Viability Of Human Gingival Fibroblasts Research Articles

Influence of titanium nanoparticles on cytotoxicity and inflammatory cytokines expression in gingival fibroblasts - An in vitro study

Background Despite the success of titanium (Ti) implants in dental rehabilitation, emerging evidence implicates the release of titanium oxide nanoparticles (TiO2NPs) from the implant surface as a potential contributor to the initiation of the pathogenic process leading to peri-implantitis and ultimately failure of the implants. However, a comprehensive investigation to elucidate the dose-dependent effects of TiO2NPs on cytotoxicity and inflammatory responses is lacking. The present study was undertaken to evaluate the effect of different concentrations of TiO2NPs on the cytotoxicity and inflammatory cytokine expression in Human Gingival Fibroblasts (HGFs). Methods HGFs were isolated from gingival tissue samples obtained from periodontally and systemically healthy subjects. Ti standard solution for ICP was diluted to create concentrations of (0.001 ppm, 0.01 ppm, 0.1 ppm, 1 ppm, 10 ppm, and 100 ppm) for cell culture media containing titanium. HGFs were then cultured in these varying concentrations for specific time periods (days 1, 3, 5, and 7) to assess cell viability. A cytotoxicity assay was performed to determine the levels and expression of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α) and anti-inflammatory cytokines (IL-10, IL-12, TGF-β) using qRT-PCR and ELISA techniques. Results Our findings demonstrate a concentration and duration-dependent decrease in HGF viability upon exposure to titanium nanoparticles. Notably, a 50% reduction in cell viability was observed at the highest concentration (100 ppm). qRT-PCR analysis revealed a significant upregulation of pro-inflammatory cytokines, particularly IL-1β and IL-6, in HGFs exposed to titanium. Interestingly, the expression of anti-inflammatory cytokines (IL-10, IL-12, and TGF-β) remained comparable or even equivalent compared to controls across different titanium concentrations. Conclusions The study revealed a concentration and duration-dependent influence on HGF viability and cytokine profile, suggesting potential cytotoxicity and modulation of the inflammatory response mediated by TiO2NPs. Further research is necessary to elucidate the underlying mechanisms and their implications for dental implant biocompatibility.

CAPE derivatives: Multifaceted agents for chronic wound healing.

Chronic wounds significantly impact the patients' quality of life, creating an urgent interdisciplinary clinical challenge. The development of novel agents capable of accelerating the healing process is essential. Caffeic acid phenethyl ester (CAPE) has demonstrated positive effects on skin regeneration. However, its susceptibility to degradation limits its pharmaceutical application. Chemical modification of the structure improves the pharmacokinetics of this bioactive phenol. Hence, two novel series of CAPE hybrids were designed, synthesized, and investigated as potential skin regenerative agents. To enhance the stability and therapeutic efficacy, a caffeic acid frame was combined with quinolines or isoquinolines by an ester (1a-f) or an amide linkage (2a-f). The effects on cell viability of human gingival fibroblasts (HGFs) and HaCaT cells were evaluated at different concentrations; they are not cytotoxic, and some proved to stimulate cell proliferation. The most promising compounds underwent a wound-healing assay in HGFs and HaCaT at the lowest concentrations. Antimicrobial antioxidant properties were also explored. The chemical and thermal stabilities of the best compounds were assessed. In silico predictions were employed to anticipate skin penetration capabilities. Our findings highlight the therapeutic potential of caffeic acid phenethyl ester (CAPE) derivatives 1a and 1d as skin regenerative agents, being able to stimulate cell proliferation, control bacterial growth, regulate ROS levels, and being thermally and chemically stable. An interesting structure-activity relationship was discussed to suggest a promising multitargeted approach for enhanced wound healing.

Silver nanoparticles incorporated dental restorative resin and its antibiofilm effect.

