Normal Human Gingival Fibroblasts Research Articles

A Combination of Curcumin and Lactobacillus rhamnosus GG Inhibits Viability and Induces Apoptosis in SCC-9 Human Oral Squamous Cell Carcinoma Cells.

The aim of this study was to evaluate the effect of curcumin combined with Lactobacillus rhamnosus GG cell-free supernatant (LGG CFS) on the proliferation and induction of apoptosis in SCC-9 oral squamous cell carcinoma (OSCC) cells. Curcumin (40 µg/ml) and 25% v/v LGG CFS (108 CFU/ml), both alone and in a combination regimen, significantly decreased the viability of SCC-9 cells and normal human gingival fibroblast (HGF) cells. Interestingly, the combination of low doses of curcumin (5 µg/ml) and 25% v/v LGG CFS (106 CFU/ml) had no effect on the HGF cells but significantly inhibited the viability of SCC-9 cells (p < 0.05). Flow cytometric analysis revealed that SCC-9 cells treated with the combination of low-dose curcumin and low-dose LGG CFS had a higher apoptotic rate than the cells in the control group and the single treatment groups (p < 0.05). The combined treatment also significantly increased the Bax/Bcl2 mRNA and protein expression in SCC-9 cells (p < 0.05) but not in HGF cells, indicating the underlying mechanism of the combination regimen. There was no significant difference in caspase-3 protein expression or the Bcl-xL/Bak and Mcl-1/Bak ratios between the treatment and control groups in both cell lines. These findings suggested that the coadministration of curcumin and LGG could exhibit anticancer effects in SCC-9 cells without causing toxicity to normal fibroblast cells.

The Effects of Injectable Platelet-Rich Fibrin and Advanced-Platelet Rich Fibrin on Gingival Fibroblast Cell Vitality, Proliferation, Differentiation.

Injectable Platelet Rich Fibrin (I-PRF) and Advanced-Platelet Rich Fibrin (A-PRF) are autologous materials derived from patients' blood and employed in periodontal regenerative surgery. Although I-PRF and A-PRF have different characteristics, their biological effects on gingival tissue fibroblasts remain unclear. This research aims to compare the in vitro capacity in inducing gene expression and proliferation of human gingival fibroblasts between A-PRF and I-PRF. Human donors undergoing dental implant surgery were sampled for normal human gingival fibroblasts (NHGFCs), followed by preparing A-PRF and I-PRF membranes. Enzyme-linked immunosorbent assay (ELISA) kit was used to assess the release of platelet-derived growth factor-AA (PDGF-AA), transforming growth factor-beta1 (TGF- β1), and insulin growth factor-1 (IGF-1) at different periods. Cell viability and proliferation of A-PRF and I-PRF were compared using CCK-8 assay. The impacts of platelet concentration on human gingival fibroblast cells (HGFCs) were evaluated by quantifying the level or amount of phosphorylated extracellular signal-regulated protein kinase (p-ERK), and Matrix metalloproteinases (MMPs), MMP-1 and MMP-3. The effects of PRF on aged human gingival fibroblast cells were examined retrospectively. Overall, A-PRF demonstrated a higher release of TGF-B1 and PDGF-AA, while I-PRF reflected higher levels of IGF-1. A significantly higher level of cell proliferation was induced by higher cell proliferation by A-PRF and I-PRF. Additionally, in comparison to I-PRF, the expression of ERK phosphorylation and MMP-1 &MMP-3 in HGFCs was demonstrated by I-PRF and A-PRF. The increase in A-PRF was time-dependent (p < 0.05). Both I-PRF and A-PRF induced a stimulatory biological impact on the proliferation of human gingiva fibroblasts, with the latter demonstrating better capacity in facilitating the release of different growth factors. A-PRF also induced higher gene expression of p-ERK, MMP-1 &MMP-3, and the proliferation of fibroblasts.

The Apoptotic Activity of Curcumin Against Oral Cancer Cells Without Affecting Normal Cells in Comparison to Paclitaxel Activity.

