Viability Of Gingival Fibroblasts Research Articles

Albiflorin Inhibits Advanced Glycation End Products-Induced ROS and MMP-1 Expression in Gingival Fibroblasts.

Periodontitis is a common complication of diabetes, with advanced glycation end products (AGEs) playing a key role in its pathogenesis. Albiflorin, a monoterpene glycoside, has shown potential anti-inflammatory and antioxidant properties. This study aims to investigate the effects of albiflorin on AGEs-induced gingival fibroblasts and its underlying mechanisms. This study aimed to evaluate the role of albiflorin in mitigating ROS production, inflammation, and MMP-1 expression in AGEs-induced gingival fibroblasts. The viability of gingival fibroblasts treated with albiflorin and AGEs was assessed using CCK-8 assays. ROS levels were measured by DCF staining, and the expression of inflammatory markers and MMP-1 was evaluated by ELISA and qPCR. The involvement of the NF-κB and Nrf2 pathways was examined by immunoblotting. Albiflorin enhanced the viability of AGEs-induced gingival fibroblasts and reduced ROS production. It also decreased the expression of IL-6, IL-8, RAGE, and MMP-1, suggesting an anti- inflammatory effect. Mechanistically, albiflorin modulated the NF-κB and Nrf2 pathways in AGEs-induced gingival fibroblasts. Albiflorin exhibited protective effects against AGEs-induced oxidative stress and inflammation in gingival fibroblasts, highlighting its potential as a therapeutic agent for periodontitis in diabetic patients. The modulation of the NF-κB and Nrf2 pathways by albiflorin provides insight into its mechanism of action.

Can Vitamin C Improve Proliferation and Viability of Smokers’ Gingival Fibroblasts on Collagen Membranes? An In Vitro Study

Periodontal regeneration using a barrier membrane can be affected by several factors, including patient-related factors (such as smoking habits), surgical techniques, and type of barrier membrane. Smoking exposure has a negative impact on the periodontium due to its direct inhibition of gingival fibroblast function. Vitamin C is widely recognized as an antioxidant that can be used to mitigate the detrimental impact of smoking products on periodontal cells. This study aimed to investigate whether vitamin C could improve the proliferation and viability of gingival fibroblasts extracted from smoking and non-smoking donors and then cultured on non-crosslinked (CopiOs Pericardium) and crosslinked (BioMend) collagen membranes. To address this aim, human gingival fibroblasts were extracted from healthy periodontium of smoker patients (Group 1) and non-smoker patients (Group 2). The cells were cultivated and subsequently subcultured in a growth medium supplemented with the required nutrients. Subsequently, the medium at passage six was supplemented with vitamin C, i.e., at the start of the experiment. An evaluation of cell proliferation and viability was carried out using cell migration assays and AlamarBlue® assays for cells grown on BioMend and CopiOs Pericardium collagen membranes. Assessment of the morphology and attachment of gingival fibroblasts to the experimental collagen membranes was conducted using scanning electron microscopy (SEM). The viability and proliferation assessments of hGFs from the migration assay were evaluated using AlamarBlue®. The results exhibited significant fluorescent intensity of gingival fibroblasts on both membrane groups (BioMend and CopiOs Pericardium) in the smoker group compared to the non-smoker group (p < 0.05), which was interpreted to be the result of hGF metabolic activity and the exclusion of any cytotoxic effects, particularly from vitamin C addition. Vitamin C positively affected cells from the smoker group with statistically significant results in the BioMend group (Wilcoxon signed-rank test of p value < 0.05; p = 0.028). SEM images revealed the crosslinking pattern of the BioMend membrane and the non-crosslinked natural tissue structure of the CopiOs Pericardium membrane, which did not change regardless of whether the cultured smoker or non-smoker hGFs were treated with vitamin C. Small numbers of attached hGFs in membrane matrices in all samples, mainly in the peripheries, were observed. It can be concluded that the addition of vitamin C to collagen membranes in vitro seems to combat the adverse effects of smoking products on gingival fibroblasts.

