periodontal parameters and putative periodontal pathogens in early-onset periodontitis

Abstract. Analysis of β-glucuronidase (βG) in the gingival crevicular fluid (GCF) provides an indication of neutrophil influx into the crevicular environment. The aim of this study was to test the hypotheses that: (1) βG is significantly elevated in individuals with early-onset periodontitis (EOP) and that βG activity correlates with disease severity: and (2) βG level may reflect the local bacterial challenge in the gingival crevice. The study subjects consisted of a sub-sample of individuals examined in the National Survey of Oral Health of United States Children, which was undertaken during the 1986/87 school year. A total of 249 individuals were selected based on presence or absence of clinical attachment loss at baseline. The individuals were examined a second time 6 years later and the clinical attachment loss was assessed, and subgingival plaque and GCF were collected. The subjects were classified into 3 types of EOP and a control group, βG activity in the GCF and the levels of 7 putative micro-organisms in the pocket were assessed. The generalized EOP group had the highest βG activity, followed by the localized and incidental EOP groups, and the controls, respectively. There was a significant increase in βG activity with the increase in probing depth. Also, sites with bleeding on probing had a significantly higher βG activity than sites without bleeding. However, the effect of gingival inflammation on βG activity was more evident in the generalized and localized EOP groups. Sites harboring high levels of one or more of the micro-organisms tended to have high βG activity. There were moderate differences between the organisms with respect to their effect on βG activity, but sites with high numbers of Porphyromonas gingivalis, Prevotella intermedia, or Treponema dentieola also had the highest βG activity. The present findings suggest that βG activity in GCF from patients with EOP can be of value in the early identification of individuals at higher risk of developing EOP, The findings also suggest that host mechanisms leading to higher βG activity in EOP represent systemic responses and are only partly related to the presence of local factors at the site-level.     

Salivary IgA, lysozyme and β-microglobulin in periodontal disease

Abstract — The concentrations of IgA, lysozyme and β-microglobulin (β₂-m) were quantitated in wax-stimulated mixed saliva from 28 patients with severe periodontitis and from 28 healthy controls. The mutual correlations between IgA, lysozyme and β₂-rn were determined. In patients with periodontitis decreased lysozyme concentrations were detected when compared with controls (P<0.05). The correlation between IgA and β₂-m concentrations was highly significant in both groups studied (P<0.0001, and P <0.002), whereas β₂-m and lysozyme concentrations were positively correlated in patients but not in controls. In addition, a significant correlation between IgA and lysozyme was found only in periodontal patients ( P <0.001).     

LL-37抗菌肽及其与牙周病的关系

牙周病是一种微生物感染导致的牙周组织炎症破坏性疾病,先天性免疫防御功能正常与否与牙周炎的发展转归息息相关。组织蛋白酶抑制素cathelicidin是一种内源性抗菌肽,人源阳离子抗菌肽(hCAP)-18和LL-37是迄今发现的唯一的cathelicidin家族成员,而前者是其前体形式,后者则是其有活性的成熟形式。本文就hCAP-18和LL-37,LL-37的功能,LL-37与牙周病的关系等研究进展作一综述。     

Crevicular interleukin-1β in moderate and severe periodontitis patients and the effect of phase I periodontal treatment

Abstract Interleukin-1β (IL-1β), a potent stimulator of bone resorption, has been implicated in the pathogenesis of periodontal destruction. However, the relationship between cytokines and periodontal disease has not been studied sufficiently to allow definitive conclusions. The aims of this study are to investigate crevicular IL-1β and the clinical status of patients with periodontitis and the effect of phase I periodontal therapy on levels of IL-1β. For this study, 130 gingival crevicular fluid (GCF) samples were harvested from non-inflamed (15) and diseased sites (115) in 11 patients with periodontitis. The gingival index (GI) and probing depth (PD) of each site was recorded initially and one month after treatment. The amount of IL-1β in the GCF was measured by enzyme-linked immunosorbent assay (ELISA) using an antibody specific for this cytokine. Before treatment, IL-1β was found in 12 of 15 non-inflamed gingival crevices and in 112 of 115 diseased pockets. The amount of IL-1β varied from 4.03 to 511.12 pg/site. The average amount of IL-1/7 from diseased sites was 3-fold greater than that from non-inflamed sites. Both total amount of IL-1β and the GCF volume, but not IL-1β concentration, were found to be correlated, positively, with GI score and PD. After therapy, 63 sites from 7 patients were re-examined, and the amount of 1L-1β in 49 of 63 sites was found to have declined. These data suggest that the amount of Crevicular IL-1β is closely associated with periodontal status. This relationship may be valuable in monitoring periodontal disease activity.     

