Impact of Metal Particles Released during Ultrasonic Scaling of Titanium Surfaces on Human Gingival Fibroblasts.

PURPOSE: Metal particles found in tissues around dental implants have been proposed to play a pathogenic role in peri-implantitis. Ultrasonic scaling has been suggested as a mechanism by which these particles can be inadvertently released into surrounding tissues. Furthermore, risk factors like diabetes can result in exacerbation of this inflammatory condition. The current study aimed to analyze metal particles released from titanium surfaces during ultrasonic scaling and their impact on pro-inflammatory cytokine production by human gingival fibroblasts. METHODS: Metal particles generated from ultrasonic scaling of titanium discs using two different tips (metal and poly-etheretherketone tips) were characterized using scanning electron microscopy and elemental analysis. Endotoxin levels and Human gingival fibroblast viability, in the presence commercial and ultrasonically generated particles were determined. Fibroblasts, cultured in high or low glucose growth medium, were incubated with commercial titanium particles or ultrasonically generated particles in the presence or absence of interluekin-1ß. Interleukin 6 and interleukin 8 production were then quantified using Enzyme linked immunosorbent assay. RESULTS: Analysis of particles after scaling of titanium discs showed significant levels of titanium particles. Commercial titanium particles and generated particles had no effect of fibroblast viability. Endotoxin levels of all particles were too low to stimulate HGF cells. IL-1ß significantly stimulated IL-6 and IL-8 production. However, commercial, and generated particles generally had no significant effect on IL- 6 and IL-8 production. CONCLUSION: Our study concluded that particles generated during ultrasonic scaling had no significant effect on viability of HGF cells and cytokine production.

Simvastatin loaded chitosan guided bone regeneration membranes stimulate bone healing.

BACKGROUND AND OBJECTIVE: Electrospun chitosan membranes (ESCM) modified with short-chain fatty acids have the ability to control the release of simvastatin (SMV), an anti-cholesterol drug with osteogenic potential, for guided bone regeneration (GBR) applications. This study evaluated in vivo osteogenic effects of rapid short release of SMV (4 weeks) vs long sustained release (8 weeks) from acetic anhydride (AA)-and hexanoic anhydride (HA)-modified ESCMs, respectively. METHODS: AA ESCMs loaded with 10 or 50 µg SMV and HA ESCMs loaded with 50 µg SMV were evaluated for biocompatibility and bone formation at 4 and 8 weeks, in 5 mm critical size rat calvarial defects, using histological evaluation and micro-CT analysis. RESULTS: No severe inflammatory response was noticed around the ESCMs. Less hydrophobic AA membranes showed signs of resorption by week 4 and were almost completely resorbed by week 8 whereas the more hydrophobic HA membranes resorbed slowly, remaining intact over 8 weeks. In micro-CT analysis, 10 µg SMV-loaded AA membranes did not show significant bone formation as compared to non-loaded AA membranes at either evaluation time points. 50 µg SMV-loaded AA membranes stimulated significantly more bone formation than non-loaded AA membranes by week 4 (%bone = 31.0 ± 5.9% (AA50) vs 18.5 ± 13.7% (AA0)) but showed no difference at week 8. HA membranes with 50 µg SMV showed significantly more bone formation as compared to corresponding non-loaded membranes by week 8 (%bone = 61.7 ± 8.9% (HA50) vs 33.9 ± 29.7% (HA0)), though such an effect was not significant at week 4. CONCLUSION: These results indicate that modified ESCMs may be used to control the release of SMV and promote bone healing in GBR applications.

Vitamin D attenuates human gingival fibroblast inflammatory cytokine production following advanced glycation end product interaction with receptors for AGE.

BACKGROUND AND OBJECTIVES: Vitamin D [1,25(OH)2 D3 or 1,25D3] is critical in musculoskeletal health, inflammation, immune response, and glucose metabolism. Patients with vitamin D deficiency may be at higher risk of diabetes and periodontitis. Diabetic patients exhibit exacerbated inflammation and more periodontal destruction. Advanced glycation end products (AGEs), formed during diabetic hyperglycemia, activate inflammatory pathways in periodontitis. Human gingival fibroblasts (HGFs) express receptors for AGEs (RAGEs) and can contribute to inflammation. OBJECTIVES: Determine whether glycated human serum albumin (G-HSA) augments HGF IL-6 and IL-8 production, and whether treatment with 1,25D3 attenuates cytokine production following stimulation with G-HSA + IL-1ß and/or IL-17. MATERIAL AND METHODS: HGFs were incubated ±G-HSA or normal human serum albumin (HSA), ±IL-1ß and/or IL-17, ±1,25D3. Cytokines were measured by ELISA. Neutralizing anti-RAGE was used to assess AGE-RAGE interaction. Endotoxin was measured using the ToxinSensor™ System. Data were expressed as mean ± standard deviation and analyzed using a one-way analysis of variance (ANOVA) and Scheffe's F procedure for post hoc comparisons. RESULTS: G-HSA or IL-1ß, but not HSA, significantly stimulated IL-6 and IL-8 production. G-HSA or HSA when combined with IL-1ß or IL-1ß + IL-17 synergistically stimulated IL-6 and IL-8. Neutralizing anti-RAGE inhibited IL-6 and IL-8 produced by cells stimulated with IL-1ß + G-HSA but not (+HSA). Synergism caused by HSA did not appear to be mediated by endotoxin since its levels in G-HSA and HSA were not sufficient to stimulate fibroblasts. Vitamin D inhibited IL-6 and IL-8 production stimulated by G-HSA or HSA + IL-1ß or IL-1ß + IL-17. CONCLUSIONS: Results suggest that the "perioprotective" effects of vitamin D are related to its ability to regulate inflammatory cytokine production by HGFs following AGE-RAGE interaction.