Selenium-Fortified Kombucha-Pollen Beverage by In Situ Biosynthesized Selenium Nanoparticles with High Biocompatibility and Antioxidant Activity.

Biogenic selenium nanoparticles (SeNPs) have been shown to exhibit increased bioavailability. Fermentation of pollen by a symbiotic culture of bacteria and yeasts (SCOBY/Kombucha) leads to the release of pollen content and enhances the prebiotic and probiotic effects of Kombucha. The aim of this study was to fortify Kombucha beverage with SeNPs formed in situ by Kombucha fermentation with pollen. Response Surface Methodology (RSM) was used to optimize the biosynthesis of SeNPs and the pollen-fermented Kombucha beverage. SeNPs were characterized by Transmission electron microscopy energy-dispersive X-ray spectroscopy (TEM-EDX), Fourier-transform infrared spectroscopy (FTIR), Dynamic light scattering (DLS), and Zeta potential. The pollen-fermented Kombucha beverage enriched with SeNPs was characterized by measuring the total phenolic content, antioxidant activity, soluble silicon, saccharides, lactic acid, and the total content of Se0. The polyphenols were identified by liquid chromatography-mass spectrometry (LC-MS). The pollen and the bacterial (nano)cellulose were characterized by scanning electron microscopy-energy dispersive X-ray spectroscopy (SEM-EDX), FTIR, and X-Ray diffraction (XRD). We also assessed the in vitro biocompatibility in terms of gingival fibroblast viability and proliferation, as well as the antioxidant activity of SeNPs and the pollen-fermented Kombucha beverage enriched with SeNPs. The results highlight their increased biological performance in this regard.

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

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. 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-1β. Interleukin 6 and interleukin 8 production were then quantified using Enzyme linked immunosorbent assay. 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-1β 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. Our study concluded that particles generated during ultrasonic scaling had no significant effect on viability of HGF cells and cytokine production.

Biologic Behavior of Pressed Lithium Disilicate Ceramic and Zirconia on Human Gingival Fibroblasts: An In Vitro Study.

This study evaluated in vitro the biologic profile of manually polished surfaces of pressed lithium disilicate (LD) ceramic compared to zirconia (Zir) in human gingival fibroblasts. Samples with a 10-mm diameter and 3-mm thickness were used. After manual polishing, the average roughness (Ra) of the samples was measured. The cell proliferation and viability of gingival fibroblasts on the surfaces were assessed at 24, 48, and 96 hours. Additionally, the morphology, cell adhesion, and type III collagen (COLIII) and vimentin (VIM) expression by fibroblasts plated onto these surfaces was analyzed. Polystyrene (Pol) was used as control for all assays. The mean Ra was 0.261 ± 0.053 μm for Zir and 0.345 ± 0.130 μm for LD. Both surfaces presented similar cell proliferation and viability (P > .05). The cell morphology demonstrated that, for both surfaces, the cells were occasionally spindle-shaped, parallel to the direction of the grooves. Compared to Pol, an upregulation of COLIII and VIM gene expression was observed by fibroblasts cultured on Zir and LD at all time points (P < .05). The characteristics presented by LD and Zir surfaces after manual polishing protocol were similar and had biologically favorable performances, thus suggesting LD as a suitable alternative to Zir in the peri-implant region for esthetic purposes.

