Effects of tobacco smoke on the secretion of interleukin-1beta,tumor necrosis factor-alpha,and transforming growth factor-beta from peripheral blood mononuclear cells

Alterations of the host response caused by short-term exposure to high levels of smoke during the act of smoking (acute smoke exposure) as well as long-term exposure to lower levels of tobacco substances in the bloodstream of smokers (chronic smoke exposure) may play a role in the pathogenesis of periodontal diseases in smokers. In this study, we examined the secretion of three cytokines [interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, and transforming growth factor (TGF)-beta] from mononuclear blood cells from current smokers and non-smokers exposed to in vitro tobacco smoke (which may be comparable to in vivo acute smoke exposure) and mononuclear blood cells from current smokers not exposed to further in vitro smoke (which may be comparable to chronic smoke exposure). Peripheral blood mononuclear cells were isolated from eight healthy current smokers and eight healthy non-smokers, plated in culture wells, exposed in vitro for 1-5 min to cigarette smoke in a smoke box system or not exposed (baseline controls), and then incubated without further smoke exposure for another 24 h. Supernatants from each well were then collected and assayed for the concentrations of the three cytokines by enzyme-linked immunosorbent assay (ELISA). At baseline, mean IL-1beta levels were higher in smokers than in non-smokers (mean: 10.6 vs. 5.9 pg/ml, anova: P < 0.05). In both smokers and non-smokers, secreted levels of IL-1beta increased from 0 to 5 min of in vitro smoke exposure (mean: 5.9-9.9 pg/ml, t-test: P < 0.05 for non-smokers only) with levels in smokers higher than in non-smokers (P > 0.05). Mean TNF-alpha levels increased from 0 to 2 min of smoke exposure and decreased from 2 to 5 min in smokers and non-smokers, with higher levels in non-smokers than smokers at all time-points (P > 0.05). Mean TGF-beta levels were higher in smokers than in non-smokers at all time-points (mean: 180.5 vs. 132.0 pg/ml, P < 0.05 at 5 min only) with no significant alteration of the pattern of secretion with cigarette smoke exposure. These observed alterations in the secretion of cytokines from mononuclear blood cells in smokers, relative to non-smokers, and with in vitro smoke exposure may play a role in the pathogenesis of periodontal diseases in smokers.     

Saliva and Serum Levels of B‐Cell Activating Factors and Tumor Necrosis Factor‐α in Patients With Periodontitis

Background: B‐lymphocytes play a central and critical role in the adaptive immune response against invading pathogens. This study evaluates saliva and serum levels of APRIL (a proliferation‐inducing ligand), B‐cell activating factor (BAFF), tumor necrosis factor‐α (TNF‐α), interleukin (IL)‐6, and IL‐10 in patients with chronic periodontitis (CP) or aggressive periodontitis (AgP) and periodontally healthy individuals. Methods: Twenty‐five patients with AgP, 20 patients with CP, and 20 periodontally healthy individuals were included. Smoking status was recorded, and all individuals were divided into non‐smokers and smokers. Saliva and serum samples were collected before clinical periodontal measurements. APRIL, BAFF, TNF‐α, IL‐6, and IL‐10 levels in serum and saliva samples were determined by enzyme‐linked immunosorbent assay. Statistical analysis was performed using multivariate analysis of variance and bivariate correlation. Results: Serum and saliva levels of TNF‐α, APRIL, BAFF, IL‐6, and IL‐10 were similar in CP and AgP groups. Serum levels of TNF‐α, APRIL, and BAFF and saliva levels of BAFF were significantly higher in periodontitis groups than healthy controls (P <0.05). Non‐smokers with CP or AgP had lower levels of saliva TNF‐α and APRIL and serum APRIL and IL‐6 than smokers with CP or AgP (P <0.05). Saliva APRIL and serum TNF‐α and IL‐6 levels were significantly higher in healthy smokers than healthy non‐smokers (P <0.05). Clinical periodontal parameters correlated positively with TNF‐family cytokines and negatively with IL‐10 (P <0.05). Conclusions: Within the limits of this study, it may be suggested that elevated salivary and serum TNF‐α, APRIL, and BAFF in patients with periodontitis may contribute to the dominance of B cells in periodontitis lesions. Moreover, higher levels in healthy smokers than non‐smoking counterparts may play a role in detrimental effects of smoking on periodontal tissues.     