Interleukin-1β stimulates collagenase production by cultured human periodontal ligament fibroblasts

To examine the effects of interleukin-lβ (IL-1β) on collagenase production by human periodontal ligament fibroblasts (PLF) and gingival fibroblasts (GF) in Culture, collagenase activity in conditioned media was determined using a novel procedure that circumvented interference by enzyme inhibitors. Fibroblasts obtained from five paired periodontal ligament and gingival tissues were cultured for two weeks, and then incubated for a further 72 h in α-MEM supplemented with various concentrations of IL-1β (0 to 1250 pg/ml). The conditioned media from individual cultures were harvested and treated with dithiothreitol to inactivate TIMPs, and then with APMA, to activate the latent collagenase. Collagenase activity was measured fluorometrically using FITC-collagen as a substrate. IL-lβ induced a ∼2.4 to 5.2-fold increase in collagenase activity in PLF compared to a ∼1.4 to 2.2-fold increase in GF. These results are in contrast to previous studies in which collagenase activity was measured in the presence of TIMPs, and indicate that PLF are more sensitive to IL-1β than GF. Since both PLF and GF are present in periodontal lesions, it is possible that collagenase secretion stimulated by exposure to inflammatory cell products such as IL-lβ may participate in the destruction of collagen fibers involved in periodontal attachment.     

Host β-globin gene fragments in crevicular fluid as a biomarker in periodontal health and disease

Thaweboon B, Laohapand P, Amornchat C, Matsuyama J, Sato T, Nunez PP, Uematsu H, Hoshino E. Host β‐globin gene fragments in crevicular fluid as a biomarker in periodontal health and disease. J Periodont Res 2009; doi: 10.1111/j.1600‐0765.2008.01197.x. © 2009 John Wiley & Sons A/S Background and Objective: Leukocytes and epithelium are the first line of defense in preventing bacterial invasion into periodontium. Some of these cells die in gingival crevicular fluid, whereupon their DNA is spilled out. The present study was designed to investigate the profile of host β‐globin gene fragments in the gingival crevicular fluid of various periodontal conditions. Material and Methods: Gingival crevicular fluid from 40 teeth with chronic periodontitis, 30 with gingivitis and 22 that were clinically healthy were centrifuged (3000 g , 10 min). The supernatant (cell‐free gingival crevicular fluid) was centrifuged again (13,000 g , 10 min), resulting in the pellet and the supernatant as debris and debris‐free fractions, respectively. Specific primers for amplifying 110 bp, 536 bp and 2 kb amplicons of human β‐globin gene were used to investigate host DNA by quantitative and qualitative polymerase chain reaction. Results: The periodontitis group showed the largest amount of host β‐globin gene fragments, while the healthy group had the lowest. In the debris and debris‐free fractions, the 536 bp and 2 kb amplicons were more often detected in the periodontitis group than in the other groups. Interestingly, the presence of 2 kb amplicon in the debris fraction could be used to discriminate periodontitis from gingivitis and healthy groups because we found it in 85% of periodontitis samples but only in 13% of gingivitis samples, and it was absent in the healthy group. Conclusion: This study shows the different DNA profiles of cell‐free gingival crevicular fluid in periodontal health and disease. It suggests that the quantity and quality of host DNA are dependent on the disease conditions. Therefore, the β‐globin gene fragments in cell‐free gingival crevicular fluid may be a potential biomarker of periodontal disease progression.     

The gingival crevicular fluid Interleukin-1β and tumour necrosis factor-α levels in patients with rapidly progressive periodontitis

Cytokines are believed to play an important role in the pathogenesis of periodontal diseases. In the present study, gingival crevicular fluid (GCF) levels of two important cytokines, interleukin 1-/3 (IL-1β) and tumour necrosis factor-α (TNF-α) and, in addition, serum IL-1β levels, were determined in patients with severe and rapid periodontal breakdown by use of ELISA. While IL-1β was detected in all of the GCF samples studied, TNF-α could only be detected in about half the samples. The mean GCF IL-1β level was 38.45 ± 13.99 pg/mL, and the mean TNF-α level was 3.20 ± 1.39 pg/mL, respectively. The GCF IL-1β levels also presented a strong positive correlation with the mean pocket depths. Although weak, both of the cytokines also presented correlations with the presence of bleeding on probing. Additionally GCF samples contained increased IL-1β levels when compared with the serum samples suggesting local production mechanisms. The findings of the present study suggest that these cytokines may be involved in the pathogenesis of periodontal diseases (IL-1β being more significant), and also may help in defining the active phase of periodontal breakdown.     