Formulation &amp; Evaluation of Myrrh Toothpaste

Objectives: To formulate and measure the cytotoxicity level of organic myrrh toothpaste. Materials and Methods: Myrrh extract was prepared by the freeze drying process. Toothpaste was formulated by mixing specified amounts of myrrh extract, hydroxypropyl methylcellulose (HPMC) polymer, sodium lauryl sulphate (SLS), mint, and sucralose with deionized distilled water. MTT test was performed using concentrations of 100, 200, 300 and 400 mg/ml to assess the effect of myrrh paste on the gingival fibroblasts viability at intervals of 24 and 48hrs. The results were analysed by using SPSS version 27. Results: The formulated myrrh toothpaste has a homogeneous consistency as the extracted myrrh successfully dissolved completely with the other components. Cell viability test showed that myrrh paste concentrations of 100 - 300mg/ml were effective in maintaining the rate of fibroblasts growth after 24 and 48 hours as compared to the control samples. Cell growth rate was suppressed in the test samples treated with paste concentration of 400mg/ml. The results of the study imply that 300mg/ml is the safe and optimal concentration for fibroblasts growth, whereas concentrations ≥ 400mg/ml are intolerable and might be suppressing the proliferation of fibroblasts. Conclusion: Myrrh toothpaste concentration at 300mg/ml is the safe and optimal concentration for fibroblasts growth as depicted in the results, suggesting that at the optimal formulation, myrrh extract may not be toxic to the soft tissues and myrrh toothpaste may be useful for oral health care. Further clinical investigations are recommended to obtain the clinical efficacy data of the organic myrrh toothpaste.

Physicochemical, Antibacterial Properties, and Compatibility of ZnO-NP/Chitosan/β-Glycerophosphate Composite Hydrogels.

In this study we aimed to develop novel ZnO-NP/chitosan/β-glycerophosphate (ZnO-NP/CS/β-GP) antibacterial hydrogels for biomedical applications. According to the mass fraction ratio of ZnO-NPs to chitosan, mixtures of 1, 3, and 5% ZnO-NPs/CS/β-GP were prepared. Using the test-tube inversion method, scanning electron microscopy and Fourier-transform infrared spectroscopy, the influence of ZnO-NPs on gelation time, chemical composition, and cross-sectional microstructures were evaluated. Adding ZnO-NPs significantly improved the hydrogel's antibacterial activity as determined by bacteriostatic zone and colony counting. The hydrogel's bacteriostatic mechanism was investigated using live/dead fluorescent staining and scanning electron microscopy. In addition, crystal violet staining and MTT assay demonstrated that ZnO-NPs/CS/β-GP exhibited good antibacterial activity in inhibiting the formation of biofilms and eradicating existing biofilms. CCK-8 and live/dead cell staining methods revealed that the cell viability of gingival fibroblasts (L929) cocultured with hydrogel in each group was above 90% after 24, 48, and 72 h. These results suggest that ZnO-NPs improve the temperature sensitivity and bacteriostatic performance of chitosan/β-glycerophosphate (CS/β-GP), which could be injected into the periodontal pocket in solution form and quickly transformed into hydrogel adhesion on the gingiva, allowing for a straightforward and convenient procedure. In conclusion, ZnO-NP/CS/β-GP thermosensitive hydrogels could be expected to be utilized as adjuvant drugs for clinical prevention and treatment of peri-implant inflammation.

Novel Titanium Nanospike Structure Using Low-Energy Helium Ion Bombardment for the Transgingival Part of a Dental Implant.

