In Vitro Cytotoxicity Analysis of Bioceramic Root Canal Sealers on Human Gingival Fibroblast Cells

ABSTRACT@#This study evaluated the cytotoxicity of four bioceramic root canal sealers (bioceramic sealers): GuttaFlow Bioseal (GB), MTA Fillapex, CeraSeal Bioceramic root canal sealer (CS), and iRoot SP root canal sealer (iRSP). The viability of human gingival fibroblast (HGF) cells was used to evaluate the cytotoxicity of these bioceramic sealers. HGF cells were cultured and exposed to bioceramic sealer extracts for 24 hours, 48 hours and 72 hours at 37°C in an incubator humidified with 5% CO2. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide or MTT assay was conducted to determine cell viability at each incubation period and compared among all bioceramic sealers. The Kruskal-Wallis test revealed statistically significant differences between the positive control group and MTA Fillapex, MTA Fillapex and GB, and between GB and iRSP with p < 0.05. However, no statistical differences were found in cell viability for each material across all the incubation periods. GB was the least cytotoxic bioceramic sealer with cell viability exceeding 90% throughout the 72-hour incubation followed by CS, iRSP, and MTA Fillapex with non-cytotoxicity after 72-hour incubation, mild cytotoxicity after 72-hour incubation, and mild cytotoxicity after 72-hour incubation, respectively. However, iRSP showed moderate cytotoxicity, and MTA Fillapex was severely cytotoxic (< 30% cell viability) after 24-hour incubation.

Cell proliferation assessment of human gingival fibroblasts treated with Clinacanthus nutans using alamarBlue assay

@#Clinacanthus nutans (C. nutans), a well-known ethnopharmacological plant consumed for its medicinal purposes by Southeast Asian communities. C. nutans is said to possess antipyretic, inflammatory, antiedemic as well as analgesic properties and used traditionally in treating various skin ailments, Herpes infection, cancer and diabetes. The young leaves of this C. nutans are consumed in Malaysia for maintaining health. In this study, the proliferative activity of human gingival fibroblast cells (HGF-1, ATCC®CRL-2014™, USA) treated with the ethanol extract obtained from C. nutans leaves at three different concentrations (250, 125 and 62.5 µg/ml) was compared with the untreated cells using alamarBlue assay. The proliferative activity of HGF-1 using alamarBlue assay showed that the cells treated with 62.5 μg/ml of ethanolic extract of C. nutans leaves exhibited increased proliferation compared to the other groups and hence does not exhibit any cytotoxicity on HGF-1.

Biological Interaction Between Human Gingival Fibroblasts and Vascular Endothelial Cells for Angiogenesis: A Co-culture Perspective

Advancement in cell culture protocols, multidisciplinary research approach, and the need of clinical implication to reconstruct damaged or diseased tissues has led to the establishment of three-dimensional (3D) test systems for regeneration and repair. Regenerative therapies, including dental tissue engineering, have been pursued as a new prospect to repair and rebuild the diseased/lost oral tissues. Interactions between the different cell types, growth factors, and extracellular matrix components involved in angiogenesis are vital in the mechanisms of new vessel formation for tissue regeneration. In vitro pre-vascularization is one of the leading scopes in the tissue-engineering field. Vascularization strategies that are associated with co-culture systems have proved that there is communication between different cell types with mutual beneficial effects in vascularization and tissue regeneration in two-dimensional or 3D cultures. Endothelial cells with different cell populations, including osteoblasts, smooth muscle cells, and fibroblasts in a co-culture have shown their ability to advocate pre-vascularization. In this review, a co-culture perspective of human gingival fibroblasts and vascular endothelial cells is discussed with the main focus on vascularization and future perspective of this model in regeneration and repair.

