Human Gingival Fibroblast Cells Research Articles

Effect of photodynamic therapy based on indocyanine green on expression of apoptosis-related genes in human gingival fibroblast cells

BackgroundPeriodontal diseases refer to inflammation of the gingiva, induction of apoptosis in human gingival fibroblast cells, destruction of the surrounding tissues, and early bone loss resulting in infections due to the pathogenic activity of the microorganisms and the host immune inflammatory responses. Recent investigations have suggested that antimicrobial photodynamic therapy (aPDT) can be an adjunct treatment therapy for periodontal infections. AimTo prove the lack of side effects of PDT on periodontal tissues, we investigated the expression of BAX and BCL-2 genes that are involved in apoptosis after the PDT on human gingival fibroblast (HGF) cells. Materials and methodsIn this study the effect of PDT based on indocyanine green (ICG) as a photosensitizer with the diode laser were tested on the expression of BAX and BCL-2 genes in monolayers of HGF cells. The effects of PDT on the expression of BAX and BCL-2 genes were evaluated by real-time quantitative reverse transcription PCR. ResultsThe results of the genes expression analysis revealed that ICG-PDT at concentrations 1000μg/mL, induced the significant expression of BAX in HGF cells; however, the laser irradiation as well as ICG showed no significant effects on the expression of these genes. Treatment with ICG alone, laser irradiation and ICG-PDT caused no observable BCL-2 gene expression changes between the tested and control groups. ConclusionOur findings indicate that ICG-PDT at 1000μg/mL of ICG with the exposure time of 60s for the diode laser would appear to be an inducer of apoptosis in HGF at transcriptome level.

Layer-Number Dependent Antibacterial and Osteogenic Behaviors of Graphene Oxide Electrophoretic Deposited on Titanium

Graphene oxide has attracted widespread attention in the biomedical fields due to its excellent biocompatibility. Herein we investigated the layer-number dependent antibacterial and osteogenic behaviors of graphene oxide in biointerfaces. Graphene oxide with different layer numbers was deposited on the titanium surfaces by cathodal electrophoretic deposition with varied deposition voltages. The initial cell adhesion and spreading, cell proliferation, and osteogenic differentiation were observed from all the samples using rat bone mesenchymal stem cells. Both Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus were used to investigate the antibacterial effect of the modified titanium surfaces. Cocultures of human gingival fibroblasts (HGF) cells with Escherichia coli and Staphylococcus aureus were conducted to simulate the conditions of the clinical practice. The results show that the titanium surfaces with graphene oxide exhibited excellent antibacterial and osteogenic effects. Increasing the layer-number of graphene oxide resulted in the augment of reactive oxygen species levels and the wrinkling, which led to the antibacterial and osteogenic effects, respectively. Compared to pure titanium surface in the cells-bacteria coculture process, the modified titanium surfaces with graphene oxide exhibited higher surface coverage percentage of cells.

In Vitro Comparison of Biological Effects of Coe-Pak and Reso-Pac Periodontal Dressings.

ABSTRACTObjectivesThe purpose of the present study was to compare the cytotoxicity of Reso-Pac and Coe-Pak periodontal dressing.Material and MethodsAccording to ISO-10993-12:2012, 1-, 3- and 7-day extracts of the two periodontal dressings were prepared in cell culture medium and exposed to the two cultured cell lines. Cell viability and proliferation at 24 h and 72 h following exposure were evaluated using quantitative MTT assay.ResultsThe results showed a significant (P < 0.05) reduction in the viability of cells exposed to the 3- and 7-day Coe-Pak extracts at 24 h and 72 h compared to the control group (no exposure to the extract). Reso-Pac extracts slightly decreased cell viability compared to the control group. Understudy materials showed greater cytotoxicity against human osteoblast-like compared to the human gingival fibroblast cells. No significant (P > 0.05) difference was found in the viability of cells exposed to undiluted (100%) one-day extract and diluted (50%) extract of both understudy materials at 24 h and 72 h after exposure.ConclusionsBased on the results, Reso-Pac periodontal dressing has less cytotoxicity than Coe-Pak.

