RESUMO
OBJECTIVES: To investigate the processes associated with the excessive production of collagen I in hereditary gingival fibromatosis (HGF). MATERIALS AND METHODS: Three HGF subjects and five controls were enrolled in the study. Histomorphological and immunohistological analyses were performed on gingival tissues. The expression of heat-shock protein 47 (HSP47), collagen I, transforming growth factor-ß1 (TGF-ß1), connective tissue growth factor (CTGF), matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) by gingival fibroblasts isolated from HGF and controls was analysed using qRT-PCR, Western blotting and ELISA. RESULTS: Considerable accumulation of fibrotic fibrils and increased synthesis of HSP47 were noted in HGF gingival tissues. The synthesis of collagen I, HSP47, TGF-ß1, CTGF and TIMP-1 was significantly elevated in HGF gingival fibroblasts compared with controls, while the production of MMP-1 was decreased. CONCLUSIONS: We report that fibrosis in HGF gingival tissues is associated with increased synthesis of HSP47. This finding was confirmed by an in vitro study, where excessive production of collagen I was associated with increased synthesis of HSP47, TGF-ß1 and CTGF by HGF gingival fibroblasts. Moreover, the shift in the TIMP-1/MMP-1 ratio identifies increased synthesis of TIMP-1 as one of the processes associated with collagen I overproduction in HGF fibroblasts.
Assuntos
Colágeno Tipo I/metabolismo , Fibromatose Gengival/metabolismo , Fibromatose Gengival/patologia , Proteínas de Choque Térmico HSP47/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Adolescente , Adulto , Células Cultivadas , Criança , Fator de Crescimento do Tecido Conjuntivo/genética , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Feminino , Fibroblastos , Fibromatose Gengival/genética , Expressão Gênica , Gengiva/citologia , Proteínas de Choque Térmico HSP47/genética , Humanos , Masculino , Metaloproteinase 1 da Matriz/metabolismo , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismoRESUMO
BACKGROUND: Although several studies have discussed some of the molecular and cellular changes associated with hereditary gingival fibromatosis (HGF), its pathogenesis is still largely unclear. This study was directed to detect and outline the degree of relationship between the immunophenotyped macrophages (M2) expressing CD163 and TGF-ß1 in patients with gingival overgrowth due to HGF. METHODS: Biopsies from 20 patients suffering from HGF and 20 normal control subjects were harvested, histologically and immunohistochemically stained then, analyzed and statistically compared and correlated for CD163 immunoexpression and TGF-ß1. RESULTS: All HGF specimens expressed TGF-ß1 by most of the connective tissue fibroblasts, with statistically high significant mean of area % (2.61 ± 0.41) compared to normal controls (0.11 ± 0.06; P = .001). All control specimens revealed negligible CD163 immunostaining of the few inflammatory cells found with a mean area of % (0.69 ± 0.12), while the specimens of HGF cases showed statistically significant higher CD163 expression (3.39 ± 0.75) at (P = .007). A statistically significant higher mean % of M2 cells expressing CD163 in relation to the total number of the inflammatory cells was revealed in HGF (34.46 ± 2.04) compared to the control group (16.36 ± 2.39; P-value ≤ .05). Moderate correlation between CD163 and TGF-ß1 was detected in HGF (r = .451; P-value < .05). CONCLUSIONS: CD163 and TGF-ß1 were clearly expressed in HGF cases compared to healthy control patients, with significant correlation. In HGF, the increase in CD 163-positive cells was specific and not dependent on the chronic gingival inflammation.
Assuntos
Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Fibromatose Gengival/metabolismo , Receptores de Superfície Celular/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Adulto , Estudos de Casos e Controles , Feminino , Fibroblastos/metabolismo , Humanos , Macrófagos/metabolismo , MasculinoRESUMO
Familial growth hormone deficiency provides an opportunity to identify new genetic causes of short stature. Here we combine linkage analysis with whole-genome resequencing in patients with growth hormone deficiency and maternally inherited gingival fibromatosis. We report that patients from three unrelated families harbor either of two missense mutations, c.347G>T p.(Arg116Leu) or c.1106C>T p.(Pro369Leu), in KCNQ1, a gene previously implicated in the long QT interval syndrome. Kcnq1 is expressed in hypothalamic GHRH neurons and pituitary somatotropes. Co-expressing KCNQ1 with the KCNE2 ß-subunit shows that both KCNQ1 mutants increase current levels in patch clamp analyses and are associated with reduced pituitary hormone secretion from AtT-20 cells. In conclusion, our results reveal a role for the KCNQ1 potassium channel in the regulation of human growth, and show that growth hormone deficiency associated with maternally inherited gingival fibromatosis is an allelic disorder with cardiac arrhythmia syndromes caused by KCNQ1 mutations.
