Human Gingival Fibroblasts (human + gingival_fibroblast)

Distribution by Scientific Domains


Selected Abstracts


Biocompatibility of various root canal filling materials ex vivo

INTERNATIONAL ENDODONTIC JOURNAL, Issue 8 2008
R. Scotti
Abstract Aim, To evaluate the biocompatibility of a resin-based endodontic filler (RealSeal) using the indirect cytotoxicity test. Methodology, Human gingival fibroblasts were cultured ex vivo. Pellets of the materials to be tested were incubated for 24, 48, and 72 h at 37 °C under sterile conditions to obtain their eluates. The fibroblasts were exposed to either diluted (50%) or undiluted eluates for 24 h. A culture medium with foetal calf serum was added to the control wells. Cell viability was estimated by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide method. The data concerning cell viability were statistically analyzed using one-way anova test and Bonferroni multiple comparisons test. Results, Eluates obtained after 24 h of incubation with the resin filler did not reduce cellular viability. An increase in cellular viability, as compared with control cells, was observed in the gutta-percha group. The undiluted eluate from the polyether material was cytotoxic, causing an 82 ± 4% decrease in cellular viability. Eluates obtained after 48 h of incubation with the resin filler increased cellular viability, whereas the polyether significantly reduced viability. Gutta-percha did not cause any detectable change. After 72 h of incubation the eluate of the resin filler caused an increase in cellular viability, as did gutta-percha, whereas polyether caused a significant decrease. Conclusions, RealSeal resin filler was nontoxic in this laboratory model. Further investigations are necessary to verify its usefulness in clinical applications. [source]


Role of D1 and E Cyclins in Cell Cycle Progression of Human Fibroblasts Adhering to Cementum Attachment Protein,

JOURNAL OF BONE AND MINERAL RESEARCH, Issue 6 2001
Takayoshi Yokokoji
Abstract Cementum attachment protein (CAP) is a collagenous protein present in the matrix of tooth cementum that mediates preferential attachment of some mesenchymal cell types, and CAP binding capacity is related to mineralizing tissue-forming capacity in culture. We have examined if adhesion to surfaces containing CAP as the only attachment protein permits human fibroblasts to escape G1 arrest and synthesize DNA, and if adhesion to CAP modulates the levels of cyclins D1 and E. Human gingival fibroblasts (HGFs) were serum-starved, trypsinized, and added to plates coated with CAP or bovine serum albumin (BSA). Cells were then exposed to either 10% fetal bovine serum (FBS) or to cementum-derived growth factor (CGF), an insulin-like growth factor I (IGF-I)-like molecule sequestered in tooth cementum, plus epidermal growth factor (EGF). DNA synthesis was measured as [3H]thymidine uptake, and cyclin D1 and E levels were determined by Western analysis. Cyclin E-dependent kinase (Cdk) activity was assessed in terms of H1 kinase activity in immunoprecipitates of cyclin E. Cells adhering to CAP synthesized DNA, whereas on BSA they remained unattached and did not synthesize DNA. Protein levels of cyclin D1 were higher in cells adhering to CAP in the absence and presence of growth factors. Cyclin E levels were not affected by adhesion alone, but they increased in the presence of growth factors. Cyclin E-associated kinase activity was higher in cells adherent on CAP, and it increased further in the presence of growth factors. Our results indicate that adhesion to CAP increases cyclin D1 levels and cyclin E-associated Cdk activity, and that these increases contribute to cell cycle progression. We previously observed that the signaling reactions induced during adhesion are characteristic of the CAP; together these observations indicate that specific matrix components present in the local environment can contribute to recruitment and differentiation of specific cell types for normal homeostasis and wound healing. [source]


Nicotine inhibits myofibroblast differentiation in human gingival fibroblasts

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2005
Yiyu Fang
Abstract Cigarette smoking has been suggested as a risk factor for several periodontal diseases. It has also been found that smokers respond less favorably than non-smokers to periodontal therapy. Previous work in our lab has shown that nicotine inhibits human gingival cell migration. Since myofibroblasts play an important role in wound closure, we asked if nicotine affects gingival wound healing process by regulating myofibroblast differentiation. Human gingival fibroblasts (HGFs) from two patients were cultured in 10% fetal bovine serum cell culture medium. Cells were pretreated with different doses of nicotine (0, 0.01, 0.1, and 1 mM) for 2 h, and then incubated with transforming growth factor beta (TGF-,1) (0, 0.25, 0.5, and 1 ng/ml) with or without nicotine for 30 h. The expression level of ,-smooth muscle actin (,-SMA), a specific marker for myofibroblasts, was analyzed by Western blots, immunocytochemistry, and real-time polymerase chain reaction (real-time PCR). Phosphorylated p38 mitogen-activated protein kinase (Phospho-p38 MAPK) activity was analyzed by Western blots. TGF-,1 induced an increase of ,-SMA protein and mRNA expression, while nicotine (1 mM) inhibited the TGF-,1-induced expression of ,-SMA but not ,-actin. Nicotine treatment down-regulated TGF-,1-induced p38 MAPK phosphorylation. Our results demonstrated for the first time that nicotine inhibits myofibroblast differentiation in human gingival fibroblasts in vitro; supporting the hypothesis that delayed wound healing in smokers may be due to decreased wound contraction by myofibroblasts. © 2005 Wiley-Liss, Inc. [source]


Effect of triclosan on interferon-, production and major histocompatibility complex class II expression in human gingival fibroblasts

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 10 2000
Manal Mustafa
Abstract Background, aims: The effect of triclosan (2,4,4,-trichloro-2,-hydroxyl-diphenyl ether) on the production of interferon-, (IFN-,) and the expression of major histocompatibility complex (MHC) class II antigen was studied in human gingival fibroblasts isolated from 4 individuals. Methods/Results: AII cell lines demonstrated high IFN-, production in 24-h cultures of human gingival fibroblasts stimulated by phytohemagglutinin (PHA) (5 ,g/ml). Human gingival fibroblasts showed a high expression of MHC class II when stimulated with 500 and 1000 pg/ml rIFN-, in 7-day cultures. Treatment of the cells with triclosan (0.5 ,g/ml) reduced both IFN-, production and MHC class II expression in human gingival fibroblast cultures. Similar inhibitory effects on IFN-, production and MHC class II expression were observed when the anti-inflammatory agent dexamethazone (1 ,M) was used. Conclusion: The present study further supports the view that the agent has an anti-inflammatory effect in addition to its antibacterial capacity. [source]


Synergistic enhancement of collagenous protein synthesis by human gingival fibroblasts exposed to nifedipine and interleukin-1-beta in vitro

JOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 1 2000
R. B. Johnson
Abstract: Gingival overgrowth commonly occurs coincident to therapy with calcium channel blockers. The biologic mechanism for this condition is unknown; however, many clinicians suggest that poor oral hygiene may contribute to development of the overgrowth. This study tests the hypothesis that collagenous protein synthesis by gingival fibroblasts is synergistically enhanced when they are exposed to both nifedipine (N) and the pro-inflammatory cytokine, interleukin-1-beta, a cytokine expressed in inflamed gingiva. Human gingival fibroblasts were isolated from biopsies of normal gingiva and cells separated into two groups. Group 1 was exposed to media containing 0, 5, 50, or 500 pg/ml IL-1-beta, or 10,7 M N for 7 days; Group 2 was exposed to those concentrations of IL-1-beta +10,7 M N. [3H]-proline was added to the medium for the final 24 h. Cells and matrix were harvested and radioactivity determined by liquid scintillation analysis. Means (d.p.m./103 cells) were compared by factorial ANOVA and Scheffè comparisons. Collagenous protein synthesis was significantly reduced by 5 pg/ml IL-1-beta +10,7 M N and enhanced by 500 pg/ml IL-1-beta +10,7 M N as compared to N or IL-1-beta alone. Thus, patients may be more susceptible to gingival overgrowth coincident to nifedipine therapy as a result of the synergistic enhancement of connective tissue synthesis by these agents. [source]


Cigarette smoke condensate affects the collagen-degrading ability of human gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 6 2009
W. Zhang
Background and Objective:, Cigarette smoke condensate, the particulate matter of cigarette smoke, is composed of thousands of chemicals, including nicotine. Cigarette smoking is a risk factor for periodontal disease. This study investigated the influence of cigarette smoke condensate on the collagen-degrading ability of human gingival fibroblasts and its mechanism. Material and Methods:, Human gingival fibroblasts were exposed for 72 h to various concentrations of total particulate matter cigarette smoke condensate. Cell proliferation and cytotoxicity were evaluated using water-soluble tetrazolium-1 and lactate dehydrogenase, respectively. The collagen-degrading ability of human gingival fibroblasts was evaluated in collagen-coated six-well plates. Conditioned media and membrane extracts were collected for zymography and western blot analyses of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Results:, Cell proliferation decreased and cytotoxicity increased in human gingival fibroblasts with increasing concentrations of cigarette smoke condensate. Cell proliferation decreased by more than 50% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 200 ,g/mL, and cytotoxicity increased to more than 30% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 400 ,g/mL. Cigarette smoke condensate increased the collagen-degrading ability of human gingival fibroblasts, especially at a concentration of 100 ,g/mL (1.5-fold increase, p < 0.05) compared with the control. Cigarette smoke condensate increased the production of proMMP-1, proMMP-2, MMP-14 and TIMP-1, and decreased the production of TIMP-2, in conditioned media. Furthermore, compared with the control group, cigarette smoke condensate increased the production of MMP-2, MMP-14 and TIMP-2 in membrane extracts, especially at concentrations of 50,100 ,g/mL. Conclusion:, Cigarette smoke condensate affects human gingival fibroblast proliferation and is toxic at total particulate matter cigarette smoke condensate concentrations of , 400 ,g/mL. Cigarette smoke condensate can increase the collagen-degrading ability of human gingival fibroblasts by altering the production and localization of MMPs and TIMPs. [source]


Epidermal growth factor stimulates urokinase-type plasminogen activator expression in human gingival fibroblasts.

JOURNAL OF PERIODONTAL RESEARCH, Issue 6 2004
Possible modulation by genistein, curcumin
Background:, Regulation of the extracellular matrix turnover is a crucial process in wound healing and the progress of periodontal disease. It has been proposed that urokinase-type plasminogen activator (uPA), under the control of growth factors or cytokines, provides the proteolytic potential to the accomplishment of these cellular events. Epidermal growth factor (EGF) is one of the growth factors that has been shown to be active in uPA regulation. Methods:, In this study, we have assessed the effect of EGF on uPA expression in primary cultures of human gingival fibroblasts. We also studied the signaling pathways involved in this process and the role of the dietary phytoestrogens curcumin and genistein as potential modulators of this response. Results:, Human gingival fibroblasts expressed a basal uPA activity, which was inhibited by genistein, but not by curcumin. After treatment with 10 ng/ml EGF, uPA production was strongly stimulated. Exposure to genistein and curcumin inhibited EGF-stimulated urokinase production, although only genistein showed a statistically significant inhibitory response. Using more specific inhibitors, we found that the mitogen-activated extracellular kinase and c-Jun N-terminal kinase (JNK) inhibitors PD98059 and SP600125 also blocked the EGF-dependent stimulatory effect. On the other hand, SB203580, inhibitor of the p38 member of mitogen-activated protein kinase family, did not alter this response. In accordance to these findings, EGF stimulated a potent activation of JNK and a mild activation of extracellular signal-regulated kinases 1/2. Finally, EGF stimulated the phosphorylation of its receptor and tyrphostin (AG1478), curcumin and genistein were able to inhibit this stimulatory effect. Conclusions:, These results indicate that EGF constitutes a strong stimuli on uPA expression in human gingival fibroblasts. Our data also shows that EGF-stimulated uPA production involves the activation of the extracellular signal-regulated kinases 1/2 and JNK signaling pathways and might be modulated by the natural phytoestrogens curcumin and genistein. [source]


Contrasting responses of human gingival and periodontal ligament fibroblasts to bacterial cell-surface components through the CD14/Toll-like receptor system

MOLECULAR ORAL MICROBIOLOGY, Issue 1 2003
J. Hatakeyama
We compared human periodontal ligament fibroblasts with human gingival fibroblasts isolated from the same donor to examine interleukin-8 (IL-8) responses of the cells to Salmonella lipopolysaccharide, a water-soluble peptidoglycan from Staphylococcus epidermidis and the synthetic muramyldipeptide, with special reference to the possible involvement of the CD14/Toll-like receptor (TLR) system of the cells in the responses. Human gingival fibroblasts expressed CD14 on their surfaces and strongly expressed CD14 mRNA, while human periodontal ligament fibroblasts showed considerably lower levels of expression in both respects. Both cells expressed mRNA of TLR-related molecules, i.e. TLR2, TLR4, MD-2 and MyD88, although human periodontal ligament fibroblasts expressed TLR2 more strongly than human gingival fibroblasts. Human gingival fibroblasts exhibited a stronger IL-8 response than human periodontal ligament fibroblasts to lipopolysaccharide, while human periodontal ligament fibroblasts exhibited a response comparable to, or slightly stronger than, that of human gingival fibroblasts to S. epidermidis peptidoglycan and muramyldipeptide. The IL-8 responses of both cells to lipopolysaccharide and S. epidermidis peptidoglycan were completely inhibited by antihuman CD14 monoclonal antibody (MAb). The responses of both cells to lipopolysaccaride were significantly inhibited by antihuman TLR4 MAb, while those to S. epidermidis peptidoglycan were inhibited by antihuman TLR2 MAb. In contrast, muramyldipeptide activated both types of cells in a TLR2- and TLR4-independent manner, although the activities of muramyldipeptide on human gingival fibroblasts, but not human periodontal ligament fibroblasts, were significantly inhibited by anti-CD14 MAb. [source]