Tumor Necrosis Factor‐α and Interleukin (IL)‐1β, IL‐6, and IL‐8 Impair In Vitro Migration and Induce Apoptosis of Gingival Fibroblasts and Epithelial Cells,Delaying Wound Healing

Background: Multiple factors affect oral mucosal healing, such as the persistence of an inflammatory reaction. The present study evaluates effects of tumor necrosis factor (TNF)‐α and interleukin (IL)‐1β, IL‐6, and IL‐8 on epithelial cells (ECs) and human gingival fibroblasts (GFs) in vitro. Methods: GFs and ECs were seeded in 96‐well plates (1 × 10⁴ cells/well) in plain culture medium (Dulbecco’s modified Eagle’s medium [DMEM]) containing 1% antibiotic/antimycotic solution and 10% fetal bovine serum, and incubated for 24 hours. Both cell lines were exposed for 24 hours to the following cytokines: 1) TNF‐α (100 ng/mL); 2) IL‐1β (1 ng/mL); 3) IL‐6 (10 ng/mL); and 4) IL‐8 (10 ng/mL). All cytokines were diluted in serum‐free DMEM. Control cultures were exposed only to serum‐free DMEM. Effects of exposure to inflammatory cytokines were determined by means of: 1) apoptosis (anexin V); 2) cell migration (wound healing assay); 3) inflammatory cytokine synthesis (enzyme‐linked immunosorbent assay). Data were analyzed by Kruskal–Wallis and Mann–Whitney U tests (α = 0.05). Results: Increased apoptosis rates were noted when cells were exposed to inflammatory cytokines, except ECs exposed to IL‐1β. Cell migration was negatively affected by all inflammatory cytokines for both cell lines. ECs and GFs exposed to IL‐6 and IL‐8 significantly increased synthesis of TNF‐α and IL‐1β. Conclusions: Demonstrated results indicate negative effects of tested inflammatory cytokines on ECs and GFs, inducing apoptosis and impairing cell migration. These results can justify delayed oral mucosa healing in the presence of inflammatory reaction.     

Comparison of Peri‐Implant Soft Tissue Parameters and Crestal Bone Loss Around Immediately Loaded and Delayed Loaded Implants in Smokers and Non‐Smokers: 5‐Year Follow‐Up Results

Background: The aim of this study is to compare peri‐implant soft tissue parameters (plaque index [PI], bleeding on probing [BOP], and probing depth [PD] ≥4 mm) and crestal bone loss (CBL) around immediately loaded (IL) and delayed loaded (DL) implants in smokers and non‐smokers. Methods: Thirty‐one patients with IL implants (16 smokers and 15 non‐smokers) and 30 patients with DL implants (17 smokers and 13 non‐smokers) were included. Personal data regarding age, sex, and duration and daily frequency of smoking were gathered using a questionnaire. Peri‐implant PI, BOP, and PD ≥4 mm were recorded, and mesial and distal CBL was measured on standardized digital radiographs. Multiple group comparisons were performed using the Bonferroni post hoc test (P <0.05). Results: All implants replaced mandibular premolars or molars. Mean scores of PI (P <0.05) and PD ≥4 mm (P <0.05) were statistically significantly higher in smokers compared with non‐smokers in patients with IL and DL dental implants. The mean score of BOP (P <0.05) was statistically significantly higher in non‐smokers compared with smokers in both groups. CBL (P <0.05) was statistically significantly higher in smokers compared with non‐smokers in both groups. There was no statistically significant difference in PI, BOP, PD ≥4 mm, and total CBL among smokers with IL and DL implants. Conclusions: Tobacco smoking enhances peri‐implant soft tissue inflammation and CBL around IL and DL implants. Loading protocol did not show a significant effect on peri‐implant hard and soft tissue status in healthy smokers and non‐smokers.     

Orthodontic Force Increases Interleukin‐1β and Tumor Necrosis Factor‐α Expression and Alveolar Bone Loss in Periodontitis

Background: The present study aims to evaluate the effects of orthodontic movement (OM) on the periodontal tissues of rats with ligature‐induced periodontal disease. Methods: Eighty‐eight rats were divided into four groups: 1) negative control (sham operated); 2) periodontal disease; 3) OM; and 4) periodontal disease followed by OM (OMP). Rats were sacrificed 3 hours or 1, 3, or 7 days after OM commencement. Bone volume fraction (BVF) and bone mineral density (BMD) were assessed in hemimaxillae by microcomputed tomography analysis. Expression of the proinflammatory cytokines interleukin (IL)‐1β and tumor necrosis factor (TNF)‐α were evaluated in gingival samples by quantitative polymerase chain reaction and enzyme‐linked immunosorbent assay, and in the furcation region by immunohistochemistry analysis (IHC). Results: The OMP group had lower BVF and BMD levels compared to the other groups at day 7 (P <0.05). Maximum messenger ribonucleic acid expression of both cytokines was observed in the OMP group at day 1 (P <0.05). In the same period, all proteins were expressed in high levels for all test groups compared to the control group. The number of cells positive for IL‐1β and TNF‐α by IHC was highest in the OMP group at day 1, with progressive reduction thereafter. Conclusion: The results suggest that OM acts synergistically with periodontal disease in periodontal breakdown through upregulation of proinflammatory cytokines.     