Aim(s): The aim of the study was to fabricate a nanospike surface on a titanium alloy surface using a newly established method of low-energy helium ion bombardment. Various methods to achieve nanospike formation on titanium have been introduced recently, and their antibacterial properties have been mainly investigated with respect to Escherichia coli and Staphylococcus aureus. Oral pathogens such as Porphyromonas gingivalis play an important role in the development of peri-implantitis. For that reason, the antibacterial properties of the novel, nanostructured titanium surface against P. gingivalis were assessed, and a possible effect on the viability of gingival fibroblasts was evaluated. Materials and Methods: Helium sputtering was employed for developing titanium surfaces with nanospikes of 500 nm (ND) in height; commercially available smooth-machined (MD) and sandblasted and acid-etched titanium disks (SLA) were used as controls. Surface structure characterization was performed through scanning electron microscopy (SEM) and atomic force microscopy (AFM). Following incubation with P. gingivalis, antibacterial properties were determined via conventional culturing and SEM. Additionally, the viability of human gingival fibroblasts (HGFs) was tested through MTT assay, and cell morphology was assessed through SEM. Results: SEM images confirmed the successful establishment of a nanospike surface with required heights, albeit with heterogeneity. AFM images of the 500 nm nanospike surface revealed that the roughness is dominated by large-scale hills and valleys. For frame sizes of 5 × 5 μm and smaller, the average roughness is dominated by the height of the titanium spikes. ND successfully induces dysmorphisms within P. gingivalis cultures following the incubation period, while conventional culturing reveals a 17% and 20% reduction for ND compared to MD and SLA, respectively. Moreover, the nanospike surfaces did not affect the viability of human growth fibroblasts despite their sharp surface. Conclusion(s): This study successfully developed a novel titanium-nanospike-based structuration technique for titanium surfaces. In addition, the nanospikes did not hinder gingival fibroblast viability. Enhanced antimicrobial effects for such a novel nanospike-based resurfacing technique can be achieved through further optimizations for nanospike spacing and height parameters.

Long-term exposure to butyric acid induces excessive production of matrix metalloproteases in human gingival fibroblasts

ObjectiveThe purpose of this study was to clarify the relationship between bacteria-induced butyric acid and periodontal disease progression. DesignNormal human gingival fibroblasts were exposed to butyric acid (0, 1, 5, 10, and 15 mM) adjusted to a pH of 7.2–7.4 using sodium hydroxide for 0–96 h and cell viability was evaluated. In addition, the effects of butyric acid on the production of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in gingival fibroblasts were analyzed by real-time RT-PCR, ELISA, western blotting, and stromelysin zymography. ResultsButyric acid reduced the viability of gingival fibroblasts in a concentration- and time-dependent manner. Furthermore, butyric acid promoted production of MMP-1, MMP-3, and MMP-10 in gingival fibroblasts and suppressed TIMP-2 protein production. ConclusionsButyric acid promoted overproduction of MMPs, resulting in a disruption of the balance between MMPs and TIMPs expression in gingival fibroblasts. Our study suggests that the butyric acid produced by causative bacteria stimulates excessive MMP expression in periodontal tissue, leading to destruction of the tissue.

Effect of photobiomodulation with 810 and 940 nm diode lasers on human gingival fibroblasts.

The growth and proliferation of gingival fibroblasts are important in the process of oral wound healing, and photobiomodulation (PBM) might be able to modify this process. The aim of the current study was to evaluate the biomodulatory effect of a single session of laser PBM by means of 810 nm and 940 nm diode lasers alone and their combined application with different fluencies on human gingival fibroblasts (HGFs). Cells were provided by the Pasteur Institute, the National Cell Bank of Iran (NCBI) (C-165). Laser irradiation was carried out using 810 nm, 940 nm and 810 nm + 940 nm in the continuous wave (CW) mode, 100 mW, and energy densities of 0.5, 1.5 and 2.5 J/cm2. Cell viability was evaluated at 24 h with the MTT assay. Trypan blue staining was used to evaluate proliferation 24, 48 and 72 h after laser therapy. Propidium iodine was used to stain DNA and the cell nucleus. Laser irradiation (810 nm, 0.5 J/cm2) increased the viability of gingival fibroblasts, while this dose had an inhibitory effect with 940 nm. No positive effect on cell viability was found with other settings at 24 h. The viability results were not statistically different from those of the control in the dual wavelength group. At all single-laser irradiation doses, the cell proliferation results were lower as compared to the control at 48 and 72 h. The dual wavelength group results were significantly better than those of the control for the 1.5 J/cm2 and 2.5 J/cm2 energy densities (p < 0.001). Propidium iodine staining showed no negative effect of laser irradiation on the cell nucleus in any of the groups. Although a single irradiation dose of 810 nm, 0.5 J/cm2, resulted in a positive effect on cell viability at 24 h, no statistically significant stimulatory effect on viability and proliferation was observed for the other single wavelength group. When a combination of the 2 wavelengths was used, better results were observed as compared to the control, which needs to be further investigated in future studies.

