transforming-growth-factor-beta and Gingival-Overgrowth

Gingival overgrowth is a side effect of certain medications. The most fibrotic drug-induced lesions develop in response to therapy with phenytoin, the least fibrotic lesions are caused by cyclosporin A, and the intermediate fibrosis occurs in nifedipine-induced gingival overgrowth. Fibrosis is one of the largest groups of diseases for which there is no therapy but is believed to occur because of a persistent tissue repair program. During connective tissue repair, activated gingival fibroblasts synthesize and remodel newly created extracellular matrix. Proteins such as transforming growth factor (TGF), endothelin-1 (ET-1), angiotensin II (Ang II), connective tissue growth factor (CCN2/CTGF), insulin-like growth factor (IGF), and platelet-derived growth factor (PDGF) appear to act in a network that contributes to the development of gingival fibrosis. Since inflammation is the prerequisite for gingival overgrowth, mast cells and its protease enzymes also play a vital role in the pathogenesis of gingival fibrosis. Drugs targeting these proteins are currently under consideration as antifibrotic treatments. This review summarizes recent observations concerning the contribution of TGF-β, CTGF, IGF, PDGF, ET-1, Ang II, and mast cell chymase and tryptase enzymes to fibroblast activation in gingival fibrosis and the potential utility of agents blocking these proteins in affecting the outcome of drug-induced gingival overgrowth.

Topics: Animals; Connective Tissue Growth Factor; Endothelin-1; Fibroblasts; Gingival Overgrowth; Humans; Models, Biological; Platelet-Derived Growth Factor; Transforming Growth Factor beta

Cyclosporine A, an extremely effective immunosuppressant, is also associated with various untoward effects, including gingival overgrowth. Despite intense clinical and laboratory investigation, the cellular-molecular mechanism through which cyclosporine A simultaneously acts as a selective immunosuppressant while it elicits a connective tissue reaction in the gingiva remains poorly understood. In recent years, cellular and molecular biologic techniques have elucidated a variety of growth factors that control connective tissue homeostasis. Two growth factors known to be major elements in wound repair and connective tissue homeostasis are platelet-derived growth factor and transforming growth factor-beta 1. Increased gingival levels of these factors may be responsible for promoting fibroblastic proliferation and fibroblastic production of extracellular matrix constituents in overgrown gingival tissues. Expression of these factors has recently been shown to be upregulated in these tissues. The results of these recent studies may provide a foundation for understanding the molecular mechanism involved in the pathogenesis of cyclosporine A-induced gingival overgrowth.

Topics: Cyclosporine; Fibroblasts; Gingival Overgrowth; Humans; Immunosuppressive Agents; Interleukin-1; Platelet-Derived Growth Factor; Transforming Growth Factor beta; Up-Regulation

To identify the possible biological roles of keratinocyte growth factor (KGF), connective tissue growth factor (CTGF) and transforming growth factor-β (TGF-β) in cyclosporine-A (CsA) and phenytoin (PNT)-induced gingival overgrowth (GO) and to correlate them with each other.. Sixty adult male albino rats were selected and divided into 3 equal groups. Group I rats received no treatment. Group II rats were administrated CsA for 7 weeks. Group III were administrated PNT for the same period. Rats were euthanized at the end of the experiment and routine tissue processing was carried out. The obtained specimens were stained with H&E, KGF, CTGF and TGF-β antibodies.. One-way MANOVA test for KGF, CTGF and TGF-β revealed an overall significant difference between the different groups (P<0.001). LSD post hoc test for multiple comparisons revealed a significant difference between each two groups. Two-tailed Pearson correlation for group II revealed non-significant weak positive correlations between KGF & CTGF and between CTGF & TGF-β. Non-significant weak negative correlation was found between KGF & TGF-β. Meanwhile, group III revealed non-significant weak positive correlation between KGF & TGF-β and between CTGF & TGF-β. Significant moderate positive correlation was found between KGF & CTGF.. The findings of the present study indicated that KGF, CTGF and TGF-β have biological roles in progression of CsA- and PNT- induced GO. KGF plays a greater role in CsA- induced GO than in PNT- induced GO. Meanwhile, CTGF and TGF-β play a role in PNT- induced GO greater than in CsA- induced GO.

Topics: Animals; Connective Tissue Growth Factor; Cyclosporine; Disease Models, Animal; Epithelium; Fibroblast Growth Factor 7; Fibroblasts; Gingiva; Gingival Overgrowth; Male; Phenytoin; Random Allocation; Rats; Transforming Growth Factor beta

Drug-induced gingival overgrowth is caused by the antiseizure medication phenytoin, calcium channel blockers, and ciclosporin. Characteristics of these drug-induced gingival overgrowth lesions differ. We evaluate the ability of a mouse model to mimic human phenytoin-induced gingival overgrowth and assess the ability of a drug to prevent its development. Lovastatin was chosen based on previous analyses of tissue-specific regulation of CCN2 production in human gingival fibroblasts and the known roles of CCN2 in promoting fibrosis and epithelial to mesenchymal transition. Data indicate that anterior gingival tissue overgrowth occurred in phenytoin-treated mice based on gross tissue observations and histomorphometry of tissue sections. Molecular markers of epithelial plasticity and fibrosis were regulated by phenytoin in gingival epithelial tissues and in connective tissues similar to that seen in humans. Lovastatin attenuated epithelial gingival tissue growth in phenytoin-treated mice and altered the expressions of markers for epithelial to mesenchymal transition. Data indicate that phenytoin-induced gingival overgrowth in mice mimics molecular aspects of human gingival overgrowth and that lovastatin normalizes the tissue morphology and the expression of the molecular markers studied. Data are consistent with characterization of phenytoin-induced human gingival overgrowth in vivo and in vitro characteristics of cultured human gingival epithelial and connective tissue cells. Findings suggest that statins may serve to prevent or attenuate phenytoin-induced human gingival overgrowth, although specific human studies are required.

