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Ligament Cells (ligament + cell)
Kinds of Ligament Cells Selected AbstractsAscorbic Acid Induces Collagenase-1 in Human Periodontal Ligament Cells but Not in MC3T3-E1 Osteoblast-Like Cells: Potential Association Between Collagenase Expression and Changes in Alkaline Phosphatase Phenotype,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 1 2003Momotoshi Shiga Abstract Ascorbic acid (AA) enhances osteoblastic differentiation by increasing collagen accumulation, which in turn, results in increased alkaline phosphatase (AP) expression in some osteogenic cells. However, in other cells, including human periodontal ligament (PDL) cells, additional osteoinductive agents are required for this response. To understand the potential basis for the maintenance of the AP phenotype of PDL cells exposed to AA, we examined the modulation of the tissue-degrading matrix metalloproteinases (MMPs) and their inhibitors by AA in short-term cell cultures. Early passage PDL cells in serum-free medium were exposed to AA for 5 days. The samples were analyzed for MMPs and their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs), AP, collagen I(,1), and osteocalcin. We found that AA dose-dependently increased the expression of collagenase-1, and minimally TIMP-1, but not stromelysin-1 or TIMP-2. Additionally, AA caused substantial increases in levels of type I collagen. AA was unable to increase AP activity or osteocalcin messenger RNA in PDL cells. However, the cells retained the ability to show a significantly greater AP expression in high- versus low-density cultures, and increased osteocalcin as well as AP levels when cultured in the presence of dexamethasone. Moreover, in cells exposed to dexamethasone, increases in AP and osteocalcin were accompanied by a repression of collagenase-1 expression. In contrast to PDL cells, AA did not induce collagenase but produced a significant increase in AP expression in MC3T3-E1 cells. These findings provide the first evidence that AA, by modulating both collagen and collagenase-1 expression in PDL cells, most likely contributes to a net matrix remodeling response in these cells. Furthermore, the relationship between changes in collagenase expression and alterations in AP activity in PDL and MC3T3-E1 cells suggests a potential role for collagenase in modulating the AP phenotype of cells with osteoblastic potential. [source] In vitro viability, mitogenicity and clonogenic capacity of periodontal ligament cells after storage in four media at room temperatureDENTAL TRAUMATOLOGY, Issue 2 2000M. Ashkenazi Abstract , The choice of storage medium for preserving traumatically avulsed teeth is important for the success of future replantation. The objective of this study was to compare the effectiveness of four recommended storage media (Hank's balanced salt solution [HBSS], culture medium, , minimal essential medium [,-MEM], and ViaSpan) to preserve cultured periodontal ligament fibroblasts (PDLF) at room temperature (22°C). PDLF were obtained from explants of extracted healthy human teeth. Plates with confluent PDLF were soaked in the various media for 2, 8 and 24 h at room temperature. A control group was incubated with culture medium at 37°C. After incubation, viability of the cells was determined by trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic capacity (by culturing one cell/well). Viability of PDLF stored up to 24 h was comparable in all tested media, and the differences were limited to 1%,3%. PDLF stored for up to 24 h in various media had statistically comparable mitogenicity to the control group. After 8 h of storage, the differences were limited to 2%,9%, except for the ,-MEM group which had 23%,29% lower mitogenic capacity compared to the control group. Increasing the storage time up to 24 h further decreased the mitogenicity of the cells by 22%,47%. The highest mitogenicity after 24 h of storage was found in PDLF stored in culture medium or HBSS, and the lowest in ,-MEM. PDLF stored for 2,8 h in various media had a comparable clonogenic capacity to the control group. However, after 24 h, the cells' clonogenic ability dropped by 14%,66%. A similar trend of reduction was noted in the mitogenic and clonogenic capacity, although it was statistically significant only in the clonogenic capacity. Culture medium and ViaSpan, followed by HBSS, were the most effective in preserving the clonogenic capacity of PDLF after 24 h of storage. The lowest clonogenic capacity after 24 h of storage was in the ,-MEM group (66%, P<0.0025). In conclusion, culture medium, followed by HBSS and ViaSpan, was the most effective media for preserving the viability, mitogenicity and clonogenic capacity of PDLF stored for up to 24 h at room temperature. The lowest functional abilities were found in PDLF stored in ,-MEM. [source] Expression of Osterix in mechanical stress-induced osteogenic differentiation of periodontal ligament cells in vitroEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 3 2008Yanhong Zhao Osterix (Osx) is an osteoblast-specific transcription factor required for the differentiation of pre-osteoblasts into functional osteoblasts. This study sought to examine the changes of Osx expression in periodontal ligament cells (PDLC) subjected to mechanical force, and to investigate whether Osx is involved in the mechanical stress-induced differentiation of PDLC. Human PDLC were exposed to centrifugal force for 1,12 h. Real-time polymerase chain reaction (PCR), western blot, and immunofluorescence assays were used to examine the mRNA and protein expression of Osx and its subcellular localization. Furthermore, PDLC were transfected with the expression vector pcDNA3.1 flag-Osx and subjected to mechanical force for 6 h. The changes in alkaline phosphatase (ALP) activity and in the expression of core-binding factor alpha1 (Cbfa1), ALP, osteopontin, bone sialoprotein, osteocalcin, and collagen I were measured. After the application of mechanical force, Osx was upregulated in a time-dependent manner at both mRNA and protein levels, and Osx protein was translocated from the cytosol into the cell nuclei. Overexpression of Osx did not affect the expression of Cbfa1, but it significantly enhanced the ALP activity and the mRNA expression of all the aforementioned osteogenic marker genes, all of which