Prostaglandin production by human gingival fibroblasts inhibited by triclosan in the presence of cetylpyridinium chloride

Background: the effect of triclosan plus the cationic detergent cetylpyridinium chloride (CPC) was evaluated for prostaglandin inhibition in human gingival fibroblasts. Since triclosan has previously been shown to inhibit proinflammatory cytokine induced prostaglandin E-2 (PGE(2)) production, we wanted to determine if triclosan, in the presence of CPC, could enhance these effects.Methods: Initial studies determined that both triclosan and CPC were cytotoxic to human gingival fibroblasts in concentrations exceeding 1.0 mu g/ml for either agent longer than 24 hours in a tissue culture. Therefore, subsequent studies measuring prostaglandin biosynthesis and cyclooxygenase (COX)-1 and COX-2 mRNA expression were performed in concentrations and times that did not significantly affect cell viability.Results: PGE2 biosynthesis was dose dependently inhibited by both triclosan and triclosan and CPC when challenged by tumor necrosis factor (TNF)-alpha or interleukin (IL)-1 beta. At pharmacologically relevant concentrations, triclosan and CPC inhibited ILAP-induced PGE(2) production to a greater extent than triclosan alone (P = 0.02). Moreover, enhanced COX-2 mRNA repression was observed with triclosan and CPC in comparison to triclosan alone in IL-1 beta and TNF-alpha stimulated cells. No effect on COX-I gene expression was observed. Further analysis of cell signaling mechanisms of triclosan and CPC indicates that nuclear factor-kappa B (NF-kappa B) and not p38 mitogen-activated protein kinase (MAPK) signaling may be impaired in the presence of triclosan and CPC.Conclusion: This study indicates that triclosan and CPC are more effective at inhibiting PGE(2) at the level of COX-2 gene regulation, and this combination may offer a potentially better anti -inflammatory agent in the treatment of inflammatory lesions in the oral cavity.

Titanium surfaces with nanotopography modulate cytokine production in cultured human gingival fibroblasts

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Toll-like receptor 2 knockdown modulates interleukin (IL)-6 and IL-8 but not stromal derived factor-1 (SDF-1/CXCL12) in human periodontal ligament and gingival fibroblasts

Background: Fibroblasts are now seen as active components of the immune response because these cells express Toll-like receptors (TLRs), recognize pathogen-associated molecular patterns, and mediate the production of cytokines and chemokines during inflammation. The innate host response to lipopolysaccharide (LPS) from Porphyromonas gingivalis is unusual inasmuch as different studies have reported that it can be an agonist for Toll-like receptor 2 (TLR2) and an antagonist or agonist for Toll-like receptor 4 (TLR4). This study investigates and compares whether signaling through TLR2 or TLR4 could affect the secretion of interleukin (IL)-6, IL-8, and stromal derived factor-1 (SDF-1/CXCL12) in both human gingival fibroblasts (HGF) and human periodontal ligament fibroblasts (HPDLF). Methods: After small interfering RNA-mediated silencing of TLR2 and TLR4, HGF and HPDLF from the same donors were stimulated with P. gingivalis LPS or with two synthetic ligands of TLR2, Pam2CSK4 and Pam3CSK4, for 6 hours. IL-6, IL-8, and CXCL12mRNA expression and protein secretion were evaluated by quantitative polymerase chain reaction and enzymelinked immunosorbent assay, respectively. Results: TLR2 mRNA expression was upregulated in HGF but not in HPDLF by all the stimuli applied. Knockdown of TLR2 decreased IL-6 and IL-8 in response to P. gingivalis LPS, or Pam2CSK4 and Pam3CSK4, in a similar manner in both fibroblasts subpopulations. Conversely, CXCL12 remained unchanged by TLR2 or TLR4 silencing. Conclusion: These results suggest that signaling through TLR2 by gingival and periodontal ligament fibroblasts can control the secretion of IL-6 and IL-8, which contribute to periodontal pathogenesis, but do not interfere with CXCL12 levels, an important chemokine in the repair process.