The anti-oxidant effects of melatonin derivatives on human gingival fibroblasts

ObjectivesAim of this in vitro study was to evaluate the anti-oxidant activity of indole ring modified melatonin derivatives as compared with melatonin in primary human gingival fibroblast (HGF) cells. MethodsAnti-oxidant activity of melatonin (MLT), acetyl-melatonin (AMLT) and benzoyl-melatonin (BMLT) was evaluated by5 standard methods as follows: 2, 2-diphenyl-1-picrylhydrazyl (DPPH); ferric ion reducing antioxidant power (FRAP); superoxide anion scavenging; nitric oxide (NO) scavenging; and thiobarbituric acid reactive substances (TBARs).Evaluation of cellular antioxidant activity (CAA) and protectivity against H2O2 induced cellular damage was performed via MTT assay in HGF cells. ResultsAccording to the standard anti-oxidant assays, the antioxidant power of AMLT and BMLT were slightly less than MLT in FRAP and superoxide scavenging assays. In the NO scavenging and TBARs assays, BMLT and AMLT were more potent than MLT, whereas DPPH assays demonstrated that MLT was more potent than others. BMLT and AMLT had more potent anti-oxidant and protective activities against H2O2in HGF cells as compared with MLT. ConclusionsMLT derivatives demonstrated different anti-oxidant activities as compared with MLT, depending upon assays. These findings imply that N-indole substitution of MLT may help to improve hydrogen atom transfer to free radicals but electron transfer property is slightly decreased. Anti-oxidant and protective effects of melatonin derivatives (AMLT and BMLT) on human gingival fibroblasts imply the potential use of these molecules as alternative therapeutics for chronic inflammatory oral diseases.

Calprotectin Induces IL-6 and MCP-1 Production via Toll-Like Receptor 4 Signaling in Human Gingival Fibroblasts.

Calprotectin, a heterodimer of S100A8 and S100A9 molecules, is associated with inflammatory diseases such as inflammatory bowel disease. We have reported that calprotectin levels in gingival crevicular fluids of periodontitis patients are significantly higher than in healthy subjects. However, the functions of calprotectin in pathophysiology of periodontitis are still unknown. The aim of this study is to investigate the effects of calprotectin on the productivity of inflammatory cytokines in human gingival fibroblasts (HGFs). The HGFs cell line CRL-2014® (ATCC) were cultured, and total RNAs were collected to examine the expression of TLR2/4 and RAGE mRNA using RT-PCR. After the cells were treated with S100A8, S100A9, and calprotectin, supernatants were collected and the levels of IL-6 and MCP-1 were measured using ELISA methods. To examine the intracellular signals involved in calprotectin-induced cytokine production, several chemical inhibitors were used. Furthermore, after the siRNA-mediated TLR4 down-regulated cells were treated with S100A8, S100A9, and calprotectin, the levels of IL-6 and MCP-1 were also measured. HGFs showed greater expression of TLR4 mRNA, but not TLR2 and RAGE mRNA compared with human oral epithelial cells. Calprotectin increased significantly the production of MCP-1 and IL-6 in HGFs, and the cytokine productions were significantly suppressed in the cells treated with MAPKs, NF-κB, and TLR4 inhibitors. Furthermore, calprotectin-mediated MCP-1 and IL-6 production were significantly suppressed in TLR4 down-regulated cells. Taken together, calprotectin induces IL-6 and MCP-1 production in HGFs via TLR4 signaling that involves MAPK and NF-κB, resulting in the progression of periodontitis. J. Cell. Physiol. 232: 1862-1871, 2017. © 2016 Wiley Periodicals, Inc.

Cytotoxicity assessment of three endodontic sealing cements used in periapical surgery. In vitro study

Introduction: Presently there are many retrofilling materials in the market, nevertheless, little is known about their toxicity on gingival fibroblasts. Objective: To assess cytotoxicity of three materials to human gingival fibroblasts and L929 mouse fibroblasts cell line. Material and methods: EndoSequence® BC RRMTM (ERRM; root repair material), white MTA Angelus® (MTA) and intermediate restoration material (IRM®) conditioned media were obtained when materials were freshly mixed, at setting time and after 1, 24 and 72 hours of setting time. Cell morphology was assessed with light microscopy and cell viability was assessed through mitochondrial metabolic activity with 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT). Statistical analysis was conducted with ANOVA. Results: We found that ERRM material did not exhibit cytotoxic effects on used fibroblast, nevertheless, MTA and IRM® respectively exhibited moderate and severe cytotoxicity, thus indicating the materials were not fully harmless. Conclusion: Bioceramic cements like ERRM could be considered the most compatible retrofilling-materials.