Dental restoration materials are susceptible to bacterial biofilm formation, which damages the restorations and causes oral health problems. Therefore, to overcome this, silver nanoparticles (AgNPs) are studied widely due to their antimicrobial, anti-inflammatory and healing properties. The purpose of this study was to develop a strategy for incorporating AgNPs onto the surface of bisacrylic resin (Bis) to evaluate its antibiofilm effects using Streptococcus sanguinis and Actinomyces naeslundii. AgNPs with an average size of 25 nm at two different concentrations were dispersed on the Bis surface (Bis-AgNPs) by mechanical deposition. Ag release was quantified until 7 days of incubation. Bacterial growth was assessed using a viability assay kit and observed using confocal microscopy. The biofilm biomass was quantified using arbitrary fluorescence units. Cell viability was evaluated using an MTT assay. The results showed that Bis-AgNPs significantly inhibited biofilm formation along with a significant difference in the viability of human gingival fibroblasts. The quantification confirmed a decrease in Ag release over time, and elemental mapping showed AgNP penetration up to 10 µm from the surface. Therefore, it was concluded that Bis-AgNPs presented enhanced antibiofilm properties, even at a concentration with no adverse effects. Therefore, this nanocomposite may be a promising alternative for biofilm control in temporary restorative materials.

In Vitro Effect of Photodynamic Therapy With Curcumin in Combination With Photobiomodulation Therapy by 660 nm on the Viability of Human Gingival Fibroblasts.

Introduction: This study aimed to assess the effect of repeated irradiations of 660 nm photobiomodulation therapy (PBMT) after photodynamic therapy (PDT) with curcumin on the viability of human gingival fibroblasts (HGFs). Methods: In this in vitro, experimental study, HGFs were cultured and assigned to five groups: One control group with no intervention and four experimental groups of PDT with curcumin, PBMT (660 nm laser irradiation) immediately after PDT, PBMT immediately and 24 hours after PDT and PBMT immediately and 24 hours and 48 hours after PDT. Cell viability was assessed after 1, 4, and 7 days using the methyl thiazolyl tetrazolium (MTT) assay. Data were analyzed by one-way ANOVA. Results: On day 1, the control group had no significant difference with one-time (P=1.00), two-time (P=1.00), and three-time (P=0.88) laser irradiation groups. On day 4, the difference between the control and one-time (P<0.001), two-time (P<0.001) and three-time (P=0.02) laser irradiation groups was statistically significant, suggesting more cell viability in test groups. On day 7, the three-time laser irradiation group showed significant cell viability compared to the other two test groups but not with the control group (P=0.98). Conclusion: PBMT with 660 nm laser irradiation after PDT with curcumin would increase the viability of HGFs by increasing the frequency of irradiation.

Development of Biologically Active Phytosynthesized Silver Nanoparticles Using Marrubium vulgare L. Extracts: Applications and Cytotoxicity Studies.

Metal nanoparticle phytosynthesis has become, in recent decades, one of the most promising alternatives for the development of nanomaterials using "green chemistry" methods. The present work describes, for the first time in the literature, the phytosynthesis of silver nanoparticles (AgNPs) using extracts obtained by two methods using the aerial parts of Marrubium vulgare L. The extracts (obtained by classical temperature extraction and microwave-assisted extraction) were characterized in terms of total phenolics content and by HPLC analysis, while the phytosynthesis process was confirmed using X-ray diffraction and transmission electron microscopy, the results suggesting that the classical method led to the obtaining of smaller-dimension AgNPs (average diameter under 15 nm by TEM). In terms of biological properties, the study confirmed that AgNPs as well as the M. vulgare crude extracts reduced the viability of human gingival fibroblasts in a concentration- and time-dependent manner, with microwave-assisted extracts having the more pronounced effects. Additionally, the study unveiled that AgNPs transiently increased nitric oxide levels which then decreased over time, thus offering valuable insights into their potential therapeutic use and safety profile.