Until now, chemotherapy, which has a series of side effects, has been the most widely employed treatment for different types of cancer. However, bioactive products have been utilized as alternative medicines for tumors due to their bioactivities with low or no side effects in normal cells. This research reported for the first time that curcumin (CUR) and paclitaxel (PTX) have significant anti-cancer activity against normal human gingival fibroblast (HGF) and tongue squamous cell carcinoma fibroblast (TSCCF) cell lines. The results showed that CUR (13.85µgmL-1) and PTX (8.17µgmL-1) significantly inhibited TSCCF cell viability, with no significant effect on normal HGF cells. SEM showed morphological changes in cells treated with CUR and PTX, especially with TSCCF cells, compared to HGF normal cells. For TSCCF, the results showed the highest necrosis was achieved with CUR (58.8%) and PTX (39%) as compared to the control (2.99%). For normal HGF cells, the highest early and late apoptosis was achieved with PTX. Further, DCFH-DA analyses showed no significant ROS stimulation in TSCCF and HGF cell lines treated with CUR and PTX. The 1H NMR analysis results show the presence of methoxy and hydroxyl groups and aromatic hydrogens in the CUR structure. In conclusion, the results confirmed that CUR is more specific to the oral cancer cells but not normal cells by inducing apoptosis in a dose- and time-dependent manner, with decreased TSCCF cell viability, and the cytotoxicity of CUR and PTX is not through the ROS pathway.

Phytochemical Analysis and Anticancer Properties of Drimia maritima Bulb Extracts on Colorectal Cancer Cells.

Cancer is a worldwide health problem and is the second leading cause of death after heart disease. Due to the high cost and severe side effects associated with chemotherapy treatments, natural products with anticancer therapeutic potential may play a promising role in anticancer therapy. The purpose of this study was to investigate the cytotoxic and apoptotic characteristics of the aqueous Drimia maritima bulb extract on Caco-2 and COLO-205 colorectal cancer cells. In order to reach such a purpose, the chemical composition was examined using the GC-MS method, and the selective antiproliferative effect was determined in colon cancer cell lines in normal gingival fibroblasts. The intracellular ROS, mitochondrial membrane potential, and gene expression changes in selected genes (CASP8, TNF-α, and IL-6 genes) were assessed to determine the molecular mechanism of the antitumor effect of the extract. GC-MS results revealed the presence of fifty-seven compounds, and Proscillaridin A was the predominant secondary metabolite in the extract. The IC50 of D. maritima bulb extract on Caco-2, COLO-205, and the normal human gingival fibroblasts were obtained at 0.9 µg/mL, 2.3 µg/mL, and 13.1 µg/mL, respectively. The apoptotic effect assay indicated that the bulb extract induced apoptosis in both colon cancer cell lines. D. maritima bulb extract was only able to induce statistically significant ROS levels in COLO-205 cells in a dose-dependent manner. The mitochondrial membrane potential (MMP) revealed a significant decrease in the MMP of Caco-2 and COLO-205 to various concentrations of the bulb extract. At the molecular level, RT-qPCR was used to assess gene expression of CASP8, TNF-α, and IL-6 genes in Caco-2 and COLO-205 cancer cells. The results showed that the expression of pro-inflammatory genes TNF-α and IL-6 were upregulated. The apoptotic initiator gene CASP8 was also upregulated in the Caco-2 cell line and did not reach significance in COLO-205 cells. These results lead to the conclusion that D. maritima extract induced cell death in both cell lines and may have the potential to be used in CRC therapy in the future.

Analysis and Evaluation of the Effects of Melatonin on Oral Keratinocytes: A Pilot Study