Interleukin‐1 β, interleukin‐12 and interleukin‐18 levels in gingival fluid and serum of patients with gingivitis and periodontitis

Introduction: Cytokines are of major importance in periodontal disease progression. Interleukin‐12 (IL‐12) stimulates interferon‐γ production by T helper type 1 (Th1) cells while IL‐18 induces Th1 responses when present with IL‐12 but Th2 responses in the absence of IL‐12. IL‐1β has been correlated with periodontal disease destruction. This study determined the local concentrations of these cytokines in sites of gingivitis and periodontitis. Methods: Gingival crevicular fluid was collected from two sites in each of 10 gingivitis patients and from two gingivitis sites and two periodontitis sites from each of 10 periodontitis patients. Serum samples were also collected. IL‐1β, biologically active IL‐12 p70, the IL‐12 p40 subunit and IL‐18 concentrations were determined by enzyme‐linked immunoabsorbent assay. Results: IL‐1β and IL‐18 concentrations were higher in the gingival crevicular fluid from periodontitis patients than in that from gingivitis patients; IL‐18 concentrations were higher than those of IL‐1β. Very little IL‐12, either p40 or p70, was detected in the gingival crevicular fluid samples. In the serum, very low levels of cytokines were found. The level of serum IL‐12 p40, however, was higher than in the fluid from periodontitis sites of periodontitis patients. Conclusion: The local production of IL‐1β and IL‐18 in the gingival crevicular fluid increased with increasing inflammation and IL‐18 was the predominant cytokine at both gingivitis and periodontitis sites. Very little IL‐12 was detected with levels decreasing with increasing inflammation. These results suggest that there is an association between severity of periodontal disease and levels of IL‐1, IL‐12 and IL‐18.     

Significance of elevated gingival crevicular fluid tumor necrosis factor‐α and interleukin‐8 levels in chronic hemodialysis patients with periodontal disease

Da? A, F?rat ET, Kadiro?lu AK, Kale E, Y?lmaz ME. Significance of elevated gingival crevicular fluid tumor necrosis factor‐α and interleukin‐8 levels in chronic hemodialysis patients with periodontal disease. J Periodont Res 2010; 45: 445–450. © 2010 John Wiley & Sons A/S Background and Objective: The prevalence of chronic renal disease in industrialized countries is increasing, and chronic renal disease and periodontitis can have significant, reciprocal effects. The aim of this study was to evaluate the associations between specific clinical parameters and the levels of tumor necrosis factor‐α (TNF‐α) and interleukin‐8 (IL‐8) in the gingival crevicular fluid of hemodialysis (HD) patients with periodontal disease. Material and Methods: Forty‐three HD patients and 43 systemically healthy subjects were enrolled in this study. Plaque index (PI), gingival index (GI) and pocket depth were used to determine periodontal status. Venous blood samples were obtained from each patient in the morning before the dialysis session and analyzed to determine the levels of inflammatory, biochemical and hematological parameters. Gingival crevicular fluid was collected from all subjects, and the levels of TNF‐α and IL‐8 were determined in the gingival crevicular fluid samples. Results: The following results were obtained from HD patients and controls: TNF‐α (pg/mL), 31.40 ± 1.46 and 3.06 ± 0.15 (p < 0.001); IL‐8 (pg/mL), 90.98 ± 94.03 and 35.05 ± 16.86 (p < 0.001); PI, 1.69 ± 1.02 and 0.04 ± 0.02 (p < 0.001); GI, 0.82 ± 0.06 and 0.04 ± 0.02 (p < 0.001); and pocket depth, 2.23 ± 0.63 and 1.51 ± 0.05 (p < 0.001), respectively. In addition, there were positive correlations between TNF‐α and PI (r = 0.642, p < 0.001), between TNF‐α and GI (r = 0.565, p < 0.001), between TNF‐α and pocket depth (r = 0.522, p < 0.001), between IL‐8 and PI (r = 0.402, p = 0.002), between IL‐8 and GI (r = 0.396, p = 0.002), and between IL‐8 and pocket depth (r = 0.326, p = 0.012). There were negative correlations between albumin and PI (r = ?0.491, p < 0.001), albumin and GI (r = ?0.406, p < 0.001), albumin and pocket depth (r = ?0.464, p < 0.001) and albumin and CRP (r = ?0.467, p = 0.002) and between the gingival crevicular fluid levels of TNF‐α and IL‐8, TNF‐α and hemoglobin (r = ?0.745, p < 0.001; r = ?0.285, p < 0.05) (respectively). Conclusion: The levels of TNF‐α and IL‐8 in gingival crevicular fluid were significantly higher in HD patients than in controls. There were strong, positive correlations between clinical periodontal parameters and the levels of inflammatory cytokines in gingival crevicular fluid from the HD patients.     

