人牙周膜成纤维细胞在无血清培养液中的生长特性

目的：探讨人牙周膜成纤维细胞在无血清培养液中的生长特性。方法：用倒置显微镜和MTT法观察人牙周膜成纤维细胞在无血清培养液中的生长和增殖变化。结果：人牙周膜成纤维细胞在无血清培养条件下可以生长的增殖，但与含血清培养液相比，其增殖速率变缓，分化明显。结论：无血清培养液可应用于人牙周膜成纤维细胞的体外培养和实验研究中。     

水蛭素对牙龈成纤维细胞碱性成纤维细胞生长因子和转化生长因子-β1表达的影响

目的 观察水蛭素对人牙龈成纤维细胞（HGFs）碱性成纤维细胞生长因子（bFGF）及转化生长因子-β1（TGF-β1）表达变化的规律,探讨水蛭素影响牙龈改建的可能作用机制。方法 体外培养并鉴定HGFs,利用不同浓度的水蛭素分别作用于正常（对照组）和受长期机械外力作用后增生的HGFs（实验组）,通过实时定量聚合酶链反应法和免疫细胞化学法检测TGF-β1及bFGF的表达。结果 未加入水蛭素时,受长期机械外力作用后,实验组促进HGFs增殖胶原合成的TGF-β1表达升高,而抑制胶原合成的bFGF表达降低（P<0.05）。加入水蛭素干预增生的HGFs后,可正向调节bFGF表达,而负向调节TGF-β1的表达（P<0.05）。结论 外力作用干扰了HGFs胶原合成与降解之间的平衡,水蛭素可能通过调节这一平衡而促进牙龈改建过程。     

Characterization of human gingival keratinocytes cultured in a serum-free medium

Primary cultures of keratinocytes were established from gingival tissue explanted on the surface of type I collagen gels and fed a serum-containing medium. Cells could be routinely subcultured for at least five passages in a basal nutrient medium (MCDB 153) containing low calcium (0.1 mM), and supplemented with ethanolamine, phosphoethanolamine, hydrocortisone, insulin, epidermal growth factor and protein of bovine pituitary extract. Cells seeded at low densities doubled exponentially in number every 24-30 h and formed a confluent monolayer within 10-14 days. Phase-contrast light and transmission electron microscopy showed that the keratinocyte cultures had features typical of epithelial cells, including desmosomes and perinuclear tonofilament bundles. Immunofluorescent microscopy showed the presence of specific keratin proteins in basal cells of proliferating cultures. Gel electrophoresis of the insoluble cytosolic proteins of gingival and skin keratinocytes showed several differences. Suspension of dividing gingival keratinocytes in 1.3% methylcellulose medium induced greater than 50% cross-linked envelopes, suggesting the existence of a terminal differentiation pathway in gingival basal cells. Clonal growth experiments showed that both insulin and epidermal growth factor were required for optimal clonal growth. The growth of subcultures was arrested and the unstratified epithelial monolayer induced to form a stratified sheet by replacing the growth medium with basal MCDB 153 medium depleted of growth factors and containing 2 mM calcium. Sheets of stratified gingival epithelium formed on and later released from the dish by enzymatic treatment may be suitable for a variety of experimental and clinical uses.     

人血小板源性生长因子-B基因转染犬牙龈成纤维细胞的瞬时表达检测

目的 检测携带人血小板源性生长因子-B(hPDGF-B)基因的真核表达质粒在Beagle犬牙龈成纤维细胞内的表达.方法 扩增和鉴定携带hPDGF-B基因的质粒EX-A0380-M03,脂质体介导的方法转染Bealge犬牙龈成纤维细胞,RT-PCR、免疫细胞化学、ELISA以及Western blot检测hPDGF-BB...     