Regulation of type I plasminogen activator inhibitor in human gingival fibroblasts with cyclosporine A

ORAL DISEASES, Issue 4 2010
Y-C Ho
Oral Diseases (2010) 16, 396,401 Objectives:, Cyclosporine A (CsA) is used as an immunosuppressive agent and its prominent side effect is the induction of gingival overgrowth. Type I plasminogen activator inhibitor (PAI-1) has shown to play an important role in CsA-induced gingival overgrowth. However, little is known about whether factors can modulate CsA-induced PAI-1 expression. Methods:, Cytotoxicity, reverse transcriptase-polymerase chain reaction, and enzyme-linked immunosorbent assay were used to investigate the effects of Human gingival fibroblasts (HGFs) exposed to CsA. In addition, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, interlukin-1,, tumor necrosis factor-,, mitogen-activated protein kinase kinase (MEK) inhibitor U0126, signal-regulated protein kinase (ERK) inhibitor PD98059 and cell-permeable glutathione precursor N -acetyl- L -cysteine (NAC) were added to test how they modulated the effects of CsA-induced PAI-1 expression. Results:, The concentration of CsA higher than 500 ng ml,1 demonstrated cytotoxicity to HGFs (P < 0.05). Periodontal pathogens as well as proinflammatory cytokines were found to increase the CsA-induced PAI-1 mRNA and protein expression (P < 0.05). Pharmacological agents NAC, U0126, and PD98059 were found to decrease the CsA-induced PAI-1 mRNA and protein expression (P < 0.05). Conclusions:, Cyclosporine A (CsA) may predispose to gingival overgrowth under inflammatory environments. The regulation of PAI-1 expression induced by CsA might be critically related with the intracellular glutathione and the ERK-MAPK pathway. [source]


Capsular polysaccharide from Actinobacillus actinomycetemcomitans inhibits IL-6 and IL-8 production in human gingival fibroblast

JOURNAL OF PERIODONTAL RESEARCH, Issue 2 2003
Yuko Ohguchi
We previously reported that a capsular polysaccharide (CP) from Actinobacillus actinomycetemcomitans Y4 induces bone resorption in a mouse organ culture system and osteoclast formation in mouse bone marrow cultures. However, the effects of A. actinomycetemcomitans Y4 CP on human gingival fibroblasts (HGF) are still unclear. The present study was undertaken to test the hypothesis that A. actinomycetemcomitans Y4 CP alters the production of inflammatory cytokines, such as interleukin-6 (IL-6) and IL-8 by HGF. When HGF were cultured with various concentrations of Y4 CP for 24 h, IL-6 and IL-8 production decreased in a concentration-dependent manner. Y4 CP (100 ,g/ml) suppressed the release of IL-6 from 9.09 ± 0.08 ng/ml to 0.34 ± 0.21 ng/ml (P < 0.01) and IL-8 production decreased from 3.76 ± 0.03 ng/ml to 0.09 ± 0.01 ng/ml (P < 0.01). Y4 CP suppressed 70,80% of the release of IL-6 and IL-8 from HGF stimulated with Y4 lipopolysaccharide (LPS), too. Interestingly, anti- A. actinomycetemcomitans Y4 CP completely inhibited the effect of A. actinomycetemcomitans Y4 CP on IL-6 and IL-8 production from HGF. These results indicate that Y4 CP inhibits the release of IL-6 and IL-8 from HGF, suggesting that A. actinomycetemcomitans Y4 modulates the inflammatory response in periodontitis. Remarkably, this inhibitory effect was reversed by specific anti- A. actinomycetemcomitans Y4 CP suggesting an important relationship between the organism and the humoral host response. [source]


The anti-inflammatory mechanism of 635 nm light-emitting-diode irradiation compared with existing COX inhibitors

LASERS IN SURGERY AND MEDICINE, Issue 7 2007
Wonbong Lim PhD
Abstract Background and Objectives Inhibition of cyclooxygenase (COX) and prostaglandin E2 (PGE2) protects cells against cell injury in specific pathophysiological situations: inflammation and oxidative stress. Although the anti-inflammatory effects have been reported in clinical fields for specific wavelength irradiation during wound healing, the physiological mechanism has not been clarified yet. The aim of the present study is to investigate the anti-inflammatory mechanism of 635 nm light-emitting-diode (LED) irradiation compared with existing COX inhibitors. Study Design/Materials and Methods The present study investigated anti-inflammatory effects of 635 nm irradiation on PGE2 release, COX and phospholipase A2 (PLA2) expression, and reactive oxygen species (ROS) dissociation in arachidonic acid (AA)-treated human gingival fibroblast (hGF). These results were compared with their existing COX inhibitors: indomethacin and ibuprofen. The PGE2 release was measured by enzyme immunoassay, the COX expression was measured by western blot and reverse transcriptase polymerase chain reaction (RT-PCR), and ROS level was measured by flow cytometry, laser scanning confocal microscope and RT-PCR. Results Results showed that 635 nm irradiation and existing COX inhibitors inhibit expression of COX and PGE2 release. Unlike indomethacin and ibuprofen, 635 nm irradiation leads to a decrease of ROS levels and mRNA expression of cytosolic phospholipase A2 (cPLA2) and secretary phospholipase A2 (sPLA2). Conclusion Taken together, 635 nm irradiation, unlike indomethacin and ibuprofen, can directly dissociate the ROS. This inhibits cPLA2, sPLA2, and COX expression, and results in the inhibition of PGE2 release. Thus, we suggest that 635 nm irradiation inhibits PGE2 synthesis like COX inhibitor and appears to be useful as an anti-inflammatory tool. Lesers Surg. Med. 39:614,621, 2007. © 2007 Wiley-Liss, Inc. [source]


Prostaglandin E2 inhibits the proliferation of human gingival fibroblasts via the EP2 receptor and Epac

