• Journal of Periodontal and Implant Science
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• v.34 no.1
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• pp.205-221
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• 2004

Periodontal ligament(PDL) cells have been known as playing an important roles in periodontal regeneration and gingival fibroblasts are also important to periodontal regeneration by forming connective tissue attachment. There were rare studies about the gene expression patterns of PDL cells and gingival fibroblasts, therefore in this study, we tried cDNA microarray-based gene expression monitoring to explain the functional differences of PDL cells and gingival fibroblasts in vivo and to confirm the characteristics of PDL cells. Total RNA were extracted from PDL cells and gingival fibroblasts of same person and same passages, and mRNA were isolated from the total RNA using Oligotex mRNA midi kit(Qiagen) and then fluorescent cDNA probe were prepared. And microarray hybridization were performed. The gene expression patterns of PDL cells and gingival fibroblasts were quite different. About 400 genes were expressed more highly in the PDL cells than gingival fibroblasts and about 300 genes were more highly expressed in the gingival fibroblasts than PDL cells. Compared growth factor- and growth factor receptor-related gene expression patterns of PDL cells with gingival fibroblasts, IGF-2, IGF-2 associated protein, nerve growth factor, placental bone morphogenic protein, neuron-specific growth- associated protein, FGF receptor, EGF receptor-related gene and PDGF receptor were more highly expressed in the PDL cells than gingival fibroblasts. The results of collagen gene expression patterns showed that collagen type I, type III, type VI and type VII were more highly expressed in the PDL cells than gingival fibroblasts, and in the gingival fibroblasts collagen type V, XII were more highly expressed than PDL cells. The results of osteoblast-related gene expression patterns showed that osteoblast specific cysteine-rich protein were more highly expressed in the PDL cells than gingival fibroblasts. The results of cytoskeletal proteins gene expression patterns showed that a-smooth muscle actin, actin binding protein, smooth muscle myosin heavy chain homolog and myosin light chain were more highly expressed in the PDL cells than gingival fibrobalsts, and ${\beta}-actin$, actin-capping protein(${\beta}$ subunit), actin- related protein Arp3(ARP) and myosin class I(myh-1c) were more highly expressed in the gingival fibroblasts than PDL cells. Osteoprotegerin/osteoclastogenesis inhibitory factor(OPG/OCIF) was more highly expressed in the PDL cells than gingival fibroblasts. According to the results of this study, PDL cells and gingival fibroblasts were quite different gene expression patterns though they are the fibroblast which have similar shape. Therefore PDL cells & gingival fibroblasts are heterogeneous populations which represent distinct characteristics. If more studies about genes that were differently expressed in each PDL cells & gingival fibroblasts would be performed in the future, it would be expected that the characteristics of PDL cells would be more clear.

• Journal of Korean society of Dental Hygiene
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• v.12 no.4
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• pp.733-739
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• 2012

Objectives : To Compare the degree of survival rate of gingival fibroblasts, which is concerned with teeth adherence based on the type of avulsed tooth's storage solution. Methods : Different media gingival fibroblasts were stored in Dulbecco's modified Eagle's medium(DMEM), Hank's balanced salt solution(HBSS), milk, saline, and green tea in for 1, 2, 3 hours. And, MTT assay was conducted to compare survival rate of human gingival fibroblasts. Results : 1. The survival rate of gingival fibroblasts in DMEM and HBSS was higher than thoes in other storage media( Milk> Saline> Green tea). 2. The survival rate of gingival fibroblasts in milk, saline and green tea decreased as time passed. 3. Because of low osmotic pressure, green tea showed decrease of survival rate of gingival fibroblasts. Conclusion : DMEM and HBSS were the most effective storage media for gingival fibroblast. Among milk, saline, green tea, milk is most effective storage media for keeping gingival fibroblasts. Milk is recommended for storage media of avulsed tooth for keeping viability of cells.