Topics: Amino Acid Oxidoreductases; Animals; Cadherins; Connective Tissue Growth Factor; Gingiva; Gingival Overgrowth; Humans; Lovastatin; Male; Mice; Mice, Inbred BALB C; Phenytoin; Transforming Growth Factor beta

The aim of the study was to investigate the effect of cyclosporine A (CsA) on TGF-β/Smad3 signaling in rat gingival fibroblasts and to explore the mechanism of CsA induced gingival overgrowth.. Healthy Sprague-Dawley rat gingival fibroblasts were cultured with different concentrations of CsA and the cell proliferations were assessed with CCK-8 assay. The mRNA levels of TGF-β1, Smad3 and collagen I were measured by real-time PCR. The protein level of TGF-β1, Smad3, p-Smad3 and collagen I were determined using western blot and immumofluorescence. Cell migration ability was detected by cell wound scratch assay. The data was analyzed with SPSS 20.0 software package.. The use of 200 ng/mL CsA stimulated proliferation and migration of gingival fibroblasts. The mRNA levels of TGF-β1 and collagen I were significantly promoted after CsA exposure. The protein syntheses of TGF-β1, p-Smad3 and Collagen I were also increased by CsA stimulation.. CsA may activate TGF-β/Smad3 signaling pathway, thus promoting the proliferation and migration of rat gingival fibroblasts as well as collagen accumulation, which eventually lead to gingival overgrowth.

Topics: Animals; Cell Proliferation; Collagen; Cyclosporine; Enzyme Inhibitors; Fibroblasts; Gingiva; Gingival Overgrowth; Rats; Rats, Sprague-Dawley; RNA, Messenger; Smad3 Protein; Transforming Growth Factor beta

Gingival overgrowth was reported as a side effect after chronic administration of several drugs, which, despite their different pharmacological effect, seem to have the gingival mucosa as a secondary target. The thickness of the gingival epithelium and fibrosis in the lamina propria are unspecific changes that together determine the enlargement of the gingival mucosa, but the molecular mechanisms responsible for the imbalance of collagen synthesis/breakdown are still uncertain. The aim of our study was to assess the role of TGF-β1-CTGF pathway in the activation of cells with a fibrilogenetic phenotype responsible for the gingival fibrosis developed after chronic administration of dihydropyridine calcium channel blockers.. Fragments of gingival tissue collected from patients clinically diagnosed with gingival overgrowth after chronic administration of nifedipine and amlodipine were processed for paraffin embedding. Serial sections were used for routine staining Masson and Gömöri's silver impregnation in order to reveal collagen accumulation and for immunohistochemical reactions to label TGF-β1, CTGF, Ki67 and α-SMA.. Routine histological staining for collagen revealed the presence of gingival fibrosis and a change between type I collagen/type III collagen ratio. Regardless of the drug involved, many slides showed extended TGF-β1 positive areas, mainly in the profound - spinous and basal - layers, but also in some cells from the subjacent connective tissue. CTGF exposed intense positive reaction in the basal and parabasal layers, but also in resident cells from the connective tissue. Ki67 immunolabeling did not reveal an increased fibroblast proliferation in the lamina propria. We noticed the presence of a small number of myofibroblasts in the lamina propria.. These findings suggest that TGF-β1-CTGF axis is activated in dihydropyridine calcium channel blockers-induced gingival overgrowth and exerts a different control on the activation of fibroblasts with a synthetic phenotype. These results also have implications for better understanding mechanisms of fibrosis and the future use of this pathogenic pathway as a therapeutic target in order to limit gingival fibrosis.

Topics: Adult; Amlodipine; Calcium Channel Blockers; Calcium Channels, L-Type; Connective Tissue Growth Factor; Female; Gingiva; Gingival Overgrowth; Humans; Immunohistochemistry; Male; Nifedipine; Signal Transduction; Transforming Growth Factor beta

Gingival enlargement is a fibrotic condition that can arise from systemic administration of the dihydropyridine calcium channel blocker nifedipine. Periostin, a transforming growth factor-beta (TGF-β)-inducible matricellular protein, has been associated with fibrosis in numerous tissues, but its expression has never been examined in nifedipine-influenced gingival enlargement (NIGE). The objective of this study was to assess if periostin up-regulation is associated with NIGE and whether nifedipine induces periostin expression in gingival fibroblasts. In NIGE tissue (n = 6), periostin is overexpressed in the gingival connective tissue compared with healthy control tissue (n = 6). The transcription factor p-SMAD2/3, which is associated with canonical TGF-β signaling, localizes to the nuclei in both HGFs and oral epithelial cells in NIGE tissues, but not in control healthy tissue. In vitro culture of HGFs with 30 and 100 ng/mL of nifedipine significantly increased periostin mRNA and protein levels, which correlated with increased levels of active TGF-β and increased phosphorylation and nuclear localization of SMAD3. Blocking of canonical TGF-β signaling through inhibition of the TGF-β receptor I with SB431542 significantly reduced nifedipine-induced SMAD3 phosphorylation and periostin expression. Our results demonstrate that nifedipine up-regulates periostin in HGFs in a TGF-β-dependent manner.