Limosilactobacillus reuteri supernatant attenuates inflammatory responses of human gingival fibroblasts to LPS but not to elevated glucose levels.

We investigated the invitro effect of Limosilactobacillus reuteri DSM 17938 supernatant on the inflammatory response of human gingival fibroblasts (HGF) challenged by lipopolysaccharide (LPS) or elevated glucose levels. HGF were exposed to LPS (1 μg/mL), glucose (5, 12 mM or 25 mM), and dilutions of supernatant prepared from L. reuteri DSM 17938 (0.5 × 107, 1.0 × 107, 2.5 × 107, and 5.0 × 107 CFU/mL). After 24 h cell viability and levels of cytokines (IL-1β, IL-6 and IL-8) and TLR-2 were determined. None of the tested L. reuteri (DSM 17938) supernatant concentrations reduced the viability of HGF. Supernatant concentrations (2.5 × 107 and 5 × 107 CFU/mL) significantly (p < .05) decreased the production of IL-1β, IL-6, IL-8, and TLR-2 in the presence of LPS. In contrast, inflammatory markers were not reduced by L. reuteri supernatant in the presence of glucose. Glucose concentrations of 12 mM and 24 mM still lead to an elevated production of the investigated biochemical mediators. While L. reuteri (DSM 17938) supernatant attenuates the inflammatory response of HGF to LPS in a dose-dependent manner, elevated glucose levels suppress this action. These invitro results support the overall anti-inflammatory efficacy of L. reuteri supplementation in plaque-associated periodontal inflammations.

Efficacy of a mouthwash containing ε-poly-L-lysine, funme peptides and domiphen in reducing halitosis and supragingival plaque: a randomized clinical trial

ObjectiveTo evaluate the antibacterial effectiveness of a combination of ε-poly-L-lysine (ε-PL), funme peptide (FP) as well as domiphen against oral pathogens, and assess the efficacy of a BOP® mouthwash supplemented with this combination in reducing halitosis and supragingival plaque in a clinical trial.Materials and methodsThe minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the compound against Fusobacterium nucleatum, Porphyromonas gingivalis, Streptococcus mutans, and Aggregatibacter actinomycetemcomitans were determined by the gradient dilution method. Subsequently, the CCK-8 assay was used to detect the toxicity of mouthwash on human gingival fibroblastst, and the effectiveness in reducing halitosis and supragingival plaque of the mouthwash supplemented with the combination was analyzed by a randomized, double-blind, parallel-controlled clinical trial.ResultsThe combination exhibited significant inhibitory effects on tested oral pathogens with the MIC < 1.56% (v/v) and the MBC < 3.13% (v/v), and the mouthwash containing this combination did not inhibit the viability of human gingival fibroblasts at the test concentrations. The clinical trial showed that the test group displayed notably lower volatile sulfur compounds (VSCs) at 0, 10, 24 h, and 7 d post-mouthwash (P < 0.05), compared with the baseline. After 7 days, the VSC levels of the and control groups were reduced by 50.27% and 32.12%, respectively, and notably cutting severe halitosis by 57.03% in the test group. Additionally, the Plaque Index (PLI) of the test and control group decreased by 54.55% and 8.38%, respectively, and there was a significant difference in PLI between the two groups after 7 days (P < 0.01).ConclusionsThe combination of ε-PL, FP and domiphen demonstrated potent inhibitory and bactericidal effects against the tested oral pathogens, and the newly formulated mouthwash added with the combination exhibited anti-dental plaque and anti-halitosis properties in a clinical trial and was safe.Trial registrationThe randomized controlled clinical trial was registered on Chinese Clinical Trial Registry (No. ChiCTR2300073816, Date: 21/07/2023).

Evaluation of Adhesion and Viability of Human Gingival Fibroblasts on Strontium-Coated Titanium Surfaces: an in vitro Study.