Improvement Effects of Dentifrice Containing Plant Extracts on Periodontal Disease / 日本補完代替医療学会誌

Effects of three plant extracts (Hordeum vulgare L., Apocynum Venetum L., Brasenia schreberi J.F.G mel.) on human gingival fibroblasts were examined. As a result, we observed the promoting effect of the extract of Hordeum vulgare L. and the extract of Apocynum Venetum L. respectively on FGF2 and FGF7 production. Moreover, the mixture of the three plant extracts showed the effect of improving the changes in type I collagen gene expression and matrix metalloproteinase 1 gene expression by LPS addition. Next, a dentifrice containing the three plant extracts was subjected to human efficacy trials. We measured periodontal pocket depth, attachment level, bleeding on probing and saliva TNFα as an indicator of periodontal disease. The results suggest that the dentifrice formulated with the three plant extracts were effective for the improvement of periodontal disease.

Effects of TiO2 NPs coating-titanium for fibroblast adhesion / Efectos del recubrimiento de NPs de TiO2 sobre placas de titanio para la adhesión de fibroblastos

The objective of this study was to determine the effects of coating nanoparticles of titanium dioxide (TiO2 NPs) and irradiation -UV on plates of titanium (Ti) for the adhesion and proliferation of human gingival fibroblasts (HGF). A total of 15 Ti plates were divided into three groups (n = 5); (i) control Ti, (ii) experimental: Ti+TiO2 NPs, (iii) experimental: Ti+TiO2 NPs+UV. The plates were analyzed with atomic force microscopy (AFM) and the roughness (Ra and Rmax) was determined. UV irradiation was performed for 20 min. HGF were subcultured in DMEM+10 % fetal bovine serum (FBS) at 37 °C with 5 % CO2. 2x106 cells/mL were inoculated on the plates and incubated for 1 h and washed with phosphate buffer saline (PBS). In the case of cell proliferation, cells were incubated for further 24 h more. Cell viability was determined with the MTT method, the formazan was dissolved with dimethylsulfoxide (DMSO) and analyzed at 540 nm. Experiments were performed of three independent experiments and data were analyzed by Kruskall-Wallis and multiple comparison of Mann-Whitney test. The surface topography of samples corresponded as follow: Ti (Ra= 0,492 µm y Rms= 0.640 µm), Ti+NPs TiO2, (Ra= 0.55 µm y Rms= 0.714 µm), respectively. The coating with TiO2 NPs significantly (p <0.05) increased the adhesion and proliferation of HGF compared with the group. The modification of Ti plates by coated with TiO2 NPs significantly increased adhesion and proliferation of HGF with the formation of a hydrophilic surface which favors the humectancy. This treatment may be reported here convenient to accelerate osseointegration of dental implants based titanium.

El objetivo fue determinar los efectos del recubrimiento con nanopartículas de dióxido de titanio (TiO2 NPs) e irradiación UV sobre placas de titanio (Ti) para la adhesión y proliferación de fibroblastos gingivales humanos (FGH). Un total de 15 placas de Ti se dividieron en tres grupos (n= 5); (i) control Ti, (ii) experimental Ti+NPs TiO2, (iii) experimental: Ti+NPs TiO2+UV. Las placas fueron analizadas en microscopía de fuerza atómica (MFA) y se determinó la rugosidad (Ra y Rmax). La irradiación con UV se realizó durante 20 min. FGH fueron subcultivados en DMEM+10 % de suero fetal bovino a 37 °C con 5 % de CO2. 2x106 células/mL fueron inoculadas sobre las placas e incubadas durante 1 h, se lavaron con solución salina de buffer fosfato. En el caso de la proliferación celular, las células se incubaron por 24 h más. La viabilidad celular se determinó con el método de MTT, el formazan fue disuelto con dimetilsulfoxido y se analizó a 540 nm. Los experimentos se realizaron a partir de tres experimentos independientes y los datos se analizaron por Kruskall-Wallis y por comparación múltiple de Mann-Whitney. La topografía de la superficie de las muestras correspondio de la siguiente manera: Ti (Ra= 0,492 µm y Rms= 0,640 µm), Ti+NPs TiO2, (Ra= 0,55 µm y Rms= 0,714 µm), respectivamente. El recubrimiento con NPs TiO2 aumentó significativamente la adhesión y proliferación de HGF en comparación con el grupo de Ti control (p <0,05). La modificación de la superficie de las placas de Ti recubiertas con NPs TiO2 aumentó significativamente la adhesión y proliferación de HGF con la formación de una superficie hidrófila que favorece la humectancia. Este tratamiento aquí informado tal vez sea un método conveniente para acelerar el proceso de la osteointegración de los implantes dentales a base de titanio.