Objective: Twenty to 30 percent of children in cross-sectional studies have significant bedtime problems or night waking. Melatonin, a synthetic form of the hormone produced by the pineal gland as a biomarker of the circadian system, is a commonly used nonprescription pharmacologic treatment for sleep disorders in children. Many studies have demonstrated the effect of melatonin supplementation on sleep duration and sleep quality, which can improve overall systemic health and disease prevention. However, despite the growing number of studies demonstrating the effects of melatonin to improve disordered sleep, no available studies have evaluated the effects of melatonin on normal oral tissues. Based upon this lack of knowledge, the primary objective of this study is to evaluate any potential effects of melatonin on normal oral cells and tissues within the physiologically relevant (supplementation) range.&#x0D; Methods: Normal oral keratinocytes (OKF4) and human gingival fibroblasts (HGF-1) were obtained and cultured for this study. Melatonin was administered in 96-well growth assays at supplement-equivalent physiologic concentrations at the low, mid and high range (1, 5 and 10 ug/uL) to determine any effects on cellular growth and proliferation. Changes in cellular viability and expression of cell cycle and apoptosis-related pathways were also evaluated.&#x0D; Results: Curvilinear U-shaped dose responses were observed in OKF cells under melatonin administration, ranging from -11.4% (low), to a maximum of -13.6% (mid) and -5.0% (high) compared with non-treated controls, p=0.029. Dose-responses among HGF-1 cells ranged from +12.1% (low), +17.4% (mid), and +5.0% (high), p=0.021. No changes in cellular viability were observed between control and experimental cells. However, qPCR screening of total RNA revealed significant changes in cell cycle related pathways, including c-myc, GAPDH and P53 but no changes in any apoptosis-related pathways, including Bcl-2, Bax, caspase-3, caspase-8 and caspase-9.&#x0D; Conclusions: This study demonstrated that melatonin does affect growth but not viability among these cell lines, which was found to be dose-dependent. These results suggest that melatonin may have some limited effects on oral tissues that may influence wound healing and repair but may not affect normal physiologic function or other cellular pathways. In agreement with other pediatric literature supporting the safety of melatonin use, this pilot study does not reveal any deleterious effects that would caution against its use in children or adults.

Different Effects of Reversible Electroporation-Mediated Fe3O4@AuNPs Delivery as Radiosensitizing Agents on Cancer Cells versus Normal Cells

Aim: In the present study, we sought to enhance the efficacy of radiotherapy via increasing the cellular uptake of Fe3O4@AuNPs as radiosensitizing agents using reversible electroporation, attaining a higher sensitivity for cancer cells to therapy-induced damage. Methods: The KB human nasopharyngeal carcinoma cell line was treated with different concentrations of gold-coated magnetic nanoparticles. The cells then received electroporation (750[Formula: see text]V/cm, 8 pulses for a duration 100[Formula: see text][Formula: see text]s with 100[Formula: see text]ms intervals) and radiotherapy (6MV X-ray, 2[Formula: see text]Gy). Control groups were carefully considered to assess the pure effect of both single and combinational therapeutic protocols. The MTT test and flow cytometry assay using Annexin V-PI kit were performed to evaluate potential effects on cell survival rate and to determine the induced level of apoptosis. Additionally, cellular uptake of nanoparticles was assessed and quantified using ICP-OES analysis. Results: Our study showed that nanoparticles and the applied procedure of electroporation had no considerable effects when utilized separately, but their combined application induced significant cell death ([Formula: see text]45%) and apoptosis ([Formula: see text]17%). Besides, the application of Fe3O4@AuNPs in the presence of electric field, 24[Formula: see text]h before the radiotherapy of tumor cells enhanced the cancer cell death ([Formula: see text]53%) as well as apoptosis rate ([Formula: see text]33%). The same scenario was also observed with normal HGF human gingival fibroblasts, confirmed by significantly lower levels of cell death and apoptosis. Conclusion: It can be concluded that electric fields at low voltage levels may significantly and selectively enhance the entry of nanoparticles into cancer cells, thereby accentuating the sensitivity of these cells to nanoparticle-mediated radiation therapy.

MiR-195 Inhibits Proliferation and Enhances Apoptosis of OSCC Cells via Targeting TLR4.

The aim of this research was to assess the function of microribonucleic acid (miR)-195 in the apoptosis and proliferation of oral squamous cell carcinoma (OSCC) cells as well as its action mechanism. The downstream target protein of miR-195 was predicted using the biological software. A quantitative polymerase chain reaction (qPCR) was implemented to examine the changes in expressions of miR-195 and its target protein toll-like receptor 4 (TLR4) in OSCC cell lines (TSCCA, Tca8223, Tb3.1, and CAL-27) and normal adult human gingival fibroblasts (HGFs), and the relation between their expressions was assessed. The expressions of phosphorylated proteins in nuclear factor-κB (NF-κB) pathway were determined through western blotting. miR-195 was expressed at a noticeably lower level in four OSCC cells than in HGFs, and the lowest level appeared in CAL-27 cells. Compared with miR-195 control, the miR-195 mimic could obviously raise the expression of miR-195. In CAL-27 cells with high expression of miR-195, the proliferation was inhibited and the apoptosis was evidently enhanced. OSCC cells exhibited evidently reduced protein and mRNA expression of TLR4, and miR-195 expression was inversely associated with TLR4 expression. It was uncovered from the dual-luciferase reporter assay that cells with wild-type TLR4 had prominently weakened luciferase activity relative to cells with mutant-type TLR4, revealing that the direct target of miR-195 is TLR4. The NF-κB pathway was impeded in cells that lowly expressed TLR4. miR-195 blocks the NF-κB pathway via inhibiting the expression of TLR4 in OSCC cells, thereby exerting an antitumor effect.

