5 polymorphisms in the transforming growth factor-beta 1 gene (TGF-beta 1) in adult periodontitis

OBJECTIVES: Transforming growth factor-beta (TGF-beta) represents a family of polypeptide growth factors, involved in inflammation and regulation of immune responses. The purpose of this study was to determine whether polymorphisms in the TGF-beta 1 gene may confer susceptibility to adult periodontitis. MATERIAL AND METHODS: We studied 90 patients with adult periodontitis together with 108 unrelated subjects. 3 polymorphisms located in the 5'region at positions -988 (C/A), -800 (G/A) and -509 (C/T) and 2 polymorphisms located at codons 10 (L10P) and 25 (R25P) of exon 1 were investigated by PCR methods. RESULTS: There was no statistically-significant difference in genotype or allele frequency distributions between patients and reference group for the -800G/A, -509C/T, L10P and R25P polymorphisms (p>0.05 in all cases). The -988 A polymorphism was present neither in our patients nor in unrelated subjects. Upon stratification for smoking status no significant differences were found in the TGF-beta 1 genotype or allele frequencies either between adult periodontitis smokers compared to control smokers, or between periodontitis non-smokers and control non-smokers. CONCLUSION: These data indicate that the mentioned polymorphisms of the TGF-beta 1 gene do not influence susceptibility to adult periodontitis. There was no association between any polymorphisms in the TGF-beta 1 gene, severity of periodontitis and the smoking status in our study.     

Behavior of human dental pulp cells exposed to transforming growth factor-beta1 and acidic fibroblast growth factor in culture

The development of methods for regenerative endodontic procedures requires an understanding of the factors regulating the development of odontoblasts from adult cell populations such as pulpal cell lines. In this study, we exposed cultures of human pulp cells (7th passage) to growth factors including transforming growth factor-beta1 (TGF-beta1, at 1 or 5 ng/mL), acidic fibroblast growth factor (aFGF, 5 ng/mL), or a combination of the 2 growth factors and evaluated cellular morphology and markers of cell phenotype including alkaline phosphatase activity, osteocalcin, bone sialoprotein (BSP), and dentin sialophosprotein (DSPP). The mean number of nucleoli in the 1 ng/mL TGF-beta1 group was significantly higher than with 5 ng/mL aFGF. Alkaline phosphatase activity was significantly greater with 1 ng/mL TGF-beta1 versus 5 ng/mL TGF-beta1 + 5 ng/mL aFGF (P < .05). Osteocalcin mRNA was expressed in all samples. The cells exposed to 1 ng/mL TGF-beta1 were stimulated; however, exposure to growth factors for 8 days was not sufficient for expression of BSP and DSPP mRNA. Cells treated with 1 ng/mL TGF-beta1 exhibited higher activity, whereas 5 ng/mL aFGF-treated cells were inhibited. Although osteocalcin was observed in all cultures, suggestive of the potential for odontoblast formation, under the present conditions, the exposure to TGF-beta1 and aFGF was not sufficient to induce expression of the dentin matrix components BSP and DSPP.     

Upregulation of transforming growth factor-beta1 and vascular endothelial growth factor gene and protein expression in cyclosporin-induced overgrown edentulous gingiva in rats

BACKGROUND: 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. METHODS: 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). RESULTS: 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. CONCLUSIONS: 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.     

Gingival fibroblast response to cyclosporin A and transforming growth factor β1

This study investigates a potential role for TGFβ₁ in the pathogenesis of cyclosporin A-induced gingival overgrowth (CsA-OG). TGFβ₁ was localized immunohistochemically in the connective tissue of both normal gingiva and CsA-OG. Intense staining for TGFβ₁ 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 TGFβ₁ (ii) 500 ng/ml CsA, or (iii) 500 ng/ml CsA and 1 ng/ml TGFβ₁. In monolayer culture TGFβ₁ 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 TGFβ₁ when compared with the maximal stimulation produced by TGFβ₁ alone. In gel, TGFβ₁ 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 TGFβ₁ in gel cultures reduced protein and collagen production by overgrown cell strains compared with TGFβ¹ alone. It is concluded that the cellular activity of gingival fibroblasts is dependant on culture conditions and that fibroblasts derived from overgrown gingival tissue are more responsive to TGFβ₁ than normal gingival fibroblasts when cultured in type I collagen gel.     