Activation of acetylcholine receptor elicits intracellular Ca2+ mobilization, transient cytotoxicity, and induction of RANKL expression

Acetylcholine receptors (AChR) including muscarinic and nicotinic AChR are widely expressed and mediate a variety of physiological cellular responses in neuronal and non-neuronal cells. Notably, a functional cholinergic system exists in oral epithelial cells, and nicotinic AChR (nAChR) mediates cholinergic anti-inflammatory responses. However, the pathophysiological roles of AChR in periodontitis are unclear. Here, we show that activation of AChR elicits increased cytosolic Ca²⁺ ([Ca²⁺]ᵢ), transient cytotoxicity, and induction of receptor activator of nuclear factor kappa-B ligand (RANKL) expression. Intracellular Ca²⁺ mobilization in human gingival fibroblast-1 (hGF-1) cells was measured using the fluorescent Ca²⁺ indicator, fura-2/AM. Cytotoxicity and induction of gene expression were evaluated by measuring the release of glucose-6-phosphate dehydrogenase and RT-PCR. Activation of AChR in hGF-1 cells by carbachol (Cch) induced [Ca²⁺]ᵢ increase in a dose-dependent manner. Treatment with a high concentration of Cch on hGF-1 cells caused transient cytotoxicity. Notably, treatment of hGF-1 cells with Cch resulted in upregulated RANKL expression. The findings may indicate potential roles of AChR in gingival fibroblast cells in bone remodeling.

The study of cytocompatibility of Co-Cr alloy and Ti alloy / 天津医药

Objective To investigate the cytocompatibility of the liching liquids of Co-Cr alloy and Ti alloy on human gingival fibroblasts (HGF). Methods The HGF were treated in vitro with leaching liquids of Co-Cr alloy and Ti alloy, respectively. The DMEM cell medium containing 10% fetal bovine serum was served as a negative control. The viability of HGF treated by two dental alloys were evaluated by means of MTT, and the contents of intracellular reduced glutathione (rGSH) were assayed by kits. The tumor necrosis factor-α(TNF-α) contents were determined in the culture supernatant by ELISA in two groups. The effects of these alloys on the expression of caspase-3 were examined by real time-PCR method. Results Compared with the control group, HGF treated with Co-Cr alloy leaching liquids showed a lower viability ( P ＜0.05), while Ti alloy leaching liquid promoted the proliferation of HGF. In Co-Cr alloy group, the rGSH content was significantly decreased (P＜0.05), while TNF-α content was significantly increased (P＜0.05) compared with control group. There were no significant differences in rGSH and TNF-α contents between the Ti alloy group and control group (P＞0.05). The expression of caspase-3 was significantly higher in Co-Cr alloy group than that of control group (P＜0.05), while there was no significant difference in the expression of caspase-3 between Ti alloy group and control group. Conclusion Results suggest that Co-Cr alloy possesses cytotoxicity, while there is better cell compatibility for Ti alloy.