Assuntos
Fibromatose Gengival/genética , Hormônio do Crescimento Humano/deficiência , Canal de Potássio KCNQ1/genética , Mutação de Sentido Incorreto , Adolescente , Hormônio Adrenocorticotrópico/metabolismo , Adulto , Alelos , Substituição de Aminoácidos , Animais , Arritmias Cardíacas/genética , Criança , Pré-Escolar , Feminino , Fibromatose Gengival/metabolismo , Humanos , Canal de Potássio KCNQ1/química , Canal de Potássio KCNQ1/metabolismo , Masculino , Herança Materna/genética , Camundongos , Pessoa de Meia-Idade , Modelos Moleculares , Linhagem , Mapas de Interação de Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Adulto JovemRESUMO
Virus-mediated gene delivery shows promise for the treatment of chronic pain. However, viral vectors have cytotoxicity. To avoid toxicities and limitations of virus-mediated gene delivery, we developed a novel nonviral hybrid vector: HIV-1 Tat peptide sequence modified with histidine and cysteine residues combined with a cationic lipid. The vector has high transfection efficiency with little cytotoxicity in cancer cell lines including HSC-3 (human tongue squamous cell carcinoma) and exhibits differential expression in HSC-3 (â¼45-fold) relative to HGF-1 (human gingival fibroblasts) cells. We used the nonviral vector to transfect cancer with OPRM1, the µ-opioid receptor gene, as a novel method for treating cancer-induced pain. After HSC-3 cells were transfected with OPRM1, a cancer mouse model was created by inoculating the transfected HSC-3 cells into the hind paw or tongue of athymic mice to determine the analgesic potential of OPRM1 transfection. Mice with HSC-3 tumors expressing OPRM1 demonstrated significant antinociception compared with control mice. The effect was reversible with local naloxone administration. We quantified ß-endorphin secretion from HSC-3 cells and showed that HSC-3 cells transfected with OPRM1 secreted significantly more ß-endorphin than control HSC-3 cells. These findings indicate that nonviral delivery of the OPRM1 gene targeted to the cancer microenvironment has an analgesic effect in a preclinical cancer model, and nonviral gene delivery is a potential treatment for cancer pain.
Assuntos
Dor do Câncer/terapia , Carcinoma de Células Escamosas/complicações , Terapia Genética/métodos , Receptores Opioides mu/metabolismo , Neoplasias da Língua/complicações , Animais , Dor do Câncer/metabolismo , Dor do Câncer/patologia , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Fibromatose Gengival/genética , Fibromatose Gengival/metabolismo , Vetores Genéticos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Camundongos , Receptores Opioides mu/genética , Neoplasias da Língua/genética , TransfecçãoRESUMO
Gingival fibromatosis is a progressive enlargement of the gingiva. It may hinder oral cavity hygiene and result in underlying bone loss. The long-term benefits of surgery cannot be predicted. On the other hand, alternative, efficient and non-invasive methods are not available at present. The aim of this study was to test the inhibitory effects of a chimeric IgG variant on collagen fibril formation in the cell culture of gingival fibroblasts taken from a patient with hereditary gingival fibromatosis with a high propensity for recurrence. Gingival biopsies were collected from the mandibular gingiva and used for histological evaluation as well as to establish a fibroblast culture. A histological evaluation was made in haematoxylin-eosin and Heidenhain's trichrome stained tissue sections. The inhibitory effect of a chimeric antibody on collagen fibril formation was determined in fibroblast cultures by using a collagen-specific Western blot and immunofluorescent staining. A histological evaluation revealed epithelial acanthosis with singular elongated rete pegs extending into the underlying connective tissue stroma that consisted of locally abundant, irregular collagen bundles. Based on observations with an in vitro model we conclude that a chimeric anti-collagen antibody efficiently inhibits collagen fibril accumulation in cell culture derived from diffuse, hereditary gingival fibromatosis that is characterized by a high propensity for recurrence (high proliferation index). Employing cell cultures from standardized group of patients with recurrent hereditary gingival fibromatosis as well as standarizing relevant 3D (tissue-like) models will be crucial for further tests of the antibody.