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2009
Evgeny Weinberg
Abstract Elevated levels of prostaglandins such as PGE2 in inflamed gingiva play a significant role in the tissue destruction caused by periodontitis, partly by targeting local fibroblasts. Only very few studies have shown that PGE2 inhibits the proliferation of a gingival fibroblast (GF) cell line, and we expanded this research by using primary human GFs (hGFs) and looking into the mechanisms of the PGE2 effect. GFs derived from healthy human gingiva were treated with PGE2 and proliferation was assessed by measuring cell number and DNA synthesis and potential signaling pathways were investigated using selective activators or inhibitors. PGE2 inhibited the proliferation of hGFs dose-dependently. The effect was mimicked by forskolin (adenylate cyclase stimulator) and augmented by IBMX (a cAMP-breakdown inhibitor), pointing to involvement of cAMP. Indeed, PGE2 and forskolin induced cAMP generation in these cells. Using selective EP receptor agonists we found that the anti-proliferative effect of PGE2 is mediated via the EP2 receptor (which is coupled to adenylate cyclase activation). We also found that the effect of PGE2 involved activation of Epac (exchange protein directly activated by cAMP), an intracellular cAMP sensor, and not PKA. While serum increased the amount of phospho-ERK in hGFs by ,300%, PGE2 decreased it by ,50%. Finally, the PGE2 effect does not require endogenous production of prostaglandins since it was not abrogated by two COX-inhibitors. In conclusion, in human gingival fibroblasts PGE2 activates the EP2,cAMP,Epac pathway, reducing ERK phosphorylation and inhibiting proliferation. This effect could hamper periodontal healing and provide further insights into the pathogenesis of inflammatory periodontal disease. J. Cell. Biochem. 108: 207,215, 2009. © 2009 Wiley-Liss, Inc. [source]


Nicotine inhibits myofibroblast differentiation in human gingival fibroblasts

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2005
Yiyu Fang
Abstract Cigarette smoking has been suggested as a risk factor for several periodontal diseases. It has also been found that smokers respond less favorably than non-smokers to periodontal therapy. Previous work in our lab has shown that nicotine inhibits human gingival cell migration. Since myofibroblasts play an important role in wound closure, we asked if nicotine affects gingival wound healing process by regulating myofibroblast differentiation. Human gingival fibroblasts (HGFs) from two patients were cultured in 10% fetal bovine serum cell culture medium. Cells were pretreated with different doses of nicotine (0, 0.01, 0.1, and 1 mM) for 2 h, and then incubated with transforming growth factor beta (TGF-,1) (0, 0.25, 0.5, and 1 ng/ml) with or without nicotine for 30 h. The expression level of ,-smooth muscle actin (,-SMA), a specific marker for myofibroblasts, was analyzed by Western blots, immunocytochemistry, and real-time polymerase chain reaction (real-time PCR). Phosphorylated p38 mitogen-activated protein kinase (Phospho-p38 MAPK) activity was analyzed by Western blots. TGF-,1 induced an increase of ,-SMA protein and mRNA expression, while nicotine (1 mM) inhibited the TGF-,1-induced expression of ,-SMA but not ,-actin. Nicotine treatment down-regulated TGF-,1-induced p38 MAPK phosphorylation. Our results demonstrated for the first time that nicotine inhibits myofibroblast differentiation in human gingival fibroblasts in vitro; supporting the hypothesis that delayed wound healing in smokers may be due to decreased wound contraction by myofibroblasts. © 2005 Wiley-Liss, Inc. [source]


Ex vivo bone morphogenetic protein-2 gene delivery using gingival fibroblasts promotes bone regeneration in rats

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 3 2010
Joong-Ho Shin
Shin J-H, Kim K-H, Kim S-H, Koo K-T, Kim T-I, Seol Y-J, Ku Y, Rhyu I-C, Chung C-P, Lee Y-M. Ex vivo bone morphogenetic protein-2 gene delivery using gingival fibroblasts promotes bone regeneration in rats. J Clin Periodontol 2009; 37: 305,311. doi: 10.1111/j.1600-051X.2009.01522.x. Abstract Aim: The aim of the present study was to investigate bone regeneration following ex vivo bone morphogenetic protein-2 (BMP-2) gene delivery using human gingival fibroblasts (HGFs) in rat calvarial defects. Materials and Methods: An 8 mm craniotomy defect was created in Sprague,Dawley rats. The animals were divided into four groups: (1) non-grafted group, the defect was left empty; (2) collagen matrix group, the defect was filled with collagen matrix only; (3) HGF group, the defect was filled with non-transduced HGFs on collagen matrix; (4) BMP-2/HGF group, the defect was filled with BMP-2 gene-transduced HGFs on collagen matrix. Animals were sacrificed at 2 and 4 weeks after surgery, and micro-computed tomographic and histologic observations were performed. Results: The BMP-2/HGF group showed promoted osseous healing of calvarial defects, as compared with the other groups. At both 2 and 4 weeks, regenerated bone area was significantly greater in the BMP-2/HGF group than the other three groups. Quite a few number of transplanted HGFs were observed within the regenerated bone tissues. Conclusions: The results of this study suggest that ex vivo BMP-2 gene delivery induces prominent bone regeneration in vivo and HGFs may be useful as target cells for ex vivo gene therapy. [source]


Transforming growth factor- , stimulates Interleukin-11 production by human periodontal ligament and gingival fibroblasts

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 3 2006
R. Yashiro
Abstract Background: Transforming growth factor (TGF)- , is a potent multifunctional polypeptide, abundant in the bone matrix. Interleukin (IL)-11 is a pleiotropic cytokine with effects on multiple cell types. The present study was performed to evaluate the regulatory effects of TGF- , on IL-11 production by human periodontal ligament cells (PDL) and human gingival fibroblasts (HGF). Material and Methods: The expression of TGF- , receptor in PDL and HGF were observed using flow cytometry. PDL and HGF were stimulated with TGF- , with or without protein kinase C (PKC) inhibitors and activator. IL-11, bone morphogenetic protein-2 (BMP-2) and TGF- , mRNA expression was quantified by real-time polymerase chain reaction (PCR). IL-11 production was measured using enzyme-linked immunosorbent assay. Results: PDL and HGF expressed both TGF- , receptor I and TGF- , receptor II on the cell surfaces. IL-11 mRNA expression and IL-11 production were augmented by TGF- , in both PDL and HGF, with higher values in PDL. PKC inhibitors partially suppressed TGF- , -induced IL-11 production in PDL and HGF, whereas activator enhanced it. TGF- , mRNA and BMP-2 mRNA expression were up-regulated by TGF- , in PDL. Conclusion: These results suggest that PDL produce IL-11 in response to TGF- ,. [source]


Triclosan reduces microsomal prostaglandin E synthase-1 expression in human gingival fibroblasts