Mammalian cell response and bacterial adhesion on titanium healing abutments: effect of multiple implantation and sterilization cycles.

Multiple implantations of the implant healing abutment (IHA) could adversely impact its surface properties in vivo. Furthermore, the effect of sterilization and reuse of the IHA on soft tissue viability and bacterial contamination has not been extensively studied. The goal of this study was to perform an in vitro analysis of mammalian cell viability and bacterial adhesion on the surfaces of retrieved IHA after single and multiple implantations and repetitive cycles of sterilization. IHA surface morphology was studied using optical microscopy. Cell viability of gingival fibroblasts (HGF-1) and oral keratinocytes (HOKg) in indirect contact with IHAs was assessed for 3 and 7 days. Immersion in bacterial culture was performed with a polyculture of Streptococcus species for 3 days and Streptococcus species with Fusobacterium nucleatum for 7 days. IHAs exhibited signs of surface damage even after a single exposure to the oral cavity. Fibroblasts did not show a significant preference towards control IHAs over used IHAs, whereas keratinocytes exhibited a significant decrease in viability when exposed to IHAs after multiple implantation cycles as compared with controls. Adherent bacterial count increased with increasing number of IHA implantations for both polycultures. Reusing of IHAs in vivo promoted surface degradation in addition to adversely impacting host cell viability and oral bacterial attachment in vitro. These findings show IHA reuse might potentially affect its clinical performance. Careful consideration should be taken when reusing IHAs in patients because this practice can result in permanent surface changes that might affect soft tissue integration during the healing period and promote bacterial colonization.

Evaluation of the In Vitro Oral Wound Healing Effects of Pomegranate (Punica granatum) Rind Extract and Punicalagin, in Combination with Zn (II).

Pomegranate (Punica granatum) is a well-established folklore medicine, demonstrating benefits in treating numerous conditions partly due to its antimicrobial and anti-inflammatory properties. Such desirable medicinal capabilities are attributed to a high hydrolysable tannin content, especially punicalagin. However, few studies have evaluated the abilities of pomegranate to promote oral healing, during situations such as periodontal disease or trauma. Therefore, this study evaluated the antioxidant and in vitro gingival wound healing effects of pomegranate rind extract (PRE) and punicalagin, alone and in combination with Zn (II). In vitro antioxidant activities were studied using DPPH and ABTS assays, with total PRE phenolic content measured by Folin–Ciocalteu assay. PRE, punicalagin and Zn (II) combination effects on human gingival fibroblast viability/proliferation and migration were investigated by MTT assay and scratch wounds, respectively. Punicalagin demonstrated superior antioxidant capacities to PRE, although Zn (II) exerted no additional influences. PRE, punicalagin and Zn (II) reduced gingival fibroblast viability and migration at high concentrations, but retained viability at lower concentrations without Zn (II). Fibroblast speed and distance travelled during migration were also enhanced by punicalagin with Zn (II) at low concentrations. Therefore, punicalagin in combination with Zn (II) may promote certain anti-inflammatory and fibroblast responses to aid oral healing.

Cytotoxicity and Gene Expression Changes of a Novel Homeopathic Antiseptic Oral Rinse in Comparison to Chlorhexidine in Gingival Fibroblasts.