Topics: Benzamides; Calcium Channel Blockers; Cell Adhesion Molecules; Cell Culture Techniques; Cell Nucleus; Connective Tissue; Dioxoles; Epithelial Cells; Fibroblasts; Gingiva; Gingival Overgrowth; Humans; Nifedipine; Phosphorylation; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta; Up-Regulation

Glycosaminoglycans (GAGs) play important roles in cell behavior and have the ability to bind and modulate cytokines. Using primary cultured fibroblasts from hereditary gingival fibromatosis (HGF), normal gingiva (NG), and NG treated with cyclosporin-A (NGc) we show changes in the expression and structural characteristics of GAGs as well as in the expression of enzymes involved in their biosynthesis and degradation. In addition, we show the over-expression of TGF-beta1 and TGF-beta type II receptor in HGF and NGc. There is an increase in the GAGs retained in the cellular fraction, and the fine structure of galactosaminoglycans show a decrease in alpha-l-iduronic acid content in HGF and NGc. Elevated extracellular levels of low molecular weight hyaluronan (HA) are found in HGF due to increase in the expression of HA synthase 3 and hyaluronidases 1 and 2. The results bring new insights to the accumulation of extracellular matrix related to TGF-beta over-expression.

Topics: Cells, Cultured; Cyclosporine; Fibroblasts; Fibromatosis, Gingival; Gingiva; Gingival Overgrowth; Glycosaminoglycans; Humans; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta; Up-Regulation

Cyclosporin A (CsA) induces gingival overgrowth (GO) in patients who seem to be prone to this disorder. It is still impossible to determine which patients will develop GO. Patients treated with the new immunosuppressive drug tacrolimus seem not to have GO. The aims of this study were to investigate transforming growth factor-beta1 (TGF-beta1) gene polymorphisms in renal transplant recipients treated with CsA or tacrolimus and to establish an association between these polymorphisms and TGF-beta1 plasma concentration and the incidence of GO.. The examined group consisted of 134 renal transplant recipients. Ninety-two underwent CsA treatment (50 with and 42 without GO), and 42 underwent tacrolimus treatment. Age, gender, time after transplantation, calcineurin inhibitor total dosage, number of teeth, and sulcus bleeding index were analyzed. TGF-beta1 plasma levels were estimated in 60 CsA- and 30 tacrolimus-treated patients. Two biallelic polymorphisms of the TGF-beta1 gene were studied at codon 10 (at position +869) and at codon 25 (at position +915) in patients from the examined group and in 108 healthy volunteers (the control group).. The distribution of the high, intermediate, and low TGF-beta1 producer phenotypes was comparable in all the studied groups and in the healthy controls. The high producer phenotype was more frequent in patients with GO. TGF-beta1 levels in the CsA group showed correlation with the phenotypes. The lowest incidence of GO was observed in the 10C/C genotype, whereas the highest was observed in the 10T/C genotype.. High and intermediate TGF-beta1 producer phenotypes and heterozygous genotype 10T/C might be considered risk factors for GO in patients treated with CsA.

Topics: Adult; Codon; Cyclosporine; Female; Genotype; Gingival Overgrowth; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Polymorphism, Genetic; Statistics, Nonparametric; Tacrolimus; Transforming Growth Factor beta; Transforming Growth Factor beta1

Cyclosporine A (CsA) is a widely used immunosuppressant but with significant side-effects, such as gingival overgrowth. This study investigates how CsA induces gingival proliferation and shows the effects of the CsA-associated signaling messengers, IL-6 and TGF-beta1, on gingival proliferation. CsA increased both IL-6 and TGF-beta1 levels. In addition to CsA, an IL-6 or TGF-beta1 treatment also induced gingival fibroblast proliferation. Inhibiting the cytokine resulted in the suppression of CsA-induced proliferation. MAPKs and PI3K are known to be involved in cell proliferation. Therefore, the effect of CsA on the kinase activities was examined. The results showed that both p38 MAPK and PI3K are essential for gingival fibroblast proliferation. TGF-beta1 and IL-6 and their associated signaling transduction may be novel bona fide molecular targets for the prevention of gingival overgrowth in CsA-treated patients. (. MAPK, mitogen-activated protein kinase; P13K, phosphatidylinositol 3-kinase.)