IL‐1β and PGE2 levels are increased in the saliva of children with Langerhans cell histiocytosis

Langerhans cell histiocytosis (LCH) is a rare disorder mainly of children, whose pathogenesis is still unknown. Some studies have demonstrated that LCH lesions produce different cytokines abnormally that may be relevant to the pathogenesis of the disease. The purpose of this study was to investigate interleukin‐1β (IL‐1β) and prostaglandin E2 (PGE₂) levels in saliva from children with different clinical subtypes of LCH. We studied 29 children with LCH: seven unifocal (Group I), seven multifocal (Group II), 15 multisystemic (Group III) and 12 healthy volunteers (Group IV). Salivary IL‐1β and PGE₂ levels were significantly higher in LCH than in normal children. A multi‐comparison test showed significantly (P < 0.001) higher levels of both IL‐1β and PGE₂ in saliva from Group III compared with Groups II and I. A significant correlation (r = 0.05) between IL‐1β and PGE₂ concentrations in saliva from each group was determined. Our findings demonstrated an association between high concentrations of salivary IL‐1β and PGE₂ and advanced stages of the disease. This allows us to suggest that the abnormal amount of these factors in saliva may serve as a risk marker for disease progression.     

Saliva and Serum Levels of Pentraxin‐3 and Interleukin‐1β in Generalized Aggressive or Chronic Periodontitis

Background: Pentraxin‐3 (PTX3) is a multifactorial protein involved in immunity and inflammation, which is rapidly produced and released by several cell types in response to inflammatory signals. The aim of the present study is to evaluate saliva, serum levels of PTX3, interleukin (IL)‐1β in patients with generalized chronic periodontitis (CP) or aggressive periodontitis (AgP), and periodontally healthy individuals. Methods: A total of 94 participants (25 patients with AgP, 25 patients with CP, and 44 periodontally healthy individuals matched with AgP and CP groups) were recruited. Saliva and serum samples were collected. Clinical periodontal measurements were recorded. PTX3, IL‐1β levels in serum, and saliva samples were determined by enzyme‐linked immunosorbent assay. Data were tested statistically using Kruskal‐Wallis, Mann‐Whitney U, and Spearman ρ rank test. Results: Serum and saliva data were similar in CP and AgP groups. Saliva levels of IL‐1β were significantly higher in the AgP and CP groups than controls (P <0.05). Salivary PTX3 levels were similar in the CP and control groups. Significantly higher salivary concentrations of PTX3 were detected in the AgP group than the control group (P <0.05). Saliva PTX3 levels correlated with plaque index and bleeding on probing in the CP group (P <0.05). Serum and saliva PTX3 levels correlated with those of IL‐1β in the AgP group (P <0.05). Conclusions: It may be suggested that PTX3 is related with periodontal tissue inflammation. Its salivary concentrations may have a diagnostic potential. Additional intervention and follow‐up studies coupling PTX3 concentrations with microbiologic analysis would better clarify its role in periodontal diseases.     

Effects of interleukin‐1β and tumor necrosis factor‐α on macrophage inflammatory protein‐3α production in synovial fibroblast‐like cells from human temporomandibular joints

Background

Interleukin‐1β (IL‐1β) and tumor necrosis factor‐α (TNF‐α) are key mediators of the intracapsular pathological conditions of the temporomandibular joint (TMJ). Therefore, the gene expression profiles in synovial fibroblast‐like cells (SFCs) from patients with internal derangement of the TMJ were examined after they were stimulated with IL‐1β or TNF‐α to determine which genes were altered.

Methods

Ribonucleic acid was isolated from SFCs after IL‐1β or TNF‐α treatment. Gene expression profiling was performed using oligonucleotide microarray analysis. On the basis of the results of this assay, we investigated the kinetics of macrophage inflammatory protein‐3α (MIP‐3α) gene expression using PCR, and protein production in TMJ SFCs stimulated by IL‐1β or TNF‐α using an ELISA. Inhibition experiments were performed with MAPK and NFκB inhibitors. SFCs were stimulated with IL‐1β or TNF‐α after treatment with inhibitors. The MIP‐3α levels were measured using an ELISA.