Most available antiseptic solutions available today have strong antibacterial effects, however most also possess major cytotoxic effects on human gingival tissues. The VEGA Oral Care Recovery Kit (StellaLife), previously evaluated in clinical studies, consists of 16 active ingredients that are monographed in the Homeopathic Pharmacopeia of United States (HPUS) and recognized for their accelerated healing properties (reduction in post-op pain). The aim of this study was to compare VEGA to chlorhexidine (CHX) in vitro on gingival fibroblast viability, survival at various concentrations, migration assay, proliferation activity, expression of both regenerative growth factors as well as inflammatory markers, and collagen synthesis. A 10-fold dilution of standard CHX (0.02%) led to cell death, whereas cell viability was significantly better in the VEGA group for all tested parameters. Furthermore, VEGA also induced significantly greater fibroblast migration and proliferation. CHX negatively impacted the cellular inflammatory response of gingival fibroblasts, and also led to a reduction in collagen synthesis (50% decrease). Findings from the present study provide support from basic laboratory experiments that validate the previous clinical studies supporting the use of the VEGA oral rinse on its superior biocompatibility and wound healing properties when compared to CHX.

Effects of chlorhexidine, ethylenediaminetetraacetic acid, and sodium hypochlorite on cell viability of human gingival fibroblasts in vitro

Introduction: The aim of this in vitro study was to evaluate the cytotoxic effects of chlorhexidine (CHX), ethylenediaminetetraacetic acid (EDTA), and sodium hypochlorite (NaOCl) irrigants on human gingival fibroblasts. Materials and Methods: Gingival fibroblasts were cultured in Dulbecco Modified Eagle Medium for 24 h. Then, the cells were exposed for 1 min to different concentrations of CHX, EDTA, and NaOCl and the cell viability was assessed using 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide tetrazolium reduction assay. The percentage of gingival fibroblast viability was compared to control (100% viability). Results: All concentrations of CHX, EDTA, and NaOCl were cytotoxic to gingival fibroblasts compared to control group and this effect was dose-dependent (P 0.05). Conclusion: The cytotoxic effect of CHX, EDTA, and NaOCl on gingival fibroblasts was dose-dependent. Further studies are needed to assess and optimize the safety and efficacy of these irrigants in vivo.

Activity of chitosan antifungal denture adhesive against common Candida species and Candida albicans adherence on denture base acrylic resin

Statement of problemCandida adherence to the denture base is an important cause of denture stomatitis. In addition, infections with drug-resistant Candida have become more prevalent, especially among elderly and immunocompromised patients. Thus, alternative safe antifungal agents for oral applications are needed. PurposeThe purpose of this in vitro study was to investigate the activity of chitosan, a natural biopolymer, against common oral Candida species and its efficacy in inhibiting C albicans adherence to denture-base acrylic resin. Material and methodsThe minimum fungicidal concentrations (MFCs) of 5 types of chitosan against 6 species of Candida and 10 C albicans clinical isolates were determined by broth and agar dilution, respectively. N-succinyl chitosan (NSC), low- and high-molecular-weight water-soluble chitosan (LMWC and HMWC), and oligomer and polymer shrimp-chitosan were examined. NSC and HMWC, as pure gel and as a mixture with carboxymethylcellulose (CMC), were applied to acrylic resin disks, incubated with C albicans for 24 hours, and washed, and adherent cells were collected for colony count. The effects of HMWC on human gingival fibroblasts after 1 and 24 hours of treatment were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The retention force of HMWC gel was measured by using a universal testing machine. The Kruskal-Wallis and Mann-Whitney U tests were used to compare the antiadherence activity (α=.05). ResultsHMWC had the highest antifungal activity against most Candida species tested and C albicans clinical isolates. HMWC gel completely inhibited C albicans adherence to denture base acrylic resin (P<.001). CMC denture adhesive significantly increased C albicans adherence (P<.001), but adding 2×MFC HMWC into CMC reduced the adherence, although this was not statistically significant (P=.06). HMWC at 1×MFC and 2×MFC showed no toxic effect on gingival fibroblast viability and proliferation. Moreover, the retention force provided by HMWC gel was sufficient for use as a denture adhesive (>5000 Pa). ConclusionsHigh-molecular-weight, water-soluble chitosan is a biocompatible biopolymer that could inhibit C albicans adherence and that showed properties suitable for development into an antifungal denture adhesive.