Topics: Blotting, Northern; Cell Count; Cell Proliferation; Cells, Cultured; Cyclosporine; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Gingiva; Gingival Overgrowth; Humans; Immunoblotting; Immunoprecipitation; Immunosuppressive Agents; Interleukin-6; MAP Kinase Kinase 4; p38 Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinases; Signal Transduction; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor (TGF)-beta1 is involved in the pathogenesis of both drug-induced gingival overgrowth and hereditary gingival fibromatosis. Ribozymes enzymatically cleave target mRNAs and are expected to be utilized as the basis of novel nucleic acid-based therapies. We designed a chimeric DNA-RNA ribozyme targeting TGF-beta1 mRNA and examined its effect on growth of gingival fibroblasts in culture.. Chimeric DNA-RNA hammerhead ribozyme with sequence complementary to the loop structure of human TGF-beta1 mRNA was used. We evaluated transfer of the chimeric ribozyme by hemagglutinating virus of Japan (HVJ)-envelope into cultured human gingival fibroblasts in vitro and rat gingival tissues in vivo. We then examined effects of the chimeric ribozyme to TGF-beta1 on proliferation and DNA synthesis in human gingival fibroblasts. We also examined effects of the chimeric ribozyme to TGF-beta1 on expression of TGF-beta1, type IV collagens, and fibronectin mRNAs and expression of TGF-beta1 protein in human gingival fibroblasts.. Chimeric ribozyme was sufficiently distributed into human fibroblasts in vitro and rat gingivae in vivo. Chimeric ribozyme to TGF-beta1 significantly inhibited expression of TGF-beta1, type IV collagen, and fibronectin mRNAs and TGF-beta1 protein in human gingival fibroblasts. Mismatch ribozyme had no effect on expression of these molecules. Chimeric ribozyme to TGF-beta1 also significantly inhibited proliferation and DNA synthesis in gingival fibroblasts.. Chimeric DNA-RNA ribozyme targeting TGF-beta1 may be a useful gene therapy agent for treatment of gingival hyperplasia.

Topics: Adolescent; Animals; Blotting, Western; Cell Proliferation; Cells, Cultured; Extracellular Matrix Proteins; Female; Fibroblasts; Gene Expression; Gingiva; Gingival Overgrowth; Humans; Male; Rats; Rats, Wistar; RNA, Catalytic; Transforming Growth Factor beta; Transforming Growth Factor beta1

Cyclosporin (CsA) and tacrolimus (FK-506) are immunosuppressive drugs that specifically inhibit T-cell activation via calcineurin inhibition. Gingival overgrowth is a common side effect following the administration of CsA. The severity of gingival overgrowth seen in patients taking FK-506 is less than that observed with CsA. Little is known about the involvement of saliva in drug-induced gingival overgrowth. The purpose of this study was to investigate the salivary contents of tumor growth factor beta1 (TGF-beta1), epidermal growth factor (EGF), and interleukin-6 (IL-6) as well as the hystometry of gingival tissue obtained from rats treated with either FK-506 or CsA.. For 30 or 60 days rats received daily subcutaneous injection doses of either CsA or FK-506 (10 mg/kg). The concentrations of TGF-beta1, EGF, and IL-6 in saliva were determined by enzyme-linked immunosorbent assay, and after histological processing, the oral epithelium and connective tissue were assessed at the region of the lower first molars.. The levels of TGF-beta1, EGF, and IL-6 in saliva were not significantly altered by any of the treatments after 30 days. After 60 days of treatment with CsA, gingival overgrowth and significant increase in salivary TGF-beta1, EGF, and IL-6 concentrations were observed; no statistically significant changes were induced by FK-506.. Within the limits of this experimental study, it can be concluded that CsA, but not FK-506, induced gingival overgrowth associated with an increase of the salivary levels of the cytokines TGF-beta1, EGF, and IL-6.

Topics: Animals; Cyclosporine; Cytokines; Epidermal Growth Factor; Gingival Overgrowth; Immunosuppressive Agents; Interleukin-6; Male; Rats; Rats, Sprague-Dawley; Saliva; Tacrolimus; Transforming Growth Factor beta; Transforming Growth Factor beta1

To examine the effects of cyclosporin A (CsA) on the expression of growth factors in induced gingival overgrowth with limited contributing factors arising from local inflammation caused by bacterial plaque, this study of gingival overgrowth was designed on the edentulous ridge of rats.. After a 3-week healing period following maxillary molar extractions, 16 five-week-old male Sprague-Dawley rats were assigned to CsA and control groups. Animals in the CsA group were fed 30 mg/kg CsA daily, whereas the control rats received a mineral oil vehicle instead. After 4 weeks, all animals were sacrificed, and the morphology of edentulous ridges was recorded by dental impression. The gingivae on the left-hand side were dissected and stored for mRNA analysis, whereas the gingivae on the right-hand side were fixed in 4% paraformaldehyde for immunohistochemistry (IHC) analysis of transforming growth factor-beta1 (TGF-beta1), platelet-derived growth factor beta (PDGF-beta), insulin-like growth factor-1 (IGF-1), and vascular endothelial growth factor (VEGF).. The edentulous gingivae were enlarged and the body weights were reduced in the CsA-treated animals compared to controls. The mRNA expressions of TGF-beta1, IGF-1, and VEGF were higher in the gingivae of the CsA group than in the control group. In addition, a greater mRNA expression (7.21-fold) of VEGF was demonstrated in the CsA group than in the control group by real-time polymerase chain reaction (PCR). The percentages of cells staining positive for TGF-beta1 and VEGF were significantly greater in the CsA rats than in the control rats.. Greater mRNA expression and positive staining for TGF-beta1 and VEGF were observed in the edentulous gingivae of rats that received CsA. Therefore, CsA may upregulate TGF-beta1 and VEGF gene expression and protein secretion in CsA-induced gingival overgrowth.