• Journal of Periodontal and Implant Science
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• v.26 no.3
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• pp.681-688
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• 1996

Periodontal regeneration requires the migration and proliferation of gingival fibroblasts and periodontal ligament cells. These cellular events are influenced and regulated by growth factors and some drugs. The purpose of this study is to examine the effect of Rhizoma coptidis and Centella asiatica extracts on human gingival fibroblasts. Gingival fibroblasts were primarily cultured from extracted premolar with non-periodontal diseases. Cells were cultured with ${\alpha}-MEM$ at $37^{\circ}C$, 5% $CO_2$, 100% humidity incubator for 2 or 3 days, as a measure of cell proliferation potential, it was examined that the DNA synthesis using $[^3H]-thyrnidine$ incorporation, the cell numbers (with or without dye), and cell viabilities. Rhizoma coptidis is increased the proliferation of gingival fibroblasts at concentration of $10^{-9}g/ml$, but Centella asiatica is decreased the proliferation at all concentrations. This study demonstrated that Rhizoma coptidis is a potential mitogen for human gingival fibroblasts in vitro, and we can expect the usefulness of this drug in periodontal regeneration.

• Journal of Periodontal and Implant Science
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• v.26 no.1
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• pp.176-187
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• 1996

The ability of fibroblasts attached to teeth is paramount important in reestablishing the lost connective tissue attachment after periodontal therapy. The migration and proliferation of periodontal ligament cells are desired goal of periodontal regeneration therapy. PDGF is well known to regulate the cell activity of mesenchymal origin cell. Tobacco contains a complex mixture of substance including nicotine, various nitrosamines, trace elements, and variety of poorly characterized substances. Human gingival fibroblasts and periodontal ligament cells were cultured from extracted tooth for non-periodontal reason. Cultured human gingival fibroblasts and periodontal ligament cells in vitro were treated with PDGF, nicotine in time dependent manner. Cellular activities were determined by MTT assay. The purpose of this study was to determine the effects of Nicotine and PDGF, respectively and the effect of PDGF presence of nicotine on human gingival fibroblasts and periodontal ligament cells. The results were as follows : 1. In the cell activities of human gingival fibroblasts and periodontal ligament cells were similar or decreased to control value at 1st day. At 2nd day, cellular activities of both group were increased to control value. At 3rd day, cellular activities of both group were returned to the control value. 2. In the cell activities of PDGF on human gingival fibroblasts and periodontal ligament cells, cell activities significantly increase from control group on periodontal ligament cells compared to gingival fibroblast group at 3rd day. 3. In the cell activities of PDGF and nicotine combined application on human gingival fibroblasts and periodontal ligament cells, it seems likely that the nicotinic effect of gingival fibroblasts were higher than periodontal ligament cells and the PDGF effect of periodontal ligament cells were higher than gingival fibroblasts. This results suggested that PDGF might stimulate the selective growth on periodontal ligament cells.

• Journal of Periodontal and Implant Science
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• v.31 no.1
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• pp.135-148
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• 2001

Gingival fibroblasts are major cellular component of gingiva. However, the molecular mechanisms of senescence of human gingival fibroblasts are unknown. Human fibroblasts undergo replicative senescence in vitro after a limited number of population doublings. A reduced rate of proliferation is a prominent phenomenon observed in senescent fibroblasts. This phenomenon is controled by cell cycle regulatory proteins. The purpose of present study was to investigate the effect of replicative senescence on cell cycle progression and to find out its molecular mechanisms in human gingival fibroblasts. Replicative senescence of gingival fibroblasts were induced by subsequent cultures that were repeated up to 18 passage. In the present study, I examined change of cell proliferation, cell activity, cell viability and cell cycle progression during the replicative process. Also, I examined expression of cell cycle regulatory proteins which was estimated by western blot analysis. Cell proliferation, cell activity and cell viability of gingival fibroblasts were notably decreased with increase of population doubling level(PDL). S phase was decreased and G1 phase was increased with increase of PDL. Western blot analysis showed that levels of P16, p21 and p53 of senescent gingival fibroblasts(PDL41, PDL58) were higher than young fibroblasts(PDL27) and cdk4 were lower than young fibroblasts(PDL27). In conclusion, these results suggest that proliferative function of human gingival fibroblasts may be decreased by replicative senescence and its molecular mechanisms may be activatied with p16, p21, p53 and pRB, and repressed wtih cdk4.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.28 no.3
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• pp.169-174
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• 2002