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 1 2005
M. Mustafa
Abstract Objective: The effect of triclosan (2,4,4,-trichloro-2,-hydroxydiphenyl ether) on the expression of cyclooxygenase-2 (COX-2) and microsomal prostaglandin E synthase-1 (mPGES-1) and on the translocation of the nuclear factor- ,B (NF- ,B) in relation to prostaglandin E2 (PGE2) production was investigated in human gingival fibroblasts challenged with tumor necrosis factor , (TNF,). Methods: Fibroblasts were established from gingival biopsies obtained from six children. COX-2 mRNA and protein expression was quantified using mRNA quantitation and enzyme immunometric assay kits. mPGES-1 mRNA was analysed by RT-PCR, mPGES-1 protein and NF-,B translocation by immunoblotting. PGE2 was determined by radioimmunoassay. Results: The cytokine TNF, enhanced the expression of mRNA as well as the protein levels of both COX-2 and mPGES-1 and subsequently the production of PGE2 in gingival fibroblasts. Treatment of gingival fibroblasts with triclosan (1 ,g/ml) significantly reduced the stimulatory effect of TNF, (10 ng/ml) on the expression of mPGES-1 at both the mRNA and the protein level by an average of 21% and 43%, respectively, and subsequently the production of PGE2 (p<0.01). Triclosan did not, however, affect the translocation of NF- ,B or the expression of COX-2 in TNF,- stimulated cells. Conclusion: The results show that triclosan reduces the augmented biosynthesis of PGE2 by inhibiting the mRNA and the protein expression of mPGES-1 in gingival fibroblasts. This finding may partly explain the anti-inflammatory effect of the agent previously reported in clinical studies. [source]


Effect of triclosan on interferon-, production and major histocompatibility complex class II expression in human gingival fibroblasts

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 10 2000
Manal Mustafa
Abstract Background, aims: The effect of triclosan (2,4,4,-trichloro-2,-hydroxyl-diphenyl ether) on the production of interferon-, (IFN-,) and the expression of major histocompatibility complex (MHC) class II antigen was studied in human gingival fibroblasts isolated from 4 individuals. Methods/Results: AII cell lines demonstrated high IFN-, production in 24-h cultures of human gingival fibroblasts stimulated by phytohemagglutinin (PHA) (5 ,g/ml). Human gingival fibroblasts showed a high expression of MHC class II when stimulated with 500 and 1000 pg/ml rIFN-, in 7-day cultures. Treatment of the cells with triclosan (0.5 ,g/ml) reduced both IFN-, production and MHC class II expression in human gingival fibroblast cultures. Similar inhibitory effects on IFN-, production and MHC class II expression were observed when the anti-inflammatory agent dexamethazone (1 ,M) was used. Conclusion: The present study further supports the view that the agent has an anti-inflammatory effect in addition to its antibacterial capacity. [source]


Areca nut extract-treated gingival fibroblasts modulate the invasiveness of polymorphonuclear leukocytes via the production of MMP-2

JOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 1 2009
Hsuan-Hsuan Lu
Background:, Areca nut chewing is associated with an increase in the incidence of oral neoplastic or inflammatory diseases. Aberrations in matrix metalloprotease (MMP) expression are associated with the pathogenesis of oral diseases. This study investigated the potential effects of areca nut extract (ANE) on human gingival fibroblasts and the consequential impacts on inflammatory pathogenesis. Methods:, Analyses of senescence marker, cell viability, changes of the cell cycle, and cell granularity in gingival fibroblasts together with an assessment of the invasiveness of polymorphonuclear (PMN) leukocytes after treatment with the supernatant of ANE-treated gingival fibroblasts were performed to characterize the phenotypic impacts. Western blotting and gelatin zymography were used to assay the expression and activity of MMP-2. Results:, Chronic subtoxic (<10 ,g/ml) ANE treatment resulted in premature growth arrest, appearance of senescence-associated ,-galactosidase activity and various other senescence-associated phenotypes in gingival fibroblasts. Gingival fibroblasts established from older individuals had a higher propensity to become ANE-induced senescent gingival fibroblasts. An activation of MMP-2 was identified in senescent cells. PMN leukocytes treated with the supernatant of ANE-induced senescent cells exhibited a significant increase in invasiveness, which was abrogated by both a MMP-2 blocker and a MMP-2 nullifying antibody. Conclusions:, This study provides evidence whereby MMP-2 secreted from ANE-induced senescent gingival fibroblasts would facilitate the invasiveness of PMN leukocytes, which could be associated with the oral inflammatory process in areca chewers. [source]


Synergistic enhancement of collagenous protein synthesis by human gingival fibroblasts exposed to nifedipine and interleukin-1-beta in vitro

JOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 1 2000
R. B. Johnson
Abstract: Gingival overgrowth commonly occurs coincident to therapy with calcium channel blockers. The biologic mechanism for this condition is unknown; however, many clinicians suggest that poor oral hygiene may contribute to development of the overgrowth. This study tests the hypothesis that collagenous protein synthesis by gingival fibroblasts is synergistically enhanced when they are exposed to both nifedipine (N) and the pro-inflammatory cytokine, interleukin-1-beta, a cytokine expressed in inflamed gingiva. Human gingival fibroblasts were isolated from biopsies of normal gingiva and cells separated into two groups. Group 1 was exposed to media containing 0, 5, 50, or 500 pg/ml IL-1-beta, or 10,7 M N for 7 days; Group 2 was exposed to those concentrations of IL-1-beta +10,7 M N. [3H]-proline was added to the medium for the final 24 h. Cells and matrix were harvested and radioactivity determined by liquid scintillation analysis. Means (d.p.m./103 cells) were compared by factorial ANOVA and Scheffè comparisons. Collagenous protein synthesis was significantly reduced by 5 pg/ml IL-1-beta +10,7 M N and enhanced by 500 pg/ml IL-1-beta +10,7 M N as compared to N or IL-1-beta alone. Thus, patients may be more susceptible to gingival overgrowth coincident to nifedipine therapy as a result of the synergistic enhancement of connective tissue synthesis by these agents. [source]


Cytotoxic effects of dental resin liquids on primary gingival fibroblasts and periodontal ligament cells in vitro

JOURNAL OF ORAL REHABILITATION, Issue 12 2004
Y.-L. Lai
summary, Cytotoxic effects of resin liquids of three in situ relining dental polymers, AlikeTM, Kooliner, and Tokuso Rebase, and their major components, methyl methacrylate (MMA), isobutyl methacrylate (IBMA), and 1,6-hexanediol dimethacrylate (1,6-HDMA) were investigated. The concentrations of major monomers in these resin liquids were determined by high-performance liquid chromatography. Cellular viability of human gingival fibroblasts (GF) and periodontal ligament (PDL) cells were evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide assay. Moreover, patterns of cell death were analysed using annexin V/propidium iodide staining with flow cytometry. The results indicated that AlikeTM liquid contained 91·3% MMA, Kooliner liquid contained 94·5% IBMA, and Tokuso Rebase liquid contained 65·8% 1,6-HDMA. All materials examined had cytotoxic effects on GF and PDL cells in dose-dependent manners. Tokuso Rebase liquid appeared to be the most cytotoxic among the various resin liquids examined. The effects of Kooliner and Tokuso Rebase liquids may have resulted from IBMA and 1,6-HDMA, respectively. Furthermore, the majority of treated cells died from necrosis; whereas a small portion of cells died from apoptosis. In conclusion, the results demonstrated that these liquid forms of dental polymers and their major monomers cause cytotoxic reactions. The direct relining procedure that cures these materials in situ should be used cautiously. [source]