Assuntos
Colágeno/metabolismo , Fibroblastos/efeitos dos fármacos , Fibromatose Gengival/metabolismo , Imunoglobulina G/farmacologia , Células Cultivadas , Colágeno/imunologia , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibromatose Gengival/patologia , HumanosRESUMO
OBJECTIVE: The purpose of the present study was to investigate the expression of the α2-integrin subunit and heat shock protein 47 (Hsp47) in two families with isolated gingival fibromatosis (GF) form and one family with GF associated with dental abnormalities and normal gingival (NG). STUDY DESIGN: Immunohistochemistry was performed with antibodies against α2-integrin and Hsp47 in specimens from two unrelated families with hereditary gingival fibromatosis (Families 1 and 2) and from one family with a gingival fibromatosis-associated dental abnormality (Family 3); NG samples were used for comparison. The results were analysed statistically. RESULTS: Immunoreactivity for α2-integrin and Hsp47 was observed in the nucleus of epithelial cells of both the basal and suprabasal layer and a more discreet signal was noted in connective tissue in all study samples. Hsp47 showed higher immunoreactivity in Family 2 compared with the other families (p ≤ 0.05). Despite the markup α2-integrin was higher in Family 3 there was no statistically significant difference between the families studied (p ≥ 0.05). CONCLUSIONS: Our results confirmed the heterogeneity of GF, such that similar patterns of expression of the condition may show differences in the expression of proteins such as Hsp47. Although no difference in α2-integrin expression was observed between GF and NG groups, future studies are necessary to determine the exact role of this protein in the various forms of GF and whether it contributes to GF pathogenesis.
Assuntos
Fibromatose Gengival/complicações , Fibromatose Gengival/metabolismo , Proteínas de Choque Térmico HSP47/biossíntese , Integrina alfa2/biossíntese , Anormalidades Dentárias/complicações , Anormalidades Dentárias/metabolismo , Estudos Transversais , Fibromatose Gengival/genética , Fibromatose Gengival/imunologia , Humanos , Anormalidades Dentárias/imunologiaRESUMO
Gingival enlargement is common among patients and can be caused by a variety of etiological factors. The most common reason is poor oral hygiene and high bacterial load that leads to gingival inflammation and enlargement. Other implicated factors include systemic drugs, such as phenytoin (Dilantin) taken by epileptic patients, calcium channel blockers such as nifedipine (Procardia) and verapamil (Calan) for the treatment of hypertension, arrhythmia and angina. Another class of medication associated with gingival enlargement is immunosuppressive agents given to organ-transplant patients to prevent rejection of the new element such as cyclosporine. Some enlargements could be associated with other conditions such as puberty, pregnancy or diabetes or be a symptom of a systemic disease (leukemia, Wegener's granulomatosis or sarcoidosis). In rare cases the cause for the enlargement is genetic and termed hereditary gingival fibromatosis (HGF). HGF is a genetic disorder characterized by a progressive enlargement of the gingiva. Histologically, the gingiva is characterized by an accumulation of dense fibrous connective tissue. This is believed to be due to an imbalance between synthesis and degradation of extracellular matrix composed mainly of collagen molecules or due to an alteration in fibroblast proliferation. Different pathogenic mechanisms have been proposed and examined over the years but no precise process has been identified. The main objective of this paper is to discuss this genetic anomaly and support it with clinical cases of a mother and her two children. It will focus on the clinical and histologic characteristics of HGF as well as known biologic and genetic features and treatment modalities.
Assuntos
Fibromatose Gengival/genética , Adulto , Criança , Colágeno Tipo I/metabolismo , Matriz Extracelular/patologia , Feminino , Fibromatose Gengival/metabolismo , Fibromatose Gengival/patologia , Fibromatose Gengival/cirurgia , Genes Dominantes , Gengivectomia , Humanos , Masculino , Metaloproteinases da Matriz/metabolismoRESUMO
INTRODUCTION: Gingiva fibromatosis is a relatively rare condition characterized by diffuse enlargement of the gingiva, which is caused by expansion and accumulation of the connective tissue. OBJECTIVE: The aim of the present study was to investigate proliferative and apoptotic biomarker expression in normal gingiva and two forms of gingival fibromatosis. METHODS: Archived tissue specimens of hereditary gingival fibromatosis, gingival fibromatosis and dental abnormality syndrome and normal gingiva were subject to morphological analysis and immunohistochemical staining. The results were analyzed statistically. RESULTS: Proteins associated with proliferation were found in the nuclei of epithelial cells from the basal and suprabasal layers, whereas apoptotic proteins were detected in the cytoplasm of the upper layers of the epithelium. Increased expressions of minichromosome maintenance proteins 2 and 5 were observed in the gingival fibromatosis and dental abnormality syndrome samples. In contrast, geminin expression was higher in normal gingiva samples. No difference in the expression of apoptotic proteins was observed among the groups. CONCLUSION: Our findings support a role for augmented proliferation of epithelial cells within the overgrown tissues associated with gingival fibromatosis or dental abnormality syndrome. However, our data suggest that different biological mechanisms may account for the pathogenesis of different types of gingival fibromatosis.