Applying multifunctional coatings employing strontium (Sr) ions on titanium (Ti) surfaces is a useful and biocompatible method to improve osseointegration and prevent tissue infections through antimicrobial activity. Nonetheless, the effectiveness of Sr coating on the adhesion and viability of human gingival fibroblasts (HGFs) to Ti surfaces remains unclear. The study aimed to evaluate the effect of Sr coating on the adhesion and viability of HGFs to Ti surfaces. The Ti wafers were divided into two groups based on Sr coating: uncoated Ti (control) and Sr-coated Ti. The Magnetron sputtering technique was used for Sr coating on Ti surfaces. The HGFs were seeded onto the surfaces and cultured for 48 and 96 hours before the cell adhesion and viability of the attached HGFs were assessed. The adhesion of HGFs was analyzed using the attached cell numbers at 48 h and 96 h, and the morphology at 24 h and 72 h. The cytotoxic effect on HGFs was assessed after 24 and 72 hours of incubation using cell viability assay. Student's t-test was used for statistical analysis. The number of cells attached to Sr-coated surfaces was significantly greater than those attached to uncoated Ti surfaces after 48 hours (P<0.0001) and 96 hours (P=0.0002). Sr-coated and uncoated Ti surfaces were not cytotoxic to HGFs, with the cell viability ranging from 92% to 105% of the untreated control HGFs. There were no significant differences in cell viability between Sr-coated and uncoated Ti surfaces at 24 hours (P=0.3675) and 72 hours (P=0.0982). Sr-coated Ti surfaces induce adhesion of HGFs compared to uncoated Ti surfaces. Further, Sr-coated and uncoated Ti surfaces show no cytotoxic effect on the attached HGFs.

Nicotine reduces cell viability and induces oxidative stress in human gingival fibroblasts

BackgroundNicotine, as the main component of cigarettes, is known to interfere with the proliferation of human gingival fibroblasts (HGFs) and can trigger oxidative stress. This study aimed to analyze the impact of nicotine on viability, expression of the antioxidant Nrf2, levels of the product of oxidative stress malondialdehyde (MDA), and the migration capacity of HGFs. MethodsAn experimental laboratory study used fibroblasts isolated from healthy human gingiva. The cells were grouped into the non-treatment control group (NTC), the solvent control (SC), and the treatment groups, exposed to nicotine at various concentrations for twenty-four hours. Cell viability was assesed using the cell counting kit-8 (CCK-8), Nrf2 expression was examined using ELISA, MDA level was measured using an MDA kit, and migration capacity was assessed using a scratch assay. Statistical analysis used one-way Anova or Kruskal-Wallis test. A p-value of &lt;0.05 was expressed statistically significant. ResultsThe Cell viability was substantially reduced in the nicotine group compared to the untreated group, accompanied by changes in cell morphology. In contrast, Nrf2 expression increased significantly (p=0.010) in the 5 mM nicotine group compared with the control group. The MDA levels were not significantly distinct across groups (p=0.056). Cell migration was delayed significantly in the 5 mM nicotine group at 72 hours after scratching compared to the control group. ConclusionNicotine decreased HGFs viability and increased Nrf2 expression significantly in a dose-dependent manner. Nicotine at 5 mM concentration did not alter MDA levels but delayed cell migration.

N-acetylcysteine as a novel methacrylate-based resin cement component: effect on cell apoptosis and genotoxicity in human gingival fibroblasts