increased further under mechanical stress. These results suggest that Osx might play an important role in the mechanical stress-induced osteogenic differentiation of PDLC and therefore be involved in alveolar bone remodeling during orthodontic therapy. [source] Osterix is a key target for mechanical signals in human thoracic ligament flavum cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Dongwei Fan Mechanical stress is considered to be an important factor in the progression of thoracic ossification of the ligament flavum (TOLF). To elucidate the mechanism underlying mechanical stress-induced TOLF, we investigated the effect of stretching on cultured flavum ligament cells derived from TOLF and non-TOLF patients. We found that the mRNA expression of alkaline phosphatase (ALP), osteocalcin, Runx2, and osterix, but not that of Dlx5 and Msx2, was significantly increased by stretching in TOLF cells. In addition, the effect seems to be finely tuned by stretching-triggered activation of distinct mitogen-activated protein kinase cascades. Specifically, a p38 specific inhibitor, SB203580, significantly inhibited stretching-induced osterix expression as well as ALP activity, whereas a specific inhibitor of ERK1/2, U0126, prevented stretching-induced Runx2 expression. We showed that overexpression of osterix resulted in a significant increase of ALP activity in TOLF cells, and osterix-specific RNAi completely abrogated the stretching-induced ALP activity, indicating that osterix plays a key role in stretching-stimulated osteogenic effect in TOLF cells. These results suggest that mechanical stress plays important roles in the progression of TOLF through induction of osteogenic differentiation of TOLF cells, and our findings support that osterix functions as a molecular link between mechanostressing and osteogenic differentiation. J. Cell. Physiol. 211: 577,584, 2007. © 2007 Wiley-Liss, Inc. [source] Biological mediators and periodontal regeneration: a review of enamel matrix proteins at the cellular and molecular levelsJOURNAL OF CLINICAL PERIODONTOLOGY, Issue 2008Dieter D. Bosshardt Abstract Background: Despite a large body of clinical and histological data demonstrating beneficial effects of enamel matrix proteins (EMPs) for regenerative periodontal therapy, it is less clear how the available biological data can explain the mechanisms underlying the supportive effects of EMPs. Objective: To analyse all available biological data of EMPs at the cellular and molecular levels that are relevant in the context of periodontal wound healing and tissue formation. Methods: A stringent systematic approach was applied using the key words "enamel matrix proteins" OR "enamel matrix derivative" OR "emdogain" OR "amelogenin". The literature search was performed separately for epithelial cells, gingival fibroblasts, periodontal ligament cells, cementoblasts, osteogenic/chondrogenic/bone marrow cells, wound healing, and bacteria. Results: A total of 103 papers met the inclusion criteria. EMPs affect many different cell types. Overall, the available data show that EMPs have effects on: (1) cell attachment, spreading, and chemotaxis; (2) cell proliferation and survival; (3) expression of transcription factors; (4) expression of growth factors, cytokines, extracellular matrix constituents, and other macromolecules; and (5) expression of molecules involved in the regulation of bone remodelling. Conclusion: All together, the data analysis provides strong evidence for EMPs to support wound healing and new periodontal tissue formation. [source] Transforming growth factor- , stimulates Interleukin-11 production by human periodontal ligament and gingival fibroblastsJOURNAL OF CLINICAL PERIODONTOLOGY, Issue 3 2006R. 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] Autocrine growth factors in human periodontal ligament cells cultured on enamel matrix derivativeJOURNAL OF CLINICAL PERIODONTOLOGY, Issue 2 2001Staale P. Lyngstadaas Abstract Objective: Enamel extracellular matrix proteins in the form of the enamel matrix derivative EMDOGAIN® (EMD) have been successfully employed to mimic natural cementogenesis to restore fully functional periodontal ligament, cementum and alveolar bone in patients with severe periodontitis. When applied to denuded root surfaces EMD forms a matrix that locally facilitates regenerative responses in the adjacent periodontal tissues. The cellular mechanism(s), e.g. autocrine growth factors, extracellular matrix synthesis and cell growth, underlying PDL regeneration with EMD is however poorly investigated. Material and Methods: Human periodontal ligament (PDL) cells were cultured on EMD and monitored for cellular attachment rate, proliferation, DNA replication and metabolism. Furthermore, intracellular cyclic-AMP levels and autocrine production of selected growth factors were monitored by immunological assays. Controls included PDL and epithelial cells in parallel cultures. Results: PDL cell attachment rate, growth and metabolism were all significantly increased when EMD was present in cultures. Also, cells exposed to EMD showed increased intracellular cAMP signalling and autocrine production of TGF-,1, IL-6 and PDGF AB when compared to controls. Epithelial cells increased cAMP and PDGF AB secretion when EMD was present, but proliferation and growth were inhibited. Conclusion: Cultured PDL cells exposed to EMD increase attachment rate, growth rate and metabolism, and subsequently release several growth factors into the medium. The cellular interaction with EMD generates an intracellular cAMP signal, after which cells secrete TGF-,1, IL-6 and PDGF AB. Epithelial cell growth however, is inhibited by the same signal. This suggest that EMD favours mesenchymal cell growth over epithelium, and that autocrine growth factors released by PDL cells exposed to EMD contribute to periodontal healing and regeneration in a process mimicking natural root development. [source] Cytotoxic effects of dental resin liquids on primary gingival fibroblasts and periodontal ligament cells in vitroJOURNAL OF ORAL REHABILITATION, Issue 12 2004Y.