Assuntos
Proteínas de Ciclo Celular/análise , Células Epiteliais/química , Fibromatose Gengival/metabolismo , Proteínas Nucleares/análise , Anormalidades Dentárias/metabolismo , Biomarcadores/análise , Estudos de Casos e Controles , Estudos Transversais , Células Epiteliais/patologia , Feminino , Fibromatose Gengival/genética , Fibromatose Gengival/patologia , Geminina , Humanos , Imuno-Histoquímica , Masculino , Componente 2 do Complexo de Manutenção de Minicromossomo , Anormalidades Dentárias/genética , Anormalidades Dentárias/patologia , Proteína X Associada a bcl-2/análiseRESUMO
INTRODUCTION: Gingiva fibromatosis is a relatively rare condition characterized by diffuse enlargement of the gingiva, which is caused by expansion and accumulation of the connective tissue. OBJECTIVE: The aim of the present study was to investigate proliferative and apoptotic biomarker expression in normal gingiva and two forms of gingival fibromatosis. METHODS: Archived tissue specimens of hereditary gingival fibromatosis, gingival fibromatosis and dental abnormality syndrome and normal gingiva were subject to morphological analysis and immunohistochemical staining. The results were analyzed statistically. RESULTS: Proteins associated with proliferation were found in the nuclei of epithelial cells from the basal and suprabasal layers, whereas apoptotic proteins were detected in the cytoplasm of the upper layers of the epithelium. Increased expressions of minichromosome maintenance proteins 2 and 5 were observed in the gingival fibromatosis and dental abnormality syndrome samples. In contrast, geminin expression was higher in normal gingiva samples. No difference in the expression of apoptotic proteins was observed among the groups. CONCLUSION: Our findings support a role for augmented proliferation of epithelial cells within the overgrown tissues associated with gingival fibromatosis or dental abnormality syndrome. However, our data suggest that different biological mechanisms may account for the pathogenesis of different types of gingival fibromatosis.
Assuntos
Feminino , Humanos , Masculino , Proteínas de Ciclo Celular/análise , Células Epiteliais/química , Fibromatose Gengival/metabolismo , Proteínas Nucleares/análise , Anormalidades Dentárias/metabolismo , Biomarcadores/análise , Estudos de Casos e Controles , Estudos Transversais , Células Epiteliais/patologia , Fibromatose Gengival/genética , Fibromatose Gengival/patologia , Imuno-Histoquímica , Anormalidades Dentárias/genética , Anormalidades Dentárias/patologia , /análiseRESUMO
Connective tissue growth factor (CCN2/CTGF) is not normally expressed in gingival fibroblasts, but is induced by the potent profibrotic cytokine TGFß and is overexpressed in gingival fibrosis. Since CCN2 is a marker and mediator of fibrosis, targeting CCN2 expression in gingival fibroblasts may provide new insights into the future development of novel therapeutic opportunities to treat oral fibrosis. Herein we used real-time polymerase chain-reaction, Western blot, and indirect immunofluorescence analysis to evaluate whether SB-431542, a specific pharmacological inhibitor of TGFß type I receptor (ALK5), blocks the ability of TGFß to induce CCN2 mRNA and protein expression in human gingival fibroblasts. Our results indicate that CCN2 mRNA and protein are induced by TGFß in gingival fibroblasts in a SB-431542-sensitive fashion. These results suggest that blocking ALK5 may be useful in blocking the profibrotic effects of TGFß in gingival fibroblasts.
Assuntos
Fator de Crescimento do Tecido Conjuntivo/antagonistas & inibidores , Fibroblastos/efeitos dos fármacos , Gengiva/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/farmacologia , Benzamidas/farmacologia , Western Blotting , Técnicas de Cultura de Células , Células Cultivadas , Fator de Crescimento do Tecido Conjuntivo/genética , Dioxóis/farmacologia , Fibroblastos/citologia , Fibromatose Gengival/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Gengiva/citologia , Humanos , Reação em Cadeia da Polimerase/métodos , RNA Mensageiro/antagonistas & inibidores , RNA Ribossômico 18S/antagonistas & inibidores , Receptor do Fator de Crescimento Transformador beta Tipo I , Fatores de Tempo , Fator de Crescimento Transformador beta/antagonistas & inibidoresRESUMO
BACKGROUND: Juvenile hyaline fibromatosis and infantile systemic hyalinosis are two rare autosomal recessive diseases arising from mutation in the capillary morphogenesis factor-2 gene. They are characterized by accumulation of hyaline material, in the skin in the first instance and in other organs in the second. We describe a case of juvenile hyaline fibromatosis. CASE REPORT: A 2-year-old girl presented gingival hyperplasia, skin papules, subcutaneous nodules and joints and bones lesion. A diagnosis of juvenile hyaline fibromatosis was suggested and this was confirmed by histopathology and genetic analyses. The patient presented frequent episodes of diarrhoea, which is evocative of infantile systemic hyalinosis. DISCUSSION: This case clearly illustrates the wide phenotypic range of juvenile hyaline fibromatosis. Diagnosis must be made as soon as possible to avoid cosmetic and functional handicap.