Results

Macrophage inflammatory protein‐3α was the gene most upregulated by IL‐1β‐ or TNF‐α stimulation. The mRNA and protein levels of MIP‐3α increased in response to IL‐1β in a time‐dependent manner. In contrast, during TNF‐α stimulation, the MIP‐3α mRNA levels peaked at 4 h, and the protein levels peaked at 8 h. In addition, the IL‐1β‐ and TNF‐α‐stimulated MIP‐3α production was potently reduced by the MAPK and NFκB signaling pathway inhibitors.

Conclusion

Interleukin‐1β and TNF‐α increased the MIP‐3α production in SFCs via the MAPK and NFκB pathways. These results suggest that the production of MIP‐3α from stimulation with IL‐1β or TNF‐α is one factor associated with the inflammatory progression of the internal derangement of the TMJ.     

Periodontitis and Endothelial Dysfunction: Periodontal Clinical Parameters and Levels of Salivary Markers Interleukin‐1β, Tumor Necrosis Factor‐α, Matrix Metalloproteinase‐2, Tissue Inhibitor of Metalloproteinases‐2 Complex,and Nitric Oxide

Background: Periodontitis has been associated with a greater risk for atherosclerotic cardiovascular disease (ACD). Endothelial dysfunction (ED) is a parameter of early ACD, and its association with periodontitis has rarely been investigated to date. The objective of this study is to evaluate the association between periodontitis and ED by means of periodontal clinical parameters and salivary markers interleukin (IL)‐1β, tumor necrosis factor (TNF)‐α, nitric oxide (NO), and matrix metalloproteinase (MMP)‐2 and tissue inhibitor of metalloproteinases (TIMP)‐2 complex. Methods: Forty‐seven individuals were divided into two groups: 1) 24 individuals with chronic periodontitis (CP); and 2) 23 individuals without CP. Periodontal examinations of bleeding on probing (BOP), probing depth (PD), and clinical attachment level (CAL) were performed. ED was evaluated by means of flow‐mediated dilatation (FMD) of the brachial artery. Salivary concentrations of IL‐1β, TNF‐α, and MMP‐2/TIMP‐2 complex were assessed using enzyme‐linked immunosorbent assay. NO determination was based on the reaction of Griess. Results: Individuals with CP presented higher occurrence of ED than individuals without CP (P = 0.03 after reactive hyperemia; P = 0.05 after sublingual nitrate). A significant association among the production of MMP‐2/TIMP‐2 complex with the presence of CP (P = 0.008) and periodontal parameters PD, CAL, and BOP was identified. Concentration of salivary markers IL‐1β, TNF‐α, and NO was similar in individuals with and without CP. A significant positive correlation between NO and ED was also identified. Conclusions: Periodontitis was positively associated with ED, expressed by a smaller percentage of FMD of the brachial artery and higher salivary levels of MMP‐2/TIMP‐2 complex. Additionally, salivary levels of NO were significantly associated with better functioning of the vascular endothelium.     

Tumor Necrosis Factor‐α and Porphyromonas gingivalis Lipopolysaccharides Decrease Periostin in Human Periodontal Ligament Fibroblasts

Background: Periostin is a matricellular protein essential for tissue integrity and maturation and is believed to have a key function as a modulator of periodontal ligament (PDL) homeostasis. The aim of this study is to evaluate whether periodontal disease‐associated pathogen‐related virulence factors (endotoxins/lipopolysaccharides [LPS]) and proinflammatory cytokines alter the expression of periostin in PDL cells. Methods: Human PDL cultures were exposed to inflammatory mediators (tumor necrosis factor‐α [TNF‐α]), bacterial virulence factors (Porphyromonas gingivalis LPS) or a combination in a biomechanically challenged environment. Culture conditions were applied for 24 hours, 4 days, and 7 days. Periostin and TGF‐β inducible gene clone H3 (βIGH3) mRNA expression from cell lysates were analyzed. Periostin and βIGH3 proteins were also detected and semiquantified in both cell lysates and cell culture supernatants by Western blot. In addition, periostin localization by immunofluorescence was performed. Analysis of variance and Fisher tests were used to define the statistical differences among groups (P <0.05). Results: In a mechanically challenged environment, periostin protein was more efficiently incorporated into the matrix compared to the non‐loaded controls (higher levels of periostin in the supernatant in the non‐loaded group). Interestingly, chronic exposure to proinflammatory cytokines and/or microbial virulence factors significantly decreased periostin protein levels in the loaded cultures. There was greater variability on βIGH3 levels, and no particular pattern was clearly evident. Conclusions: Inflammatory mediators (TNF‐α) and bacterial virulence factors (P. gingivalis LPS) decrease periostin expression in human PDL fibroblasts. These results support a potential mechanism by which periostin alterations could act as a contributing factor during periodontal disease progression.     