Preparation and Characterization of Novel Schiff Base Derived From 4-Nitro Benzaldehyde and Its Cytotoxic Activities.

Normal drugs exhibit activities against both normal and cancer cells. Furthermore, cancer cells may develop resistance to these drugs that alternative treatment must be explored. The main objective of this study was to examine the anticancer activity of Schiff base against Tongue Squamous Cell Carcinoma Fibroblasts (TSCCF) and normal human gingival fibroblasts (NHGF) and to propose its mechanism. A Novel Schiff base ligand was synthesized from the reaction of 5-C-2-4-NABA (5-chloro-2-((4-nitrobenzylidene) amino) benzoic acid). These Schiff bases possessed azomethine group (-HC=N-) and aromatic group (CH) as analyzed by Fourier transforms infrared (FTIR) spectroscopy and UV-Vis spectra. The in vitro cytotoxicity screening assay suggested promising activity against TSCCF with IC50 of 446.68 µg/mL, but insignificant activity against NHGF cells (IC50 of 977.24 µg/mL) after 72 h. The evidence of apoptotic induction was supported by DAPI staining of apoptotic nuclei with reduced cell numbers, suggesting that Schiff base could induce apoptotic bodies in cancer cells being observed. Based on the Schiff base structure, the anti-cancer mechanism may be attributed to the -HC=N- azomethine group. For the first time, our findings highlighted the anticancer activities of the new Schiff base against oral cancer cell lines.

IN VITRO CYTOTOXIC EVALUATION AND APOPTOTIC EFFECTS OF DILLAPIOLE ON HUMAN NASAL SQUAMOUS CELL CARCINOMA

Dillapiole is one of the major compounds in Peperomia pellucida. Dillapiole exerts cytotoxicity effect on several cancer cells such as breast and colon with a minimal effect on normal cells. However, its effect on cancers in head and neck region remains ambiguous. Therefore, in the current study, the cytotoxicity effect of dillapiole on human nasal squamous cell carcinoma, RPMI 2650 and the underlying mechanism were investigated. Normal human gingival fibroblast, HGnF cells was used as comparison. Cisplatin and untreated cells were used as positive and negative controls, respectively. Cell cytotoxicity was determined using WST-1 assay; and validated by trypan blue exclusion assays. Cell death mechanism was determined using Annexin V-FITC detection kit and analyzed by flow cytometer. The cytotoxic effect of dillapiole on RPMI 2650 cells was shown significantly increased with the increasing amount of dillapiole from 25.93 ± 6.39 % to 83.87 ± 8.43 % (p&lt;0.05, n=3) with the respective IC50 and IC75 of 46 µM and 125 µM. None of both IC50 and IC75 were obtained for HGnF cells up to 150 µM. Loss of normal shape, cytoplasm shrinkage and reduction in cell volume were detected in RPMI 2650 cells after dillapiole treatment at respective IC50 (46 µM) and IC75 (125 µM), indicating apoptosis. These findings indicate that dillapiole cytotoxicity effect is more selective towards RPMI 2650 compared to HGnF cells and the mechanism of death was through the induction of apoptosis.

Biological effects of exosome derived from Cal27 on normal human gingival fibroblasts.