MyD88 or TRAM knockdown regulates interleukin (IL)-6, IL-8, and CXCL12 mRNA expression in human gingival and periodontal ligament fibroblasts

Background: In a previous report, it was shown that Toll-like receptor (TLR) 2 knockdown modulates interleukin (IL)-6 and IL-8 but not the chemokine CXCL12, an important mediator with inflammatory and proangiogenic effects, in human gingival fibroblasts (HGF) and human periodontal ligament fibroblasts (HPDLF). This study investigates whether knocking down two important TLR adaptor molecules, such as myeloid differentiation protein 88 (MyD88) and TRIF-related adaptor molecule (TRAM), could affect mRNA expression of IL-6, IL-8, and CXCL12 in HGF and HPDLF. Methods: After small interfering (si) RNA-mediated silencing of MyD88 or TRAM, HGF and HPDLF were stimulated with Porphyromonas gingivalis (Pg) lipopolysaccharide (LPS) or two synthetic ligands of TLR2 (Pam2CSK4 and Pam3CSK4) for 6 hours. IL-6, IL-8, and CXCL12 mRNAs were evaluated by quantitative polymerase chain reaction. Results: Knockdown of MyD88 or TRAM partially impaired the IL-8 mRNA upregulation in both fibroblast subpopulations. Similarly, IL-6 upregulation was partially prevented by siMyD88 or siTRAM in HGF stimulated with Pg LPS, as well as in both fibroblast subtypes challenged with Pam2CSK4. Conversely, constitutive CXCL12 mRNA levels were upregulated by MyD88 or TRAM knockdown in non-stimulated cells. Conclusions: These results suggest that TLR adaptor molecules knockdown, such as MyD88 or TRAM, can decrease IL-6 and IL-8 mRNA and increase CXCL12 mRNA expression in HGF and HPDLF. This can be an important step for better understanding the mechanisms that control the inflammatory cytokine and chemokine expression, which in turn contributes to periodontal pathogenesis.

Effect of LPS treatment on the viability and chemokine synthesis by epithelial cells and gingival fibroblasts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Cytotoxic Effects of Zoledronic Acid on Human Epithelial Cells and Gingival Fibroblasts

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

In vitro wound healing improvement by low-level laser therapy application in cultured gingival fibroblasts

The aim of this study was to determine adequate energy doses using specific parameters of LLLT to produce biostimulatory effects on human gingival fibroblast culture. Cells (3 10 4 cells/cm 2) were seeded on 24-well acrylic plates using plain DMEM supplemented with 10 fetal bovine serum. After 48-hour incubation with 5 CO2 at 37C, cells were irradiated with a InGaAsP diode laser prototype (LASERTable; 780 3 nm; 40mW) with energy doses of 0.5, 1.5, 3, 5, and 7J/cm 2. Cells were irradiated every 24h totalizing 3 applications. Twenty-four hours after the last irradiation, cell metabolism was evaluated by the MTT assay and the two most effective doses (0.5 and 3J/cm 2) were selected to evaluate the cell number (trypan blue assay) and the cell migration capacity (wound healing assay; transwell migration assay). Data were analyzed by the Kruskal-Wallis and Mann-Whitney nonparametric tests with statistical significance of 5. Irradiation of the fibroblasts with 0.5 and 3J/cm 2 resulted in significant increase in cell metabolism compared with the nonrradiated group (P 0.05). Both energy doses promoted significant increase in the cell number as well as in cell migration (P 0.05). These results demonstrate that, under the tested conditions, LLLT promoted biostimulation of fibroblasts in vitro. Copyright © 2012 Fernanda G. Basso et al.