Major surface protein complex of Treponema denticola induces the production of tumor necrosis factor α, interleukin‐1β, interleukin‐6 and matrix metalloproteinase 9 by primary human peripheral blood monocytes

Gaibani P, Caroli F, Nucci C, Sambri V. Major surface protein complex of Treponema denticola induces the production of tumor necrosis factor α, interleukin‐1β, interleukin‐6 and matrix metalloproteinase 9 by primary human peripheral blood monocytes. J Periodont Res 2010; 45: 361–366. © 2010 John Wiley & Sons A/S Background and Objective: Treponema denticola is a micro‐organism that is involved in the pathogenesis of periodontitis. Major surface protein complex (MSPc), which is expressed on the envelope of this treponeme, plays a key role in the interaction between T. denticola and gingival cells. The peptidoglycan extracted from T. denticola induces the production of a large variety of inflammatory mediators by macrophage‐like cells, suggesting that individual components of T. denticola cells induce the inflammatory response during periodontal disease. This study was designed to demonstrate that MSPc of T. denticola stimulates release of proinflammatory mediators in primary human monocytes. Material and Methods: Primary human monocytes were separated from the blood of healthy donors and incubated for up to 24 h with varying concentrations of MSPc. The production of tumor necrosis factor α (TNF‐α), interleukin‐1β (IL‐1β), interleukin‐6 (IL‐6) and matrix metalloproteinase 9 (MMP‐9) was measured at different time points with commercially available enzyme‐linked immunosorbent assays. Results: T. denticola MSPc induced the synthesis of TNF‐α, IL‐1β, IL‐6 and MMP‐9 in a dose‐ and time‐dependent manner. Similar patterns of TNF‐α, IL‐1β and IL‐6 release were observed when cells were stimulated with 100 and 1000 ng/mL of MSPc. The production of MMP‐9 was significant only when cells were treated with 1000 ng/mL of MSPc. Conclusion: These results indicate that T. denticola MSPc, at concentrations ranging from 100 ng/mL to 1.0 μg/mL, activates a proinflammatory response in primary human monocytes.     

Interleukin‐6 Family of Cytokines in Crevicular Fluid of Renal Transplant Recipients With and Without Cyclosporine A–Induced Gingival Overgrowth

Background: Interleukin (IL)‐6 family of cytokines, including IL‐6, oncostatin M (OSM), leukemia inhibitory factor (LIF), and IL‐11, have fibrogenic features. The current study determines gingival crevicular fluid (GCF) levels of fibrosis‐related IL‐6–type cytokines in cyclosporine A (CsA)–induced gingival overgrowth (GO). Methods: Eighty non‐smokers were included (40 CsA‐medicated renal transplant patients with GO [GO + ; n = 20] or without GO [GO?; n = 20], 20 individuals with gingivitis, and 20 healthy participants). Probing depth and plaque, papilla bleeding, and hyperplastic index scores were recorded. GCF samples were obtained from the mesio‐buccal aspects of two teeth. GCF IL‐6, IL‐1β, OSM, LIF, and IL‐11 levels were analyzed by enzyme‐linked immunosorbent assay. Results: The GO+ and GO? groups had higher IL‐6 total amounts than the healthy group (P <0.008). IL‐1β total amounts in the GO+ group were significantly higher than in both the healthy and GO? groups (P <0.008). OSM total amount was elevated in the GO+ and GO? groups compared with both the gingivitis and healthy groups (P <0.008). All groups had similar LIF and IL‐11 total amounts (P >0.008). Moderate positive correlations were detected among IL‐6, IL‐1β, OSM, and IL‐11 total amount in GCF and clinical parameters (P <0.05). Conclusions: IL‐6 and OSM increases in GCF as a result of CsA usage or an immunosuppressed state irrespective of the severity of inflammation and the presence of GO. The IL‐6 family of cytokines might not be directly involved in biologic mechanisms associated with CsA‐induced GO. Lack of an association between assessed IL‐6 cytokines and CsA‐induced GO might indicate distinct effects of these cytokines on fibrotic changes of different tissues.     