BackgroundN-acetylcysteine (NAC) reduces the cytotoxicity and genotoxicity induced by monomers leached from dental composite resins. Herein, we investigated the effects of methacrylate-based resin cement used in dental implant restoration on apoptosis and genotoxicity, as well as the antiapoptotic and antigenotoxic capabilities of its component, NAC.MethodsThe antioxidant NAC (0.1 or 1 wt.%) was experimentally incorporated into the methacrylate-based dental resin cement Premier®. The Premier® + NAC (0.1 or 1 wt.%) mixture was subsequently immersed into Dulbecco’s modified Eagle’s medium for 72 h, and used to treat human gingival fibroblasts (HGFs). The viability of HGFs was determined using the XTT assay. The formation of deoxyribonucleic acid (DNA) double-strand breaks (DNA-DSBs) was determined using a γ-H2AX assay. Reactive oxygen species (ROS), apoptosis, necrosis, and cell cycles were detected and analyzed using flow cytometry.ResultsThe eluate of Premier® significantly inhibited HGF proliferation in vitro by promoting a G1-phase cell cycle arrest, resulting in cell apoptosis. Significant ROS production and DNA-DSB induction were also found in HGFs exposed to the eluate. Incorporating NAC (1 wt.%) into Premier® was found to reduce cell cytotoxicity, the percentage of G1-phase cells, cell apoptosis, ROS production, and DNA-DSB induction.ConclusionIncorporating NAC (1 wt.%) into methacrylate-based resin cement Premier® decreases the cell cytotoxicity, ROS production, and DNA-DSBs associated with resin use, and further offers protective effects against the early stages of cell apoptosis and G1-phase cell cycle arrest in HGFs. Overall, our in vitro results indicate that the addition of NAC into methacrylate-based resin cements may have clinically beneficial effects on the cytotoxicity and genotoxicity of these materials.

The dynamin inhibitor, dynasore, prevents zoledronate-induced viability loss in human gingival fibroblasts by partially blocking zoledronate uptake and inhibiting endosomal acidification.

For treatment of medication-related osteonecrosis of the jaw, one proposed approach is the use of a topical agent to block entry of these medications in oral soft tissues. We tested the ability of phosphonoformic acid (PFA), an inhibitor of bisphosphonate entry through certain sodium-dependent phosphate contransporters (SLC20A1, 20A2, 34A1-3) as well as Dynasore, a macropinocytosis inhibitor, for their abilities to prevent zoledronate-induced (ZOL) death in human gingival fibroblasts (HGFs). MTT assay dose-response curves were performed to determine non-cytotoxic levels of both PFA and Dynasore. In the presence of 50 μM ZOL, optimized PFA and Dynasore doses were tested for their ability to restore HGF viability. To determine SLC expression in HGFs, total HGF RNA was subjected to quantitative real-time RT-PCR. Confocal fluorescence microscopy was employed to see if Dynasore inhibited macropinocytotic HGF entry of AF647-ZOL. Endosomal acidification in the presence of Dynasore was measured by live cell imaging utilizing LysoSensor Green DND-189. As a further test of Dynasore's ability to interfere with ZOL-containing endosomal maturation, perinuclear localization of mature endosomes containing AF647-ZOL or TRITC-dextran as a control were assessed via confocal fluorescence microscopy with CellProfiler™ software analysis of the resulting photomicrographs. 0.5 mM PFA did not rescue HGFs from ZOL-induced viability loss at 72 hours while 10 and 30 μM geranylgeraniol did partially rescue. HGFs did not express the SLC transporters as compared to the expression in positive control tissues. 10 μM Dynasore completely prevented ZOL-induced viability loss. In the presence of Dynasore, AF647-ZOL and FITC-dextran co-localized in endosomes. Endosomal acidification was inhibited by Dynasore and perinuclear localization of both TRITC-dextran- and AF647-ZOL-containing endosomes was inhibited by 30 μM Dynasore. Dynasore prevents ZOL-induced viability loss in HGFs by partially interfering with macropinocytosis and by inhibiting the endosomal maturation pathway thought to be needed for ZOL delivery to the cytoplasm.

A Fibrin-Based Human Multicellular Gingival 3D Model Provides Biomimicry and Enables Long-Term In Vitro Studies.