Evaluation of biocompatibility and degradation of chitosan nanofiber membrane crosslinked with genipin.

Chitosan, a natural polysaccharide, has demonstrated potential as a degradable biocompatible guided bone regeneration membrane. This study aimed to evaluate the in vivo biocompatibility and degradation of chitosan nanofiber membranes, with and without genipin crosslinking as compared with a commercial collagen membrane in rat model. Chitosan nanofiber membranes, with and without genipin crosslinking, and collagen membrane (control) were implanted subcutaneously in the backs of 30 rats. The membranes were analyzed histologically at 2, 4, 8, 12, 16, and 20 weeks. Sections were viewed and graded by a blinded pathologist using a 4-point scoring system (0 = absent, 1 = mild, 2 = moderate, and 3 = severe) to determine the tissue reaction to the membranes and to observe membrane degradation. There was no statistically significant difference in histological scores among chitosan and collagen membranes at different time points. Absence or minimal inflammation was observed in 57-74% of the membranes across all groups. Most chitosan membranes persisted for 16-20 weeks, whereas most collagen membranes disappeared by resorption at 12-16 weeks. The general tissue response to chitosan nanofiber membranes with and without genipin crosslinking, was similar to that of control commercial collagen membrane. However, the chitosan membranes exhibited slower degradation rates than collagen membranes.

Vitamin D and its impact on oral health--an update.

Vitamin D has been shown to regulate musculoskeletal health by mediating calcium absorption and mineral homeostasis. Evidence has demonstrated that vitamin D deficiency may place subjects at risk for not only low mineral bone density/osteoporosis and osteopenia but also infectious and chronic inflammatory diseases. Studies have shown an association between alveolar bone density, osteoporosis and tooth loss and suggest that low bone mass may be a risk factor for periodontal disease. Several recent reports demonstrate a significant association between periodontal health and the intake of vitamin D. An emerging hypothesis is that vitamin D may be beneficial for oral health, not only for its direct effect on bone metabolism but also due to its ability to function as an anti-inflammatory agent and stimulate the production of anti-microbial peptides.

Profiling gingival crevicular fluid from smoking and non-smoking chronic periodontitis patients.

BACKGROUND: Smoking is a major risk factor for the development and progression of chronic periodontitis (CP). Gingival crevicular fluid (GCF) derived from these patients contains many proteins that could serve as important diagnostic indicators. A protein chip technology called Surface Enhanced Laser Desorption/Ionization-Time of Flight-Mass Spectrometry (SELDI-TOF-MS) has recently been developed to facilitate protein profiling of complex biological mixtures. The hypothesis to be tested was SELDI-TOF-MS could distinguish between GCF of CP patients by smoking status and pocket depth. METHODS: GCF samples collected before treatment from sixteen CP patients (eight smokers and eight non-smokers) at both shallow and deep sites were evaluated by SELDI-TOF-MS. Spectral fingerprints were constructed for both cohorts and analyzed by a two-way ANOVA according to smoking status and pocket depth. Significance threshold was set at p < 0.05. Mean molecular weight (MW) peaks and intensities were also analyzed. RESULTS: The spectral fingerprints were significantly different between the two cohorts when analyzed by ANOVA according to smoking status (p < 0.0001) but not pocket depth (p = 0.9876). Also, the mean intensity of many individual MW peaks were determined to be significantly different between the two cohorts. Several peaks ranging in MW from 11-14 kDa were only detected in the GCF obtained from smokers. CONCLUSIONS: This study has demonstrated that profiling of GCF by SELDI-TOF-MS can distinguish between CP smokers and non-smokers. Moreover, over-expressed proteins in GCF from smokers may serve as biomarkers for this high risk patient population.

Augmented transforming growth factor-beta1 in gingival crevicular fluid of smokers with chronic periodontitis.