LPS from P. gingivalis Negatively Alters Gingival Cell Mitochondrial Bioenergetics.

Objective Oral inflammatory pathologies are linked to increased oxidative stress, thereby partly explaining their relevance in the etiology of systemic disorders. The purpose of this work was to determine the degree to which LPS from Porphyromonas gingivalis, the primary pathogen related to oral inflammation, altered gingival mitochondrial function and reactive oxygen species generation. Methods Human gingival fibroblast (HGF-1) cells were treated with lipopolysaccharide of P. gingivalis. Mitochondrial function was determined via high-resolution respirometry. P. gingivalisMitochondrial function was determined via high-resolution respirometry. Results LPS-treated HGF-1 cells had significantly higher mitochondrial complex IV and higher rates of mitochondrial respiration. However, this failed to translate into greater ATP production, as ATP production was paradoxically diminished with LPS treatment. Nevertheless, production of the reactive H2O2 was elevated with LPS treatment. Conclusions LPS elicits an increase in gingival cell mitochondria content, with a subsequent increase in reactive oxygen species production (i.e., H2O2), despite a paradoxical reduction in ATP generation. These findings provide an insight into the nature of oxidative stress in oral inflammatory pathologies.

The comparison of antioxidant capacity and cytotoxic, anticarcinogenic,and genotoxic effects of Fe@Au nanosphere magnetic nanoparticles

Magnetic gold nanoparticles are used in various biomedical, biochemistry, and biotechnology applications due to their controllable size distribution, long-term stability, reduced toxicity, and biocompatibility. Different coating materials, such as proteins, carbohydrates, lipids, and polyphenols, are applied to enhance the biocompatibility of nanoparticles. In this study, the effects of surface coatings of core-shell structured Fe@Au nanosphere magnetic nanoparticles with regard to antioxidant capacity and cytotoxic, anticarcinogenic, and genotoxic properties were investigated. The obtained results demonstrated that avidin-coated Fe@Au nanospheres had higher antioxidant capacities than uncoated nanospheres. Neither avidin-coated nor uncoated nanoparticles had a cytotoxic effect on normal cells (human gingival fibroblast cell line, HGF-1). In addition, they had anticarcinogenic effects on human cervical carcinoma (HeLa), human breast adenocarcinoma (MCF-7), and human colorectal adenocarcinoma (CCL-221). The genotoxic effects of nanoparticles were also evaluated with DNA tail damage ratio.

EFFECTS OF LIGHT-EMITTING DIODE (LED 640NM) ON HUMAN GINGIVAL FIBROBLASTS: A COMPARATIVE IN VITRO STUDY

The light-emitting diodes (LEDs) have been applied in oral surgery for tissue stimulation and wound healing. Several Authors have highlighted that fibroblasts subjected to phototherapy have an increased viability, proliferation, biomodulation of inflammatory cytokines and genes expression. It remains to be determined which are the best irradiation parameters (energy, wavelength, power) for each type of cell in order to obtain the best bio-stimulation. The aim of this study was to investigate the effects of LED irradiation on primary human gingival fibroblast cells (HGF) on DSP, ELN, HAS1, ELANE, HYAL1, RPL13 genes activation using Real Time PCR. These genes activation is directly connected with elastin protein production and HGF functionality. Human gingival tissue biopsies were obtained from three healthy patients during extraction of teeth. The gingival pieces were fragmented with a scalpel and transferred in culture dishes for allow the cells growth. Human gingival fibroblasts at the second passage were seeded on multiple 6-well plates and were stimulated with three different light-emitting diodes (LEDs) fixture. After irradiation, the cells were trypsinized, harvested and lysed for RNA extraction. Genes expression was quantified using Real Time PCR. We didn't found significant differences in genes activation of HGF of the three different LEDs. The LED irradiation seems to be directly correlated with the elastin and hyaluronoglucosaminidase 1 genes activation that are directly connected with proteins production and HGF functionality. HGF show an increased deposition of elastin as well as enhanced expression of collagen type I, which is the main protein related to the synthesis and of the collagen-rich matrix.