Antimicrobial activity against periodontopathogenic bacteria, antioxidant and cytotoxic effects of various extracts from endemic Thermopsis turcica

Objective: To investigate the in vitro antimicrobial potential of Thermopsis turcica Kit Tan, Vural&Küçüködük against periodontopathogenic bacteria, its antioxidant activity and cytotoxic effect on various cancer cell lines. Methods: In vitro antimicrobial activities of ethanol, methanol, ethyl acetate (EtAc), n-hexane and water extracts of Thermopsis turcica herb against periodontopathogenic bacteria, Aggregatibacter actinomycetemcomitans ATCC 29523 and Porphyromonas gingivalis ATCC 33277 were tested by agar well diffusion, minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Antioxidant properties of the extracts were evaluated by 1,1-diphenyl-2-picryl-hydrazyl radical scavenging activity and β-carotene bleaching methods. Amounts of phenolic contents of the extracts were also analysed by using the Folin-Ciocalteu reagent. Additionally, cytotoxic activity of the extracts on androgen-insensitive prostate cancer, androgen-sensitive prostate cancer, chronic myelogenous leukemia and acute promyelocytic leukemia human cancer cell lines were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Human gingival fibroblast cells were used as a control. Results: Our data showed that EtAc extract had the highest antimicrobial effect on Aggregatibacter actinomycetemcomitans (MIC: 1.562 mg/mL, MBC: 3.124 mg/mL) and Porphyromonas gingivalis (MIC: 0.781 mg/mL, MBC: 1.562 mg/mL). In antioxidant assays, EtAc extract exhibited also the highest radical scavenging activity [IC

Antimicrobial activity against periodontopathogenic bacteria, antioxidant and cytotoxic effects of various extracts from endemic Thermopsis turcica

Objective: To investigate the in vitro antimicrobial potential of Thermopsis turcica Kit Tan, Vural &Kü?ük?dük against periodontopathogenic bacteria, its antioxidant activity and cytotoxic effect on various cancer cell lines.Methods: In vitro antimicrobial activities of ethanol, methanol, ethyl acetate (EtAc), n-hexane and water extracts of Thermopsis turcica herb against periodontopathogenic bacteria, Aggregatibacter actinomycetemcomitans ATCC 29523 and Porphyromonas gingivalis ATCC 33277 were tested by agar well diffusion, minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Antioxidant properties of the extracts were evaluated by 1,1-diphenyl-2-picryl-hydrazyl radical scavenging activity and β-carotene bleaching methods. Amounts of phenolic contents of the extracts were also analysed by using the Folin-Ciocalteu reagent. Additionally, cytotoxic activity of the extracts on androgen-insensitive prostate cancer, androgen-sensitive prostate cancer, chronic myelogenous leukemia and acute promyelocytic leukemia human cancer cell lines were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Human gingival fibroblast cells were used as a control.Results:Our data showed that EtAc extract had the highest antimicrobial effect on Aggregatibacter actinomycetemcomitans (MIC: 1.562 mg/mL, MBC: 3.124 mg/mL) and Porphyromonas gingivalis (MIC: 0.781 mg/mL, MBC: 1.562 mg/mL). In antioxidant assays, EtAc extract exhibited also the highest radical scavenging activity [IC50=(30.0±0.3) μg/mL] and the highest inhibition [(74.35±0.30)%] against lineloic acide oxidation. The amount of phenolic content of it was also the highest [(162.5±1.2) μg/mg gallic acid]. In cytotoxic assay, only ethanol [IC50=(80.00±1.21) μg/mL] and EtAc extract [IC50=(70.0±0.9) μg/mL] were toxic on acute promyelocytic leukemia cells at 20-100 μg/mL (P<0.05). However, no toxic effect was observed on human gingival fibroblast cells.Conclusions:According to our findings, owing to its antioxidant and cytotoxic potential, EtAc extract might include anticancer agents for acute promyelocytic leukemia.