The synthesis of 5α - dihydrotestosterone from androgens by human gingival tissues and fibroblasts in culture in response to TGIF-β and PDGF

The effects of TGF-β and PDGF on the metabolic conversion of 14C-testosterone by human gingival tissue (HGT) from 5 subjects was investigated. The metabolic conversions in response to TGF-β and PDGF were also studied in 4-6 cell-lines of cultured gingival fibroblasts, using 14C-testosterone and 14C-4-androstenedi one as substrates. Duplicate incubations of HGT were performed in Eagle's MEM + 10% FCS and optimal stimulatory concentrations of TGF-β/PDGF for 24 h. Similar incubations were performed in duplicate with cell-lines of cultured gingival fibroblasts, TGF-β PDGF, 14C-testosterone/14C-4-androstenedione in Eagle's MEM + 10% FCS. The radioactive metabolites were extracted, separated and quantified. With HGT, TGF-β and PDGF caused 2.5/12-fold increases in DHT synthesis (p< 0.1; Wilcoxon signed rank test) and 3.4/12-fold increases in 4-androstenedione formation (p<0.1) from 14C-testosterone. PDGF increased DHT and testosterone synthesis from 14C-4-androstenedione by 3-fold in gingivae (p<0.1). With cell-lines, average values of duplicate incubations showed 2.8/2-fold increases in DHT synthesis from 14C-testosterone in response to TGF-β/PDGF (p<0.1; p<0.2) and 2.4/2-fold increases in 4-androstenedione synthesis (p<0.1; p<0.2). With 14C-4-androstenedione as substrate, TGF-β/PDGF caused 1.611.9fold increases in DHT synthesis compared with controls (p<0.05; p<0.1) and 1.711.5-fold increases in testosterone formation from this substrate (p<0.05; p<0.1). Due to the strong implications of TGF-β/PDGF and anabolic androgens on matrix repair, significant increases in DHT synthesis from 2 androgenic substrates in response to TGF-β and PDGF are of particular relevance to inflammatory repair processes.     

Effects of enamel matrix derivative and transforming growth factor-beta1 on human periodontal ligament fibroblasts

AIM: The objective of this study was to evaluate the effects of enamel matrix derivative (EMD), transforming growth factor-beta1 (TGF-beta1), and a combination of both factors (EMD+TGF-beta1) on periodontal ligament (PDL) fibroblasts. MATERIAL AND METHODS: Human PDL fibroblasts were obtained from three adult patients with a clinically healthy periodontium, using the explant technique. The effects of EMD, TGF-beta1, or a combination of both were analysed on PDL cell proliferation, adhesion, wound healing, and total protein synthesis, and on alkaline phosphatase (ALP) activity and bone-like nodule formation. RESULTS: Treatment with EMD for 4, 7, and 10 days increased cell proliferation significantly compared with the negative control (p<0.05). At day 10, EMD and EMD+TGF-beta1 showed a higher cell proliferation compared with TGF-beta1 (p<0.01). Cell adhesion was significantly up-regulated by TGF-beta1 compared with EMD and EMD+TGF-beta1 (p<0.01). EMD enhanced in vitro wound healing of PDL cells compared with the other treatments. Total protein synthesis was significantly increased in PDL cells cultured with EMD compared with PDL cells treated with TGF-beta1 or EMD+TGF-beta1 (p<0.05). EMD induced ALP activity in PDL fibroblasts, which was associated with an increase of bone-like nodules. CONCLUSION: These findings support the hypothesis that EMD and TGF-beta1 may play an important role in periodontal regeneration. EMD induced PDL fibroblast proliferation and migration, total protein synthesis, ALP activity, and mineralization, while TGF-beta1 increased cellular adhesion. However, the combination of both factors did not positively alter PDL fibroblast behaviour.     