Cationic Amphiphilic Polymers with Antimicrobial Activity for Oral Care Applications: Eradication of S. mutans Biofilm.

The antibacterial and antibiofilm activities of cationic amphiphilic methacrylate polymers against cariogenic bacterium S. mutans were investigated. Cationic homopolymer PE0 and copolymer PE31 containing 31 mol % of ethyl methacrylate were synthesized by reversible addition-fragmentation chain transfer polymerization. These polymers displayed bactericidal activity toward S. mutans and prevented biofilm formation by killing the planktonic bacteria. At a concentration of 1000 μg/mL when incubated for 2 h the polymers reduced >80% of biofilm biomass. When the polymer assay solution with the biofilm was vigorously mixed using a pipet for 30 s, >50% of biofilm mass was removed at a polymer concentration of 250 μg/mL. Chlorhexidine and a cationic surfactant failed to reduce the biofilm mass at the same concentration. PE0 was the most effective in removing biofilm and did not show any significant cytotoxicity to human gingival fibroblast and periodontal ligament stem cells when incubated for 10 min.

Hydrogen-rich water achieves cytoprotection from oxidative stress injury in human gingival fibroblasts in culture or 3D-tissue equivalents, and wound-healing promotion, together with ROS-scavenging and relief from glutathione diminishment.

The aim of the present study is to investigate protective effects of hydrogen-rich water (HW) against reactive oxygen species (ROS)-induced cellular harmful events and cell death in human gingival fibroblasts (HGF) and three-dimensional (3D-) gingival tissue equivalents. HW was prepared with a magnesium stick in 600-mL double distilled water (DDW) overnight. Dissolved hydrogen was about 1460±50μg/L versus approximately 1600μg/L for the saturated hydrogen. Under cell-free conditions, HW, dose-dependently, significantly scavenged peroxyl radicals (ROO·) derived from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH). Extract from HW-treated HGF cells scavenged ROO· more markedly than that from DDW-treated cells, suggesting that HW can increase the intracellular antioxidant capacity. Hydrogen peroxide dose-dependently increased the intracellular ROS generation, which was significantly repressed by HW, both in the cytoplasm and nuclei. LIVE/DEAD staining and our original cell viability dye-extraction assay showed that HW significantly protected HGF cells from hydrogen peroxide-induced cell death. Hydrogen peroxide also diminished the contents of intracellular glutathione, which were appreciably relieved by HW-pretreatment. Additionally, HW noticeably prevented cumene hydroperoxide-induced generation of cellular ROS in epidermis parts of 3D-gingival equivalents. The in vitro scratch assay showed that HW was able to diminish physical injury-induced ROS generation and promote wound healing in HGF cell monolayer sheets. In summary, HW was able to increase intracellular antioxidative capacity and to protect cells and tissue from oxidative damage. Thus, HW might be used for prevention/treatment of oxidative stress-related diseases.

Ex vivo nonviral gene delivery of μ-opioid receptor to attenuate cancer-induced pain.

Virus-mediated gene delivery shows promise for the treatment of chronic pain. However, viral vectors have cytotoxicity. To avoid toxicities and limitations of virus-mediated gene delivery, we developed a novel nonviral hybrid vector: HIV-1 Tat peptide sequence modified with histidine and cysteine residues combined with a cationic lipid. The vector has high transfection efficiency with little cytotoxicity in cancer cell lines including HSC-3 (human tongue squamous cell carcinoma) and exhibits differential expression in HSC-3 (∼45-fold) relative to HGF-1 (human gingival fibroblasts) cells. We used the nonviral vector to transfect cancer with OPRM1, the μ-opioid receptor gene, as a novel method for treating cancer-induced pain. After HSC-3 cells were transfected with OPRM1, a cancer mouse model was created by inoculating the transfected HSC-3 cells into the hind paw or tongue of athymic mice to determine the analgesic potential of OPRM1 transfection. Mice with HSC-3 tumors expressing OPRM1 demonstrated significant antinociception compared with control mice. The effect was reversible with local naloxone administration. We quantified β-endorphin secretion from HSC-3 cells and showed that HSC-3 cells transfected with OPRM1 secreted significantly more β-endorphin than control HSC-3 cells. These findings indicate that nonviral delivery of the OPRM1 gene targeted to the cancer microenvironment has an analgesic effect in a preclinical cancer model, and nonviral gene delivery is a potential treatment for cancer pain.