Topics: Animals; Cyclosporine; Gingiva; Gingival Overgrowth; Immunohistochemistry; Insulin-Like Growth Factor I; Male; Mouth, Edentulous; Polymerase Chain Reaction; Proto-Oncogene Proteins c-sis; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation; Vascular Endothelial Growth Factor A

To determine if local, gingival crevicular fluid (GCF) levels of TGFbeta1 were altered in drug-induced gingival overgrowth.. GCF samples were collected on Periopaper strips from 45 renal transplant recipients who had been medicated with cyclosporin or cyclosporin in combination with other putative overgrowth-inducing drugs for a minimum of 6 months. Twenty-two subjects had gingival overgrowth while the other 23 patients showed no signs of gingival changes and constituted the medicated control group. Non-medicated controls consisted 20 periodontally healthy individuals who had never taken overgrowth-inducing drugs. GCF levels of TGFbeta1 and alkaline phosphatase, a marker of inflammation, were determined by enhanced chemiluminescence ELISA and enzyme activity assays, respectively.. TGFbeta1 levels in GCF from overgrowth and non-overgrowth sites in overgrowth sufferers did not differ. However, there were significant differences in median concentration (P = 0.001) and GCF levels of TGFbeta1 per sample (P = 0.05) between study groups with overgrowth patients having higher amounts per sample and lower concentrations than medicated and healthy controls. Median levels of alkaline phosphatase per GCF sample differed between site (P = 0.01) with higher levels present at overgrowth sites. Despite this, the concentration of enzyme in GCF did not differ between site or patient group.. GCF TGFbeta1 detected in overgrowth patients could reflect a higher level of gingival inflammation because of difficulties in plaque control consequent on the development of overgrowth. However, the higher local levels of total TGFbeta1 in overgrowth patients could indicate that it is a risk factor for developing gingival overgrowth.

Topics: Adult; Alkaline Phosphatase; Antihypertensive Agents; Cyclosporine; Female; Gingival Crevicular Fluid; Gingival Overgrowth; Humans; Immunosuppressive Agents; Kidney Transplantation; Male; Middle Aged; Risk Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1

In a significant number of cases (25-81%) immunosuppressant treatment with cyclosporin A (CsA) is associated with gingival overgrowth, seriously interfering with the functions of mastication and speech. In CsA-induced gingival enlargement, quantitative modifications of the extracellular matrix components occur, and collagen (COL) metabolism and matrix metalloproteinases (MMPs) have been suggested as being the main targets. Since the mechanisms at the basis of CsA-induced gingival overgrowth are not yet completely understood, our aim was to analyze the effect of CsA on COL turnover in cultured human gingival fibroblasts. Cultured human gingival fibroblasts from four healthy volunteers were incubated with CsA (800 ng/ml) or with its vehicle (VH) for variable intervals of time (24, 48, 72 h). Fibroblast morphology was studied by light and electron microscope. Collagen type I (COL-I), MMP-1, MMP-2, TIMP-1 and TGF-beta1 mRNA were determined by RT-PCR; COL-I and MMP-1 by dot blot, and MMP-2 by zymography. Our results evidenced an up-regulation of COL-I and TGF-beta1 gene expression 72 h after CsA treatment. MMP-1, MMP-2 and TIMP-1 mRNA levels are affected but not significantly. Protein analysis revealed COL-I increase at all the considered times and, 72 h after CsA treatment, reduced collagenolytic levels. Our data suggest that COL accumulation during CsA-induced gingival overgrowth may be mainly sustained by an altered COL-I degradation due to decreased MMP-1 activity. However, interindividual differences of collagenase levels after CsA treatment suggest that a genetic predisposition to develop gingival overgrowth may be relevant.

Topics: Cells, Cultured; Collagen Type I; Cyclosporine; Fibroblasts; Gingiva; Gingival Overgrowth; Humans; Immunoblotting; Immunosuppressive Agents; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Microscopy, Electron; Reverse Transcriptase Polymerase Chain Reaction; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta; Transforming Growth Factor beta1

It has been demonstrated that cyclosporin A (CyA) blocks the immune system, acts on cytoskeleton and stimulates the production of extracellular matrix (ECM) and transforming growth factor-beta1 (TGF-beta1). This cytokine, such as transforming growth factor-alpha (TGF-alpha), induces deposition of glycosaminoglycans (GAG), proteoglycans and collagen fibres in the ECM.. In this work, we examined the effect induced by CyA, TGF-beta1 and TGF-alpha on cultures of healthy and overgrown human gingival fibroblasts in order to evaluate the glycosaminoglycan, cytoskeletal changes and the behaviour of fibroblasts after concanavalin A (Con A) treatment. Moreover, we examined gingival biopsies by Alcian blue histochemical staining and electron transmission microscopy.. Total and extracellular sulphated GAG in overgrown gingiva specimens and in derived fibroblast cultures treated with CyA and cytokines were significantly higher than controls. The action of cytokines was increased (P < or = 0.01) compared with CyA with a greater effect of TGF-alpha in comparison with TGF-beta1; the electron microscopy showed ECM accumulation. The agglutinations showed the heterogeneity of fibroblast populations.. Stimulation with Con A showed that the fibroblast population had cell surface heterogeneity, and could respond in a different way to both CyA and cytokine stimulus. Moreover, increased synthesis of GAG in overgrown gingiva compared with synthesis in normal fibroblasts before CyA treatment suggests a possible genetic origin of damage. As not all CyA-treated patients develop gingival overgrowth, a genetic predisposition may explain the different responses of gingival fibroblast populations.