Assuntos
Fibromatose Gengival/patologia , Artropatias/patologia , Proteínas de Membrana/genética , Dermatopatias/patologia , Pré-Escolar , Consanguinidade , Constipação Intestinal/etiologia , Diagnóstico Diferencial , Diarreia/etiologia , Feminino , Fibromatose Gengival/diagnóstico , Fibromatose Gengival/genética , Fibromatose Gengival/metabolismo , Genes Recessivos , Humanos , Hialina/química , Artropatias/diagnóstico , Artropatias/genética , Artropatias/metabolismo , Proteínas de Membrana/deficiência , Marrocos/etnologia , Fenótipo , Receptores de Peptídeos , Prolapso Retal/etiologia , Prolapso Retal/cirurgia , Dermatopatias/diagnóstico , Dermatopatias/genética , Dermatopatias/metabolismoRESUMO
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.
Assuntos
Fibroblastos/metabolismo , Crescimento Excessivo da Gengiva/metabolismo , Glicosaminoglicanos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Regulação para Cima , Células Cultivadas , Ciclosporina/farmacologia , Fibromatose Gengival/metabolismo , Gengiva/efeitos dos fármacos , Gengiva/metabolismo , Humanos , Receptores de Fatores de Crescimento Transformadores beta/genética , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta/genéticaRESUMO
OBJECTIVE: To investigate the possible correlation between integrin alpha1, alpha2, and beta1 expression and excessive collagen synthesis in fibroblasts from 3 unrelated Chinese families with hereditary gingival fibromatosis (HGF). DESIGN: Gingival fibroblasts from three Chinese HGF patients and three healthy subjects were included. The expression of alpha1, alpha2, and beta1 integrin subunits was examined by immunohistochemistry, quantitative PCR, and flow cytometry. We also investigated the effects of transforming growth factor-beta1 (TGF-beta1) on the expression of these integrin subunits. RESULTS: Our results demonstrate that the expression of alpha2 was significantly higher in HGF fibroblasts compared with control fibroblasts (P < 0.01). No significant differences in the expression of alpha1 and beta1 were detected. Furthermore, TGF-beta1 promoted the expression of alpha1 and alpha2 in fibroblasts from both HGF patients and controls. However, it had a larger effect on the expression of alpha2 in HGF fibroblasts than in control cells. In contrast, alpha1 expression was stimulated more in control fibroblasts. CONCLUSION: The increased expression of integrin alpha2 and the increased response to TGF-beta1 of HGF fibroblasts may be related to the excessive collagen deposition in HGF patients.
Assuntos
Fibroblastos/metabolismo , Fibromatose Gengival/metabolismo , Gengiva/metabolismo , Integrina alfa2/metabolismo , Fator de Crescimento Transformador beta1/fisiologia , Adolescente , Adulto , Estudos de Casos e Controles , Células Cultivadas , Colágeno/metabolismo , Feminino , Fibromatose Gengival/genética , Regulação da Expressão Gênica/fisiologia , Gengiva/citologia , Humanos , Imuno-Histoquímica , Integrina alfa1/genética , Integrina alfa1/metabolismo , Integrina alfa2/genética , Integrina beta1/genética , Integrina beta1/metabolismo , Masculino , RNA/análise , Valores de Referência , Estatísticas não Paramétricas , Distribuição Tecidual , Adulto JovemRESUMO
OBJECTIVES: Hereditary gingival fibromatosis (HGF) is a rare benign disorder characterized by progressive fibrous overgrowth of the gingiva. The proliferation and expression of growth factors of HGF keratinocytes are abnormal. However, the exact role of keratinocytes in HGF pathogenesis is still unknown. The present study aimed to clarify the interactions between HGF keratinocytes and underlying fibroblasts in the pathogenesis of HGF. DESIGN: Gingival tissues, fibroblasts and keratinocytes from three Chinese HGF patients and three healthy subjects were collected. Histological analyses were performed by histochemical and immunohistochemical staining (Ki-67). Gingival fibroblasts were cocultured with gingival keratinocytes in an in vitro coculture system. The mRNA levels of type I collagen, MMP-1, MMP-3, and TIMP-1 were analysed in the cocultured gingival fibroblasts by reverse-transcription polymerase chain reaction (RT-PCR). The production of type I collagen and TIMP-1 was examined by ELISA. RESULTS: The number of Ki-67-positive keratinocytes in tissue sections from patients was higher than in those from controls. HGF fibroblasts cocultured with HGF keratinocytes showed an increased expression of type I collagen and TIMP-1. Transmission electron microscopy showed increased rough endoplasmic reticulum and ribosomes in cocultured HGF fibroblasts. CONCLUSIONS: These results suggest that HGF keratinocytes have an important role in HGF pathogenesis by inducing extracellular matrix (ECM) accumulation by fibroblasts.