-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] Effects of adenoviral-mediated coexpression of bone morphogenetic protein-7 and insulin-like growth factor-1 on human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 4 2010L. Yang Yang L, Zhang Y, Dong R, Peng L, Liu X, Wang Y, Cheng X. Effects of adenoviral-mediated coexpression of bone morphogenetic protein-7 and insulin-like growth factor-1 on human periodontal ligament cells. J Periodont Res 2010; 45: 532,540. © 2010 John Wiley & Sons A/S Background and Objective:, Bone morphogenetic protein-7 (BMP-7) and insulin-like growth factor-1 (IGF-1) are important in periodontal reconstruction. However, their synergistic effect in periodontal regeneration by gene delivery has not been reported. In this study, gene delivery of these two growth factors to human periodontal ligament cells (hPDLCs) was examined for its effects on cell proliferation and differentiation. Material and Methods:, Recombinant adenoviruses containing both human BMP-7 and IGF-1 cDNA created by introducing the internal ribosome entry site (IRES) sequence were used to transfer the genes into hPDLCs. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and cell cycle analysis were used to observe their effects on cell proliferation, while alkaline phosphatase activity measurement, RT-PCR and in vivo tests were conducted to investigate their effects on cell differentiation. Results:, The proliferation of hPDLCs transduced by adenoviruses coexpressing BMP-7 and IGF-1 was suppressed while their differentiation ability was enhanced. There was a synergism of BMP-7 and IGF-1 in up-regulating alkaline phosphatase activity and mRNA levels of collagen type I and Runx2. Implantation in vivo with scaffolds illustrated that the transduced cells exhibited osteogenic differentiation and formed bone-like structures. Conclusion:, The combined delivery of BMP-7 and IGF-1 genes using an IRES-based strategy synergistically enhanced differentiation of hPDLCs. It is suggested that this could be a new potential method in gene therapy for periodontal reconstruction. [source] Localization of SOST/sclerostin in cementocytes in vivo and in mineralizing periodontal ligament cells in vitroJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2010A. Jäger Jäger A, Götz W, Lossdörfer S, Rath-Deschner B. Localization of SOST/sclerostin in cementocytes in vivo and in mineralizing periodontal ligament cells in vitro. J Periodont Res 2009; doi: 10.1111/j.1600-0765.2009.01227.x. © 2009 John Wiley & Sons A/S Background and Objective:, Cementum and bone are rather similar hard tissues, and osteocytes and cementocytes, together with their canalicular network, share many morphological and cell biological characteristics. However, there is no clear evidence that cementocytes have a function in tissue homeostasis of cementum comparable to that of osteocytes in bone. Recent studies have established an important role for the secreted glycoprotein sclerostin, the product of the SOST gene, as an osteocyte-derived signal to control bone remodelling. In this study, we investigated the expression of sclerostin in cementocytes in vivo as well as the expression of SOST and sclerostin in periodontal ligament cell cultures following induction of mineralization. Material and Method:, Immunolocalization of sclerostin was performed in decalcified histological sections of mouse and human teeth and alveolar bone. Additionally, periodontal ligament cells from human donors were cultured in osteogenic conditions, namely in the presence of dexamethasone, ascorbic acid and ,-glycerophosphate, for up to 3 wk. The induction of calcified nodules was visualized by von Kossa stain. SOST mRNA was detected by real-time PCR, and the presence of sclerostin was verified using immunohistochemistry and western blots. Results:, Expression of sclerostin was demonstrated in osteocytes of mouse and human alveolar bone. Distinct immunolocalization in the cementocytes was shown. In periodontal ligament cultures, following mineralization treatment, increasing levels of SOST mRNA as well as of sclerostin protein could be verified. Conclusion:, The identification of SOST/sclerostin in cementocytes and mineralizing periodontal ligament cells adds to our understanding of the biology of the periodontium, but the functional meaning of these findings can only be unravelled after additional in vitro and in vivo studies. [source] Osteoprotegerin induces osteopontin via syndecan-1 and phosphoinositol 3-kinase/Akt in human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2009T. Yongchaitrakul Background and Objective:, Our previous study found that thrombin induced osteoprotegerin synthesis in human periodontal ligament cells. As elevated levels of osteoprotegerin can exert biological effects on various cell types, in the present study we investigated the effect of osteoprotegerin on the expression of osteopontin in human periodontal ligament cells. Material and Methods:, Cultured human periodontal ligament cells were treated with recombinant human osteoprotegerin (0,100 ng/mL) for 24,48 h. The expression of osteopontin mRNA and protein was analyzed using reverse transcription,polymerase chain reaction and western blot analyses, respectively. Phosphoinositol 3-kinase inhibitor, Akt inhibitor, heparinase, neutralizing antibody against receptor activator of nuclear factor-,B ligand (RANKL) and syndecan-1, and small interfering RNA against syndecan-1, were used to determine the mechanism involved. Results:, Osteoprotegerin up-regulated the mRNA and protein expression of osteopontin in human periodontal ligament cells in a dose-dependent manner. Addition of neutralizing antibody against RANKL attenuated the inductive effect of osteoprotegerin on osteopontin expression. In addition, the inductive effect of osteoprotegerin was abolished by phosphoinositol 3-kinase and Akt inhibitors, as well as by syndecan-1 antibody or syndecan-1 small interfering RNA. None of the inhibitors had any effect on the background level of osteopontin expression. Conclusion:, An increased level of osteoprotegerin can generate signals via a RANKL/syndecan-1/phosphoinositol 3-kinase/Akt pathway. The results also suggest that osteopontin is one of the downstream targets of the pathway mediated by osteoprotegerin in human periodontal ligament cells. Thus, in addition to counteracting RANKL in the RANKL,osteoprotegerin system, osteoprotegerin may play a role in periodontal tissue remodeling through modulation of the extracellular matrix. [source] Expression of matrix metalloproteinase-1, matrix metalloproteinase-2 