Purpose: Irradiation in the oral cancer patients causes early and late complications such as intraoral mucositis and fibrosis, with a various expression of GM-CSF and TGF-${\beta}1$. The purpose of this study was to investigate the production of GM-CSF and TGF-${\beta}1$ by the irradiated human gingival fibroblasts cultivated with lipopolysaccharide. Materials and Methods: Irradiated (total dose, 60 Gy) human gingival fibroblasts were incubated with LPS. Culture supernatants that were collected at 24, 48, and 72 hours were assessed for GM-CSF and TGF-${\beta}1$ by enzyme-linked immunosorbent assay. Results: 1. GM-CSF production in nomal gingival fibroblasts was increased with incubation time, but decreased with incubation time in irradiated gingival fibroblasts. GM-CSF production in both normal and irradiated gingival fibroblasts induced with LPS was higher than the control. 2. TGF-${\beta}1$ production in normal gingival fibroblasts was decreased after 24 hours, but, it was increased until 48 hours in irradiated gingival fibroblasts. TGF-${\beta}1$ production in normal gingival fibroblasts exposed with LPS was higher than the control. Conversely, It was lower than the control in irradiated gingival fibroblasts exposed with LPS. Conclusion: This indicates that irradiation in gingival fibroblasts may play an important role in radiation-induced intraoral mucositis and fibrosis. However, LPS decreases the production of TGF-${\beta}1$ in the irradiated gingival fibroblasts.

• Journal of Periodontal and Implant Science
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• v.47 no.2
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• pp.66-76
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• 2017

Purpose: Oral wound healing requires gingival fibroblasts to respond to local growth factors. Epigenetic silencing through DNA methylation can potentially decrease the responsiveness of gingival fibroblasts to local growth factors. In this study, our aim was to determine whether the inhibition of DNA methylation sensitized gingival fibroblasts to transforming growth factor-${\beta}1$ (TGF-${\beta}1$). Methods: Gingival fibroblasts were exposed to 5-aza-2'-deoxycytidine (5-aza), a clinically approved demethylating agent, before stimulation with TGF-${\beta}1$. Gene expression changes were evaluated using quantitative polymerase chain reaction (PCR) analysis. DNA methylation was detected by methylation-sensitive restriction enzymes and PCR amplification. Results: We found that 5-aza enhanced TGF-${\beta}1$-induced interleukin-11 (IL11) expression in gingival fibroblasts 2.37-fold (P=0.008). 5-aza had no significant effects on the expression of proteoglycan 4 (PRG4) and NADPH oxidase 4 (NOX4). Consistent with this, 5-aza caused demethylation of the IL11 gene commonly next to a guanosine (CpG) island in gingival fibroblasts. The TGF-${\beta}$ type I receptor kinase inhibitor SB431542 impeded the changes in IL11 expression, indicating that the effects of 5-aza require TGF-${\beta}$ signaling. 5-aza moderately increased the expression of TGF-${\beta}$ type II receptor (1.40-fold; P=0.009), possibly enhancing the responsiveness of fibroblasts to TGF-${\beta}1$. As part of the feedback response, 5-aza increased the expression of the DNA methyltransferases 1 (DNMT1) (P=0.005) and DNMT3B (P=0.002), which are enzymes responsible for gene methylation. Conclusions: These in vitro data suggest that the inhibition of DNA methylation by 5-aza supports TGF-${\beta}$-induced IL11 expression in gingival fibroblasts.

• Journal of Periodontal and Implant Science
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• v.25 no.2
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• pp.181-191
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• 1995