Topics: Adult; Cells, Cultured; Concanavalin A; Cyclosporine; Extracellular Matrix Proteins; Female; Fibroblasts; Fluorescent Antibody Technique; Gingiva; Gingival Overgrowth; Glycosaminoglycans; Humans; Immunosuppressive Agents; Middle Aged; Transforming Growth Factor alpha; Transforming Growth Factor beta; Transforming Growth Factor beta1

Induction of the pro-fibrotic growth factor TGF-beta1 has been suggested as a possible mechanism through which immunosuppressant drugs may induce gingival overgrowth. This study aims to investigate plasma levels of TGF-beta1 and relate them to the development and severity of gingival overgrowth in immunosuppressed transplant patients.. One hundred and thirty-two ciclosporin-treated and 13 tacrolimus-treated transplant patients and 24 drug-free control subjects underwent a full periodontal examination including a determination of the presence and severity of gingival overgrowth.. Plasma TGF-beta1 concentrations were determined by ELISA, and were found to be significantly elevated in samples from the transplant patients (mean=29.1 ng/ml) as compared with controls (mean=6.1 ng/ml, p<0.0001). There was no significant difference between the levels of plasma TGF-beta1 in the ciclosporin- and tacrolimus-treated patient groups.. Furthermore, concomitant treatment with calcium channel blockers did not influence the levels of plasma TGF-beta1 in the patients group. The relationship between gingival overgrowth, independent periodontal variables and TGF-beta1 plasma concentrations was examined using univariate and multivariate regression analyses; low TGF-beta1 plasma concentrations were found to be a risk factor for gingival overgrowth in immunosuppressed patients concomitantly receiving a calcium channel blocker.

Topics: Age Factors; Analysis of Variance; Calcium Channel Blockers; Cohort Studies; Cyclosporine; Enzyme-Linked Immunosorbent Assay; Female; Gingival Overgrowth; Humans; Immunocompromised Host; Immunosuppressive Agents; Male; Regression Analysis; Tacrolimus; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transplantation

Gingival overgrowth is a common side effect following the administration of cyclosporin A (CsA). The pathogenesis of this condition is not fully understood; however, recent studies show that CsA regulates the transcription of several cytokines including transforming growth factor-beta1 (TGF-beta1). The aim of this study was to investigate the potential role of TGF-beta1 in the pathogenesis of CsA-induced gingival overgrowth, exploring a possible autocrine stimulation of TGF-beta1 as a cellular regulator of synthesis of matrix metalloproteinases (MMPs) and its tissue inhibitors (TIMPs).. Gingival fibroblasts from human normal gingiva were incubated with increasing concentrations of CsA, cultured for 24 hours, and the expression and production of TGF-beta1 determined by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. MMP and TIMP mRNA expression levels were also analyzed by RT-PCR. To determine the effect of TGF-beta1 on the expression of MMP and TIMP by human gingival fibroblasts under CsA treatment, human gingival fibroblast cultures were treated with sense oligonucleotides (SON) or antisense oligonucleotides (AON).. CsA simultaneously stimulated TGF-beta1 expression and production and inhibited expression of MMP-1 and MMP-2 by human gingival fibroblasts, whereas CsA has a slight effect on TIMP-1 and TIMP-2 expression. AON reduced TGF-beta1 production as demonstrated by ELISA, whereas TGF-beta1 mRNA expression levels were not significantly modified. The inhibition of TGF-beta1 production by AON modulated MMP expression, demonstrating the autocrine inhibitory effect of TGF-beta1 in CsA-treated human gingival fibroblasts.. The data presented here suggest that TGF-beta1 in an autocrine fashion may contribute to a reduction of proteolytic activity of human gingival fibroblasts in CsA-induced gingival overgrowth, which favors the accumulation of extracellular matrix.

Topics: Analysis of Variance; Cells, Cultured; Cyclosporine; Down-Regulation; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Fibroblasts; Gingiva; Gingival Overgrowth; Humans; Matrix Metalloproteinase Inhibitors; Matrix Metalloproteinases; Oligonucleotides, Antisense; Peptide Chain Initiation, Translational; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Statistics, Nonparametric; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta; Transforming Growth Factor beta1