Immunohistochemical detection of hepatocyte growth factor, transforming growth factor-β and their receptors in epithelial odontogenic tumors

BACKGROUND: Tumors derived from odontogenic epithelium exhibit considerable variation and are classified into several benign and malignant entities. To clarify the role of growth factors in oncogenesis, cytodifferentiation and progression of epithelial odontogenic tumors, expression of hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-beta) and their receptors were analyzed in these tumors as well as in tooth germs. METHODS: Specimens of five tooth germs, 34 ameloblastomas, three calcifying epithelial odontogenic tumors (CEOTs), two clear cell odontogenic tumors (CCOTs), five adenomatoid odontogenic tumors (AOTs), six calcifying odontogenic cysts (COCs) and six malignant ameloblastomas were examined immunohistochemically with the use of antibodies against HGF, TGF-beta and their receptors. RESULTS: In tooth germs and epithelial odontogenic tumors, immunoreactivity for HGF and TGF-beta was detected in both epithelial and mesenchymal cells, while expression of their receptors was found only in epithelial cells. In tooth germs and main types of ameloblastomas, HGF and TGF-beta reactivity was marked in epithelial cells near the basement membrane, and their receptors were diffusely positive in most epithelial cells. In subtypes of ameloblastomas, reduced expression of HGF, c-Met and TGF-beta and increased reactivity for TGF-beta receptors were detected in keratinizing cells in acanthomatous ameloblastomas, and granular cells in granular cell ameloblastomas demonstrated little or no expression of HGF, TGF-beta or their receptors. As compared with main types of ameloblastomas, basal cell ameloblastomas showed high HGF reactivity, and desmoplastic ameloblastomas exhibited elevated reactivity for TGF-beta and its receptors. Neoplastic cells in CEOTs, AOTs and COCs showed reactivity for HGF, TGF-beta and their receptors. Elevated HGF and TGF-beta reactivity was found in pseudoglandular cells in AOTs, and high expression of their receptors was noted in ghost cells in COCs. Metastasizing ameloblastomas showed similar expression patterns of HGF, TGF-beta and their receptors to those of benign ameloblastomas, while CCOTs and ameloblastic carcinomas had increased HGF expression and low reactivity for TGF-beta and its receptors as compared with benign ameloblastomas. CONCLUSIONS: Immunohistochemical localization of HGF, TGF-beta and their receptors in tooth germs and epithelial odontogenic tumors supports the hypothesis that HGF and TGF-beta act on epithelial cells via paracrine and autocrine mechanisms. Altered expression of the agents in these epithelial odontogenic tumors, especially subtypes of ameloblastomas, AOTs and COCs, suggests that HGF and TGF-beta signaling might affect differentiation of neoplastic odontogenic epithelial cells. Activated HGF/c-Met pathway and reduced TGF-beta signaling in CCOTs and ameloblastic carcinomas may be associated with the malignant potential of these epithelial odontogenic tumors.     

Immunolocalization of platelet-derived growth factor A and B chains and PDGF-α and β receptors in human gingival wounds

Platelet-derived growth factor (PDGF) is a polypeptide growth factor which has been implicated as a major mitogen involved in wound healing. The PDGF appears to promote periodontal regeneration; however, its distribution in gingival tissues is not known and how it participates in gingival wound healing is unclear. Using highly specific antibodies we have studied the distribution of PDGF A and B chains and α- and β-PDGF receptors in healing human gingival wounds. Wounds were created by making a 0.75 mm deep incision in the papilla of healthy human gingiva and biopsies were obtained from the same site after 8 h and 1, 3, 7, 14 and 21 d. Frozen sections were immunostained with affinity purified antibodies. The results showed that both epithelium and fibrin clot manifested positive immunostaining for anti-PDGF-A and B-chain antibodies. Staining was present in unwounded and wounded epithelia, and in the fibrin clot it appeared to be more intense for the PDGF-A chain. Blood vessels in connective tissue were also positive while other areas were largely negative. No significant staining was detectable in healthy tissues for anti-PDGF-α or -β receptor antibodies. However, the wound site began to manifest positive immunostaining for anti-β-receptor antibody after 3 d of healing, became maximal at 7d, and then decreased. Our data indicate, but do not prove, that gingival epithelium may be a source of PDGF A and B chains and that the A chain may have a more prominent role to play during early stages of healing. Expression of PDGF β-receptor appears later at the wound site, indicating that the PDGF B isomer may regulate later wound healing events.     