Constituents Released from Streptococcus mutans Attenuate Arecoline-mediated Cytotoxicity in HGF Cells by Altering Intracellular Ca2+ Signaling

Streptococcus mutans (S. mutans) is a facultative anaerobic bacterium mainly found in the oral cavity and is known to contribute to tooth decay and gingivitis. Recent studies on intestinal microbiota have revealed that microorganisms forming a biofilm play important roles in maintaining tissue homeostasis through their own metabolism. However, the physiological roles of oral microorganisms such as S. mutans are still unclear. In our current study, we identified that constituents released from S. mutans (CR) reduce arecoline-mediated cytotoxicity without producing toxic effects themselves. Arecoline, as a major alkaloid of areca nut, is known to mediate cytotoxicity on oral epithelial cells and induces a sustained intracellular Ca2+ ([Ca2+]i) increase that is cytotoxic. The exposure of human gingival fibroblast (HGF) cells to CR not only inhibited the sustained [Ca2+]i increase but also the initial [Ca2+]i elevation. In contrast, CR had no effects on the gene regulation mediated by arecoline. These results demonstrate that S. mutans has physiological role in reducing cytotoxicity in HGF cells and may be considered a novel pharmaceutical candidate.

Effect of bamboo salt on human gingival fibroblasts / 대한구강보건학회지

OBJECTIVES: To evaluate the anti-inflammatory and cytotoxic activities of bamboo salt. METHODS: Cytotoxicity of bamboo salt and bay salt (0.01%, 0.1%, and 1%) was evaluated using MTT assay. In addition, secretion of the pro-inflammatory cytokines interleukin (IL)-1beta and IL-6 from human gingival fibroblasts (HGFs) was measured after application of 0.01% and 0.1% concentrations by using real-time polymerase chain reaction. RESULTS: Bamboo salt and bay salt at 1% concentration were cytotoxic to HGFs at 24 h; however, no such effect was observed at 0.01% or 0.1%. Bamboo salt showed a relatively low inhibitory effect. IL-1beta secretion was inhibited by a 0.1% solution of bamboo salt. IL-6 secretion was inhibited by both bamboo salt and bay salt at 0.1% concentration. CONCLUSIONS: The above results suggest that bamboo salt inhibits the release of IL-1beta and IL-6 from HGFs. Thus, bamboo salt may be a useful material for gingival inflammation.

Cellular viability and genetic expression of human gingival fibroblasts to zirconia with enamel matrix derivative (Emdogain(R))

PURPOSE: The objective of this study was to investigate the biologic effects of enamel matrix derivative (EMD) with different concentrations on cell viability and the genetic expression of human gingival fibroblasts (HGF) to zirconia surfaces. MATERIALS AND METHODS: Immortalized human gingival fibroblasts (HGF) were cultured (1) without EMD, (2) with EMD 25 microg/mL, and (3) with EMD 100 microg/mL on zirconia discs. MTT assay was performed to evaluate the cell proliferation activity and SEM was carried out to examine the cellular morphology and attachment. The mRNA expression of collagen type I, osteopontin, fibronectin, and TGF-beta1 was evaluated with the real-time polymerase chain reaction (RT-PCR). RESULTS: From MTT assay, HGF showed more proliferation in EMD 25 microg/mL group than control and EMD 100 microg/mL group (P<.05). HGFs showed more flattened cellular morphology on the experimental groups than on the control group after 4h culture and more cellular attachments were observed on EMD 25 microg/mL group and EMD 100 microg/mL group after 24h culture. After 48h of culture, cellular attachment was similar in all groups. The mRNA expression of type I collagen increased in a concentration dependent manner. The genetic expression of osteopontin, fibronectin, and TGF-beta1 was increased at EMD 100 microg/mL. However, the mRNA expression of proteins associated with cellular attachment was decreased at EMD 25 microg/mL. CONCLUSION: Through this short term culture of HGF on zirconium discs, we conclude that EMD affects the proliferation, attachment, and cell morphology of HGF cells. Also, EMD stimulates production of extracellular matrix collagen, osteopontin, and TGF-beta1 in high concentration levels. CLINICAL RELEVANCE: With the use of EMD, protective barrier between attached gingiva and transmucosal zirconia abutment may be enhanced leading to final esthetic results with implants.