Antimicrobial and cytotoxic activities of &lt;i&gt;Barleria prionitis&lt;/i&gt; and &lt;i&gt;Barleria grandiflora&lt;/i&gt;: A comparative study

&lt;p class="Abstract"&gt;Leaf juices as well as leaves of &lt;em&gt;Barleria prionitis&lt;/em&gt; and &lt;em&gt;Barleria grandiflora&lt;/em&gt; are being used by rural people across various regions of India in the treatment of oral ailments such as dental troubles, gum ailments, pyorrhoea, dental carries and mouth ulcers. Zone of inhibition and MIC values obtained for all the extracts suggest ethanolic extract of the herbs were more antimicrobial when compared to the aqueous extract. Results of biofilm suppression were found statistically significant (p&amp;lt;0.05) when compared to control. Cytotoxicity was evaluated by MTT assay on human gingival fibroblast and human dermal fibroblast cell lines for ethanolic extract of the herbs. CTC&lt;sub&gt;50&lt;/sub&gt; value was found to be more than 1,000 µg/mL for ethanolic extracts of both herbs. Chlorhexidine was found to be more cytotoxic with CTC&lt;sub&gt;50 &lt;/sub&gt;value of 12.5–25 µg/mL. Ethanolic extract of &lt;em&gt;B. prionitis&lt;/em&gt; and &lt;em&gt;B. grandiflora&lt;/em&gt; found significantly cytotoxic (p&amp;lt;0.05) in comparison with control.&lt;/p&gt;&lt;p class="Abstract"&gt;&lt;strong&gt;Video Clip:&lt;/strong&gt;&lt;/p&gt;&lt;p class="Abstract"&gt;&lt;a href="https://youtube.com/v/DNByJJQT3RY"&gt;Cytotoxicity assay:&lt;/a&gt; 18 min 45 sec&lt;/p&gt;&lt;p&gt; &lt;/p&gt;

Cytotoxicity Evaluation of Two Bis-Acryl Composite Resins Using Human Gingival Fibroblasts.

Bis-acryl resins are used for temporary dental restorations and have shown advantages over other materials. The aim of this work was to evaluate the in vitro cytotoxicity of two bis-acryl composite resins (Protemp 4 and Luxatemp Star), obtained at 1, 7 and 40 days after mixing the resin components, using a standardized assay employing human primary cells closely related to oral tissues. Human gingival fibroblast cell cultures were exposed for 24 h to either bis-acryl composite resins, polystyrene beads (negative control) and latex (positive control) extracts obtained after incubation by the different periods, at 37 °C under 5% CO2. Cell viability was evaluated using a multiparametric procedure involving sequential assessment (using the same cells) of mitochondrial activity (XTT assay), membrane integrity (neutral red test) and total cell density (crystal violet dye exclusion test). The cells exposed to the resin extracts showed cell viability indexes exceeding 75% after 24 h. Even when cells were exposed to extracts prepared with longer conditioning times, the bis-acryl composite resins showed no significant cytotoxic effects (p>0.05), compared to the control group or in relation to the first 24 h of contact with the products. There were no differences among the results obtained for the bis-acryl composite resins evaluated 24 h, 7 days and 40 days after mixing. It may be concluded that the bis-acryl resins Protemp 4 and Luxatemp Star were cytocompatible with human gingival fibroblasts, suggesting that both materials are suitable for use in contact with human tissues.