Carbon monoxide releasing molecule‐3 inhibits concurrent tumor necrosis factor‐α‐ and interleukin‐1β‐induced expression of adhesion molecules on human gingival fibroblasts

Song H, Zhao H, Qu Y, Sun Q, Zhang F, Du Z, Liang W, Qi Y, Yang P. Carbon monoxide releasing molecule‐3 inhibits concurrent tumor necrosis factor‐α‐ and interleukin‐1β‐induced expression of adhesion molecules on human gingival fibroblasts. J Periodont Res 2011; 46: 48–57. © 2010 John Wiley & Sons A/S Background and Objective: Carbon monoxide releasing molecule‐3 (CORM‐3) is a newly reported compound that has shown anti‐inflammatory effects in a number of cells. In this study, we aimed to investigate the influence of CORM‐3 on concurrent tumor necrosis factor‐α (TNF‐α)‐ and interleukin (IL)‐1β‐induced expression of adhesion molecules on human gingival fibroblasts (HGF). Material and Methods: HGF were cultured from the explants of normal gingival tissues. Cells were costimulated with TNF‐α and IL‐1β in the presence or absence of CORM‐3 for different periods of time. The expression of adhesion molecules, nuclear factor‐kappaB (NF‐κB) and phosphorylated p38 was studied using western blotting. RT‐PCR was applied to check the expression of the adhesion molecules at the mRNA level. The activity of NF‐κB was analysed using a reporter gene assay. Results: CORM‐3 inhibited the up‐regulation of intercellular adhesion molecule 1, vascular cell adhesion molecule 1 and endothelial leukocyte adhesion molecule in HGF after costimulation with TNF‐α and IL‐1β, which resulted in the decreased adhesion of peripheral blood mononuclear cells to these cells. Sustained activation of the NF‐κB pathway by costimulation with TNF‐α and IL‐1β was suppressed by CORM‐3, which was reflected by a reduced NF‐κB response element‐dependent luciferase activity and decreased nuclear NF‐κB‐p65 expression. CORM‐3 inhibited MAPK p38 phosphorylation in response to stimulation with proinflammatory cytokines. Conclusion: The results of this study bode well for the application of CORM‐3 as an anti‐inflammatory agent to inhibit NF‐κB activity and to suppress the expression of adhesion molecules on HGF, which suggests a promising potential for CORM‐3 in the treatment of inflammatory periodontal disease.     

Clinical and Immunoinflammatory Evaluation of One‐Stage Full‐Mouth Ultrasonic Debridement as a Therapeutic Approach for Smokers With Generalized Aggressive Periodontitis: A Short‐Term Follow‐Up Study

Background: This study aims to evaluate the effect of one‐stage full‐mouth ultrasonic debridement (OSFMUD) on clinical and immunoinflammatory parameters in smokers with generalized aggressive periodontitis (GAgP). Methods: Fourteen smoking and 14 non‐smoking patients with GAgP were selected. After initial supragingival therapy, patients were treated by OSFMUD. Full‐mouth parameters evaluated were: 1) plaque index (PI); 2) bleeding scores (BS); 3) probing depth (PD); and 4) clinical attachment level (CAL). Clinical evaluation was performed, and gingival crevicular fluid (GCF) was collected for selected sites (ss) at baseline and 1, 3, and 6 months. GCF was analyzed via enzyme‐linked immunosorbent assay for: 1) receptor activator of nuclear factor‐κ B ligand (RANKL); 2) osteoprotegerin (OPG); 3) interleukin (IL)‐6; and 4) tumor necrosis factor (TNF)‐α, whereas secreted osteoclastogenic factor of activated T‐cells (SOFAT) was evaluated by Western blotting. Results: Significant reduction (P <0.05) was observed between baseline and 6 months for: 1) PI; 2) BS; and 3) PD, with no difference between smoking and non‐smoking patients (P >0.05). Regarding CAL, only non‐smoking patients showed a significant decrease (P <0.05). Significant reduction (P <0.05) was observed in both groups for: 1) PIss; 2) PDss; 3) bleeding on probing; and 4) relative CAL. Smoking and non‐smoking patients presented significantly decreased levels of IL‐6 and TNF‐α over time (P <0.05); however, no difference was observed between groups (P >0.05). RANKL was significantly different (P <0.05) only for non‐smokers at 6 months, whereas OPG was not significant (P >0.05). SOFAT expression was significantly lower (P <0.05) after OSFMUD for non‐smokers only. Conclusion: Considering the clinical and immunoinflammatory parameters evaluated in this short‐term follow‐up study, it can be concluded that OSFMUD can be used as an alternative treatment for smokers with GAgP.     