Cytotoxic effects of gingival retraction cords on human gingival fibroblasts in vitro

JOURNAL OF ORAL REHABILITATION, Issue 4 2004
C.-M. Liu
summary, The objective of this study was to determine the cytocompatibility of three different extracts of gingival retraction cords and to compare the cytotoxic effect of these materials on human gingival fibroblasts. Gingival retraction cords impregnated with aluminium sulphate (Gingi-Aid), dl -adrenaline HCl (Gingi-Pak) and non-drug-impregnated cord (Gingi-Plain) were eluted with culture medium for 10 min and 24 h. Cytotoxicity was judged using a tetrazolium bromide reduction assay. Our data demonstrated that gingival retraction cords applied alone almost completely inhibited cell viability (P < 0·05). In addition, the results also showed that the eluates from aluminium sulphate-impregnated cord, dl -adrenaline HCl-impregnated cord and non-drug-impregnated cord were cytotoxic to primary human gingival fibroblast cultures (P < 0·05). The cell viability of incubation of gingival fibroblasts containing 10-min eluates of aluminium sulphate, dl -adrenaline HCl and non-drug-impregnated cord was 61, 21 and 70%, respectively. The cell viability of incubation of gingival fibroblasts containing 24 h eluates of aluminium sulphate, dl -adrenaline HCl and non-drug-impregnated cord was 68, 58 and 72%, respectively. It was found that dl -adrenaline HCl-impregnated gingival retraction cord was the most toxic gingival retraction cord among the materials tested in all cultures (P < 0·05). The cytotoxicity decreased in an order of dl -adrenaline HCl-impregnated cord > aluminium sulphate-impregnated cord > non-drug-impregnated cord. The extent or degree of the cytotoxicity depended on the materials tested. Gingival retraction cords have significant potential for gingival toxicity. Careful management of gingiva retraction cords would lower the risk of potential gingival tissue damage during clinical application procedure and thus increase the success of prosthodontic procedures. [source]


The upregulation of heat shock protein 47 in human gingival fibroblasts stimulated with cyclosporine A

JOURNAL OF PERIODONTAL RESEARCH, Issue 3 2010
T.-Y. Chang
Chang T-Y, Tsai C-H, Chang Y-C. The upregulation of heat shock protein 47 in human gingival fibroblasts stimulated with cyclosporine A. J Periodont Res 2010; 45: 317,322. © 2009 John Wiley & Sons A/S Background and Objective:, Heat shock protein 47 (Hsp47), a collagen-specific molecular chaperone, is involved in the processing and/or secretion of procollagen. Heat shock protein 47 is consistently and dramatically upregulated in a variety of fibrotic diseases. The aim of this study was to compare Hsp47 expression in normal gingival tissues and cyclosporine A-induced gingival overgrowth specimens and further explore the potential mechanisms that may lead to induction of Hsp47 expression. Material and Methods:, Fifteen cyclosporine A-induced gingival overgrowth specimens and five normal gingival tissues were examined by immunohistochemistry. Western blot was used to investigate the effects of cyclosporine A on the expression of Hsp47 in human gingival fibroblasts. In addition, Aggregatibacter actinomycetemcomitans, interleukin-1, (IL-1,) and mitogen-activated protein kinase kinase (MEK) inhibitor U0126 were added to seek the possible regulatory mechanisms of Hsp47 expression. Results:, A significantly higher percentage of cells positively stained for Hsp47 was noted in the cyclosporine A-induced gingival overgrowth group than in the normal gingival group (p < 0.05). Expression of Hsp47 was observed mainly in the cytoplasm of fibroblasts, endothelial cells, epithelial cells and inflammatory cells. Expression of Hsp47 was significantly higher in cyclosporine A-induced gingival overgrowth specimens with higher levels of inflammatory infiltrates (p < 0.05). Cyclosporine A upregulated Hsp47 expression in human gingival fibroblasts in a dose-dependent manner (p < 0.05). The addition of A. actinomycetemcomitans or interlukin-1, significantly increased Hsp47 expression compared with cyclosporine A alone (p < 0.05). The MEK inhibitor U0126 was found to inhibit cyclosporine A-induced Hsp47 expression (p < 0.05). Conclusion:, Expression of Hsp47 is significantly upregulated in cyclosporine A-induced gingival overgrowth specimens, and Hsp47 expression induced by cyclosporine A in fibroblasts may be mediated by the MEK signal transduction pathway. The expression of Hsp47 could be significantly enhanced by A. actinomycetemcomitans and interlukin-1,. [source]


Cigarette smoke condensate affects the collagen-degrading ability of human gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 6 2009
W. Zhang
Background and Objective:, Cigarette smoke condensate, the particulate matter of cigarette smoke, is composed of thousands of chemicals, including nicotine. Cigarette smoking is a risk factor for periodontal disease. This study investigated the influence of cigarette smoke condensate on the collagen-degrading ability of human gingival fibroblasts and its mechanism. Material and Methods:, Human gingival fibroblasts were exposed for 72 h to various concentrations of total particulate matter cigarette smoke condensate. Cell proliferation and cytotoxicity were evaluated using water-soluble tetrazolium-1 and lactate dehydrogenase, respectively. The collagen-degrading ability of human gingival fibroblasts was evaluated in collagen-coated six-well plates. Conditioned media and membrane extracts were collected for zymography and western blot analyses of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Results:, Cell proliferation decreased and cytotoxicity increased in human gingival fibroblasts with increasing concentrations of cigarette smoke condensate. Cell proliferation decreased by more than 50% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 200 ,g/mL, and cytotoxicity increased to more than 30% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 400 ,g/mL. Cigarette smoke condensate increased the collagen-degrading ability of human gingival fibroblasts, especially at a concentration of 100 ,g/mL (1.5-fold increase, p < 0.05) compared with the control. Cigarette smoke condensate increased the production of proMMP-1, proMMP-2, MMP-14 and TIMP-1, and decreased the production of TIMP-2, in conditioned media. Furthermore, compared with the control group, cigarette smoke condensate increased the production of MMP-2, MMP-14 and TIMP-2 in membrane extracts, especially at concentrations of 50,100 ,g/mL. Conclusion:, Cigarette smoke condensate affects human gingival fibroblast proliferation and is toxic at total particulate matter cigarette smoke condensate concentrations of , 400 ,g/mL. Cigarette smoke condensate can increase the collagen-degrading ability of human gingival fibroblasts by altering the production and localization of MMPs and TIMPs. [source]


Effect of transforming growth factor-beta1 on expression of the connective tissue growth factor (CCN2/CTGF) gene in normal human gingival fibroblasts and periodontal ligament cells