Expression of transforming growth factor-beta 1 (TGF-beta 1) in odontogenic cysts

AIM: To evaluate the positivity to transforming growth factor-beta 1 (TGF-beta 1) in different types of odontogenic cysts. METHODOLOGY: A total of 30 radicular cysts (RCs), 27 follicular cysts (FCs) and 28 odontogenic keratocysts (OKCs) were evaluated for immunohistochemical analysis of TGF-beta 1. TGF-beta 1 was evaluated in blood vessels, stromal cells (fibroblasts) and pluristratified squamous epithelium. TGF-beta 1 expression was determined by evaluating the number of positive elements. TGF-beta 1 expression was determined by evaluating 1000 cells in the pluristratified squamous epithelium (500 in the basal and parabasal layers, and 500 in the superficial layer) and 500 cells (the fibroblasts in the stroma) for each specimen, and counting the number of positive cells. The number of positive vessels was evaluated in 10 high power fields (HPF). The Chi-square test was used to evaluate differences between the two groups (RC + FC and OKC). A P-value <0.05 was considered to indicate statistical significance. RESULTS: A higher and statistically significant positivity was found in the basal-suprabasal epithelial layers (P=0.0011), superficial epithelium (P=0.053) and stromal cells (P=0.0002) of orthokeratotic and parakeratotic OKC as compared with RC and FC. CONCLUSIONS: These differences suggest that control of the cell cycle may be abnormal in orthokeratotic OKCs. These OKCs may have an intrinsic growth potential not present in other cyst types.     

The role of fibroblast growth factor (FGF) and type beta transforming growth factor (TGF-beta 1-beta 2-beta 3) during rat craniofacial development

Growth factors seem to be part of a complex cellular signalling language, in which individual growth factors are the equivalents of the letters that compose words. According to this analogy, informational content lies, not in an individual growth factor, but in the entire set of growth factors and others signals to which a cell is exposed. The ways in which growth factors exert their combinatorial effects are becoming clearer as the molecular mechanisms of growth factors actions are being investigated. A number of related extracellular signalling molecules that play widespread roles in regulating development in both invertebrates and vertebrates constitute the Fibroblast Growth Factor (FGF) and type beta Transforming Growth Factor (TGF beta). The latest research literature about the role and fate of these Growth factors and their influence in the craniofacial bone growth ad development is reviewed.     

The role of transforming growth factor-beta, insulin-like growth factor I, and basic fibroblast growth factor in distraction osteogenesis of the mandible.

Distraction osteogenesis is a viable method for regenerating large amounts of bone. In contrast to fracture healing, the mode of bone formation in distraction osteogenesis is primarily intramembranous ossification. The basic biology of the process is still not well understood. The growth factor cascade is likely to play an important role in distraction. This study examines the growth factor cascade in a lengthened ovine mandible model. Twenty-four animals were divided into four groups with varying rates of distraction (1, 2, 3, and 4 mm/day). A unilateral distractor at the angle of the mandible was used. The mandibles were lengthened to 24 mm and fixed for a period of 5 weeks, after which the animals were killed. The sections were probed for transforming growth factor-beta, basic fibroblast growth factor, and insulin-like growth factor I. The growth factors studied were present in all four groups. Transforming growth factor-beta, basic fibroblast growth factor, and insulin-like growth factor I were present in both the bony matrix of the sections and the cytoplasm of the cells, osteoblasts, and a small number of mesenchymal cells. The sections obtained from groups distracted at faster rates showed stronger presence of the growth factors examined by more intense staining. In fracture healing, the localization of transforming growth factor-beta in stage I of healing corresponded with the precise region of intramembranous ossification in stage II. Diffuse presence of transforming growth factor-beta throughout the lengthened region corresponded with the process of intramembranous ossification observed in distraction. In fracture healing, insulin-like growth factor I and basic fibroblast growth factor have been shown to promote proliferation and differentiation of osteoblasts from precursor cells. The intense presence of insulin-like growth factor I and basic fibroblast growth factor in the distracted region may account for osteoblast proliferation and formation from precursor mesenchymal cells. Mechanical strain has been shown to increase the expression of transforming growth factor-beta and insulin-like growth factor I. Distraction may serve as a source of mechanical strain, which may explain, in part, the expression of these growth factors, particularly in the faster groups.     