Periodontal Status and Whole Salivary Cytokine Profile Among Smokers and Never‐Smokers With and Without Prediabetes

Background: Whole salivary interleukin (IL)‐1β and IL‐6 in smokers and never‐smokers with prediabetes remains uninvestigated. The aim of this study is to assess the periodontal status and whole salivary IL‐1β and IL‐6 levels among smokers and never‐smokers with and without prediabetes (controls). Methods: Ninety‐five males (45 with prediabetes and 50 systemically healthy controls) were included. Twenty‐seven controls and 29 patients with prediabetes were smokers. Periodontal parameters (plaque index, bleeding on probing, probing depth, clinical attachment loss, and marginal bone loss) were measured, and the number of missing teeth were recorded. Fasting blood glucose (FBG) and hemoglobin A1c (HbA1c) levels were recorded. Unstimulated whole saliva samples were collected, unstimulated whole salivary flow rate (UWSFR) was determined, and IL‐1β and IL‐6 levels were measured. P values <0.05 were considered statistically significant. Results: FBG (P <0.05) and HbA1c (P <0.05) levels were higher among patients with prediabetes than controls. All patients with prediabetes were hyperglycemic. UWSFR was significantly higher among controls than among patients with prediabetes (P <0.05). Periodontal parameters and whole salivary IL‐1β and IL‐6 levels were comparable among smokers and never‐smokers with prediabetes. Among controls, periodontal parameters and whole salivary IL‐1β and IL‐6 levels were higher among smokers than never‐smokers (P <0.05). Conclusions: Among controls, periodontal inflammation was worse, and whole salivary IL‐1β and IL‐6 levels are higher in smokers than never‐smokers. Among patients with prediabetes, periodontal inflammation and whole salivary IL‐1β and IL‐6 levels were comparable between smokers and never‐smokers.     

Moderate acute exercise (70% VO2 peak) induces TGF‐β, α‐amylase and IgA in saliva during recovery

Strenuous exercise promotes changes in salivary IgA and can be associated with a high incidence of upper respiratory tract Infections. However, moderate exercise enhances immune function. The effect of exercise on salivary IgA has been well studied, but its effect on other immunological parameters is poorly studied. Thus, this study determined the effect of moderate acute exercise on immunological salivary parameters, such as the levels of cytokines (TGF‐β and IL‐5), IgA, α‐amylase and total protein, over 24 h. Ten male adult subjects exercised for 60 min at an intensity of 70% VO₂ peak. Saliva samples were collected before (‘basal’) and 0, 12 and 24 h after an exercise session. The total salivary protein was lower after 12 and 24 h than immediately after exercise, whereas α‐amylase increased at 12 and 24 h after exercise compared with basal levels. The IgA concentration was increased at 24 h after exercise relative to immediately after exercise, and there was no difference in the IL‐5 while TGF‐β concentration increased in recovery. In conclusion, 70% VO₂ peak exercise does not induce changes immediately after exercise, but after 24 h, it produces an increase in salivary TGF‐β without changing IL‐5.     

Focal adhesion kinase activation is required for TNF‐α‐induced production of matrix metalloproteinase‐2 and proinflammatory cytokines in cultured human periodontal ligament fibroblasts

Since focal adhesion kinase (FAK) was proposed as a mediator of the inflammatory response, we have investigated the role of this molecule in the release of inflammatory cytokines by cultured human periodontal ligament fibroblasts (HPDLFs), cells that are thought to be important in the patient's response to periodontal infection. Human periodontal ligament fibroblasts were stimulated by tumor necrosis factor alpha (TNF‐α) and its effects on interleukin (IL)‐6 and IL‐8 release were measured by ELISA. Expression of matrix metalloproteinase 2 (MMP‐2) protein was analysed by western blotting. The levels of IL6, IL8, and MMP2 mRNA were evaluated by real‐time PCR. Tumor necrosis factor alpha dose‐dependently induced the phosphorylation of FAK, whereas small interfering FAK (siFAK) inhibited TNF‐α‐induced FAK phosphorylation. Tumor necrosis factor alpha also stimulated the production of IL‐6, IL‐8, and MMP‐2 in a dose‐dependent manner. Knockdown of FAK significantly suppressed TNF‐α‐induced expression of IL6 and IL8 mRNA and release of IL‐6 and IL‐8 protein in HPDLFs. Similarly, MMP‐2 down‐regulation was significantly prevented by siFAK. Our results strongly suggest that knockdown of FAK can decrease the production of TNF‐α‐induced IL‐6, IL‐8, and MMP‐2 in HPDLFs. These effects may help in understanding the mechanisms that control expression of inflammatory cytokines in the pathogenesis of periodontitis.