JOURNAL OF PERIODONTAL RESEARCH, Issue 2 2009
H. Takeuchi
Background and Objective:, Connective tissue growth factor (CCN2/CTGF) plays an important role in wound healing and regulation of the extracellular matrix in periodontal tissue. However, the functional relationship between altered transforming growth factor-beta1 levels and CCN2/CTGF has not been extensively investigated in human gingival fibroblasts and periodontal ligament cells. This study investigated the effects of transforming growth factor-beta1 on the expression of the CCN2/CTGF gene in human gingival fibroblasts and periodontal ligament cells in vitro. Material and Methods:, Cells were isolated from normal periodontal tissues and cultured in Dulbecco's modified Eagle's minimal essential medium/F12 containing 10% fetal bovine serum. Subconfluent cells were maintained under serum deprivation for 24 h then treated with Dulbecco's modified Eagle's minimal essential medium/F12 containing 0.5% fetal bovine serum (control) and 0.1, 1, 5 or 10 ng/mL of transforming growth factor-beta1 for 24, 48 or 72 h. The effects of transforming growth factor-beta1 on CCN2/CTGF mRNA expression were measured by reverse transcription,polymerase chain reaction. CCN2/CTGF protein was quantitatively analyzed using enzyme-liked immunosorbent assay. Subcellular distribution of CCN2/CTGF protein in both human gingival fibroblasts and periodontal ligament cells was observed using immunofluorescence microscopy. Results:, In both human gingival fibroblasts and periodontal ligament cells, the expression of CCN2/CTGF mRNA and CCN2/CTGF protein was significantly increased, in a dose- and time-dependent manner, in the presence of transforming growth factor-beta1. Moreover, immunofluorescence analysis indicated that immunoreactivity to CCN2/CTGF showed a granular pattern of protein localization. Conclusion:, The expression of CCN2/CTGF mRNA and protein was induced by transforming growth factor-beta1 in human gingival fibroblasts and periodontal ligament cells. These results suggest that CCN2/CTGF plays an important role in wound healing and in the regeneration of periodontal tissue. [source]


Oral malodorous compound causes apoptosis and genomic DNA damage in human gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 4 2008
K. Yaegaki
Background and Objective:, Volatile sulfur compounds are the main cause of halitosis. Hydrogen sulfide is one of these volatile sulfur compounds and the principal malodorous compound in physiological halitosis. Periodontally pathogenic activities of hydrogen sulfide have been previously reported. Hydrogen sulfide induces apoptotic cell death in aorta smooth muscle cells and in other tissues. Apoptosis plays an important role in the onset and progress of periodontitis. The objective of this study was to determine whether hydrogen sulfide causes apoptosis in human gingival fibroblasts. Material and methods:, Necrotic cells were detected using a lactate dehydrogenase assay. Apoptosis was ascertained using a histone-complexed DNA fragment assay and flow cytometry. The level of caspase 3, a key enzyme in apoptotic signaling, was also measured, and the effects of hydrogen sulfide on reactive oxygen species and superoxide dismutase were assessed. DNA damage caused by hydrogen sulfide was examined by means of single-cell gel electrophoresis. Results:, After 72 h of incubation with 100 ng/mL of hydrogen sulfide, necrosis was found in less than 10% of human gingival fibroblasts, whereas apoptosis was significantly increased (p < 0.05). Superoxide dismutase activity was strongly inhibited, and reactive oxygen species production was enhanced, after 48 and 72 h of incubation. Caspase 3 activity was also increased after 72 h of incubation (p < 0.01). Tail length, percentage of DNA in tail, and tail moment, measured by single-cell gel electrophoresis, were also intensified after 72 h of incubation (p < 0.001). Conclusion:, Hydrogen sulfide caused apoptosis and DNA damage in human gingival fibroblasts. An increased level of reactive oxygen species stimulated by hydrogen sulfide may induce apoptosis and DNA strand breaks. [source]


CC Chemokine ligand 17 in periodontal diseases: expression in diseased tissues and production by human gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 4 2008
Y. Hosokawa
Background and Objective:, It has been reported that T helper 2 (Th2) cells are related to exacerbation of periodontal disease. However, it is uncertain how the migration of Th2 cells is controlled. In this study, we examined the expression of CC chemokine ligand 17 (CCL17), which is a Th2 chemokine, in periodontal tissues. Moreover, we investigated the effects of cytokines and toll-like receptor (TLR) ligands on the production of CCL17 by human gingival fibroblasts (HGFs). Material and Methods:, We used immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR) to detect CCL17 in periodontal tissues. HGFs were exposed to cytokines and TLR ligands. Expression of CCL17 was examined by RT-PCR and enzyme-linked immunosorbent assay. We used signal transduction inhibitors in some experiments. Results:, Both CCL17 and its receptor, CC chemokine receptor 4 (CCR4), were expressed in diseased periodontal tissues. A combination of tumour necrosis factor , (TNF-,) and interleukin (IL)-4/IL-13 increased CCL17 expression. Moreover, treatment of HGFs with a low dose of interferon-, (IFN-,) in combination with TNF-, and IL-4 or IL-13 had synergistic effects on the production of CCL17, whereas a high dose of IFN-, inhibited CCL17 production. Furthermore, Escherichia coli (E. coli) lipopolysaccharide (TLR4 ligand) and Pam3CSK4 (TLR2 ligand) inhibited CCL17 production by TNF-, + IL-4-stimulated HGFs, while CpG DNA (TLR9 ligand) enhanced TNF-, + IL-4 induced-CCL17 production by HGFs. Furthermore, a c-Jun NH2 terminal kinase (JNK) inhibitor, a phosphatidylinositol-3-kinase (PI3K) inhibitor and a nuclear factor ,B (NF-,B) inhibitor inhibited CCL17 production by HGFs. Conclusion:, These results suggest that the CCL17 produced by HGFs may be involved in the migration of Th2 cells into inflamed tissues, and provide evidence that CCL17 production is controlled by cytokines and TLR ligands in periodontal disease. [source]