Design and fabrication of novel chitin hydrogel/chitosan/nano diopside composite scaffolds for tissue engineering

In this research, novel porous composite scaffolds consisting of chitin, chitosan and nano diopside powder were prepared using the freeze-drying method. The prepared nanocomposite scaffolds were characterized by SEM, XRD, BET, TGA and FT-IR techniques. In addition, swelling, degradation and biomineralization capability, cell viability and cell attachment of the composite scaffolds were evaluated. The results indicated better swelling and degradation properties of such scaffolds their ability to become bioactive. Cytocompatibility of the scaffolds were assessed by MTT assay and cell attachment studies using Human Gingival Fibroblast cells. Cell viability studies demonstrated no sign of toxicity and cells were found to be attached to the pore walls within the scaffolds. These results suggested that the developed composite scaffolds could be a potential candidate for tissue engineering.

Modulation of stromal cell-derived factor 1 alpha (SDF-1\u03b1) and its receptor CXCR4 in Porphyromonas gingivalis-induced periodontal inflammation

BackgroundThe production of chemokines by tissue resident cells during inflammation is considered one of the main mechanisms involved in the formation of inflammatory infiltrates. Fibroblasts are the main resident cell type in gingival and periodontal ligament tissues, and their ability to produce chemokine stromal cell-derived factor 1 alpha (SDF-1α) and its receptor CXCR4 under stimulation by gram negative bacteria, Porphyromonas gingivalis, commonly found in periodontal infections was investigated.MethodsWestern blots were used to assess SDF-1α and CXCR4 protein expression levels in human gingival fibroblast cells (HGF-1) induced by Lipopolysaccharide (LPS) from P. gingivalis in the presence or absence of LY294002, a highly selective inhibitor of PI-3K/Akt. RT-PCR and quantitative Real-time PCR was performed using gingival mRNAs from periodontitis patients. Immunohistochemistry was performed to analyze the expression and subcellular localization of SDF-1α and CXCR4, together with NF-kβ phosphorylation, in specimens from patients with periodontitis and in an experimental rat periodontitis model.ResultsWe found that P. gingivalis LPS up-regulated SDF-1α and CXCR4 protein levels and elevated phosphorylation of the SDF-1α-responsive NF-kβ and Akt at 24 h in HGF-1 cells. SDF-1α and CXCR4 mRNA and protein expression levels were high in all patients with periodontitis. In the P. gingivalis-induced rat experimental periodontitis model, SDF-1α and CXCR4 immunoreactivity was higher in gingival and periodontal ligament tissues compared to the control.ConclusionOur data showed that PI-3K/Akt is an upstream participant in the P. gingivalis LPS-mediated induction of SDF-1α. Taken together, these results suggest that the chemokine SDF-1α and its receptor CXCR4 contribute to P. gingivalis-induced periodontal inflammation.

Biocompatibility evaluation of orthodontic composite by real-time cell analysis.

The aim of this study was to evaluate the cytotoxic effects of three different light-cured orthodontic composites. Light Bond (Reliance orthodontic products), Grengloo (Ormco corporation), and Kurasper F (Kuraray Europe GmbH) were selected for the experiment. Specimens were prepared according to the manufacturers' instructions, measuring 5 mm in diameter and 2 mm in thickness. Fibroblast cells were obtained from healthy gingival connective tissues. The composite cylinders were incubated in Dulbecco's modified Eagle's culture medium for 72 h according to ISO 10993-5 standards. The xCELLigence method was used to evaluate fibroblast cell vitality. After seeding 200 mL of the cell suspensions into the wells (20,000 cells/well) of the E-plate 96, gingival fibroblasts were treated with bioactive components released by the orthodontic composite materials and monitored every 15 min for 121 h. There were no significant differences between the human gingival fibroblast (HGF) cell indexes of the control and all testing groups (p > 0.05) at 24 and 48 h. Light Bond demonstrated statistically significant decrease in HGF index (p < 0.05) at 72 h, but there was no significant difference among the Kurasper F, Grengloo, and untreated control groups (p > 0.05). Light Bond (p < 0.001) and Grengloo (p < 0.05) groups had lower HGF cell index values when compared to untreated control group, but Kurasper F demonstrated no significant differences between the control groups at 96 h (p > 0.05). Orthodontic composite materials include biologically active components and may change oral tissue. So, biocompatible orthodontic bonding composites should be used.