Collagen-type I gels are widely used for the fabrication of 3D in vitro gingival models. Unfortunately, their long-term stability is low, which limits the variety of in vitro applications. To overcome this problem and achieve better hydrolytic stability of 3D gingival models, fibrin-based hydrogel blends with increased long-term stability in vitro are investigated. Two different fibrin-based hydrogels are tested: fibrin 2.5% (w/v) and fibrin 1% (w/v)/gelatin 5% (w/v). Appropriate numbers of primary human gingival fibroblasts (HGFs) and OKG4/bmi1/TERT (OKG) keratinocytes are optimized to achieve a homogeneous distribution of cells under the assumed 3D conditions. Both hydrogels support the viability of HGFs and the stability of the hydrogel over 28 days. In vitro cultivation at the air-liquid interface triggers keratinization of the epithelium and increases its thickness, allowing the formation of multiple tissue-like layers. The presence of HGFs in the hydrogel further enhances epithelial differentiation. In conclusion, a fibrin-based 3D gingival model mimics the histology of native gingiva in vitro and ensures its long-term stability in comparison with the previously reported collagen paralogs. These results open new perspectives for extending the period within which specific biological or pathological conditions of artificial gingival tissue can be evaluated.

Fibronectin-coated microgroove titanium surface promotes the behaviors of human gingival fibroblasts.

The present study was designed to clarify the activity of human gingival fibroblasts (HGFs) on the fibronectin (FN)-coated silanized microgroove titanium surface. The surface elemental composition of titanium discs was detected using XPS. HGFs' adhesion to the titanium discs was detected by immunofluorescence staining of vinculin. HGFs' number on the titanium discs was detected using the CCK8 assay. HGFs' secretion of type 1 collagen after five days of culturing was detected using ELISA and qPCR. HGFs could proliferate and spread well on the surface. The viability of HGFs in the experimental group was significantly more than in the control group. The HGFs in the experimental group significantly secreted more type 1 collagen than in the control group. Therefore, FN-coated can improve the morphology, viability, and type 1 collagen secretion of HGFs silanized microgroove titanium surface, which might ameliorate the efficacy of implants.

Biofilm formation on metal alloys and coatings, zirconia, and hydroxyapatite as implant materials in vivo.

Composition of implant material and its surface structure is decisive for oral biofilm accumulation. This study investigated biofilm formation on eight different materials. Eighteen healthy subjects wore intraoral splints fitted with two sets of eight materials for 24 h: zirconia [ZrO2 ]; silver-gold-palladium [AgAuPd]; titanium zirconium [TiZr]; Pagalinor [PA]; hydroxyapatite [HA]; silver-platinum [AgPt]; titanium aluminum niobium [TAN]; titanium grade4 [TiGr4]. Total biomass was stained by safranin to assess plaque accumulation while conventional culturing (CFU) was conducted to investigate viable parts of the biofilm. Cell viability of human gingival fibroblasts (HGF-1) was assessed in vitro. Statistical evaluation was performed with linear mixed-effects models to compare materials (geometric mean ratios, 95% CI), with the level of significance set at ɑ = .05. Less biofilm mass and CFU were found on noble metal alloys (AgPt, AgAuPd, and PA). Compared to AgPt, PA had 2.7-times higher biofilm mass value, AgAuPd was 3.9-times, TiGr4 was 4.1-times, TiZr was 5.9-times, TAN was 7.7-times, HA was 7.8-times, and ZrO2 was 9.1-times higher (each p < .001). Similarly, CFU data were significantly lower on AgPt, AgAuPd had 4.1-times higher CFU values, PA was 8.9-times, TiGr4 was 11.2-times, HA was 12.5-times, TiZr was 13.3-times, TAN was 16.9-times, and ZrO2 was 18.5-times higher (each p < .001). HGF-1 viability varied between 47 ± 24.5% (HA) and 94.4 ± 24.6% (PA). Noble alloys are considered as beneficial materials for the transmucosal part of oral implants, as less biofilm mass, lower bacterial counts, and greater cell viability were detected than on titanium- or zirconia-based materials.