The proliferation, migration capacity, and expression of activation-related proteins of NHGFs+Cal27-exo were determined by coculturing Cal27 exosome (Cal27-exo) with normal human gingival fibroblasts (NHGFs) to explore the effects of Cal27-exo on the activation and biological behavior of NHGFs. Cal27-exo was extracted using supercentrifugation, and exosomes were identified using Western blot, transmission electron microscopy (TEM), and particle size detection. Cal27-exo was cocultured with NHGFs to detect the uptake of Cal27-exo by NHGFs, and the proliferation and migration capacity of NHGFs+Cal27-exo were detected using CCK8 and wound healing tests, respectively. The expression levels of NHGF activation-related proteins, i.e., matrix metalloproteinase-9 (MMP-9), fibroblast-activating protein (FAP), alpha smooth muscle actin (αSMA), and transforming growth factor-β (TGF-β), were detected using real-time quantitative polymerase chain reaction (qRT-PCR). Cal27-exo was extracted u-sing supercentrifugation, and Western blot showed the positive expression levels of Alix and CD63. TEM showed that Cal27-exo had a circular double-layer vesicle. The particle size was between 30 and 150 nm. Cal27-exo labeled with PKH67 entered NHGFs after the coculture method. The wound healing test showed that the migration capacity of NHGFs+Cal27-exo was stronger after the scratch compared with that of NHGFs. CCK8 results showed that the proliferation activity of NHGFs+Cal27-exo was enhanced. qRT-PCR results showed that the MMP-9 levels of NHGFs+Cal27-exo were upregulated, whereas the TGF-β and αSMA mRNA levels of NHGFs+Cal27-exo were downregulated (P<0.05). The proliferation and migration ability of NHGFs+Cal27-exo are enhanced, and the mRNA expression of related proteins is changed. Cal27-exo can activate NHGFs, which suggests that Cal27-exo has potential significance in tumor invasion and metastasis.

Polymeric Nanocarriers with Luminescent Colloidal Nanoplatelets as Hydrophilic and Non-Toxic Two-Photon Bioimaging Agents.

IntroductionSemiconductor nanoplatelets (NPLs) are promising materials for nonlinear optical microscopy since they feature good two-photon absorption (TPA) properties, narrow photoluminescence spectra and high quantum yields of luminescence. Nevertheless, the use of semiconductor NPLs is inevitably connected with concerns about heavy metal ion toxicity and their intrinsically hydrophobic character.MethodsOur contribution focuses on the design and engineering of coloidal bionanomaterial consisting of two-dimensional highly luminescent CdSe semiconductor NPLs loaded into spherical and homogeneous polymeric nanocarriers (NCs) based on poly(ethylene oxide) and poly(propylene oxide) block co-polymer. The biocompatibility and usefulness of the NPLs-loaded polymeric NCs in two-photon induced bioimaging was demonstrated invitroby cytotoxicity and two-photon microscopic studies using eukaryotic (normal fibroblasts and cancer ovarian) cells.ResultsThe encapsulated NPLs maintain their intensive and spectrally narrow photoluminescence, as well as preserve good TPA properties, while the surrounding polymer shell imparts hydrophilic character and non-toxicity towards eukaryotic cells. Specifically, TPA cross-sections of the colloidal NCs loaded with NPLs show large values reaching up to 2.0 × 108 GM, with simultaneously two-photon brightness reaching 2.2 × 107 GM at 870 nm. MTT proliferation assay performed on cell lines treated with encapsulated NPLs revealed at least 70% viability of normal human gingival fibroblast (HGF) and cancer ovarian (MDAH-2774) cells, while the results of multiphoton imaging of murine (L-929) fibroblasts suggest that the encapsulated NPLs are capable of labelling the target cells enabling their visualization.ConclusionAs a result, we obtained water dispersible and temporally stable hydrophilic NPLs-loaded NCs that offer excellent, both one- and two-photon excited fluorescence preserving optical properties of the raw hydrophobic and colloidal NPLs. The biological responses upon eukaryotic cells indicate that the encapsulation process protects cells from the toxic influence of cadmium simultaneously preserving the unique multiphoton properties of the active cargo which opens a promising perspective for its application in multiphoton cancer bioimaging excited at the “optical transmission window” of biological tissues in near-infrared range.