and extracellular metalloproteinase inducer in human periodontal ligament cells stimulated with interleukin-1betaJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2009J. Xiang Background and Objectives: Matrix metalloproteinases (MMPs), produced by both infiltrating and resident cells of the periodontium, play important roles in physiologic and pathologic events. Both interleukin-1beta and extracellular MMP inducer can stimulate the expression of MMPs, which in turn leads to breakdown of the periodontium. However, it is currently unknown whether interleukin-1beta up-regulates MMPs through stimulating the expression of extracellular MMP inducer. The aims of this study were to investigate the effect of interleukin-1beta on the expression of MMP-1, MMP-2 and extracellular MMP inducer in human periodontal ligament cells and to evaluate whether the regulation of MMP-1 and MMP-2 by this cytokine occurred through an effect on extracellular MMP inducer expression. Material and Methods: Cultured human periodontal ligament cells were treated with varying concentrations (0.01,10 ng/mL) of interleukin-1beta at for 6, 12 and 24 h. Reverse transcription,polymerase chain reaction, enzyme-linked immunosorbent assay, gelatin zymography and western blotting were performed to measure the mRNA and protein levels of MMP-1, MMP-2 and extracellular MMP inducer. Results: Basal levels of mRNA and protein for MMP-1, MMP-2 and extracellular MMP inducer were detected in untreated human periodontal ligament cells. Interleukin-1beta significantly up-regulated the expression of MMP-1 and MMP-2 mRNA and protein (p < 0.05); however, the levels of mRNA and protein for extracellular MMP inducer were not significantly different (p > 0.05). In the culture medium, the concentration of MMP-1 was also increased significantly, but the concentration of MMP-1 was not related to the concentration of extracellular MMP inducer (R2 = 0.2538, p > 0.05). Conclusion: Interleukin-1beta up-regulated the levels of MMP-1 and MMP-2, but it did not alter the expression of extracellular MMP inducer. Expression of MMP-1 and MMP-2 might be elevated by interleukin-1beta and extracellular MMP inducer via two different signal pathways. [source] Putative signaling action of amelogenin utilizes the Wnt/,-catenin pathwayJOURNAL OF PERIODONTAL RESEARCH, Issue 3 2009M. Matsuzawa Background and Objective:, While it has long been known that amelogenin is essential for the proper development of enamel, its role has generally been seen as structural in nature. However, our new data implicate this protein in the regulation of cell signaling pathways in periodontal ligament cells and osteoblasts. In this article we report the successful purification of a recombinant mouse amelogenin protein and demonstrate that it has signaling activity in isolated mouse calvarial cells and human periodontal ligament cells. Material and Methods:, To determine the regulatory function of canonical Wnt signaling by amelogenin, we used TOPGAL transgenic mice. These mice express a ,-galactosidase transgene under the control of a LEF/TCF and ,-catenin-inducible promoter. To investigate in greater detail the molecular mechanisms involved in the ,-catenin signaling pathway, isolated osteoblasts and periodontal ligament cells were exposed to full-length recombinant mouse amelogenin and were evaluated for phenotypic changes and ,-catenin signaling using a TOPFLASH construct and the LacZ reporter gene. Results:, In these in vitro models, we showed that amelogenin can activate ,-catenin signaling. Conclusion:, Using the TOPGAL transgenic mouse we showed that amelogenin expression in vivo is localized mainly around the root, the periodontal ligament and the alveolar bone. [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 cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2009H. 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] Subculture affects the phenotypic expression of human periodontal ligament cells and their response to fibroblast growth factor-2 and bone morphogenetic protein-7,in vitroJOURNAL OF PERIODONTAL RESEARCH, Issue 5 2008S. Lossdörfer Background and Objective:, Although periodontal ligament cells display several osteoblastic traits, their phenotypic expression is still not well established. It remains a matter of debate whether they resemble a terminally differentiated cell type or an intermediate maturation state that potentially can be directed towards a fibroblastic or an osteoblastic phenotype. Material and Methods:, To explore the characteristics of periodontal ligament cells in greater detail, fourth-passage, sixth-passage and eighth-passage human periodontal ligament cells were cultured for up to 3 wk. Ki-67, alkaline phosphatase, osteocalcin, osteoprotegerin and receptor activator of nuclear factor-,B ligand (RANKL) mRNA expression was quantified by real-time polymerase chain reaction. Furthermore, the cellular response to fibroblast growth factor-2 and bone morphogenetic protein-7 was examined in first-passage and fourth-passage cells. Dermal fibroblasts (1BR.3.G) and osteoblast-like cells (MG63) served as reference cell lines. Results:, Proliferation decreased over time and was highest in fourth-passage cells. The expression of differentiation parameters, osteoprotegerin and RANKL increased with culture time and was higher in fourth-passage cells than in cells of later passages. The RANKL/osteoprotegerin ratio increased steadily until day 21. Administration of fibroblast growth factor-2 enhanced cell numbers in both passages, whereas alkaline phosphatase and osteocalcin production remained unchanged. By contrast, exposure of periodontal ligament cells to bone morphogenetic protein-7 resulted in a reduction of cell number in the first and fourth passages, whereas the production of alkaline phosphatase and osteocalcin was enhanced. In dermal fibroblasts, differentiation parameters did not respond to both stimuli. MG63 cells behaved similarly to periodontal ligament cells. Conclusion:, These results indicate that subculture affects the phenotypic expression of human periodontal ligament cells with respect to the characteristics that these cells share with osteoblasts. Furthermore, the periodontal ligament cell phenotype can be altered by fibroblastic and osteoblastic growth factors. [source] Expression of receptor