Impaired early bone formation in periodontal fenestration defects in dogs following application of insulin-like growth factor (II). Basic fibroblast growth factor and transforming growth factor beta 1

Abstract. Effects of a topically applied growth factor combination on fibroblast migration, collagen fiber formation and bone regeneration were studied in standardized periodontal defects in 4 beagle dogs. Following elevation of facial mucoperiosteal flaps, fenestration defects, 3 mm in diameter, were made through the cortical bone and into the dentin of maxillary and mandibular teeth. Collagen sponges, impregnated with 200 ng insulin-like growth factor II, 20 ng basic fibroblast growth factor and 6 ng transforming growth factor beta 1 were fitted to defects randomly in right or left quadrants and the flaps repositioned and sutured. Contralateral control defects received the collagen with vehicle only. Experimental procedures were staggered to allow observations of healing 3, 7, 10, and 14 days after surgery. Histometric analysis showed no differences in fibroblast and collagen density between control and growth factor defects. Bone regeneration was significantly greater in control than in growth factor defects 10 and 14 days after surgery. The rate of healing generally appeared more affected by intra-dog variations or procedural variations than by the growth factor combination.     

Different roles of dexamethasone on transforming growth factor-beta1-induced fibronectin and nerve growth factor expression in dental pulp cells

During pulp injury, it has been hypothesized that transforming growth factor-beta1 (TGF-beta1) is released from dentin into pulp tissue and promotes pulp tissue healing. Dexamethasone is a glucocorticoid that has been used to treat pulp injury and shown to induce differentiation of hard tissue forming cells. However, the interaction between dexamethasone and TGF-beta1 is still unknown. This study aimed to examine the effects of dexamethasone on human pulp cells in the presence of TGF-beta1. TGF-beta1 increased expression and synthesis of both fibronectin and nerve growth factor (NGF), whereas dexamethasone stimulated fibronectin synthesis but inhibited NGF expression. The application of both TGF-beta1 and dexamethasone resulted in an additional effect on fibronectin; however, dexamethasone inhibited the TGF-beta1-induced NGF expression. Dexamethasone promotes fibronectin synthesis and suppresses NGF secretion, suggesting that this reagent could be used clinically to reduce pain and promote dental pulp tissue healing.     

Controlled release of fibroblast growth factor 2 stimulates bone healing in an animal model of diabetes mellitus

PURPOSE: Bone formation and the healing of calvarial defects in mice is diminished in chemically induced type 1 diabetes. The present study investigated whether controlled local release of fibroblast growth factor 2 (FGF-2) stimulates bone defect healing in this model of diabetes. MATERIALS AND METHODS: First, in vitro release kinetics of different doses of recombinant human FGF-2 (rhFGF-2) from polyglycolate:polylactide membranes was determined over a 14-day period by incubating loaded membranes in PBS with constant shaking. The amount of FGF-2 was measured by enzyme-linked immunosorbent assay. Then, the effects of rhFGF-2-loaded and control membranes on calvarial defect healing over a 14-day healing period were determined in diabetic and nondiabetic mice. The degree of healing was determined by histomorphometric analyses of bone area percentage and by area measurements. The significance of the data was determined by statistical analyses, including analysis of variance. RESULTS: Kinetic release data in vitro showed that membranes loaded with 5 microg FGF-2 released measurable levels of growth factor for more than 14 days. Data from the in vivo study supported the previous finding that diabetes inhibits bone formation. Membranes containing rhFGF-2 significantly (P < .05) stimulated bone formation in diabetic animals to near normal levels during the healing period. CONCLUSION: FGF-2-loaded membranes may be useful in further studies aimed at developing therapeutic strategies for correcting deficient bone healing in patients with diabetes.     

rhbFGF、rhTGF-β、rhBMP-2对骨髓基质细胞体外培养增殖的影响

目的 :探讨重组人骨形成蛋白 (rhBMP 2 )、重组人转化生长因子 beta(rhTGF β1)、重组人碱性成纤维细胞生长因子 (rhbFGF)单独或联合应用对骨髓基质细胞体外培养生物特性的影响。方法 :体外培养大鼠骨髓基质细胞 ,分别用特定浓度的rhBMP 2 (2 0 0 μg/L ,简称b )、rhTGF β1(5 μg/L ,简称t)和rhbFGF(1μg/L ,简称f)单独或联合作用 ,观察它们对骨髓基质细胞增殖的影响 ,进行细胞计数测定细胞生长曲线 ,计算细胞倍增时间 ,用四唑盐比色法 (MTT)法测定细胞增殖情况。结果 :细胞计数测定显示所有各组在第 3天细胞数增加 ,对照组、t组、f组、f +t组和f +t +b组第 4天进入平顶期 ,而b组和f +b组细胞仍保持较高的增殖活性 ,细胞倍增时间f +b组最短为 3 2h ,MTT法测定结果与细胞计数法相似 ,b组和f +b组与对照组相比细胞增殖有显著差异 (P <0 .0 5 )。结论 :生长因子rhbFGF(1μg/L)和rhBMP 2 (2 0 0 μg/L)结合使用可以促进骨髓基质细胞的增殖 ,可以作为体外培养骨髓基质细胞扩增的最佳培养条件     