Drug therapy and hereditary factors are two of the main causes of gingival overgrowth (GO). Both of these forms of GO are associated with increased extracellular matrix production by fibroblasts. Transforming growth factor beta (TGF-beta) is an important mediator of wound healing and tissue regeneration, which stimulates fibroblasts to produce extracellular matrix materials. The aim of this immunohistochemical study was to determine whether there is any altered expression of TGF-beta isoforms or its receptors in tissue from patients with drug-induced GO (DIGO; n=10) and hereditary gingival fibromatosis (n=10) when compared to non-overgrowth tissue (n=10). Compared to control tissues, significantly more fibroblasts expressed TGF-beta1 in both DIGO and hereditary gingival fibromatosis tissues (P<0.03). Cells expressing TGF-beta2 were present at control levels in DIGO but were significantly reduced in hereditary gingival fibromatosis (P<0.02). By contrast, the number of TGF-beta3-positive cells was the same in overgrowth tissues and controls. However, because of differences in total fibroblast densities between groups, there was a proportional increase in TGF-beta3 as well as TGF-beta1 expressing cells within both overgrowth populations (P<0.0001). Furthermore, representation of the TGF-beta2-positive phenotype was reduced in hereditary gingival fibromatosis (P<0.01) but increased in DIGO (P<0.005) compared to controls. Absorbance measurements of the positive cell populations showed that the level of expression was significantly higher for TGF-beta1 in hereditary gingival fibromatosis (P<0.002) and significantly lower for TGF-beta3 in DIGO (P<0.03). No significant differences in the numbers of TGF-betaRI- or RII-positive cells were detected between overgrowth tissues and controls. However, there were increases in the proportion of receptor-positive cells in the total cell population analysed in overgrowth tissues (P<0.0001). These results indicate qualitative and quantitative differences in TGF-beta isoform and receptor expression by fibroblasts in gingival overgrowth that may contribute to disease pathogenesis.

Topics: Adolescent; Adult; Aged; Analysis of Variance; Case-Control Studies; Cell Count; Cyclosporine; Fibroblasts; Fibromatosis, Gingival; Gingival Overgrowth; Humans; Immunohistochemistry; Middle Aged; Protein Isoforms; Receptors, Transforming Growth Factor beta; Statistics, Nonparametric; Transforming Growth Factor beta

The aim of the present study was to investigate the level of transforming growth factor-beta 1 (TGF-beta 1) in gingival crevicular fluid (GCF) samples of cyclosporin A (CsA)-treated patients and to compare the results with control groups.. Fourteen renal transplant patients exhibiting severe CsA-induced gingival overgrowth, 10 patients with chronic gingivitis, and 10 subjects with clinically healthy periodontium were included in the study. In CsA-treated patients, GCF samples were harvested from sites exhibiting gingival overgrowth (CsA GO+) and sites not exhibiting gingival overgrowth (CsA GO-). The TGF-beta 1 levels in a total of 96 GCF samples from the 34 participants were analyzed by enzyme-linked immunosorbent assay. The results were expressed in terms of total amount (pg/2 sites) and concentration (ng/ml).. TGF-beta 1 total amounts in CsA GO+ and CsA GO- sites were similar and significantly higher than that of healthy sites (P < 0.02 and P < 0.01, respectively). The total amount of TGF-beta 1 was also higher in gingivitis sites compared to the healthy sites, but the difference was not statistically significant (P > 0.05). CsA GO+ and CsA GO- sites exhibited higher total amount and concentration of TGF-beta 1 than that of gingivitis sites, but the differences were insignificant (P > 0.05).. The results of the present study support the theory that CsA increases the synthesis of TGF-beta 1 in GCF. However, since the difference between CsA GO+ and CsA GO- sites was not statistically significant, it seems unlikely that GCF TGF-beta 1 level is the sole factor responsible for the CsA-induced gingival overgrowth. Complex interactions between various mediators of inflammation and tissue modeling are possibly involved in the pathogenic mechanisms of this side effect.

Topics: Adolescent; Adult; Confidence Intervals; Cyclosporine; Enzyme-Linked Immunosorbent Assay; Female; Gingiva; Gingival Crevicular Fluid; Gingival Overgrowth; Gingivitis; Humans; Immunosuppressive Agents; Inflammation Mediators; Kidney Transplantation; Male; Statistics, Nonparametric; Transforming Growth Factor beta; Transforming Growth Factor beta1

The purpose of this study was to determine whether the prevalence and severity of gingival overgrowth in renal transplant recipients concomitantly treated with cyclosporin and a calcium channel blocker was associated with functional polymorphisms within the signal sequence of the transforming growth factor-(TGF)beta1 gene.. The extent and severity of gingival overgrowth for 164 renal transplant recipients immunosuppressed with cyclosporin A and concomitantly taking a calcium channel blocker since transplant were entered into the study (86 in Manchester, 78 in Belfast). Two biallelic polymorphisms of the TGF-beta1 gene were studied at position +869, codon 10 (leucine to proline substitution), and position +915, codon 25 (arginine to proline substitution).. Subjects who were homozygous for proline at codon 10 had significantly higher overgrowth scores than those who were heterozygous (P= 0.03) or homozygous for leucine (P= 0.01). Subjects who were heterozygous (arginine/proline) at codon 25 had a significantly higher (P= 0.04) gingival overgrowth score than those who were homozygous for arginine. Logistic regression analysis indicated that for codon 25 independent predictors of severe gingival overgrowth were the heterozygous arginine/proline genotype (P= 0.009) and whether the individual was young (P= 0.05).. Polymorphisms in the TGF-beta1 gene influence the expression of gingival overgrowth in renal transplant recipients concomitantly treated with cyclosporin and a calcium channel blocker. The polymorphism in the TGF-beta1 gene at codon 25 represented an independent genetic determinant of severe gingival overgrowth in the susceptible subjects studied.