A long-term controlled drug-delivery with anionic beta cyclodextrin complex in layer-by-layer coating for percutaneous implants devices

This study demonstrated a drug-delivery system with anionic beta cyclodextrin (β-CD) complexes to retain tetracycline (TC) and control its release from multilayers of poly(acrylic acid) (PAA) and poly(l-lysine) (PLL) in a ten double layers ([PAA/PLL]10) coating onto titanium. The drug-delivery capacity of the multilayer system was proven by controlled drug release over 15 days and sustained released over 30 days. Qualitative images confirmed TC retention within the layer-by-layer (LbL) over 30 days of incubation. Antibacterial activity of TC/anionic β-CD released from the LbL was established against Staphylococcus aureus species. Remarkably, [PAA/PLL]10/TC/anionic β-CD antibacterial effect was sustained even after 30 days of incubation. The non-cytotoxic effect of the multilayer system revealed normal human gingival fibroblast growth. It is expected that this novel approach and the chemical concept to improve drug incorporation into the multilayer system will open up possibilities to make the drug release system more applicable to implantable percutaneous devices.

Effects of melatonin on the Bisphenol-A- induced cytotoxicity and genetic toxicity in colon cancer cell lines, normal gingival cell lines, and bone marrow stem cell lines.

Bisphenol-A (BPA) is a synthetic chemical that has widely been used in the production of polycarbonate plastic and epoxy resins in the manufacture of consumer products. The most common path of human exposure to BPA is by oral intake that involves genotoxicity, oxidative stress, endocrine disruption, mutagenicity, and carcinogenicity in both in vitro and in vivo models. Melatonin is known as a free-radical scavenger and a powerful antioxidant agent. This study aimed to investigate the effects of melatonin on viability and genetic disorders of normal Human Gingival Fibroblasts (HGF), colon cancer (MKN45), and bone marrow stem cell (MSC) lines exposed to BPA. For this purpose, MTT and Comet assays were performed to evaluate the cytotoxicity and genotoxicity properties of BPA and the role of melatonin. The results showed that BPA exposure resulted in increased oxidative stress parameters including MDA and ROS, and decreased GSH content. The current study demonstrated the cytotoxicity and genotoxicity effects of BPA and the protective role of melatonin in preventing cytotoxicity and DNA damage are induced by BPA.

Activated KCNQ1 channel promotes fibrogenic response in hereditary gingival fibromatosis via clustering and activation of Ras.

Activated potassium channels were found to be strongly correlated with gingival overgrowth (GO) phenotype as we reviewed syndromic hereditary gingival fibromatosis (HGF). Nevertheless, the functional roles of potassium channels in gingival fibrosis or gingival overgrowth remained uncovered. The aim of the present study was to explore the pathogenic role of aberrantly activated potassium channel in Hereditary Gingival Fibromatosis (HGF). Gingival tissues were collected from 9 HGF patients and 15 normal controls. Expression of KCNQ1 was detected by immunohistochemistry. Gingival fibroblasts were isolated, and outward K+ currents were detected by whole-cell patch-clamp analysis, transmembrane potential was determined by flow cytometry. Normal human gingival fibroblasts (NHGFs) were transfected with KCNQ1 adenovirus or treated with KCNQ1 selective agonist ML277 and antagonist chromanol 293B. Accumulation of Extracellular Matrix (ECM) was measured by Western blotting and Sircol Soluble Collagen Assay. Content of secreted TGF-β1 was measured by ELISA. Active RAS pull-down assay and cell immunofluorescence were utilized to verify RAS activation. KCNQ1 was upregulated in gingival tissues derived from HGF patients and HGF gingival fibroblasts presented increased outward K+ currents than NHGFs. Overexpression of KCNQ1, or KCNQ1 agonist ML277, promoted fibrotic responses of NHGFs. TGF-β1 and KCNQ1 channels formed a positive feed-back loop. ML277 generated lateral clustering and activation of Ras on plasma membrane, followed by augmented MAPK/AP-1 signaling pathway output. JNK or ERK1/2 inhibitors suppressed ML277-induced AP-1 and ECM upregulation. Activation of KCNQ1 potassium channel promoted fibrogenic responses in NHGFs via Ras/MAPK/AP-1 signaling.