Focal adhesion kinase mediates human leukocyte histocompatibility antigen class II-induced signaling in gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 6 2007
S. Yoshizawa
Background and Objective:, The role of human leukocyte antigen class II molecules on nonantigen-presenting cells has been a matter of controversy. We previously reported that human leukocyte antigen class II molecules on human gingival fibroblasts do not present antigens, but transduce signals into the cells by making a complex with antigenic peptide T-cell receptor or by stimulating cell surface human leukocyte antigen-DR molecules with human leukocyte antigen-DR antibody (L243), which mimics the formation of the human leukocyte antigen class II,antigenic peptide T-cell receptor complex, resulting in the expression of several cytokines. The aim of this study was to detect human leukocyte antigen class II-associated molecules mediating human leukocyte antigen class II-induced signals into the cells. Material and Methods:, Antibody-based protein-microarray analysis was performed to detect activated signaling molecules in gingival fibroblasts stimulated via human leukocyte antigen class II molecules. Then, we examined if these molecules structurally associate with human leukocyte antigen class II and actually transduce signals into the cells. Results:, Stimulation of human leukocyte antigen class II on gingival fibroblasts by L243 resulted in enhanced phosphorylation of focal adhesion kinase. Focal adhesion kinase was co-immunoprecipitated with human leukocyte antigen-DR by L243. Stimulation of gingival fibroblasts with L243 induced phosphorylation of focal adhesion kinase. Luteolin, a putative focal adhesion kinase inhibitor, suppressed phosphorylation of focal adhesion kinase and dose dependently inhibited human leukocyte antigen class II-induced cytokine production. Conclusion:, Focal adhesion kinase is structurally associated with human leukocyte antigen-DR and mediates human leukocyte antigen class II-induced signals in gingival fibroblasts. [source]


Identification of marker genes distinguishing human periodontal ligament cells from human mesenchymal stem cells and human gingival fibroblasts

JOURNAL OF PERIODONTAL RESEARCH, Issue 3 2007
T. Fujita
Background and Objective:, Molecular gene markers, which can distinguish human bone marrow mesenchymal stem cells from human fibroblasts, have recently been reported. Messenger RNA levels of tissue factor pathway inhibitor-2, major histocompatibility complex-DR-,, major histocompatibility complex-DR-,, and neuroserpin are higher in human bone marrow mesenchymal stem cells than in human fibroblasts. However, human bone marrow mesenchymal stem cells express less apolipoprotein D mRNA than human fibroblasts. Periodontal ligament cells are a heterogeneous cell population including fibroblasts, mesenchymal stem cells, and progenitor cells of osteoblasts or cementoblasts. The use of molecular markers that distinguish human bone marrow mesenchymal stem cells from human fibroblasts may provide insight into the characteristics of human periodontal ligament cells. In this study, we compared the molecular markers of human periodontal ligament cells with those of human bone marrow mesenchymal stem cells and human gingival fibroblasts. Material and Methods:, The mRNA expression of the molecular gene markers was analyzed using real-time polymerase chain reaction. Statistical differences were determined with the two-sided Mann,Whitney U -test. Results:, Messenger RNA levels of major histocompatibility complex-DR-, and major histocompatibility complex-DR-, were lower and higher, respectively, in human periodontal ligament cells than in human bone marrow mesenchymal stem cells or human gingival fibroblasts. Human periodontal ligament cells showed the lowest apolipoprotein D mRNA levels among the three types of cells. Conclusion:, Human periodontal ligament cells may be distinguished from human bone marrow mesenchymal stem cells and human gingival fibroblasts by the genes for apolipoprotein D, major histocompatibility complex-DR-,, and major histocompatibility complex-DR-,. [source]


Differential expression of periodontal ligament-specific markers and osteogenic differentiation in human papilloma virus 16-immortalized human gingival fibroblasts and periodontal ligament cells

JOURNAL OF PERIODONTAL RESEARCH, Issue 2 2007
S.-H. Pi
Background and Objective:, Periodontal ligament cells and gingival fibroblasts are important in the remodeling of periodontal tissue, but human papilloma virus (HPV)16-immortalized cell lines derived from human periodontal ligament cells and gingival fibroblasts has not been characterized. The purpose of this study was to establish and differentially characterize the immortalized cell lines from gingival fibroblasts and periodontal ligament by HPV16 transfection. Material and Methods:, Cell growth, cell cycle analysis, western blot for cell cycle regulatory proteins and osteogenic differentiation markers, and reverse transcription,polymerase chain reaction for periodontal ligament-specific markers were performed. Results:, Both immortalized cell lines (immortalized gingival fibroblasts and immortalized periodontal ligament cells) grew faster than primary cultured gingival fibroblasts or periodontal ligament cells. Immortalized gingival fibroblasts and immortalized periodontal ligament cells overexpressed proteins p16 and p21, and exhibited degradation of proteins pRb and p53, which normally cause cell cycle arrest in G2/M-phase. Western blotting and reverse transcription,polymerase chain reaction for periodontal ligament-specific and osteogenic differentiation marker studies demonstrated that a cell line, designated IPDL, mimicked periodontal ligament gene expression for alkaline phosphatase, osteonectin, osteopontin, bone sialoprotein, bone morphogenic protein-2, periostin, S-100A4 and PDLs17. Conclusion:, These results indicate that IPDL and immortalized gingival fibroblast cell lines consistently retain normal periodontal ligament and gingival fibroblast phenotypes, respectively, and periodontal ligament markers and osteogenic differentiation in IPDL are distinct from immortalized gingival fibroblast cells. [source]


Epidermal growth factor stimulates urokinase-type plasminogen activator expression in human gingival fibroblasts.

JOURNAL OF PERIODONTAL RESEARCH, Issue 6 2004
Possible modulation by genistein, curcumin
Background:, Regulation of the extracellular matrix turnover is a crucial process in wound healing and the progress of periodontal disease. It has been proposed that urokinase-type plasminogen activator (uPA), under the control of growth factors or cytokines, provides the proteolytic potential to the accomplishment of these cellular events. Epidermal growth factor (EGF) is one of the growth factors that has been shown to be active in uPA regulation. Methods:, In this study, we have assessed the effect of EGF on uPA expression in primary cultures of human gingival fibroblasts. We also studied the signaling pathways involved in this process and the role of the dietary phytoestrogens curcumin and genistein as potential modulators of this response. Results:, Human gingival fibroblasts expressed a basal uPA activity, which was inhibited by genistein, but not by curcumin. After treatment with 10 ng/ml EGF, uPA production was strongly stimulated. Exposure to genistein and curcumin inhibited EGF-stimulated urokinase production, although only genistein showed a statistically significant inhibitory response. Using more specific inhibitors, we found that the mitogen-activated extracellular kinase and c-Jun N-terminal kinase (JNK) inhibitors PD98059 and SP600125 also blocked the EGF-dependent stimulatory effect. On the other hand, SB203580, inhibitor of the p38 member of mitogen-activated protein kinase family, did not alter this response. In accordance to these findings, EGF stimulated a potent activation of JNK and a mild activation of extracellular signal-regulated kinases 1/2. Finally, EGF stimulated the phosphorylation of its receptor and tyrphostin (AG1478), curcumin and genistein were able to inhibit this stimulatory effect. Conclusions:, These results indicate that EGF constitutes a strong stimuli on uPA expression in human gingival fibroblasts. Our data also shows that EGF-stimulated uPA production involves the activation of the extracellular signal-regulated kinases 1/2 and JNK signaling pathways and might be modulated by the natural phytoestrogens curcumin and genistein. [source]