Effects of cytokines on gingival fibroblasts in vitro are modulated by the extracellular matrix

Fibroblast function in gingival tissue is thought to be regulated by the local cellular environment – both the extracellular matrix and soluble factors. In an attempt to artificially re-create this situation fibroblasts have been cultured within 3-dimensional collagen gels in an environment more physiologically comparable to connective tissue. Using such a model we investigated the effects of the extracellular matrix on gingival fibroblast growth and synthetic activity and on the cellular responsiveness to 4 soluble factors – epidermal growth factor (EGF), platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF- β ₁) and interleukin-1 β (IL-1 β ). Fibroblasts cultured within collagen gels showed similar growth rates, an increased production of collagen but reduced levels of hyaluronan synthesis in comparison to cells in monolayer culture. Cellular responsiveness to soluble mediators was also modulated by the collagen matrix, with a generalised reduction in response by cells embedded within the matrix. The stimulatory effects of EGF and PDGF on cell growth in monolayer over a 14-day period were only found during the initial stages of culture within gels. Similarly the stimulation of matrix production by cells induced by TGF- β ₁, on plastic was reduced or even negated when cells were cultured in collagen gels. On plastic IL-1 β significantly stimulated cell growth but had no effect on either collagen or hyaluronan production by fibroblasts. In gel cultures, this cytokine had no effect on cell proliferation, but significantly inhibited both collagen and hyaluronan synthesis. These findings further illustrate the usefulness of fibroblast-populated collagen gels as a model system for studying the modulatory effects of soluble factors and extracellular matrix molecules on fibroblast function in vitro .     

Expression of platelet-derived growth factor-AA and platelet-derived growth factor-α receptor in ameloblastomas

Background:  Platelet-derived growth factor (PDGF)-AA isoform and its receptor, PDGF-α receptor (PDGFRA) regulate tooth development and growth. We investigated the expression of both proteins in ameloblastomas, to contribute the understanding of the potential role of the PDGF/PDGFR system in this odontogenic neoplasm.
Method:  Twenty-nine specimens of ameloblastoma were analyzed for PDGF-AA and PDGFRA expression using immunohistochemistry. The proliferation activity was investigated with the MIB-1 antibody. Additionally, capillary sequencing of genomic DNA was performed to search for mutations in therapeutically relevant exons 12 and 18 of the PDGFRA gene.
Results:  PDGF-AA and PDGFRA expression were detectable in all cases with the exception of one tumor. However, protein expression levels did neither correlate with each other nor with MIB-1 expression. Unicystic ameloblastomas did not differ from solid tumors with regard to PDGF-AA, PDGFRA, and MIB-1 expression. One tumor revealed a somatic mutation of exon 12 of the PDGFRA gene.
Conclusion:  PDGF-AA and PDGFRA proteins are regularly expressed in variable levels in ameloblastomas, and somatic mutations of exon 12 and exon 18 of the PDGFRA gene are rare findings.     

Effects of connective tissue growth factor on human periodontal ligament fibroblasts

ObjectiveThe aim of this study was to evaluate the effects of different concentrations of connective tissue growth factor (CTGF) on human periodontal ligament fibroblasts(HPLFs).DesignHPLFs were cultured and identified. Then, different concentrations of CTGF (1, 5, 10, 50, 100 ng/ml) were added to the HPLF culture. Next, CCK-8 assays, alkaline phosphatase (ALP) assays, hydroxyproline determination, alizarin red staining methods, Transwell chambers and real-time PCR methods were applied to observe the effects of CTGF on the proliferation, ALP activity, synthesis of collagen, formation of mineralized nodules and migration. We also studied expression of ALP, fiber link protein (FN), integrin-binding sialoprotein (IBSP), osteocalcin (OC), and integrin beta 1 (ITGB1) mRNA by HPLFs. Statistical significance was assumed if P < 0.05 or P < 0.01.ResultsThe addition of CTGF (1, 5, 10 ng/ml) remarkably promoted the proliferation and collagen synthesis of HPLFs compared with controls. CTGF (1, 5, 10, 50 ng/ml) improved ALP activity of HPLFs, and at all concentrations, CTGF (1, 5, 10, 50, 100 ng/ml) improved the expression of ALP, FN, IBSP and ITGB1 mRNA. In addition, CTGF (1, 5, 10, 50, 100 ng/ml) promoted the migration of HPLFs, which was dose-dependent, with maximal promotion in the 10 ng/ml group (P < 0.05 or P < 0.01).ConclusionsThus, in a certain range of concentrations, CTGF can promote the biological effects, including proliferation, migration and collagen synthesis of HPLFs, to promote the differentiation of HPLFs in the process of osteogenesis.     

