Peroxiredoxin I, Platelet-derived Growth Factor A, And Platelet-derived Growth Factor Receptor Alpha Are Overexpressed In Carcinoma Ex Pleomorphic Adenoma: Association With Malignant Transformation.

Carcinoma ex pleomorphic adenoma is a rare salivary gland malignancy. It constitutes an important model for the study of carcinogenesis, as it can display the tumor in different stages of progression, from benign pleomorphic adenoma to frankly invasive carcinoma. Growth signaling pathways undergo continuous activation in human tumors, commonly as a consequence of the overexpression of ligands and receptors such as platelet-derived growth factor and platelet-derived growth factor receptor. Hydrogen peroxide is produced after platelet-derived growth factor receptor activation, and it is essential for the sequential phosphorylation cascade that drives cell proliferation and migration. By their ability to degrade hydrogen peroxide, peroxiredoxins are involved in growth factor signaling regulation and in the oxidative stress response. To verify the potential association of peroxiredoxin I, platelet-derived growth factor-A, and platelet-derived growth factor receptor-alpha with carcinoma ex pleomorphic adenoma progression, we investigated the expression of these molecules in carcinoma ex pleomorphic adenoma showing different degrees of invasion. The peroxiredoxin I, platelet-derived growth factor-A, and platelet-derived growth factor receptor-alpha proteins were present in remnant pleomorphic adenoma to only a small extent, but, collectively, they were highly expressed as soon as the malignant phenotype was achieved and remained at elevated concentrations during progression to the advanced stages of carcinoma ex pleomorphic adenoma. In addition, their locations overlapped significantly, strengthening their connection to this growth-signaling pathway. Our results indicate that carcinoma ex pleomorphic adenoma cells acquire at least 2 significant advantages relative to their normal counterparts: resistance to oxidative stress-induced apoptosis, conferred by high peroxiredoxin I concentrations, and sustained growth, reflecting platelet-derived growth factor-A and platelet-derived growth factor receptor-alpha overexpression.40390-

Collagen Type I may Influence the Expression of E-Cadherin and Beta-catenin in Carcinoma Ex-pleomorphic Adenoma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Carcinoma ex-pleomorphic adenoma (CXPA) is an aggressive salivary gland malignancy, usually derived from a long-standing or a recurrent benign tumor, the pleomorphic adenoma (PA). In the context of dynamic reciprocity, changes in the composition and structure of extracellular matrix proteins and cell surface receptors have been frequently associated with dysfunctional adhesion and invasive behavior of tumor cells. It is not fully understood if these changes are involved in the conversion of PA to CXPA. In this study, different progression stages of CXPA were investigated regarding the expression of the major extracellular matrix proteins, collagen type I, and of E-cadherin and beta-catenin, the components of adherens junctions. By immunohistochemical analysis, we have demonstrated that direct contact of tumor cells with fibrillar type I collagen, particularly near the invasive front and in invasive areas prevailing small nests of CXPA cells, could be associated with reduced expression of the E-cadherin and beta-catenin adhesion molecules and with invasive behavior of epithelial; but not of CXPA with myoepithelial component. Our results also suggested that this association could depend on the organization of collagen molecules, being prevented by high-order polymeric structures. These findings could implicate the local microenvironment in the transition from the premalignant PA to invasive CXPA.174312318Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Histological Evaluation of a New Beta-Tricalcium Phosphate/Hydroxyapatite/Poly (1-Lactide-Co-Caprolactone) Composite Biomaterial in the Inflammatory Process and Repair of Critical Bone Defects

Background: The use of biomaterials is commonplace in dentistry for bone regeneration. The aim of this study was to evaluate the performance of a new alloplastic material for bone repair in critical defects and to evaluate the extent of the inflammatory process. Methods: Forty-five New Zealand rabbits were divided into five groups according to evaluation time (7, 14, 30, 60, 120 days), totaling 180 sites with six-millimeter diameter defects in their tibiae. The defects were filled with alloplastic material consisting of poly (lactide-co-caprolactone), beta-tricalcium phosphate, hydroxyapatite and nano-hydroxyapatite (BTPHP) in three different presentations: paste, block, and membrane. Comparisons were established with reference materials, such as Bio-ossTM, Bio-oss CollagenTM, and Bio-gideTM, respectively. The samples were HE-stained and evaluated for inflammatory infiltrate (scored for intensity from 0 to 3) and the presence of newly formed bone at the periphery of the defects. Results: Greater bone formation was observed for the alloplastic material and equivalent inflammatory intensity for both materials, regardless of evaluation time. At 30 days, part of the synthetic biomaterial, regardless of the presentation, was resorbed. Conclusions: We concluded that this novel alloplastic material showed osteoconductive potential, biocompatibility, low inflammatory response, and gradual resorption, thus an alternative strategy for guided bone regeneration