Assuntos
Fibroblastos/metabolismo , Fibromatose Gengival/patologia , Queratinócitos/fisiologia , Adolescente , Adulto , Técnicas de Cultura de Células , Técnicas de Cocultura , Colágeno Tipo I/biossíntese , Colágeno Tipo I/genética , Feminino , Fibroblastos/ultraestrutura , Fibromatose Gengival/genética , Fibromatose Gengival/metabolismo , Expressão Gênica , Gengiva/metabolismo , Gengiva/ultraestrutura , Humanos , Masculino , Metaloproteinase 1 da Matriz/biossíntese , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 3 da Matriz/biossíntese , Metaloproteinase 3 da Matriz/genética , Microscopia Eletrônica , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Inibidor Tecidual de Metaloproteinase-1/biossíntese , Inibidor Tecidual de Metaloproteinase-1/genética , Adulto JovemRESUMO
Human gingival overgrowth may occur as a side effect of chronic administration of some therapeutic agents. The mechanisms responsible for the gingival tissues lesions, fibrosis and inflamation, involve an impaired balance between the production and the degradation of type I collagen. It has been demonstrated that CCN2/CTGF, a connective tissue growth factor, is highly expressed in the gingival tissues and positively correlated with the degree of fibrosis in the drug-induced gingival overgrowth. The aim of this study was to identify the presence and localization of CCN2/CTGF and CCN1/Cyr61, members of the same molecular family, in gingival tissues of cyclosporin A- and nifedipine-treated rats, by immunohistochemistry. Staining was evaluated with light microscope and the results show cellular and extracellular CTGF in nifedipin gingival overgrowth tissues with intensity of labeling higher compared to the CsA gingival overgrowth tissues or the controls. The staining for Cyr61 shows its intracellular localization with no diference of labeling intensity between drug-induced gingival overgrowth and normal tissues. Also, we were interested in the gingival TGF-â expression in those animals. We didn't find any commercial anti-rat TGF antibody and our anti-human antibody shows no cross-reactivity with rat tissues. The data from our study sustain the involvement of CTGF and Cyr61 as growth factors in the gingival tissues and the CTGF association with drug-induced gingival overgrowth.
Assuntos
Fator de Crescimento do Tecido Conjuntivo/metabolismo , Proteína Rica em Cisteína 61/metabolismo , Hiperplasia Gengival/induzido quimicamente , Hiperplasia Gengival/metabolismo , Crescimento Excessivo da Gengiva/metabolismo , Crescimento Excessivo da Gengiva/patologia , Animais , Ciclosporina/farmacologia , Modelos Animais de Doenças , Fibromatose Gengival/metabolismo , Fibromatose Gengival/patologia , Hiperplasia Gengival/patologia , Crescimento Excessivo da Gengiva/induzido quimicamente , Imuno-Histoquímica , Imunossupressores/farmacologia , Masculino , Nifedipino/farmacologia , Ratos , Ratos Wistar , Vasodilatadores/farmacologiaRESUMO
Mutation of human SOS1 is responsible for hereditary gingival fibromatosis type 1, a benign overgrowth condition of the gingiva. Here, we investigated molecular mechanisms responsible for the increased rate of cell proliferation in gingival fibroblasts caused by mutant SOS1 in vitro. Using ectopic expression of wild-type and mutant SOS1 constructs, we found that truncated SOS1 could localize to the plasma membrane, without growth factor stimuli, leading to sustained activation of Ras/MAPK signaling. Additionally, we observed an increase in the magnitude and duration of ERK signaling in hereditary gingival fibromatosis gingival fibroblasts that was associated with phosphorylation of retinoblastoma tumor suppressor protein and the up-regulation of cell cycle regulators, including cyclins C, D, and E and the E2F/DP transcription factors. These factors promote cell cycle progression from G(1) to S phase, and their up-regulation may underlie the increased gingival fibroblast proliferation observed. Selective depletion of wild-type and mutant SOS1 through small interfering RNA demonstrates the link between mutation of SOS1, ERK signaling, cell proliferation rate, and the expression levels of Egr-1 and proliferating cell nuclear antigen. These findings elucidate the mechanisms for gingival overgrowth mediated by SOS1 gene mutation in humans.