体外培养碱性成纤维细胞因子对人牙周膜成纤维细胞生物学特性的影响

目的：探讨体外培养过程中,碱性成纤维细胞生长因子（bFGF）对人牙周膜成纤维细胞（hPDLFs）生物学特性的影响。方法：体外分离培养人牙周膜成纤维细胞并鉴定,加入不同浓度bFGF（1ng／ml、10ng／ml、50ng／ml、100ng／ml）培养,MTT方法检测细胞增殖情况;并对细胞进行矿化诱导,检测细胞的碱性磷酸酶活性。结果：hPDLFs呈星形或长梭形,免疫组化波丝蛋白阳性,角蛋白阴性,证实该细胞来源可靠。bFGF在一定浓度范围内,与细胞增殖成正比,而在本实验培养条件下（10％FBS）bFGF最大效应浓度为10ng／ml。矿化诱导条件下,碱性磷酸酶活性明显增加,在联合应用bFGF的情况下,100ng／ml组碱性磷酸酶活性明显高于其他组。结论：不同浓度bFGF对人牙周膜成纤维细胞生物学行为的影响不同,在一定浓度条件下,低浓度bFGF促进人牙周膜成纤维细胞的增殖,而高浓度bFGF作用于人牙周膜成纤维细胞可能更易于分化。     

Vascular endothelial growth factor, fibroblast growth factor 2, platelet derived growth factor and transforming growth factor beta released in human dental pulp following orthodontic force

OBJECTIVE: The release of four diffusible angiogenic growth factors in human dental pulp following orthodontic force was investigated by using neutralising growth factor antibodies (NAs), individually and in four different combinations to block their effects. This study investigated if increasing the number of NAs (anti h vascular endothelial growth factor (VEGF), anti h fibroblast growth factor (FGF2), anti h platelet derived growth factor (PDGF) and anti Transforming growth factor beta (TGFbeta)) in combination resulted in a progressive reduction of the angiogenic response of the pulp. MATERIALS AND METHODS: The dental pulps from two groups of 40 premolar teeth, four teeth from each of 20 patients treated with fixed appliances for 2 weeks, were divided vertically, and sections from each half pulp co-cultured with sections of rat aorta in collagen. In one group, one of each of the four NAs, and in the other group, one of the four different NA combinations were added to the media of the co-cultures from one half of the pulp from each of the four teeth of each patient; the other half pulp co-cultures were controls. Cultures were examined daily by light microscopy for growth and number of microvessels. RESULTS: NAs significantly reduced microvessel numbers in the co-cultures when added individually (P<0.004), and in each of the four combinations (P<0.002), with a trend to progressively reduced microvessel numbers with increasing number of NAs in combination. CONCLUSIONS: Results indicated that all four angiogenic growth factors examined were released following orthodontic force application and play a role in the angiogenic response of the pulp, and that these factors may be more effective in combination.     

Down-regulation of epidermal growth factor receptor-dependent signaling by Porphyromonas gingivalis lipopolysaccharide in life-expanded human gingival fibroblasts