A high mannose concentration is well tolerated by colorectal adenocarcinoma and melanoma cells but toxic to normal human gingival fibroblast: an in vitro investigation

BackgroundThe primary cause of cancer is gene mutation which allows the growth of abnormal and damaged cells. Nutrition is one of the key factors that either increases or decreases the risk of cancer. Mannose has been found in many fruits such as oranges, apples and berries. Mannose has been linked to increase the risk factors or potential therapeutic for cancers. However, insufficient information is available on the effects of high mannose concentration on the normal and cancer cell lines. This study aimed to evaluate the viability patterns of human cancer and normal cell lines treated with mannose. Human gingival fibroblast (HGF), skin malignant melanoma (A375) and colorectal adenocarcinoma (HT29) cell lines were cultured and treated with additional mannose in three respective concentrations: 1 mg/ml, 5 mg/ml and 10 mg/ml. Then, cell viability was measured using an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide)-assay.ResultsThe HGF cells’ percentage pattern of viability showed a rapid decline of nearly 95% on the third day of treatment. A375 cells were able to survive in high mannose condition as the cell viability percentage was at the highest value on Day 5. Meanwhile, HT29 cells showed declining cell viability pattern when treated with mannose. The data exhibited significance; the p value was less than 0.001.ConclusionsHigh mannose concentration can be toxic to HGF. In addition, A375 is adaptive to mannose at all concentrations in which it shares the same pattern with the untreated group. However, the cell viability pattern for HT29 cell is declining.

Antimicrobial activity and inhibition of biofilm formation in vitro and on human dentine by silver nanoparticles/carboxymethyl-cellulose composites

ObjectiveTo evaluate the antimicrobial activity of a silver nanoparticles/carboxymethyl-cellulose (AgNPs/CMC) composite on in vitro and dentine disc heterogeneous biofilms. DesignAgNPs/CMC composite effect on normal human gingival fibroblast cells (HGF) viability was determined by the MTT reduction assay. In addition, we evaluated the antimicrobial effect of AgNPs/CMC composite on Candida albicans, Enterococcus faecalis, and Fusobacterium nucleatum growth in vitro and heterogeneous biofilms, as well as dentine disc biofilms. ResultsQuasi-spherical AgNPs/CMC composites, with a mean 22.3 nm particle-size were synthesized. They were not toxic to HGF cells at concentrations tested that were antimicrobial, however they caused significant cytotoxicity (89 %, p < 0.05) at concentrations > 15 μg/mL. In vitro, they inhibited up to 67 %, 66 %, and 96 % C. albicans, E. faecalis, and F. nucleatum growth at concentrations ranging from 1.2 μg/mL to 9.6 μg/mL, as compared with untreated control. We also demonstrated significant (p < 0.05) 58 % biofilm reduction by 4.8 μg/mL AgNPs/CMC composite on human dentine discs. ConclusionAgNPs/CMC composite showed anti biofilm activity on monocultures, heterogenous cultures, and dentine discs, resulting a potentially effective alternative to prevent and eliminate infections after endodontic treatment.

Evaluation of cytotoxicity of areca nut and its commercial products on normal human gingival fibroblast and oral squamous cell carcinoma cell lines

Consumption of areca nut products is the most common cause of oral cancers, particularly in South Asian countries. This study evaluates the cytotoxic and necrotizing effects of areca nut and its formulations on normal human gingival fibroblasts (HGF-1) and oral squamous cell carcinoma (OSCC, CAL-27) cell lines. Identification of various carcinogens and adulterants using LC-HR-ESI-MS/MS analysis was performed in the extracts of areca nut and its products. Apart from alkaloids and flavonoids, a major adulterant, saccharin was found in all the samples of chalia (one of the most common chewing products of areca nut) in the ranges between 1.697–7.170 mg/g of the sample. Cytotoxic studies showed that most of the areca nut products were found cytotoxic to HGF-1 cells while being relatively non-cytotoxic against CAL-27 cells, rather they promote the growth of cancer cells. Our findings revealed that the components of areca nut and its products were injurious to HGF-1 cells and caused necrosis, which may attenuate HGF-1 protection toward oral epithelial cells. Moreover, the non-cytotoxic effect of these products on cancer cell lines suggests further predisposal of the habitual chewers for developing oral carcinomas. This study will give a better understanding of the hazardous effects of areca nut products.