Assuntos
Fibroblastos/metabolismo , Fibromatose Gengival/metabolismo , Fase G1 , Sistema de Sinalização das MAP Quinases , Fase S , Proteína SOS1/metabolismo , Membrana Celular/genética , Membrana Celular/metabolismo , Membrana Celular/patologia , Células Cultivadas , Ciclinas/biossíntese , Fatores de Transcrição E2F/biossíntese , Proteína 1 de Resposta de Crescimento Precoce/biossíntese , Proteína 1 de Resposta de Crescimento Precoce/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/patologia , Fibromatose Gengival/genética , Fibromatose Gengival/patologia , Fase G1/genética , Humanos , Sistema de Sinalização das MAP Quinases/genética , Fosforilação , Antígeno Nuclear de Célula em Proliferação/biossíntese , Antígeno Nuclear de Célula em Proliferação/genética , Processamento de Proteína Pós-Traducional/genética , Transporte Proteico/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Fase S/genética , Proteína SOS1/antagonistas & inibidores , Proteína SOS1/genética , Regulação para Cima/genéticaRESUMO
BACKGROUND/AIM: Previously, we have shown that myofibroblasts, the main cell type associated with interstitial fibrosis, may be implicated with the gingival overgrowth observed in hereditary gingival fibromatosis (HGF) patients. The goal of this study was to determine whether transforming growth factor-beta1 (TGF-beta1) stimulates myofibroblast generation in gingival fibroblast cultures. Moreover, we analysed how interferon-gamma (IFN-gamma) interferes in this process. MATERIAL AND METHODS: Fibroblast cultures from normal gingiva and myofibroblast cells from HGF were included in this study. To determine the effects of TGF-beta1 and IFN-gamma stimulation in these cells, the expression of the specific myofibroblast marker smooth muscle isoform of alpha-actin (alpha-SMA) was examined by semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), Western blot and immunofluorescence. Enzyme-linked immunosorbent assay (ELISA) for type I collagen was performed to measure the myofibroblast activity. RESULTS: Our results demonstrated that TGF-beta1 promotes a dose- and time-dependent increase in the expression of alpha-SMA, whereas IFN-gamma blocks it and markedly prevents the fibroblast-myofibroblast switch induced by TGF-beta1 on normal gingiva cultures. IFN-gamma altered HGF myofibroblasts metabolism with a decrease of both alpha-SMA and type I collagen expression. Additionally, IFN-gamma treatment stimulated SMAD7 expression and inhibited connective tissue growth factor, which has been considered a key molecule to promote the transdifferentiation of myofibroblasts via TGF-beta1 activation. CONCLUSIONS: These findings demonstrate that TGF-beta1 induces gingival fibroblast-myofibroblast transdifferentiation, whereas IFN-gamma blocks this process. More importantly, this study suggests that IFN-gamma may be clinically effective in attenuating excessive accumulation of extracellular matrix produced by myofibroblasts in HGF.
Assuntos
Fibromatose Gengival/patologia , Gengiva/citologia , Interferon gama/fisiologia , Fator de Crescimento Transformador beta1/fisiologia , Actinas/antagonistas & inibidores , Actinas/biossíntese , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Colágeno Tipo I/antagonistas & inibidores , Colágeno Tipo I/biossíntese , Fator de Crescimento do Tecido Conjuntivo , Regulação para Baixo , Fibroblastos , Fibromatose Gengival/metabolismo , Humanos , Proteínas Imediatamente Precoces/biossíntese , Peptídeos e Proteínas de Sinalização Intercelular/biossíntese , Interferon gama/farmacologia , Músculo Liso/citologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína Smad7/metabolismo , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Regulação para CimaRESUMO
Fibromatosis is a group of disorders characterized by infiltrating fibroblastic proliferation. Among them Juvenile hyaline fibromatosis (J.H.F) is a rare, progressive, crippling autosomal recessive disorder diagnosed based on the characteristic clinicopathological findings of generalized cutaneous nodular lesions, gingival hypertrophy, flexion contractures of large joints with osteolytic lesions, and proliferating fibroblasts set within a hyalinized stroma. This disorder commonly manifests in childhood with family history of consanguinity and in siblings. A case of seven year old boy born to consanguinous parents is reported who presented with multiple subcutaneous nodules and gingival hypertrophy. Histopathological examination revealed proliferating fibroblasts embedded in an abundant homogeneous eosinophilic hyalinized matrix. The matrix showed PAS stain positivity, supporting the pathogenesis of this disorder as an inborn error of glycosaminoglycans metabolism. The differential diagnosis is discussed and the literature reviewed.