Comparative Effects of Oral and Injectable Bisphosphonates in Primary Human Gingival Fibroblasts

Bisphosphonates are effective antiresorptive agents frequently used in the treatment of different bone disorders, as osteoporosis, Paget�s disease and tumours that cause osteolysis. A major concern related to bisphosphonates therapy is represented by osteonecrosis of jaw, a serious, debilitating, and mostly, a therapy-resistant disease, reported as a frequent side effect of bisphosphonates. The present study was aimed to assess and compare the effect of four commercially available bisphosphonates, very frequently used as oral (Fosamax - F and Actonel - A) and injectable (Ossica - O and Zoledronic acid - Z) therapy on primary human gingival fibroblasts - HGF viability. Alamar blue cell viability assay was performed to assess the effect of test compounds (1.5; 2.5; 5 and 10 mM) on gingival fibroblasts viability after a 24 h interval. In these experimental conditions, injectable bisphosphonates (O and Z) proved to be safe for HGF cells, whereas oral compounds (F and A) were cytotoxic even at low concentrations.

Copper-modified Ti6Al4V alloy fabricated by selective laser melting with pro-angiogenic and anti-inflammatory properties for potential guided bone regeneration applications

Custom-made biocompatible titanium alloy mesh can be designed to facilitate the regeneration of alveolar bone defects by supporting a protected space and inhibiting bacterial infections. Copper ions are often incorporated into titanium alloy due to their high bioactivity and outstanding antibacterial properties. However, the impacts of copper-bearing alloys on peri-implanted cell behaviors have rarely been systematically explored. In the present study, a copper-bearing alloy (Ti6Al4V-6Cu) was fabricated by selective laser melting (SLM) technology. The characterization of Ti6Al4V-6Cu alloy and its effects on the behaviors of gingival fibroblasts (HGFs), human umbilical vein endothelial cells (HUVECs), osteoblasts and macrophages were evaluated and compared with Ti6Al4V. The diffraction peaks of the Ti2Cu intermetallic phase were observed in the Ti6Al4V-6Cu alloy. Adding Cu enhanced the release of Ti and Al ions. The chemical state of Cu in the Ti6Al4V-6Cu alloy may exist predominantly in Cu2O or TiCuOx. Ti6Al4V-6Cu did not affect the attachment of HGFs or the osteogenic activity of osteoblasts. Furthermore, it inhibited the activation, proliferation, and pro-inflammatory cytokine secretion of macrophages and upregulated angiogenesis-related gene expression and VEGF-A secretion of HUVECs. These results demonstrate that a Ti6Al4V-6Cu alloy was successfully fabricated that did not negatively impact the cell viability of gingival fibroblasts and osteoblasts, inhibited the inflammatory response of macrophages, and increased the angiogenesis of HUVECs. Thus, Ti6Al4V-6Cu has potential applications for the fabrication of titanium alloy mesh to promote alveolar bone regeneration.

Curcumin inhibits TGF-β1-induced connective tissue growth factor expression through the interruption of Smad2 signaling in human gingival fibroblasts.

Many fibrotic processes are associated with an increased level of transforming growth factor-β1 (TGF-β1). TGF-β1 can increase synthesis of matrix proteins and enhance secretion of protease inhibitors, resulting in matrix accumulation. Connective tissue growth factor (CTGF) is a downstream profibrotic effector of TGF-β1 and is associated with the fibrosis in several human organs. Curcumin has been applied to reduce matrix accumulation in fibrotic diseases. This study was aimed to evaluate whether curcumin could suppress TGF-β1-induced CTGF expression and its related signaling pathway involving in this inhibitory action in primary human gingival fibroblasts. The differences in CTGF expression among three types of gingival overgrowth and normal gingival tissues were assessed by immunohistochemistry. Gingival fibroblast viability in cultured media with different concentrations of curcumin was studied by MTT assay. The effect of curcumin on TGF-β1-induced CTGF expression in primary human gingival fibroblasts was examined by immunoblotting. Moreover, the proteins involved in TGF-β1 signaling pathways including TGF-β1 receptors and Smad2 were also analyzed by immunoblotting. CTGF was highly expressed in fibroblasts, epithelial cells and some of endothelial cells, smooth muscle cells, and inflammatory cells in phenytoin-induced gingival overgrowth tissues rather than in those of hereditary and inflammatory gingival overgrowth tissues. Moreover, CTGF expression in the epithelial and connective tissue layers was higher in phenytoin-induced gingival overgrowth tissues than in normal gingival tissues. Curcumin was nontoxic and could reduce TGF-β1-induced CTGF expression by attenuating the phosphorylation and nuclear translocation of Smad2. Curcumin can suppress TGF-β1-induced CTGF expression through the interruption of Smad2 signaling.