MiR-200a inhibits proliferation rate in drug-induced gingival overgrowth through targeting ZEB2

Gingival overgrowth can occur as a result of poor oral hygiene or a side effect of taking certain medications, such as cyclosporine A (CsA). It has been shown that this immunosuppressant drug induces epithelial-to-mesenchymal transition (EMT) in the gingival epithelium but the associated molecular mechanism remains to be elucidated. We first assessed the relative expression of microRNA-200a (miR-200a) in response to the CsA treatment using qRT-PCR. Next, luciferase reporter assay was applied to examine whether miR-200a was able to regulate ZEB2 and Western blot was utilized to measure the expression of ZEB2 in normal human gingival fibroblasts (HGFs). To confirm the significance of miR-200a and ZEB2 in the CsA-induced gingival overgrowth, miR-200a inhibitor and shRNA mediated knockdown of ZEB2 were used and cell proliferation in HGFs was assessed by MTT assay. The expression of miR-200a was dose-dependently downregulated following the CsA treatment. Luciferase reporter assay confirmed that ZEB2 was a direct downstream target regulated by miR-200a and ZEB2 was indeed increased after the administration of CsA. We demonstrated that knockdown of ZEB2 hampered the CsA-induced HGFs proliferation and the elevated cell proliferation due to inhibition of miR-200a was reversed by repression of ZEB2. Our results showed that insufficient miR-200a in HGFs caused by CsA administration may lead to gingival enlargement mediated by the upregulation of ZEB2. This finding supported that CsA-induced EMT contributed to the adverse effect of using CsA and miR-200a may serve as an upstream target to prevent the overgrowth of the gingiva.

Leucocyte- and platelet-rich fibrin regulates expression of genes related to early wound healing in human gingival fibroblasts.

Leucocyte- and platelet-rich fibrin (L-PRF) is a blood-derived biomaterial rich in leucocytes and platelets embedded in a high-density fibrin network that can be compressed into a membrane and used in surgical applications to stimulate tissue regeneration and wound healing, especially in oral cavity. This study aimed to determine the combined effects of the growth factors and cells present in L-PRF on fibroblasts that directly face the L-PRF membranes placed during surgical procedures. The effect of L-PRF from six donors on the expression of 84 key wound healing genes in normal human gingival fibroblasts was tested by RT-qPCR. L-PRF significantly regulated the expression of 33 fibroblast genes (39%), including interleukins, myofibroblast-, extracellular matrix- and angiogenesis-associated genes, and matrix metalloproteinase-1 and -3. L-PRF regulated fibroblast gene expression both time- and dose-dependently, and the effects were mediated by mitogen-activated protein kinases ERK1/2, JNK and p38. L-PRF also stimulated fibroblast wound closure and promoted the ability of fibroblasts to induce endothelial tube formation. L-PRF-induced gene expression changes in fibroblast were similar to those observed in early human and pig wounds. This study provides new insights into the biological mechanism by which L-PRF regulates key gingival fibroblast functions important in wound healing.

A biological active artificial saliva formulation containing flower mucilage from Ceylon Spinach (Basella alba Linn.).

Ceylon Spinach (Basella albe) is an edible perennial vine found in tropical Asia and Africa, known as vegetables containing mucilage. Its mucilage from flowers was extracted by microwaving and precipitated with 95% ethanol. Five artificial saliva formulations composing of mucilage from Ceylon Spinach, calcium chloride (CaCl2), potassium chloride (KCl) and sodium fluoride (NF) were developed. The best formulation No.5 containing 0.61% of the mucilage with the non-Newtonian pseudoplastic flow (8.9 ± 0.2 cP) and the wetting time (12.50 ± 2.24 min) similar to the normal human saliva was selected. This artificial saliva formulation exhibited biological activities including an antioxidative activity by DPPH free radical scavenging with the SC50 of 14.26 ± 2.00 mg/ml (0.05 folds of ascorbic acid), and the adhesion inhibition of S. mutans on hydroxyapatite beads at 17.01 ± 7.75%, while the natural human saliva exhibited an increase bacterial adhesion of 33.10 ± 9.70%. The safety of this formulation which gave no cytotoxicity on normal human gingival fibroblasts at 99.20 ± 21.09% cell viability was also demonstrated. The results from this study have indicated high biological activity and safety of the developed formulation containing mucilage from Ceylon Spinach which is potential to be used as artificial saliva for xerostomia patients.