Antimicrobial activity against periodontopathogenic bacteria, antioxidant and cytotoxic effects of various extracts from endemic Thermopsis turcica

<p><b>OBJECTIVE</b>To investigate the in vitro antimicrobial potential of Thermopsis turcica Kit Tan, Vural & Küçüködük against periodontopathogenic bacteria, its antioxidant activity and cytotoxic effect on various cancer cell lines.</p><p><b>METHODS</b>In vitro antimicrobial activities of ethanol, methanol, ethyl acetate (EtAc), n-hexane and water extracts of Thermopsis turcica herb against periodontopathogenic bacteria, Aggregatibacter actinomycetemcomitans ATCC 29523 and Porphyromonas gingivalis ATCC 33277 were tested by agar well diffusion, minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Antioxidant properties of the extracts were evaluated by 1,1-diphenyl-2-picryl-hydrazyl radical scavenging activity and β-carotene bleaching methods. Amounts of phenolic contents of the extracts were also analysed by using the Folin-Ciocalteu reagent. Additionally, cytotoxic activity of the extracts on androgen-insensitive prostate cancer, androgen-sensitive prostate cancer, chronic myelogenous leukemia and acute promyelocytic leukemia human cancer cell lines were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Human gingival fibroblast cells were used as a control.</p><p><b>RESULTS</b>Our data showed that EtAc extract had the highest antimicrobial effect on Aggregatibacter actinomycetemcomitans (MIC: 1.562 mg/mL, MBC: 3.124 mg/mL) and Porphyromonas gingivalis (MIC: 0.781 mg/mL, MBC: 1.562 mg/mL). In antioxidant assays, EtAc extract exhibited also the highest radical scavenging activity [IC50=(30.0±0.3) µg/mL] and the highest inhibition [(74.35±0.30)%] against lineloic acide oxidation. The amount of phenolic content of it was also the highest [(162.5±1.2) µg/mg gallic acid]. In cytotoxic assay, only ethanol [IC50=(80.00±1.21) µg/mL] and EtAc extract [IC50=(70.0±0.9) µg/mL] were toxic on acute promyelocytic leukemia cells at 20-100 µg/mL (P<0.05). However, no toxic effect was observed on human gingival fibroblast cells.</p><p><b>CONCLUSIONS</b>According to our findings, owing to its antioxidant and cytotoxic potential, EtAc extract might include anticancer agents for acute promyelocytic leukemia.</p>

Focal adhesion linker proteins expression of fibroblast related to adhesion in response to different transmucosal abutment surfaces

PURPOSE: To evaluate adherence of human gingival fibroblasts (HGFs) to transmucosal abutment of dental implant with different surface conditions with time and to investigate the roles of focal adhesion linker proteins (FALPs) involved in HGFs adhesion to abutment surfaces. MATERIALS AND METHODS: Morphologies of cultured HGFs on titanium and ceramic discs with different surface were observed by scanning electron microscopy. Biocompatibility and focal adhesion were evaluated by ultrasonic wave application and cell viability assay. FALPs expression levels were assessed by RT-PCR and western blot. RESULTS: There seemed to be little difference in biocompatibility and adhesion strength of HGFs depending on the surface conditions and materials. In all experimental groups, the number of cells remaining on the disc surface after ultrasonic wave application increased more than 2 times at 3 days after seeding compared to 1-day cultured cells and this continued until 7 days of culture. FALPs expression levels, especially of vinculin and paxillin, also increased in 5-day cultured cells compared to 1-day cultured fibroblasts on the disc surface. CONCLUSION: These results might suggest that the strength of adhesion of fibroblasts to transmucosal abutment surfaces increases with time and it seemed to be related to expressions of FALPs.