Nicotine-Induced Damage Affects Gingival Fibroblasts in the Gingival Tissue of Rats

Background: Very limited information is available from in vivo studies about whether smoking and/or nicotine affect gingival tissues in the absence of plaque. The purpose of this study is to evaluate the effect of the systemic administration of nicotine in the proliferation and counting of fibroblast-like cells in the gingival tissue of rats.Methods: Thirty adult male Wistar rats were randomly assigned into two groups to receive subcutaneous injections of a saline solution (control group = group C) or nicotine solution (group N; 3 mg/kg) twice a day. The animals were euthanized 37, 44, or 51 days after the first subcutaneous injection. Specimens were routinely processed for serial histologic sections. Five fields of view in the connective tissue adjacent to the gingival epithelium and above the alveolar bone crest of the maxillary first molar were selected for the counting of fibroblast-like cells. Data were statistically analyzed (P<0.05).Results: The intergroup analysis detected a lower number of fibroblast-like cells in group N compared to group C on days 37 (2.65 +/- 1.41 and 6.67 +/- 3.25, respectively), 44 (2.70 +/- 1.84 and 8.57 +/- 2.37, respectively), and 51(2.09 +/- 1.41 and 7.49 +/- 2.60, respectively) (P<0.05). The quantification of fibroblast-like cells showed no significant difference (P >0.05) in the intragroup analysis of control and nicotine throughout experimental periods. In the intergroup analysis, group N had reduced proliferating cell nuclear antigen positive fibroblasts compared to group C in all periods (P<0.05).Conclusion: The daily systemic administration of nicotine negatively affected, in vivo, the number and proliferation of fibroblast-like cells in the gingival tissue of rats. J Periodontol 2011;82:1206-1211.

Interleukin-4 and interleukin-13 inhibit the expression of leukemia inhibitory factor and interleukin-11 in fibroblasts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Stimulation of IL-6 Cytokines in Fibroblasts by Toll-like Receptors 2

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Reaction of the human gingival tissue to different suture materials used in periodontal surgery.

The gingival reaction to 4 different suture materials used in periodontal surgery was studied in 36 patients. The gingiva was sutured prior to surgery and biopsies were taken at 3, 7 and 14 days to observe the tissue reaction. The histological examination showed that silk caused the most intense and longest inflammatory response. Polyester and perlon provoked shorter, less intense tissue reactions than silk, and nylon caused the least inflammatory response, with earlier tissue repair.

Biostimulatory effects of low-level laser therapy on epithelial cells and gingival fibroblasts treated with zoledronic acid

Low-level laser therapy (LLLT) has been considered as an adjuvant treatment for bisphosphonate-related osteonecrosis, presenting positive clinical outcomes. However, there are no data regarding the effect of LLLT on oral tissue cells exposed to bisphosphonates. This study aimed to evaluate the effects of LLLT on epithelial cells and gingival fibroblasts exposed to a nitrogen-containing bisphosphonate - zoledronic acid (ZA). Cells were seeded in wells of 24-well plates, incubated for 48 h and then exposed to ZA at 5 μM for an additional 48 h. LLLT was performed with a diode laser prototype - LaserTABLE (InGaAsP - 780 nm ± 3 nm, 25 mW), at selected energy doses of 0.5, 1.5, 3, 5, and 7 J cm-2 in three irradiation sessions, every 24 h. Cell metabolism, total protein production, gene expression of vascular endothelial growth factor (VEGF) and collagen type I (Col-I), and cell morphology were evaluated 24 h after the last irradiation. Data were statistically analyzed by Kruskal-Wallis and Mann-Whitney tests at 5% significance. Selected LLLT parameters increased the functions of epithelial cells and gingival fibroblasts treated with ZA. Gene expression of VEGF and Col-I was also increased. Specific parameters of LLLT biostimulated fibroblasts and epithelial cells treated with ZA. Analysis of these in vitro data may explain the positive in vivo effects of LLLT applied to osteonecrosis lesions. © 2013 Astro Ltd.