Evaluation of antimicrobial photodynamic therapy with indocyanine green and curcumin on human gingival fibroblast cells: An in vitro photocytotoxicity investigation

Recent investigations have suggested that antimicrobial photodynamic therapy (aPDT) can be an alternative treatment for the management of periodontal infections. However, currently there is very limited data regarding the photocytotoxicity of this method on human gingival fibroblast (HuGu) cells. AimThe in vitro optimal concentrations of indocyanine green (ICG) and curcumin as photosensitizers (PSs) and the irradiation time of diode laser emission were evaluated by assessing the photocytotoxicity of the treatment on HuGu cells. Materials and methodMonolayers of HuGu cells were incubated with various final concentrations of ICG (500, 750, 1000, 1250, 1500, 1750, and 2000μg/ml) and curcumin (3, 4, 5, 10, and 20mM). Three exposure times of the diode laser (30s, 60s, and 2×30s irradiation with an interval of 1min between each) and one of exposure time of 5min for LED were tested; cell viability was determined using neutral red assay. Chlorhexidine (CHX) as a gold standard antimicrobial agent for periodontal disease was considered as a control group. ResultsICG and curcumin significantly reduced HuGu cell viability at concentrations below 1000μg/ml and 10mM, respectively (P<0.01). Cytotoxicity was higher when the cells were treated for 2×30s irradiation with an interval of 1min and then again exposed to the laser for 30s (2% and 0.1%). CHX demonstrated no significant reduction in HuGu cell survival. ConclusionPhotocytotoxicity is influenced by PS concentration, exposure time of PS, and time of irradiation. High doses of ICG and curcumin with lowest exposure time of light source and without cytotoxic effects may be an effective strategy for aPDT as an alternative treatment for periodontal disease.

Uniform, fast, high concentration delivery of bone marrow stromal cells and gingival fibroblasts by gas-brushing

Delivering multipotent stem cells in adequate milieu directly to site of injury is important for successful tissue regeneration. Accelerated wound healing has been reported in recent publications when large area wounds were treated with sprayed cells. However, little is known about the basic physics of cell spraying in hydrogel free medium, causes of concomitant cell damage, and whether it can be applied for craniofacial tissue regeneration. To answer some of these questions, a simple co-axial CO2 gas operated brush (g-brush) was used to deliver human bone marrow stromal (hBMSC) and mouse gingival fibroblast (ESK-1) cells at low gas flow rates and in a small volume of medium. Live/dead, metabolic activity (MTS-1) and total DNA quantification assays were utilized to evaluate cell cultures after g-brush and typical pipette deposition. Viable hBMSC and ESK-1 cells were uniformly delivered at higher cell concentrations (≥60 000), in a short period of time (15 and 9 s), and in a small fluid volume (0.25 ml). Significantly, after the hBMSC were g-brushed and cultured up to 21 d, the cell multipotency was found to be preserved and comparable to the pipetted cells. Fluid dynamic calculations highlighted importance of low interfacial shear stress (<35 Pa) and gas velocity (<160 m s−1) in delivering concentrated, viable cells. Results of this research support applicability of the g-brushing technique for the quick, large number delivery of viable cells for tissue regeneration applications.

Comparison of liposomal and 2-hydroxypropyl-β-cyclodextrin–lidocaine on cell viability and inflammatory response in human keratinocytes and gingival fibroblasts

The aim of this study was to observe the effect multilamellar liposomes (MLV) and 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) in the in-vitro effects of lidocaine in cell viability, pro-inflammatory cytokines and prostaglandin E2 release of both human keratinocytes (HaCaT) and gingival fibroblasts (HGF) cells. HaCaT and HGF cells were exposed to lidocaine 100-1 μm in plain, MLV and HP-β-CD formulations for 6 h or 24 h. The formulation effects in cell viability were measured by XTT assay and by fluorescent labelling. Cytokines (IL-8, IL-6 and TNF-α) and PGE2 release were quantified by ELISA. MLV and HP-β-CD formulations did not affect the HaCaT viability, which was significantly decreased by plain lidocaine after 24 h of exposure. Both drug carriers increased all cytokines released by HGF after 24-h exposure, and none of the carriers was able to reduce the PGE2 release induced by lidocaine. The effect of drug carrier in the lidocaine effects was dependent on the cell type, concentration and time of exposure. MLV and HP-β-CD showed benefits in improving cell viability; however, both of them showed a tendency to increase cytokine release when compared to the plain solution.