Effects of nifedipine on proliferation of gingival fibroblasts in drug-induced gingival hyperplasia / 上海交通大学学报（医学版）

Objective To investigate the changes in proliferation index (PrI) of gingival fibroblasts in nifedipine-induced gingival hyperplasia ( NIFr-HGF). Methods Gingival fibroblasts were derived from a patient with nifedipine-induced gingival hyperplasia. Cells were induced by 10 ng/mL and 1 000 ng/mL nifedipine ( low- and high-concentration drug intervention groups), respectively. Cells were harvested 18 h and 30 h after intervention, cell cycles were detected by flow cytometry, and Prls were calculated. NIFr-HGF without nifedipine induction were served as blank control. Results After induction for the same time, Prls of NIFr-HGF cell cycle of low- and high-concentration drug intervention groups were significantly higher than those of blank control group (P <0.05) , while there was no significant difference between low and high-concentration drug intervention groups (P > 0.05). In low and high-concentration drug intervention groups, Prls of NIFr-HGF cell cycle after intervention for 30 h were significantly higher than those after intervention for 18 h [(57. 54 ± 0.019)% vs (21.15 ±0.011)%, and (59.36 ±0.031)% vs (19.01 ±0.012) %, respectively] (P < 0.05). Conclusion For patients with nifedipine-induced gingival hyperplasia, PrI of NIFr-HGF cell cycle increases with time of nifedipine intervention, while is not significantly related to drug concentration.

The effect of five osteotropic factors on osteoprotegerin mRNA expression in gingival fibroblasts / 대한치주과학회지

PURPOSE: Osteoprotegerin (OPG) is a secreted glycoprotein and a member of the tumor necrosis factor (TNF) receptor family that inhibits bone resorption by suppressing osteoclastogenesis. Gingival fibroblasts (GF) play a role in periodontal disease progression, and the purpose of this experiment was to evaluate influence of osteotropic factors on the expression of osteoprotegerin mRNA in these cells. MATERIALS AND METHODS: In this experiment, the influence of osteoclastogenic factors, interleukin-1 beta (IL-1beta), TNF-alpha, prostanglandin E2 (PGE2). parathyroid hormone (PTH) and 1alpha, 25-dihydroxyvitamin D3 on the expression of osteoprotegerin mRNA in GF was studied by Northern blot hybridization. RESULTS: As expected, PGE2 tended to inhibit OPG levels and this was most prominent at 24 hours of culture with 10(-7)M of PGE2. TNF-alpha at 10ng/ml and also at 25ng/ml decreased OPG levels to almost 30% of the control at 24 hours. This contrasts with reports of increased OPG levels from osteoblast/stromal cells and gingival fibroblasts stimulated by TNF-alpha. Decrease of OPG levels with PGE2 and TNF-alphasuggests a pathway whereby these mediators exert their resorptive effects. However, OPG levels were increased almost 3-fold at 24 hours with IL-1beta(1 to 15ng/ml) and increased 1.4 fold with 24-hour treatment of 10(-7)M PTH. CONCLUSION: Increase of OPG levels suggests that these 'osteoclastogenic' factors act in more complex ways and may act to inhibit bone resorption in inflammatory periodontitis. This result supports the role of OPG as a negative feedback mechanism in osteoclastic activity.

Jatropha curcas latex inhibits the release of collagenase by gingival fibroblast.

Folkloric use of J. curcas latex among others are to cure tooth pain, bleeding gum and as anti-inflammatory drug. Collagenase is a neutral protease released by activated macrophage and also by fibroblasts in small amounts. The aim of this study was to evaluate the effect of J. curcas latex on collagenase released by fibroblasts. Four doses of J. curcas latex from 37.5-300 g/ml were added to 3 human gingival primary fibroblast cell culture. After 1 to 4 days of incubation, collagenase in the supernatant was assayed with collagen. The degradation products were then separated by SDS-PAGE and the density of ¾ A bands were measured semi quantitatively by Adobe Photo computer program. Result showed that J. curcas latex decreased collagenase released by human gingival fibroblast, and increasing dose inhibits more. It may be concluded that the latex of J. curcas inhibits the release of collagenase by human gingival fibroblast.