Background and Objective: Human gingival fibroblasts exhibit proliferative responses following epidermal growth factor exposure, which are thought to enhance periodontal regeneration in the absence of bacterial products such as lipopolysacharide. However, lipopolysaccharide challenge activates human gingival fibroblasts to release several inflammatory mediators that contribute to the immune response associated with periodontitis and attenuate wound repair. We tested the hypothesis that Porphyromonas gingivalis lipopolysaccharide‐activated signaling pathways down‐regulate epidermal growth factor receptor‐dependent events. Material and Methods: To study lipopolysaccharide/epidermal growth factor interactions in human gingival fibroblasts, we introduced the catalytic subunit of human telomerase into human gingival fibroblasts, thereby generating a more long‐lived cellular model. These cells were characterized and evaluated for lipopolysaccharide/epidermal growth factor responsiveness and regulation of epidermal growth factor‐dependent pathways. Results: Comparison of human telomerase‐transduced gingival fibroblasts with human gingival fibroblasts revealed that both cell lines exhibit a spindle‐like morphology and express similar levels of epidermal growth factor receptor, CD14 and Toll‐like receptors 2 and 4. Importantly, human telomerase‐transduced gingival fibroblasts proliferation rates are increased 5–9 fold over human gingival fibroblasts and exhibit a longer life span in culture. In addition, human telomerase‐transduced gingival fibroblasts and human gingival fibroblasts exhibit comparable profiles of mitogen‐activated protein kinase kinase (extracellular signal‐regulated kinase 1/2) activation upon epidermal growth factor or P. gingivalis lipopolysaccharide administration. Interestingly, treatment with P. gingivalis lipopolysaccharide leads to a down‐regulation of epidermal growth factor‐dependent extracellular signal‐regulated kinase 1/2, p38 and cyclic‐AMP response element binding protein phosphorylation in both cell types. Conclusion: These studies demonstrate that human telomerase‐transduced gingival fibroblasts exhibit an extended life span and recapitulate human gingival fibroblasts biology. Moreover, this system has allowed for the first demonstration of lipopolysaccharide down‐regulation of epidermal growth factor activated pathways in human gingival fibroblasts and should facilitate the analysis of signaling events relevant to the pathogenesis and treatment of periodontitis.     

Enhanced expression of vascular endothelial growth factor by periodontal pathogens in gingival fibroblasts

Vascular endothelial growth factor (VEGF) has recently attracted attention as a potent inducer of vascular permeability and angiogenesis. Aberrant angiogenesis is often associated with lesion formation in chronic periodontitis. The aim of the present study was to investigate the properties of VEGF expression in human gingival fibroblasts (HGF) culture. HGF were stimulated with lipopolysaccharide (LPS), vesicle (Ve) and outer membrane protein (OMP) from Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. HGF constitutively produced VEGF and levels were significantly enhanced (P < 0.01) by stimulation with Ve and OMP from A. actinomycetemcomitans and P. gingivalis at concentrations of 10 microg/ml or higher. On the other hand, VEGF levels were not increased by LPS stimulation. VEGF mRNA expression was also observed in Ve- and OMP-stimulated HGF. A vascular permeability enhancement (VPE) assay was performed using guinea pigs to ascertain whether supernatant from cultures of Ve- and OMP-stimulated HGF enhance vascular permeability in vivo. Supernatant from cultures of Ve- and OMP-stimulated HGF strongly induced VPE. This was markedly suppressed upon simultaneous injection of anti-VEGF polyclonal antibodies with the supernatant. Heating and protease treatment of the stimulants reduced the VEGF enhancing levels in Ve and OMP in vitro. These results suggest that Ve and OMP may be crucial heat-labile and protease-sensitive components of periodontal pathogens that enhance VEGF expression. In addition, VEGF might be associated with the etiology of periodontitis in its early stages according to neovascularization stimulated by periodontal pathogens causing swelling and edema.     

Effects of nicotine on apoptosis in human gingival fibroblasts

Aim

Cigarette smoke is a complex mixture of more than 4700 chemical compounds including free radicals and oxidants and it is a world widely known problem to health. Nicotine is the major compound of tobacco and known as the cause of gingivitis and periodontitis. It induces intracellular oxidative stress recognized as the important agent in the damage of biological molecules. The aim of this study is to clarify the cytotoxic pathway of nicotine in human gingival fibroblasts (HGFs).