HHV-6, HHV-7, HHV-8 in gingival biopsies from chronic adult periodontitis patients. A case-control study

BACKGROUND: Recent reports have suggested that various herpesviruses may be involved in the occurrence and progression of different forms of periodontal disease. OBJECTIVE: The objective of the present study was to investigate the presence of the novel herpesviruses HHV-6, HHV-7 and HHV-8 in gingival biopsies from patients affected by chronic adult periodontitis. As control, gingival biopsies from periodontally healthy subjects were analysed. MATERIALS AND METHODS: Gingival biopsies were harvested from 23 volunteers: 13 patients affected by chronic adult periodontitis (CAP) and 10 periodontally healthy subjects. Each CAP patient contributed two biopsies involving the epithelium and connective tissue facing the sulcus/periodontal pockets: one biopsy from a site having a probing pocket depth (PPD) > or =5 mm and presenting with bleeding upon probing (affected site) at the time of biopsy collection, and the other biopsy from a site with PPD< or =3 mm and without bleeding on probing (nonaffected site). After DNA extraction, nested PCR was used in herpesvirus identification. RESULTS: HHV-6 DNA sequences were detected in one non-affected site (8%) and no affected sites (0%) of CAP patients. One biopsy (10%) in healthy subjects revealed HHV-6 positivity. Tissue specimens in 10/13 CAP patients (77%) and 7/10 healthy subjects (70%) contained HHV-7 DNA. HHV-7 prevalence in affected and nonaffected sites of CAP patients was 77% and 54%, respectively. HHV-8 was detected in 7.7% of CAP patients and 0% of healthy subjects. CONCLUSIONS: Gingival tissue may act as a reservoir for HHV-7. A high prevalence of HHV-7 was detected in both periodontally diseased and healthy individuals. The prevalence of HHV-6 and -8 was similarly low in both groups. Our data do not support an association of investigated herpesvirus species with destructive periodontal disease.     

Expression of toll-like receptors 2, 4 and 9 is increased in gingival tissue from patients with type 2 diabetes and chronic periodontitis

Rojo‐Botello NR, García‐Hernández AL, Moreno‐Fierros L. Expression of toll‐like receptors 2, 4 and 9 is increased in gingival tissue from patients with type 2 diabetes and chronic periodontitis. J Periodont Res 2012; 47: 62–73. © 2011 John Wiley & Sons A/S Background and Objective: Broad evidence indicates that diabetes both increases the risk and hastens the progression of periodontal disease. Likewise, chronic inflammation or infections seem to provoke insulin resistance and thereby contribute to the development of diabetes and its complications. Innate immune responses, which appear to be altered in individuals with diabetes, are usually mediated by the recognition of pathogens through toll‐like receptors (TLRs). The constitutive expression of some TLRs has been reported in healthy human gingival tissue. Interestingly, the expression of TLRs 2 and 4 is increased with the severity of periodontal disease. Considering that the inflammatory reaction is exacerbated in individuals with diabetes and periodontitis, we suspected that the expression of some TLRs might be increased in gingival tissue in these patients. Material and Methods: In this study, we analyzed, by immunofluorescence, the expression of TLRs 2, 3, 4 and 9 in gingival tissues from healthy individuals and from periodontal patients with or without type 2 diabetes. Results: We found that the expression levels of TLRs 2, 3, 4 and 9 were higher in all periodontal patients than in healthy individuals. The expression of some TLRs was increased in subjects with periodontitis and diabetes relative to subjects with periodontitis but without diabetes; this increase in expression was found particularly in TLR2 and TLR9 in the connective tissue and in TLR4 at the epithelial region. Conclusion: These data suggest that the expression of these TLRs 2, 3, 4 and 9 in gingival tissue is higher in individuals with diabetes because its inflammatory reaction is exacerbated. Additionally, the expression of these TLRS is positively regulated with the severity of periodontal disease.     