BACKGROUND: Patients who smoke are at increased risk for chronic periodontitis (CP). Also, CP patients who smoke exhibit significantly less reduction of probing depths and gains in clinical attachment compared to non-smokers following periodontal therapy. Several studies suggest that the effects of smoking on the host response may be paramount in regulating the basal systemic inflammatory status and therapeutic outcomes in this cohort. Growth factors, specifically transforming growth factor beta1 (TGF-beta1), are critical in regulating the wound healing response by controlling cell division, differentiation, and motility. The hypothesis to be tested was that gingival crevicular fluid (GCF) TGF-beta1 production was altered in smokers compared to non-smokers with CP. METHODS: GCF was collected from smokers and non-smokers with CP, both at baseline and 1 to 2 weeks after initial therapy. GCF volume was determined using an electronic device and TGF-beta1 concentration was measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Smokers exhibited a higher mean concentration of GCF TGF-beta1 at baseline compared to non-smokers (P = 0.03). After initial therapy, smokers exhibited significantly less reduction in mean GCF volume compared to non-smokers (P = 0.04). CONCLUSIONS: Augmented constitutive production of GCF TGF-beta1 in smokers may explain the clinical appearance of fibrotic gingival tissue exhibited by this patient cohort. A diminished reduction in GCF volume in smokers following root instrumentation suggests a chronic inflammatory status in conjunction with an ineffective host response. These findings support the concept that smokers with CP display an altered local inflammatory response after initial therapy, perhaps symptomatic of colonization by residual periodontal pathogens.

Cyclooxygenase-2 inhibitors decrease interleukin-1beta-stimulated prostaglandin E2 and IL-6 production by human gingival fibroblasts.

BACKGROUND: Previous work showed that normal and aggressive periodontitis (AgP) gingival fibroblasts produce the bone-resorbing cytokine IL-6. PGE2 is important in regulating IL-6 production. Non-steroidal anti-inflammatory drugs inhibit PG synthesis via COX-1 and/or COX-2 isoenzymes and may inhibit periodontal destruction. COX-2 is induced after cellular activation (i.e., by inflammatory cytokines such as IL-1beta). Little is known about IL-1beta-stimulated AgP fibroblast IL-6 and PGE2 production and their regulation by COX inhibitors. The objective of this study was to determine the effects of COX-2 inhibitors on amounts of PGE2 and IL-6 made by IL-1beta-stimulated gingival fibroblasts. METHODS: Gingival fibroblasts (2.5 x 10(4)) from healthy or severe periodontitis patients were cultured in serum-free medium, with or without IL-1beta (10(-11)M) for 24 hours, with or without the COX-1/2 inhibitor indomethacin or the selective COX-2 inhibitors NS-398, celecoxib, or rofecoxib. PGE2 and IL-6 in culture supernatants were determined by specific enzyme-linked immunosorbent assay (ELISA)s. RESULTS: All of the COX inhibitors caused dose-dependent decreases in IL-1beta-stimulated PGE2, to a maximum of > 90% in all cell lines (P < or = 0.0001). The selective COX-2 inhibitors, but not indomethacin, caused partial (generally up to approximately 60%), dose-dependent decreases in IL-1beta-stimulated IL-6 in all cell lines (P < or = 0.003). When exogenous PGE2 was added concurrently with COX-2 inhibitors before addition of IL-1beta, IL-6 production returned to levels at or approaching that produced by cells exposed only to IL-1beta (P < or = 0.04). CONCLUSION: The results suggest that COX-2 inhibition may be useful in helping to control fibroblast production of IL-6 in patients with severe periodontitis.

Cyclooxygenase-2 Inhibitors Decrease Interleukin-1ß-Stimulated Prostaglandin E2 and IL-6 Production by Human Gingival Fibroblasts.

BACKGROUND: Previous work showed that normal and aggressive periodontitis (AgP) gingival fibroblasts produce the boneresorbing cytokine IL-6. PGE2 is important in regulating IL-6 production. Non-steroidal anti-inflammatory drugs inhibit PG synthesis via COX-1 and/or COX-2 isoenzymes and may inhibit periodontal destruction. COX-2 is induced after cellular activation (i.e., by inflammatory cytokines such as IL-1ß). Little is known about IL-1ß-stimulated AgP fibroblast IL-6 and PGE2 production and their regulation by COX inhibitors. The objective of this study was to determine the effects of COX-2 inhibitors on amounts of PGE2 and IL-6 made by IL-1ß-stimulated gingival fibroblasts. METHODS: Gingival fibroblasts (2.5 × 104 ) from healthy or severe periodontitis patients were cultured in serum-free medium, with or without IL-1ß (10-11 M) for 24 hours, with or without the COX-1/2 inhibitor indomethacin or the selective COX-2 inhibitors NS-398, celecoxib, or rofecoxib. PGE2 and IL-6 in culture supernatants were determined by specific enzyme-linked immunosorbent assay (ELISA)s. RESULTS: All of the COX inhibitors caused dose-dependent decreases in IL-1ß-stimulated PGE2 , to a maximum of >90% in all cell lines (P ≤0.0001). The selective COX-2 inhibitors, but not indomethacin, caused partial (generally up to approximately 60%), dose-dependent decreases in IL-1ß-stimulated IL-6 in all cell lines (P ≤0.003). When exogenous PGE2 was added concurrently with COX-2 inhibitors before addition of IL-1ß, IL-6 production returned to levels at or approaching that produced by cells exposed only to IL-1ß (P ≤0.04). CONCLUSION: The results suggest that COX-2 inhibition may be useful in helping to control fibroblast production of IL-6 in patients with severe periodontitis. J Periodontol 2003;74:1754-1763.