Evaluation on cytotoxicity of propolis and calcium hydroxide / 重庆医科大学学报

Objective:To evaluate the cytotoxicity of propolis and calcium hydroxide for human gingival fibroblast.Methods:Human gingival fibroblast(HGF)was incubated in culture solution containing propolis or calcium hydroxide at different concentrations.The relative growth rates(RGR)were examined by method of MTT assay and cytotoxicity was classified by the five-class method.Then the results were statistically analysed.Results:The cytotoxicities of both 0.1% to 0.6% propolis and calcium hydroxide solutions at different concentration were all graded as first or second class.There was no statistical difference between them.Conclusion:Propolis and calcium hydroxide have weak cytotoxicity for HGF and they are all safe as local application for oral diseases.

Two colorimetric assays verify that calcium sulfate promotes proliferating activity of human gingival fibroblasts / 대한치과보철학회지

STATEMENT OF PROBLEM: The role of calcium sulfate in stimulating the growth of gingival soft tissue has been reported in few studies. Such a unique property of calcium sulfate could serve as a trouble-solving broker in compensating for the lack of soft tissues in various oral surgeries. PURPOSE: The purpose of this study was to compare the proliferating activities of human gingival fibroblasts seeded on various bone graft barrier materials of calcium sulfate, collagen, and polytetrafluorethylene (PTFE). MATERIAL AND METHODS: Two calcium sulfates (CAPSET(R) and CalForma(R), Lifecore Biomedical Inc., St. Paul, Minnesota, USA), a resorbable natural collagen (Bio-Gide(R), Geistlich Pharma Ag., Wolhusen, Switzerland), and a non-resorbable PTFE (TefGen-FD(R), Lifecore Biomedical Inc., St. Paul, Minnesota, USA) served as the human gingival fibroblasts'substrates and comprised the four experimental groups, whereas the untreated floors of culture plastics were used in the control group, in this study. Cells were trypsinized, seeded, and incubated for 48 h. The proliferating activities of fibroblasts were determined by XTT and SRB assay and absorbance (optical density, OD) was measured. One-way ANOVA was used to analyze the differences in the mean OD values between the groups of CAPSET, CalForma, Bio-Gide, TefGen, and the control (p<0.05). RESULTS: From the XTT assay, the mean OD value of the control group, the highest, was significantly greater than that of any of the four experimental groups followed by CalForma, CAPSET, TefGen, and Bio-Gide. Further, the mean OD value of CalForma, was significantly greater compared to that of Bio-Gide. From the SRB assay, Calforma showed the highest mean OD value, which was significantly greater than that of any other groups, followed by the control, CAPSET, Bio-Gide, and TefGen. The mean OD values of both the control and CAPSET were significantly greater compared to that of TefGen (p<0.05). CONCLUSION: Assessment of the viability and proliferation of cultured fibroblasts seeded and incubated for 48 h on various barrier-material substrates using XTT and SRB assay showed that calcium sulfate CalForma(R) promotes the proliferating activity of human gingival fibroblasts.

The study on the effect of nanoemulsion for the prevention and treatment of gingival inflammation

We have found out the relationship of nanoemulsion containing nano vitamin C, E and propolis and gingival disease. We've confirmed effect of nanoemulsion through the experiment of in vivo and in vitro. We tested cell viability of gingival fibroblast cells by MTT assay and mRNA appearance of interleukin-1 beta, using mouse that was guided inflammation. Anti-microbacterial activity for Antibacterial effect's experiment was carried out by using S.aureus and E.coli. In addition, inflammation tissue has been observed with scanning electrical microscopy. In this study, expression of interleukin-1 beta was decreased after adding nanoemulsion containing nanovitamin C, E and propolis. We've also obtained good results from the test of Antibacterial effect against S.aureus and E.coli. Also, swelling of inflammation tissues observed by scanning electrical microscopy has gone down. In conclusion, we have gained confidence that nanoemulsion containing nano vitamin C, E and propolis has very high Antibacterial effect against bacteria in oral. And it made us guess that inflammation of gingival reduces after decreasing interleukin-1 beta. Thus, we expect that nanoemulsion containing nano vitamin C, E and propolis gives good effects to patient having gingival disease.