Chitosan-triclosan particles modulate inflammatory signaling in gingival fibroblasts.

An important goal of periodontal therapy is the modulation of the inflammatory response. To this end, several pharmacological agents have been evaluated. Triclosan corresponds to an antibacterial and anti-inflammatory agent currently used in periodontal therapy. Chitosan is a natural polymer that may act as a drug delivery agent and exerts antibacterial and anti-inflammatory activities. Therefore, an association between both molecules might be useful to prevent inflammation and tissue destruction in periodontal tissues. In the present study, we have generated chitosan-triclosan particles and evaluated their morphology, charge, biocompatibility and gene expression analysis in human gingival fibroblasts. The chitosan-triclosan particles size and Z potential were 129±47nm and 51±17mV respectively. Human gingival fibroblast viability was not affected by chitosan-triclosan. A total of 1533 genes were upregulated by interleukin (IL)-1β. On the other hand, 943 were downregulated in fibroblasts stimulated with IL-1β plus chitosan-triclosan particles. Fifty-one genes were identified as molecular targets upregulated by IL-1 β and downregulated by the chitosan-triclosan particles. The gene ontology analysis revealed that these genes were enriched in categories related to biological processes, molecular function and cellular components. Furthermore, using real-time reverse transcription-polymerase chain reaction beta-actin, fibronectin, interleukin-6 and IL-1b genes were confirmed as targets upregulated by IL-1β and downregulated by chitosan-triclosan particles. Our results show that chitosan-triclosan particles are able to modulate the inflammatory response in gingival fibroblasts. This effect might be useful in the prevention and/or treatment of inflammation in periodontal diseases.

Evaluation of cytotoxicity and pH changes generated by various dental pulp capping materials - an in vitro study.

Various materials are used in direct dental pulp capping method. Their biocompatibility and alkalizing abilities are of primary importance affecting therapeutic effects. The aim of this study was to evaluate and compare the cytotoxicity of various pulp-capping materials on human gingival fibroblasts and investigate the pH changes induced by these materials. Human gingival fibroblasts were cultured with nine direct pulp materials using culture plate inserts. The cytotoxic effects were recorded by using an MTT-based colorimetric assay after 3 and 24 h. In the second part of the experiment, the materials were inserted in dialysis tubes and transferred into plastic vials containing deionized water. The changes of the medium pH were measured after 3 and 24 h. We showed differences in cell viability of gingival fibroblasts after varied time of exposition for the tested materials. Cell viability after 24 h increased for Dycal, Biopulp, and Calcipro, and decreased for Calcipulpe, Angelus, Angelus White, and ProRoot Regular. Cell viability for ProRoot and Life did not change. Non-setting calcium hydroxide preparations followed by the MTA group and setting calcium hydroxide materials produced the highest pH. All the tested materials significantly increased pH (p < 0.0001) at 24 h. Currently used pulp capping materials varied in their cytotoxicity relative to human gingival fibroblasts and their alkalizing capacities. Since most likely pH does not affect the viability of cultured cells, further investigations are required to determine physicochemical properties of these materials and the biological activity of the dental pulp.