Water temperature for fabrication of autopolymerizing polymethyl methacrylate (PMMA) interim fixed restoration affects cytotoxicity and residual methyl methacrylate (MMA)

Background/purposeClinically, dentists are suggested to immerse autopolymerizing interim fixed restorations in hot water during fabrication. However, this suggestion, without including the best temperature, mostly comes from clinical experience instead of scientific evidence. This in vitro study evaluated the effect of water temperature on the cytotoxicity of interim partial fixed dental prostheses (FDPs) and examined its correlation with residual MMA. Materials and methodsTempron was chosen as the autopolymerizing polymethyl methacrylate material. Tempron was mixed and then soaked in water at different temperatures, except control group (Controlair) was not being soaking in water. The specimens were incubated with conditioned medium. The concentration of residual MMA was determined by liquid chromatography-tandem mass spectrometry (LC-MS-MS). The cell viability of human gingival fibroblasts (HGFs) was evaluated by MTT assay. ResultsThe 60 °C and 80 °C groups exhibited significantly higher cell viabilities than those of the other groups (P < 0.05) at 48 and 72 h. The concentration of residual MMA was highly correlated with this outcome: the higher the concentration of residual MMA detected in the eluates, the poorer the cell viability was; the longer the incubation time was, the stronger the correlation was between the concentration of residual MMA and the cell viability. ConclusionAutopolymerizing PMMA interim FDPs that are polymerized in water up to at least 60 °C could reduce cell toxicity. Higher water temperature could certainly decrease the amount of residual MMA, which is closely correlated with the outcome of cell viability.

Resolvin D1 (RvD1) Regulates Porphyromonas Gingivalis Lipopolysaccharide-induced Del-1 and Cytokine Expressions in Human Gingival Fibroblasts

Purpose: To detect the effect of Resolvin D1 (RvD1) on Developmental endothelial locus-1 (Del-1) and cytokine expressions of human gingival fibroblast cells exposed to Porphyromonas gingivalis lipopolysaccharide (P. gingivalis-LPS).&#x0D; &#x0D; Material and Methods: The effect of RvD1 on cell viability of human gingival fibroblasts exposed to P. gingivalis-LPS were determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Meanwhile, the effect of RvE1 on Del-1 and cytokine (IL-1β, IL-6, IL-8, IL-10, IL-17) expressions of human gingival fibroblasts exposed to P. gingivalis-LPS were studied by real-time PCR experiment, statistical analysis was performed using GraphPad Prism version 5 for Windows.&#x0D; &#x0D; Results: Cell viability assay results demonstrated that RvD1 concentrations upregulated cell number compared to control group at 24 and at 72 hours. While RvD1 reduced IL-6, IL-8, IL-17 mRNA expression, IL-10 and Del-1 mRNA expression increased as a time- and dose-dependent manner. Also, IL-1β was not affected by RvD1 treatments. &#x0D; &#x0D; Conclusion: The increased expression of Del-1 and IL-10 by RvD1 down-regulated the pro-inflammatory cytokine expressions induced by P. gingivalis-LPS in gingival fibroblast. RvD1 displayed regulatory effects on gingival inflammation in P. gingivalis LPS-induced cell culture experiment. In particular, results of study display that Del-1 induced by RvD1 may have therapeutic potential to modulate periodontal inflammation.