activator of nuclear factor kappa B ligand relates to inflammatory bone resorption, with or without occlusal trauma, in ratsJOURNAL OF PERIODONTAL RESEARCH, Issue 5 2007Y. Yoshinaga Background and Objective:, Receptor activator of nuclear factor kappa B ligand (RANKL) is an important factor in osteoclast differentiation, activation and survival; however, its involvement in inflammatory bone resorption, with or without occlusal trauma, is unclear. The purpose of the present study was to investigate the distribution of RANKL-expressing cells in rat periodontium during lipopolysaccharide-induced inflammation with or without occlusal trauma. Material and Methods:, Lipopolysaccharide was injected into rat gingiva of the lower left first molar to induce inflammation. In addition, the occlusal surface of the upper left first molar of rat was raised by placing a gold inlay to induce occlusal trauma in the lower left first molars. The distribution of RANKL-expressing cells was immunohistochemically observed. Results:, In the inflammatory model, many osteoclasts were observed at the apical inter-radicular septum on day 5 and they were reduced by day 10. On the other hand, in the inflammatory model with occlusal trauma, many osteoclasts were still observed on day 10. RANKL expression was similar to the changes in osteoclast number. The expression of RANKL increased in endothelial cells, inflammatory cells and periodontal ligament cells. Conclusion:, These findings clearly demonstrated that RANKL expression on endothelial cells, inflammatory cells and periodontal ligament cells is involved in inflammatory bone resorption and the expression is enhanced by traumatic occlusion. These results suggest that RANKL expression on these cells is closely involved in the increase of osteoclasts induced by occlusal trauma. [source] Mitogen-activated protein kinases mediate interleukin-1,-induced receptor activator of nuclear factor-,B ligand expression in human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 4 2007A. Oikawa Background and Objective:, Interleukin-1,-stimulated receptor activator of nuclear factor-,B ligand (RANKL) expression in human periodontal ligament cells is partially mediated by endogenous prostaglandin E2, whereas mitogen-activated protein kinases (MAPKs) are implicated in regulating various interleukin-1-responsive genes. We investigated herein the involvement of MAPKs in interleukin-1,-stimulated RANKL expression in human periodontal ligament cells. Material and Methods:, Human periodontal ligament cells were pretreated separately with specific inhibitors of MAPKs, including extracellular signal-regulated kinase, p38 MAPK and c-Jun N-terminal kinase, and subsequently treated with interleukin-1,. Following each treatment, the phosphorylation of each MAPK, the expression of RANKL, and the production of prostaglandin E2 were determined. RANKL activity was evaluated using an assay to determine the survival of prefusion osteoclasts. Results:, Interleukin-1, induced RANKL expression at the mRNA and protein levels, as well as RANKL activity in human periodontal ligament cells. Interleukin-1, also activated extracellular signal-regulated kinase, p38 MAPK, and c-Jun N-terminal kinase. Pretreatment with each MAPK inhibitor partially, but significantly, suppressed interleukin-1,-induced RANKL expression and its activity, as well as prostaglandin E2 production. Conclusion:, In human periodontal ligament cells, three types of MAPK inhibitor may abrogate RANKL expression and activity induced by interleukin-1,, directly or indirectly through partial suppression of prostaglandin E2 synthesis. In addition, extracellular signal-regulated kinase, p38 MAPK, and c-Jun N-terminal kinase signals may co-operatively mediate interleukin-1,-stimulated RANKL expression and its activity in those cells. [source] Identification of marker genes distinguishing human periodontal ligament cells from human mesenchymal stem cells and human gingival fibroblastsJOURNAL OF PERIODONTAL RESEARCH, Issue 3 2007T. 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 cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2007S.-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] Aging stimulates cyclooxygenase-2 expression and prostaglandin E2 production in human periodontal ligament cells after the application of compressive forceJOURNAL OF PERIODONTAL RESEARCH, Issue 1 2007Kotoe Mayahara Background and Objectives:, Some clinical studies show that alveolar crestal bone loss is higher in adults than in young patients during orthodontic treatment, but the causes of such a phenomenon have not been elucidated. It is known that prostaglandin E2 (PGE2) is a proinflammatory agent and one of the potent osteoclast-inducing factors, and is produced by human periodontal ligament cells in response to orthodontic force. The aim of this study was to investigate age-related change in the biosynthetic capacity of PGE2 and its regulatory gene, cyclooxygenase 2 (COX-2) from periodontal ligament cells in response to mechanical stress. Methods:, Compressive force of 2 g/cm2 was applied for 3,48 h to periodontal ligament cells obtained from human donors aged 9,50 years, and COX-2 mRNA expression in and PGE2 production by the periodontal ligament cells in response to the compressive force were examined. Results:, Application of a compressive force of 2 g/cm2 for 3,48 h significantly stimulated these factors in both time- and age-dependent manners. Furthermore, these increases were dramatically larger in periodontal ligament cells obtained from donors over the age of 35. Conclusions:, Periodontal ligament cells obtained from old donors have significantly greater COX-2 expression and PGE2 production in response to compressive force than those from younger donors. The turning point of aging, where significantly larger amounts of theses factors begin production, appears to be around the age of 35. These results may be positively related to the acceleration of alveolar crestal bone loss during orthodontic treatment in adult patients. [source] Identification of genes related to mechanical stress in human periodontal ligament cells using microarray analysisJOURNAL OF PERIODONTAL RESEARCH, Issue 1 2007R. M. S. De Araujo Background and Objective:, Differential expression of genes in human periodontal ligament (PDL) under mechanical