Methods

Human gingival fibroblasts stimulated by nicotine were used as an in vitro model. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect cell viability and reactive oxygen species (ROS) generation was assessed with 2,7-dichlorofluoroscein diacetate (DCF-DA). Morphological change was detected by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) nick end labelling (TUNEL) assay, stained with 4,6-diamidino-2-phenylindole (DAPI). To delineate the roles of extracellular signal-regulated kinase (ERK), P38 and c-Jun N-terminal kinase (JNK), Western blot and caspase-3 (CASP3) activity assay were performed.

Results

Exposure of the human gingival fibroblasts to nicotine reduced cell viability by time and dose dependent and increased the generation of ROS. It also showed morphological evidence of increased apoptosis, resulted in transient activation of JNK and ERK concomitant with activation of P38, and stimulated apoptosis as evidenced by CASP3 activation and Poly ADP ribose polymerase (PARP) cleavage.

Conclusion

These results suggest that nicotine induces apoptosis through the ROS generation and CASP3 dependent pathways in HGFs.     

Effect of phenytoin on interleukin-1β production in human gingival fibroblasts challenged to tumor necrosis factor αin vitro

Effects and interaction of tumor necrosis factor α (TNFα) and the antiepileptic drug phenytoin (PHT) on interleukin-1β (IL-1β) production as well as on prostaglandin E₂ (PGE₂) formation were studied in gingival fibroblasts in vitro. TNFα, in contrast to PHT, dose-dependently stimulated the production of cell-associated IL-1β. The stimulatory effect of TNFα on IL-1β production was accompanied by enhanced PGE₂ formation. When PHT and TNFα were added simultaneously, the drug potentiated the stimulatory effect of TNFα on both IL-Iβ production and PGE₂ formation. The major PHT metabolite, p-HPPH, did not affect IL-1β production, either alone or in combination with TNFα. The production of IL-1β induced by TNFα and the combination of TNFα and PHT was further enhanced in the presence of the prostaglandin endoperoxide (PGH) synthase inhibitors, indomethacin and flurbiprofen. The PHT-mediated enhancement of TNFα-induced IL-1β production and PGE₂ formation in gingival fibroblasts may be an important link in the pathogenesis of gingival overgrowth induced by PHT.     

Synthesis of proinflammatory cytokines by human gingival fibroblasts in response to lipopolysaccharides and interleukin-1β

We have examined the ability of gingival fibroblasts (GF) to participate in inflammatory response and function as accessory immune cells. The accessory immune function of GF cells was evaluated by their ability to elaborate proinflammatory cytokines following stimulation with lipopolysaccharides and interleukin-1β (IL-β). Using three separate clonally derived and characterized human gingival fibroblast (GF) cell lines, we demonstrate that LPS from Actinobacillus actinomycetemcomitans (Aa) and Escherichia coli (Ec) induce mRNA and synthesis of proinflammatory cytokines, IL-1β, IL-6 and IL-8. IL-1β activation of GF cells showed that IL-1β not only induces the expression of IL-6, IL-8 and TNF-α, but also acts in an autocrine manner on GF cells and induces IL-1βexpression. Furthermore, the continuous presence of IL-1β in GF cell cultures did not down regulate the response of GF cells to IL-1β. Pretreatment of GF cells with IL-lβ resulted in the enhanced synthesis of TNF-α in response to additional IL-lβ. These findings indicate that GF cells, in addition to providing structural support, may also function as accessory immune cells and play an important role in the initial inflammatory reaction as well as in the amplification of immune response.     

牛血小板衍化生长因子对人牙周膜成纤维细胞的生物学效应

目的：观察在牛血小板衍化生长因子作用下人牙周膜成纤维细胞ＤＮＡ和胶原蛋白合成的情况。方法：采用体外细胞培养技术和核素掺入法。结果：２０ｎｇ／ｍｌ～６０ｎｇ／ｍｌ牛血小板衍化生长因子可明显促进人牙周膜成纤维细胞ＤＮＡ合成，４０ｎｇ／ｍｌ浓度使细胞ＤＮＡ合成在２４ｈ达最高峰；对细胞的胶原蛋白合成无明显促进作用。结论：牛血小板衍化生长因子对人牙周膜成纤维细胞ＤＮＡ合成有促进作用，在牙周组织的创伤修复中可能起重要作用。