Topics: Adult; Age Factors; Alleles; Analysis of Variance; Arginine; Calcium Channel Blockers; Chi-Square Distribution; Codon; Confidence Intervals; Cyclosporine; DNA; Female; Gene Expression Regulation; Genotype; Gingival Overgrowth; Heterozygote; Homozygote; Humans; Immunosuppressive Agents; Kidney Transplantation; Leucine; Logistic Models; Male; Odds Ratio; Polymorphism, Genetic; Proline; Transforming Growth Factor beta; Transforming Growth Factor beta1

Drug-induced gingival overgrowth is a known side effect of certain chemotherapeutic agents used for the treatment of systemic disorders. The pathogenesis and mechanisms responsible for this condition are not fully understood. This study assesses for the presence and localization of connective tissue growth factor (CTGF) in drug-induced gingival overgrowth tissues. CTGF immunostaining was compared with sections stained with transforming growth factor (TGF)-beta1 and CD31 antibodies in order to investigate possible pathogenic mechanisms.. Gingival overgrowth samples were obtained from patients undergoing therapy with phenytoin (n = 9), nifedipine (n = 4), cyclosporin A (n = 5), and control tissues from systemically healthy donors (n = 9). Tissue sections were subjected to peroxidase immunohistochemistry and were stained with CTGF and TGF-beta1 polyclonal primary antibodies. Possible relationships between CTGF staining and angiogenesis were also studied using an anti-CD31 antibody as a marker for endothelial cells. Staining was analyzed by computer-assisted quantitative and semiquantitative methodology at 5 defined sites in all samples based on the location of specific landmarks including epithelium and underlying connective tissues.. Cellular and extracellular CTGF content in phenytoin gingival overgrowth tissues was significantly (P<0.05) higher compared to the other gingival overgrowth tissues and the controls. Higher CTGF staining in phenytoin gingival overgrowth tissues was accompanied by an increased abundance of fibroblasts and connective tissue fibers. No strong association of CTGF staining with TGF-beta1 or CD31 staining was found.. The data from the present study show significantly higher CTGF staining in phenytoin-induced gingival overgrowth tissues compared to controls, cyclosporin A-, or nifedipine-induced gingival overgrowth. Moreover, semiquantitative analyses of histologic samples support the concept that the phenytoin overgrowth tissues are fibrotic. These associations suggest a possible role for CTGF in promoting development of fibrotic lesions in phenytoin-induced gingival overgrowth.

Topics: Adult; Antibodies; Anticonvulsants; Calcium Channel Blockers; Carrier Proteins; Coloring Agents; Connective Tissue; Connective Tissue Growth Factor; Cyclosporine; Endothelium, Vascular; Epithelium; Female; Fibroblasts; Fibrosis; Gingival Overgrowth; Growth Substances; Humans; Image Processing, Computer-Assisted; Immediate-Early Proteins; Immunoenzyme Techniques; Immunosuppressive Agents; Intercellular Signaling Peptides and Proteins; Leukocytes, Mononuclear; Male; Middle Aged; Mitogens; Neovascularization, Pathologic; Nifedipine; Phenytoin; Platelet Endothelial Cell Adhesion Molecule-1; Statistics, Nonparametric; Transforming Growth Factor beta; Transforming Growth Factor beta1

This study investigates a potential role for TGF beta 1 in the pathogenesis of cyclosporin A-induced gingival overgrowth (CsA-OG). TGF beta 1 was localized immunohistochemically in the connective tissue of both normal gingiva and CsA-OG. Intense staining for TGF beta 1 was detected at the tips of the dermal papillae of the overgrown gingiva. In addition, fibroblasts derived from healthy gingiva and fibroblasts derived from CsA-OG were cultured both as monolayers or embedded in a 3D-collagen gel. Fibroblast activity was monitored in terms of protein and collagen production in the presence of (i) 1 ng/ml TGF beta 1, (ii) 500 ng/ml CsA, or (iii) 500 ng/ml CsA and 1 ng/ml TGF beta 1. In monolayer culture TGF beta 1 significantly increased protein and collagen production in all cell strains (p < 0.05); however, there was no difference in response between fibroblasts from overgrown and healthy tissue. The production of both protein and collagen was significantly lower in the presence of the combination of CsA and TGF beta 1 when compared with the maximal stimulation produced by TGF beta 1 alone. In gel, TGF beta 1 significantly elevated matrix production by all overgrown cell strains (p < 0.05) but had little or no effect on the normal cell strains. The combination of CsA and TGF beta 1 in gel cultures reduced protein and collagen production by overgrown cell strains compared with TGF beta 1 alone. It is concluded that the cellular activity of gingival fibroblasts is dependent on culture conditions and that fibroblasts derived from overgrown gingival tissue are more responsive to TGF beta 1 than normal gingival fibroblasts when cultured in type I collagen gel.

Topics: Adult; Analysis of Variance; Cells, Cultured; Collagen; Coloring Agents; Connective Tissue; Culture Media; Cyclosporine; Epithelium; Extracellular Matrix; Fibroblasts; Gels; Gingiva; Gingival Overgrowth; Humans; Immunohistochemistry; Immunosuppressive Agents; Male; Protein Biosynthesis; Proteins; Transforming Growth Factor beta