stress, such as orthodontic force, is thought to be involved in the remodeling of PDL cells and periodontal tissues. However, little is known about the genes expressed in PDL cells under mechanical stress. Material and Methods:, We employed microarray analysis to assess, in a comprehensive manner, the gene expression profiles in PDL cells compressed by a static force using an in vitro three-dimensional culture system. Six genes were selected and validated by quantitative real-time polymerase chain reaction analysis, consistent with the microarray data. Results:, The microarray data revealed that 108 of 30,000 genes tested were differentially expressed by mechanical force loading. Among them, 85 genes were up-regulated by mechanical stress, while 23 genes were down-regulated, judging by the thresholds of a two-fold increase/decrease compared with the controls. Thirty-two of the up-regulated and eight of the down-regulated genes, well-characterized in protein function, were involved in numerous biological processes including cell communication, cell signaling, cell cycle, stress response, and calcium release. However, several genes differentially expressed in our microarray data have not been well defined as stress-response molecules. Conclusion:, Our microarray is the first to show the gene profile in PDL cells caused by mechanical stress; however, further studies to clarify the physiological function of these molecules in PDL cells are required. [source] Chemically modified tetracyclines stimulate matrix metalloproteinase-2 production by periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 5 2006M. M. Bildt Background and Objective:, The purpose of this study was to investigate the effects of chemically modified tetracyclines (CMTs) on the production of gelatinases [matrix metalloproteinase (MMP)-2 and -9] by human periodontal ligament (PDL) cells, and on the activity of recombinant gelatinases. Material and Methods:, Human PDL cells were cultured with CMT-1, -3, -5, -7 or -8 in concentrations of 0, 1, 5, 10, 20, 50, 100, 200 and 500 µm. Gelatin zymography was used to determine MMP-2 and -9 production of the cells. The amount of DNA present in the cultures was analyzed using a fluorescent assay. The cytotoxicity of the CMTs was also determined. Recombinant human MMP-2 and -9 were incubated with the CMTs (0,500 µm) and their activity was analyzed using an internally quenched fluorogenic substrate. Results:, MMP-2 production was stimulated up to sevenfold by CMT-1, -3, -7 and -8 at low concentrations (10,200 µm). No significant amounts of MMP-9 were produced. In contrast, MMP-2 and -9 activity was reduced by ,,10,40-fold at higher concentrations (200,500 µm). CMT-5 had no effect on the production or on the activity of MMP-2 and -9. Only CMT-3 and -8 had cytotoxic effects on the PDL cells at the highest concentrations. Conclusion:, Surprisingly, CMTs are able to stimulate MMP-2 production at relatively low concentrations. However, at higher concentrations they exert a much stronger inhibitory effect on gelatinase activity. A possible stimulatory effect of CMTs on MMP production should be considered in their clinical use. [source] Retinoic acid is a potential negative regulator for differentiation of human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2005Natsuko Shibuya Background and objectives:, Retinoic acid (RA) exerts a wide variety of effects on development, cellular differentiation and homeostasis in various tissues. However, little is known about the effects of RA on the differentiation of periodontal ligament cells. In this study, we investigated whether RA can affect the dexamethasone-induced differentiation of periodontal ligament cells. Methods and results:, Human periodontal ligament cells were differentiated via culturing in the presence of dexamethasone, ascorbic acid, and ,-glycerophosphate for mineralized nodule formation, as characterized by von Kossa staining. Continuous treatment with all- trans -RA inhibited the mineralization in a dose-dependent manner, with complete inhibition over 1 µm RA. Other RA analogs, 9- cis -RA and 13- cis -RA, were also effective. Furthermore, addition of RA for just the first 4 days completely inhibited the mineralization; however, as RA was added at later stages of culture, the inhibitory effect was diminished, suggesting that RA had a phase-dependent inhibition of mineralization. RA receptor (RAR)-, agonist (AM-580), but not retinoid X receptor agonist (methoprene acid), inhibited the mineralization, and reverse transcription,polymerase chain reaction analysis revealed that RAR-, was expressed on the cells, suggesting that RAR-, was involved in the inhibitory mechanism. This inhibition was accompanied by inhibition of alkaline phosphatase activity; however, neither expression of platelet-derived growth factor (PDGF) receptor-,, PDGF receptor-,, or epidermal growth factor (EGF) receptor, nor phosphorylation of extracellular signal-regulated kinases triggered by PDGF-ascorbic acid or PDGF-BB was changed, as assessed by flow cytometry or western blot analyses. Conclusions:, These findings suggest that RA is a potential negative regulator for differentiation of human periodontal ligament cells. [source] Behavior of human periodontal ligament cells on CO2 laser irradiated dentinal root surfaces: an in vitro studyJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2004V. Pant Objective:, The aim of this study was to investigate the in vitro attachment behavior of human periodontal ligament fibroblasts on periodontally involved root surface after conditioning with CO2 laser and to compare its efficacy with chemical conditioning agents, namely tetracycline hydrochloride, citric acid, hydrogen peroxide (H2O2) and EDTA, using scanning electron microscopy. Methods:, A total of 84 scaled and root-planed specimens from periodontally involved single-rooted human teeth showing hopeless prognosis were selected and assigned to two groups. One group was lased with a CO2 laser (from 5 cm at 3 W for 0.8, 1.0 and 1.2 s), and the other group was treated with either tetracycline hydrochloride (2.5%), citric acid (saturated solution, pH 1), H2O2 (6%) or EDTA (5%; pH 7.4) for 3 min. The specimens were then seeded with human periodontal ligament fibroblasts, incubated for either 12 h or 24 h, and then the cell attachment