The effect of tetracycline fiber therapy on β-glucuronidase and interleukin-1β in crevicular fluid

Abstract Treatment with the tetracycline HCL-containing (Actisite®) fiber has been shown to improve clinical measures of periodontitis, as well as reduce the number of sites infected with putative periodontal pathogens. In this study, we examined the effect of the tetracycline fiber on biochemical mediators of the host's inflammatory response in gingival crevicular fluid (GCF). The total amount of the lysosomal enzyme β-glucuronidase (βG), considered a marker of primary granule release from polymorphonuclear leukocytes and interleukin-1β, a cytokine with important proinflammatory effects, were examined in GCF. Patients with localized recurrent periodontitis were followed over a 16 week period. Treated teeth (Tx), teeth adjacent to treated teeth (ADJ) and control teeth (Cx) were studied. Following fiber therapy, the Tx teeth displayed statistically significant reductions in mean probing depth, depth of the deepest site and bleeding on probing over the 16 weeks of the trial. Significant reduction in the depth of the deepest site was also seen for the ADJ teeth over 16 weeks. Total βG in GCF was reduced for the Tx teeth comparing baseline to 16 weeks, but no significant changes were observed for the ADJ or Cx teeth. Prior to treatment, total βG for the Tx teeth was 211±49 U (mean±standard error), versus 146±174 U for the ADJ teeth and 121±33 U for the Cx teeth. 16 weeks treatment, the mean values for these 3 categories of teeth were comparable (Tx=95±20 U. ADJ = 93±42 U and Cx=103±29 U). For the Tx teeth, the maximum reduction in total βG following therapy occurred at 6 weeks (65%). Total IL-1β was significantly reduced for the Tx teeth at 3 and 6 weeks, but rebounded at 16 weeks. In contrast to what was seen for βG, the maximum reduction in total IL-1β for the Tx teeth was observed at 3 weeks (68%). These data suggest that host mediators associated with increased risk for active disease are reduced following tetracycline fiber therapy. Future studies will determine the relative importance of a reduced microbial challenge versus a tetracycline-mediated direct modification of the host response to account for the reduction in the host inflammatory response in GCF following tetracycline fiber therapy.     

Five parameters of gingival crevicular fluid from eight surfaces in periodontal health and disease

Volume and amounts of myeloperoxidase (MPO), lactoferrin (LF), aryl sulfatase (AS) and lactate dehydrogenase (LDH) were measured in gingival crevicular fluid (GCF) collected from the mesial and distal proximal surfaces of the premolars and first and second molars of 3 subject groups. Group assignment was based on subject mean gingival index (GI) and probing depth (PD) of sampled sites as follows: healthy, GI less than or equal to 0.5, PD less than or equal to 3.0; disease 1, GI greater than or equal to 1.0, PD greater than or equal to 3.0 mm; disease 2, PD greater than or equal to 4.0 mm. Attachment loss (ATL) of most sites in the 3 groups was: healthy, 0-1 mm; disease 1, 1-2 mm; and disease 2, 4-9 mm. GCF volume differed among surfaces and teeth in each of the 3 groups. The greater amount of GCF collected from posterior locations was not related to the GI and PD. Differences with sampling location in amounts of GCF constituents were restricted to MPO and LF. Most of these differences (greater amounts at posterior sites) were associated with more severe disease. Variability in amount and composition of GCF collected from different sites, therefore, should be considered in experiments which include quantitation of GCF parameters. The ratio of MPO in disease group 2 to disease group 1 was greater than similar ratios for GCF volume and LF, AS and LDH. The quantity of MPO was the only measure which differed between the 2 disease groups at all surfaces. MPO thus appears to have the greatest potential, among the measured parameters, to serve as a marker for advanced periodontal disease.     

β-Hemolytic streptococci and other β-hemolytic organisms in apical periodontitis and severe marginal periodontitis

Abstract— Thirteen teeth with necrotic pulps and apical periodontitis and nine severe periodontal pockets were cultured for presence of β-hemolytic streptococci and other β-hemolytic organisms. Samples were dispersed and plated on two non-selective and one selective growth media and incubated anaerobically and in 10% CO₂ in air. A total of 59 β-hemolytic colonies were purified and identified. Eight β-hemolytic streptococcal isolates were obtained from three of the severe marginal periodontitis sites. All were identified as belonging to the Streptococcus sanguis group. No β-hemolytic streptococci were detected in apical periodontitis samples. Twenty obligately anaerobic isolates were detected, all of which were known periodontal and endodontic pathogens. Isolates from apical periodontitis sites were identified as Propionibactrium acnes, Actinomyces naeslundii, Actinomyces odontolyticus and Peptostreptococcus micros , while severe marginal periodontal sites contained the same species with the addition of Actinomyces viscosus and Actinomyces meyeri . Of 19 staphylococci and micrococci, Staphylococcus epidermidis was the predominant isolate in both apical periodontitis and severe marginal periodontitis sites. However, less commonly known organisms such as Staphylococcus cohnii and Micrococcus sp . were identified in severe marginal periodontitis sites. The isolation of Bacillus sp. (12 isolates) in one severe marginal periodontitis and two apical periodontitis subjects was especially interesting, warranting consideration of this organism as a legitimate isolate and potential pathogen in oral disease.