Effects of metformin on human gingival fibroblasts: an in vitro study

ObjectiveTo investigate the effects of metformin (MF) treatment on the matrix metalloproteinases (MMPs) and proinflammatory cytokines production from lipopolysaccharide (LPS) - stimulated human gingival fibroblasts (HGFs).MethodsHGFs were obtained from subcultures of biopsies from clinically healthy gingival tissues of patients undergoing oral surgeries. Cell cytotoxicity assay was used to determine the effect of different concentrations of MF on viability of HGFs. HGFs were then incubated and treated with different concentrations of MF and Porphyromonas gingivais (Pg) LPS. MMP-1, MMP-2, MMP-8, MMP-9, IL-1β, and IL-8 expression analysis was performed using xMAP technology (Luminex 200, Luminex, Austin, TX, USA). Student’s t-test for a single sample was used to compare the mean values of the study groups with the control value. A p-value of <0.05 and 95% confidence intervals were used to report the statistical significance and precision of mean values.ResultsConcentrations of 0.5, 1- and 2-mM MF had a minimal non-significant cytotoxic effect on the HGFs and caused statistically significant reduction of MMP-1, MMP-2, MMP-8 and IL-8 expressed by the LPS-stimulated HGFs.ConclusionThe results of the present study confirm that MF suppresses MMP-1, MMP-2, MMP-8 and IL-8 in LPS-stimulated HGFs suggesting an anti-inflammatory effect of MF and potential adjunct therapeutic role in the treatment of periodontal diseases.

Advanced co-culture model: Soft tissue cell and bacteria interactions at the transgingival dental implant interface.

To better simulate and understand the clinical situation in which tissue cells and bacteria compete for settlement on an implant surface, the aim was to develop an improved transgingival co-culture model. For this model human gingival fibroblasts (HGF) were seeded on different titanium surfaces in the presence of the early colonizer Streptococcus gordonii or mixed oral bacteria. Subsequently adhesion and viability of HGF cells was analyzed. Simultaneous co-culture showed no decrease in the viability of HGF cells at early stages compared to the control group. However, a moderate impact on HGF viability (76±23%) was observed after 4h of co-culture, which then significantly decreased after 5h (21±2%) of co-cultivation, resulting in cell death and detachment from the surface. Further experiments including saliva pre-treatment of smooth and structured titanium surfaces with Streptococcus gordonii or mixed oral bacteria suggested a cell-protective property of saliva. Our study revealed that during simultaneous co-culture of cells and bacteria, which resembles the clinical situation the closest, the viability of gingival cells is considerably high in the early phase, suggesting that increasing initial cell adhesion rather than antibacterial functionality is a major goal and a relevant aspect in the development and testing of transgingival implant and abutment surface modifications.

Induction of Migration and Collagen Synthesis in Human Gingival Fibroblasts Using Periodontal Ligament Stem Cell Conditioned Medium.

This study aimed to examine the effect of periodontal ligament stem cell conditioned medium (PDLSC-CM) on human gingival fibroblast (HGF) migration and collagen synthesis. To assess cell viability, we extracted PDLSC-CM, and the total derived protein concentration was adjusted to 12.5 to 200 µg/mL, followed by treatment with HGFs. The viability of HGFs was observed for 24 hours using the MTT assay. Cell migration was monitored for 24 to 48 hours by wound healing and Boyden chamber assays. Collagen synthesis from HGFs was examined by picrosirius red dye and real-time polymerase chain reaction (PCR) to measure collagen type I and III gene expression for 7 to 10 days. A comparison among the groups was assessed using a one-way analysis of variance (ANOVA) and Bonferroni post hoc test, with the exception of the cell viability assay, which was subjected to Welch's test and Dunnett's T3 post hoc test. HGF viability was significantly enhanced by 12.5, 25, and 50 µg/mL PDLSC-CM. The HGFs treated with 50 µg/mL PDLSC-CM promoted cell migration as shown by wound healing and Boyden chamber assays. At this concentration, collagen synthesis increased at 10 days. Collagen type I gene expression increased by 1.6-fold (p < 0.001) and 4.96-fold (p < 0.001) at 7 and 10 days, respectively. Collagen type III gene expression showed an increase of 1.76-fold (p < 0.001) and 6.67-fold (p < 0.001) at the same time points. Our study suggested that a low concentration of PDLSC-CM at 50 µg/mL has given an amelioration of HGFs providing for periodontal wound healing and periodontal regeneration, particularly migration and collagen synthesis.

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

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 &lt; 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 (&lt; 30% cell viability) after 24-hour incubation.