behavior was observed. Results:, CO2 laser irradiation for 1.0 s was found to be the most efficient, showing consistently good cell attachment with the highest mean value (15.00 ± 3.41 cells/10,000 µm2 after incubation for 12 h and 29.17 ± 2.04 cells/10,000 µm2 after 24 h), followed by irradiation for 0.8 s (13.11 ± 3.04 cells/10,000 µm2 after incubation for 12 h and 22.91 ± 7.10 cells/10,000 µm2 after 24 h). Charring was observed following irradiation for 1.2 s. Amongst chemical conditioning agents, citric acid was found to be the most efficient, with a mean cell attachment of 17.82 ± 2.16 cells/10,000 µm2 after incubation for 12 h and 23.62 ± 1.94 cells/10,000 µm2 after 24 h. EDTA and H2O2 did not do well in the study. Conclusion:, The results suggest that CO2 laser irradiation for 1.0 s may promote comparatively better attachment of periodontal ligament fibroblast on dentinal root surfaces than the conventional chemical conditioning agents used in the study. [source] Growth hormone regulates osteogenic marker mRNA expression in human periodontal fibroblasts and alveolar bone-derived cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 4 2003H. R. Haase Background:, Growth hormone (GH) is a potent regulator of bone formation. The proposed mechanism of GH action is through the stimulation of osteogenic precursor cell proliferation and, following clonal expansion of these cells, promotion of differentiation along the osteogenic lineage. Objectives:, We tested this hypothesis by studying the effects of GH on primary cell populations of human periodontal ligament cells (PLC) and alveolar bone cells (ABC), which contain a spectrum of osteogenic precursors. Methods:, The cell populations were assessed for mineralization potential after long-term culture in media containing ,-glycerophosphate and ascorbic acid, by the demonstration of mineral deposition by Von Kossa staining. The proliferative response of the cells to GH was determined over a 48-h period using a crystal violet dye-binding assay. The profile of the cells in terms of osteogenic marker expression was established using quantitative reverse transcriptase polymerase chain reaction (RT-PCR) for alkaline phosphatase (ALP), osteopontin, osteocalcin, bone sialoprotein (BSP), as well as the bone morphogenetic proteins BMP-2, BMP-4 and BMP-7. Results:, As expected, a variety of responses were observed ranging from no mineralization in the PLC populations to dense mineralized deposition observed in one GH-treated ABC population. Over a 48-h period GH was found to be non-mitogenic for all cell populations. Quantitative reverse transcriptase polymerase chain reaction (RT-PCR) BSP mRNA expression correlated well with mineralizing potential of the cells. The change in the mRNA expression of the osteogenic markers was determined following GH treatment of the cells over a 48-h period. GH caused an increase in ALP in most cell populations, and also in BMP expression in some cell populations. However a decrease in BSP, osteocalcin and osteopontin expression in the more highly differentiated cell populations was observed in response to GH. Conclusion:, The response of the cells indicates that while long-term treatment with GH may promote mineralization, short-term treatment does not promote proliferation of osteoblast precursors nor induce expression of late osteogenic markers. [source] Activation of MMP-2 by Porphyromonas gingivalis in human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2003Kassara Pattamapun It has been reported that matrix metalloproteinase (MMP) produced by host cells plays a major role in periodontal tissue destruction. In addition, secreted virulence factors from Porphyromonas gingivalis can alter MMP secretion and cause activation in host cells that lead to the tissue degradation. In this study, we examine the effects of P. gingivalis supernatant on matrix metalloproteinase-2 (MMP-2) activation in human periodontal ligament (HPDL) cells. Cultures of HPDL cells were treated with P. gingivalis supernatant for 48 h and the level of MMP-2 activation was monitored by gelatin zymography. The profound activation of MMP-2 was seen only in the treated group. The activation of MMP-2 was inhibited by MMP inhibitors phenanthroline and EDTA, but not serine protease or cysteine protease inhibitors. To study the correlation between the expression of membrane-type-1 matrix metalloproteinase (MT1-MMP) and the activation of MMP-2, the level of MT1-MMP was analyzed. The results from reverse-transcription polymerase chain reaction (RT-PCR) and Western analysis indicated that P. gingivalis supernatant up-regulated the expression of MT1-MMP in both transcription and translation levels within 48 h. These results suggest that P. gingivalis supernatant can activate MMP-2 in HPDL cells and the mechanism of activation may involve the increased amount of MT1-MMP. It is possible that the activation of MMP-2 by P. gingivalis plays a role in the process of chronic periodontitis. [source] Basic fibroblast growth factor induces the expression of matrix metalloproteinase-3 in human periodontal ligament cells through the MEK2 mitogen-activated protein kinase pathwayJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2003Atsushi Shimazu Basic fibroblast growth factor (bFGF, FGF-2) is one of the potent mitogens for periodontal ligament (PDL) cells. However, the role of bFGF on the matrix metalloproteinase-3 (MMP-3) expression in PDL cells is unknown. In this study, the effect of bFGF on MMP-3 expression in PDL cells and the mechanism of this process were examined. Human PDL cells were exposed to bFGF at various concentrations (0.01,10 ng/ml) in monolayer cultures. bFGF increased [3H]thymidine incorporation and suppressed proteoglycan synthesis concentration-dependently. However, similar concentration ranges of bFGF increased the release of the cell-associated proteoglycans into the medium. Furthermore, bFGF increased MMP-3 mRNA levels concentration-dependently as examined by reverse transcription-polymerase chain reaction (RT-PCR). Induction of MMP-3 after the stimulation with bFGF was observed as early as 12 h with maximal at 24 h. Thereafter, the MMP-3 mRNA level gradually decreased until 72 h. Cycloheximide blocked the induction of MMP-3 by bFGF, indicating the requirement of de novo protein synthesis for this