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。     

Temporal and spatial patterns of transforming growth factor-beta 1 expression in developing rat molars

Regulatory peptides of the TGF-beta family affect various aspects of embryonic development. Recent immunolocalization and in situ hybridization studies have demonstrated a specific time- and tissue-dependent expression of TGF-beta 1 in the developing mouse embryo. The purpose of this study was to evaluate the distribution of TGF-beta 1 within rat molars at different stages of development, using a well-characterized antibody, highly specific for TGF-beta 1, and immunohistochemical methods of detection. TGF-beta 1 was immunolocalized intensely within the ectodermally derived stellate reticulum and the mesenchyme of the dental papilla at the bell stage of development. Marked immunostaining was also evident in the papillary layer and the reduced dental organ subjacent to ameloblasts in the differentiation and secretory phases of amelogenesis. During the formation of coronal tissues and in the pre-eruptive phase, immunoreactive TGF-beta 1 was localized conspicuously within the dental follicle overlying the tooth germ. This temporospatial pattern of expression of TGF-beta 1 appears to correlate with specific events in morphogenesis, histogenesis and cytodifferentiation during tooth development.     

Gene expression of transforming growth factor-beta 3 and tissue inhibitor of metalloproteinase type 1 during membranous bone healing in rats

A number of growth factors have been implicated in fracture repair. Transforming growth factor-beta 3 (TGF-beta 3) is believed to be involved in osteoblast proliferation, chemotaxis, and collagen synthesis. The collagens act as the scaffolding for new bone matrix formation, whereas tissue inhibitors of metalloproteinases (TIMPs) may help regulate matrix remodeling in bone repair. Despite their hypothesized integral role in fracture repair, the temporal expression of these molecules in membranous bone fracture healing remains unknown. The objective of this study was to assess the temporal pattern of TGF-beta 3 and TIMP type 1 (TIMP-1) expression in rat mandibular fracture healing. Twenty-eight adult male Sprague-Dawley rats underwent a mandibular osteotomy, and the healing regenerate was harvested on postoperative days 3, 5, 7, 9, 23, and 37. Total cellular ribonucleic acid was isolated, and Northern analysis was performed. TGF-beta 3 expression was downregulated dramatically 3 days after the osteotomy and remained less than 20% of control levels throughout repair. In marked contrast, TIMP-1 gene expression, low during early repair, increased more than twofold over control at later time points. Understanding the temporal pattern of gene expression during membranous fracture healing has important clinical implications because elucidating these mechanisms may lead to appropriate biomolecular approaches to augment membranous bone fracture healing.     

Immunohistochemical localization of fibroblast growth factor-1 (FGF-1), FGF-2 and fibroblast growth factor receptor-1 (FGfR-1) in pleomorphic adenoma of the salivary glands

Fibroblast growth faclor-1 (FGF-l) and FGF-2 are heparin-binding polypeplides that are potent mitogens for neoplastic cells. In this study, fibroblast growth factor-1 (FGF-l), FGF-2, and fibroblast growth factor receptor-1 (FGfR-1) were immunohistochemically analyzed in 10 patients with pleomorphic adenoma of the salivary gland by using specific monoclonal antibodies. The tumor tissues were histopathologically classified as: tubular, solid, myxoid or chondroid. Both FGF-1 and FGF-2 were immunohistochemically identified in the tumor cells of all histological types. In addition, immunoreactive FGF-2 was also found in the basement membrane of tubular type tumor cells. Conversely. FGfR-1-positive tumor cells were essentially confined to the tubular and solid areas of tumors. Tumor cells in the myxoid and chondroid areas were FGfR-1 immunonegative. These results suggest that the co-expression of FGF and its receptor appears to be related to the proliferative activity of tumor cells in the tubular and solid areas, whereas loss of FGF receptor expression may be associated with the differentiation of tumor cells into myxoid and chondroid tissue types.