Assuntos
Fibroma/patologia , Neoplasias Cutâneas/patologia , Criança , Diagnóstico Diferencial , Fibroma/genética , Fibroma/metabolismo , Fibromatose Gengival/genética , Fibromatose Gengival/metabolismo , Fibromatose Gengival/patologia , Genes Recessivos , Humanos , Hialina/metabolismo , Masculino , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismoRESUMO
Gingival overgrowth is a side effect of certain medications and occurs in non-drug-induced forms either as inherited (human gingival fibromatosis) or idiopathic gingival overgrowth. The most fibrotic drug-induced lesions develop in response to therapy with phenytoin; the least fibrotic lesions are caused by cyclosporin A; and intermediate fibrosis occurs in nifedipine-induced gingival overgrowth. Connective tissue growth factor (CTGF/CCN2) expression is positively related to the degree of fibrosis in these tissues. The present study has investigated the hypothesis that CTGF/CCN2 is expressed in human gingival fibromatosis tissues and contributes to this form of non-drug-induced gingival overgrowth. Histopathology/immunohistochemistry studies showed that human gingival fibromatosis lesions are highly fibrotic, similar to phenytoin-induced lesions. Connective tissue CTGF/CCN2 levels were equivalent to the expression in phenytoin-induced gingival overgrowth. The additional novel observation was made that CTGF/CCN2 is highly expressed in the epithelium of fibrotic gingival tissues. This finding was confirmed by in situ hybridization. Real-time polymerase chain reaction (PCR) analyses of RNA extracted from drug-induced gingival overgrowth tissues for CTGF/CCN2 were fully consistent with these findings. Finally, normal primary gingival epithelial cell cultures were analysed for basal and transforming growth factor beta1 (TGF-beta1) or lysophosphatidic acid-stimulated CTGF/CCN2 expression at protein and RNA levels. These data indicate that fibrotic human gingival tissues express CTGF/CCN2 in both the epithelium and connective tissues; that cultured gingival epithelial cells express CTGF/CCN2; and that lysophosphatidic acid further stimulates CTGF/CCN2 expression. These findings suggest that interactions between epithelial and connective tissues could contribute to gingival fibrosis.
Assuntos
Células do Tecido Conjuntivo/química , Fibromatose Gengival/metabolismo , Proteínas Imediatamente Precoces/análise , Peptídeos e Proteínas de Sinalização Intercelular/análise , Adulto , Células Cultivadas , Fator de Crescimento do Tecido Conjuntivo , Células Epiteliais/química , Fibroblastos/química , Fibroblastos/patologia , Fibrose , Gengiva/química , Gengiva/patologia , Humanos , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Lisofosfolipídeos/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
OBJECTIVE: To investigate possible associations between platelet-derived growth factor (PDGF), PDGF receptor expression and macrophages in drug-induced and hereditary gingival overgrowth. MATERIALS AND METHODS: Tissues from patients with drug-induced gingival overgrowth (DIGO) (n = 10) and hereditary gingival fibrosis (n = 10) were studied and compared with 'control' gingiva (n = 10). Expression of PDGF and its alpha and beta receptors was investigated immunohistochemically and by RT-PCR. Macrophages were identified by immunostaining for CD68. RESULTS: PDGF isoforms and receptors were detected in most cells within all specimens. There were no differences in the numbers of macrophages, or fibroblasts expressing PDGF or receptors, between groups. The level of PDGF expression by fibroblasts, determined by absorbance measurements, was similar between groups for PDGF A. Significantly lower levels of total PDGF and the receptors were detected in drug-induced overgrowth compared to those in hereditary fibrosis (P < 0.004) and control specimens (P < 0.034). All specimens expressed mRNA for PDGF A, PDGF B and alpha and beta receptors. CONCLUSIONS: These data do not support a pivotal role for macrophage-derived PDGF B in the pathogenesis of DIGO. They suggest that fibroblasts in drug-induced lesions have a lowered capacity to produce, and respond to, PDGF, a property not shared by fibroblasts associated with hereditary fibrosis.