stimulation. Furthermore, MMP-3 expression induced by bFGF was abrogated by U0126, a specific inhibitor of MEK1/2 and ERK1/2 in mitogen-activated protein (MAP) kinase pathway, not by PD98059, a specific inhibitor of MEK1. In addition, bFGF up-regulated the phosphorylated ERK1/2 in 5 min with the maximal at 20 min as examined by Western blotting, and U0126 inhibited the ERK1/2 phosphorylation induced by bFGF. These findings suggest that bFGF induces MMP-3 expression in PDL cells through the activation of the MEK2 in MAP kinase pathway. bFGF stimulation on MMP-3 synthesis may be involved in the control of the cell-associated proteoglycans in PDL cells during periodontal regeneration and degradation. [source] Expression of receptor activator of NF-kappa B ligand and osteoprotegerin in culture of human periodontal ligament cellsJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2002Tomokazu Hasegawa The receptor activator of NF-kappa B ligand (RANKL) and its decoy receptor, osteoprotegerin (OPG), are the important proteins implicated in osteoclastogenesis. In this study, we investigated the expressions of RANKL and OPG in cultured human periodontal ligament (PDL) cells and their roles in osteoclastogenesis. Northern blotting revealed that the OPG mRNA was down-regulated remarkably by application of 10,8 m one-alpha, 25-dihydroxyvitamin D3[1,25-(OH)2D3] and 10,7 m dexamethasone (Dex). In contrast, RANKL mRNA was up-regulated by the same treatment. Western blotting demonstrated decrease of OPG by the application of 1,25-(OH)2D3 and Dex. Tartrate-resistant acid phosphatase-positive multinuclear cells were markedly induced when the PDL cells were cocultured with mouse bone marrow cells in the presence of an anti-OPG antibody together with 1,25-(OH)2D3 and Dex. These results indicate that PDL cells synthesize both RANKL and OPG and that inactivation of OPG may play a key role in the differentiation of osteoclasts. [source] Phenotypic comparison of periodontal ligament cells in vivo and in vitroJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2001P. Lekic The mammalian periodontal ligament contains heterogeneous populations of connective tissue cells, the precise function of which is poorly understood. Despite close proximity to bone and the application of high amplitude physical forces, cells in the periodontal ligament (PL) are capable of expressing regulatory factors that maintain PL width during adult life. The study of PL homeostasis and PL cell differentiation requires culture and phenotypic methods for precise characterization of PL cell populations, in particular those cells with an inherently osteogenic program. Currently it is unknown if cells cultured from the PL are phenotypically similar to the parental cells that are present in the tissues. We have compared the phenotype of cells in vivo with cells derived from the PL and expanded in vitro to assess the general validity of in vitro models for the study of phenotypic regulation in vivo. Rat PL cells were isolated by either scraping the root of the extracted first mandibular molars (Group A), or by scraping the alveolar socket following extraction of first mandibular molars (Group B), or by obtaining a mixture of cells after disaggregating a block of tissue consisting of first mandibular molar, PL and the surrounding alveolar bone (Group C). Cultured cells at confluence were fixed and immunostained for ,-smooth muscle actin (,-SMA), osteopontin (OPN), alkaline phosphatase (AP), or bone sialoprotein (BSP). For in vivo assessments, frontal sections of rat first mandibular molar were immunostained for ,-SMA, OPN, AP and BSP. We examined osteogenic differentiation of cultured PL cell cultures by bone nodule-forming assays. In vivo and at all examined sites, >68% of PL cells were immunostained for AP; ,50% and ,51% for OPN and ,-SMA (p=0.3), respectively, while only ,8% were positively stained for BSP (p<0.01). Analysis of cultured PL cells in Groups A, B and C showed 54%, 53% and 56% positive staining for ,-SMA respectively; 51%, 56%, 54% for OPN; 66%, 70%, 69% for AP and 2.2%, 1.4% and 2.8% for BSP. The mean percentage of PL cells in situ stained for the different markers was similar to that of cultured PL cells (Group A,Group B,Group C in situ for p>0.2) except for BSP which was 3 to 4 fold higher in vivo(p<0.01). PL cell cultures treated with dexamethasone showed mineralized tissue formation for all groups (A, B, C), but no mineralized tissue formation was detected in the absence of dexamethasone. As PL cells express quantitatively similar phenotypes in vitro and in vivo, we conclude that the in vitro models used here for assessment of PL cell differentiation appear to be appropriate and are independent of the cell sampling method. Further, dexamethasone-dependent progenitors are present both on the root and bone-related sides of the PL. [source] Cyclic stretching force-induced early apoptosis in human periodontal ligament cellsORAL DISEASES, Issue 3 2008W Zhong Objective:, Human periodontal ligament (PDL) cells occur changes in morphology and express relative protein by stretching force. However, whether stretching force, especially excessive stretching force, induces PDL cell apoptosis is not yet clearly understood. In the present study we investigated the relationship between early apoptosis and stretching force in human PDL cells in vitro. Materials and methods:, The human PDL cells were obtained from healthy premolars. After three to five passages, the cells were stretched by strain 1%, 10% and 20% for 30 min, 1 h, 6 h and 12 h, then early apoptosis were detected through annexin fluorescein isothiocyanate (V-FITC) binding by flow cytometry and confocal laser scanning microscopy. Results:, The experiments indicated that human PDL apoptotic cells in the early stage increased in a time- and force-dependent manner in response to stretching strain within 6 h, and then apoptosis decreased at 12 h. Human PDL cells which stretched inclined parallel to each other and aligned their long axis perpendicular to the stretching force vector, but in the centre of the disc, cells showed minimal deformation and unidirectional alignment of PDL cells. Conclusion:, The overall results suggested that stretching force not only influenced morphology but also induced early apoptosis in human PDL cells. [source] | |||||||||||||||||||||||||