The ability of fibroblasts attach to teeth is of paramount imporance in re-establishing the lost connective tissue attachment after periodontal therapy. Tobacco contains a complex mixture of substances including nicotine. various nitrousamines, trace elements. and a variety of poorly characterized substances. The effects of nicotine on fibroblasts have reported an altered morphology and attachment of fibroblasts to substrates and disturbances in protein synthesis and secretion. This study examined the effect of nicotine, a major component of the particulate phase of tobacco smoke, on human gingival fibroblasts and periodontal ligament cells attachment to tissue culture surfaces and cellular activity of human gingival fibroblasts and periodontal ligament cells. Pooled human gingival fibroblasts made from extraction of 3rd molar were utilized between passage 4 and 5 and plated in 96 well plate at 20,000 cells per well. Cell number were determined using 3-(4,5-dimethylthiazole-2-y)2,5-diphenyltetrazolium bromide(MTI) , which is reflection of mitochondrial dehydrogenase activity. The concentration of nicotine used were 0.025, 0.05, 0.1, 0.2 and $0.4{\mu}M$, the average serum concentration for a smoker being approximately $0.1{\mu}M$. The results were as follows : 1. Attachment effects of nicotine on human gingival fibroblasts and periodontal ligament cells Excepts of $0.4{\mu}M$, the effects on attachment with increasing numbers of cells attaching with increasing nicotine concentrations, compared to control group. But over the 60min, return to control value. 2. The effect of cellular activity on human gingival fibroblasts and periodontal ligament cells. The cellular activity of human gingival fibroblasts and periodontal ligament cells were similar or decrease to control value at 1st incubation day. At 2nd incubation day, 0.05, 0.1, 0.2, $0.4{\mu}M$ concentrations were statistically different from control value on gingival fibroblasts group. But at 3rd incubation day, cellular activities of all experimental group were significantly decrease than control group.

• Journal of Periodontal and Implant Science
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• v.32 no.1
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• pp.235-247
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• 2002

The aim of this study was to evaluate the effects of chitosan coating on the attachment, proliferation, functional and morphological change of human gingival fibroblasts. Primary culture of human gingival fibroblasts were grown in Dulbecco's modified Eagle's medium with 10% fetal bovine serum and 1% antibiotics. In experimental group, cells were inoculated in the multiwell plates coated with chitosan in concentration of 0.02, 0.2, and 2 mg/ml. Cell counting and MTT assay were done after 0.5, 1.5, 3, 6 and 24 hours of incubation to evaluate the cell attachment, and then after 2 and 7 days of culture to evaluate the cell proliferation. The alkaline phosphatase activity was measured after 4 and 7 days of culture and the ability to produce mineralized nodules was evaluated after 21 days of culture. The results were as follows : The morphology of cells on the chitosan-coated well was round or spheric. Round cells were aggregated since 6 hours of culture and showed nodule-like appearance after 24 hours of culture and did not achieved confluency at 7 days. The attachment of gingival fibroblasts was inhibited by chitosan coating with a tendency of dose dependent pattern. But, cellular activity of unit cell was higher than control. The proliferation of gingival fibroblasts was inhibited by chitosan coating at 2 mg/ml(P<0.01), while the cell proliferation at 0.02, 0.2 $mg/m{\ell}$ was comparable to the control well. Total alkaline phosphatase activity was inhibited by chitosan coating and decreased in the course of time. While ALP activity of unit cell was the highest at 2mg/ml after 4 days of culture. Finally, gingival fibroblasts produced the mineralized nodule at 2 mg/ml. In summary, the attachment, proliferation, and alkaline phosphatase activity of gingival fibroblasts were influenced differently by the concentration of coated chitosan. From this study, it could be used as the matrix of tissue engineering for gingiva without inhibition on proliferation of gingival fibroblasts using chitosan at the optimal concentration (0.02mg/ml).

• International Journal of Oral Biology
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• v.40 no.1
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• pp.27-33
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• 2015

In the present study, we investigated the effect of staurosporine on the formation of cellular processes in human gingival fibroblasts and rat astrocytes. Staurosporine caused a rapid induction of process formation in human gingival fibroblasts and rat astrocytes in a concentration dependent manner. The process formation of human gingival fibroblasts and rat astrocytes was prevented by the pretreatment with N-acetylcysteine, suggesting that staurosporine-induced ROS production was responsible for the process formation. Colchicine, a microtubule depolymerizing agent, inhibited the staurosporine-induced process formation, whereas cytochalasin D, an actin filament breakdown agent, failed to suppress the formation of cellular processes. This result indicated that polymerization of microtubule, and not actin filament, was responsible for the formation of cellular processes induced by staurosporine. In support of this hypothesis, Western blot analysis was conducted using anti-tubulin antibody, and the results showed that the amount of polymerized microtubule was increased by the treatment with staurosporine while that of depolymerized beta-tubulin in soluble fraction was decreased. These results indicate that staurosporine induces ROS-mediated, microtubule-dependent formation of cellular processes in human gingival fibroblasts and rat astrocytes.