Staining of esthetic brackets by plaque disclosing solutions

AIM: To evaluate the staining of esthetic orthodontic brackets by plaque disclosing solutions. METHODS: Two types of brackets manufactured by GAC/DENTSPLY(r) were evaluated: ceramic (n=30) and polycarbonate (n=30). The brackets were divided into 6 groups. Two control groups (n=6) were immersed in absolute ethanol: GI - ceramic brackets and GII - polycarbonate brackets. Four experimental groups (n=12) were immersed in different plaque disclosing solutions: GIII (ceramic brackets) and GIV (polycarbonate brackets) were immersed in Replak(r); GV (ceramic brackets) and GVI (polycarbonate brackets) were immersed in Replasul "S"(r). Relative quantitative analysis of the influence of plaque disclosing tablets on bracket staining was performed using reflectance spectrophotometry of stain deposition. Exploratory analysis of the data was performed using Analysis of Variance (ANOVA) in a 2x2 factorial setup (bracket x immersion) with additional treatments (controls). RESULTS: The results demonstrated that the ceramic brackets presented the highest amount of staining when Replasul "S"(r) was used (pd"0.05). However, when Replak(r) was used, no statistically significant difference was found in comparison with the control group (p>0.05). For polycarbonate brackets, staining was detected for both disclosing solutions (p>0.05). CONCLUSIONS: The disclosing solutions caused stain formation on polycarbonate brackets and, under the tested conditions, use of Replak(r) on ceramic brackets did not cause staining

Collagen Type I may Influence the Expression of E-Cadherin and Beta-catenin in Carcinoma Ex-pleomorphic Adenoma

Carcinoma ex-pleomorphic adenoma (CXPA) is an aggressive salivary gland malignancy, usually derived from a long-standing or a recurrent benign tumor, the pleomorphic adenoma (PA). In the context of dynamic reciprocity, changes in the composition and structure of extracellular matrix proteins and cell surface receptors have been frequently associated with dysfunctional adhesion and invasive behavior of tumor cells. It is not fully understood if these changes are involved in the conversion of PA to CXPA. In this study, different progression stages of CXPA were investigated regarding the expression of the major extracellular matrix proteins, collagen type I, and of E-cadherin and beta-catenin, the components of adherens junctions. By immunohistochemical analysis, we have demonstrated that direct contact of tumor cells with fibrillar type I collagen, particularly near the invasive front and in invasive areas prevailing small nests of CXPA cells, could be associated with reduced expression of the E-cadherin and beta-catenin adhesion molecules and with invasive behavior of epithelial; but not of CXPA with myoepithelial component. Our results also suggested that this association could depend on the organization of collagen molecules, being prevented by high-order polymeric structures. These findings could implicate the local microenvironment in the transition from the premalignant PA to invasive CXPA.FAPESPCNP

In vitro influence of the extracellular matrix in myoepithelial cells stimulated by malignant conditioned medium

In order to investigate the role of myoepithelial cell and tumor microenvironment in salivary gland neoplasma, we have performed a study towards the effect of different extracellular matrix proteins (basement membrane matrix, type I collagen and fibronectin) on morphology and differentiation of benign myoepithelial cells from pleomorphic adenoma cultured with malignant cell culture medium from squamous cell carcinoma. We have also analyzed the expression of alpha-smooth muscle actin (alpha-SMA) and FGF-2 by immunofluorescence and qPCR. Our immunofluorescence results, supported by qPCR analysis, demonstrated that alpha-SMA and FGF-2 were upregulated in the benign myoepithelial cells from pleomorphic adenoma in all studied conditions on fibronectin substratum. However, the myoepithelial cells on fibronectin substratum did not alter their morphology under malignant conditioned medium stimulation and exhibited a stellate morphology and, occasionally focal adhesions with the substratum. In summary, our data demonstrated that the extracellular matrix exerts an important role in the morphology of the benign myoepithelial cells by the presence of focal adhesions and also inducing increase FGF-2 and alpha-SMA expression by these cells, especially in the fibronectin substratum. (C) 2011 Elsevier Ltd. All rights reserved.FAPESP/Brazil [2008/58722-3, 2008/58721-7

In vitro influence of the extracellular matrix in myoepithelial cells stimulated by malignant conditioned medium

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)In order to investigate the role of myoepithelial cell and tumor microenvironment in salivary gland neoplasma, we have performed a study towards the effect of different extracellular matrix proteins (basement membrane matrix, type I collagen and fibronectin) on morphology and differentiation of benign myoepithelial cells from pleomorphic adenoma cultured with malignant cell culture medium from squamous cell carcinoma. We have also analyzed the expression of alpha-smooth muscle actin (alpha-SMA) and FGF-2 by immunofluorescence and qPCR. Our immunofluorescence results, supported by qPCR analysis, demonstrated that alpha-SMA and FGF-2 were upregulated in the benign myoepithelial cells from pleomorphic adenoma in all studied conditions on fibronectin substratum. However, the myoepithelial cells on fibronectin substratum did not alter their morphology under malignant conditioned medium stimulation and exhibited a stellate morphology and, occasionally focal adhesions with the substratum. In summary, our data demonstrated that the extracellular matrix exerts an important role in the morphology of the benign myoepithelial cells by the presence of focal adhesions and also inducing increase FGF-2 and alpha-SMA expression by these cells, especially in the fibronectin substratum. (C) 2011 Elsevier Ltd. All rights reserved.482102109Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Staining of esthetic brackets by plaque disclosing solutions

evaluate the staining of esthetic orthodontic brackets by plaque disclosing solutions. Methods: Two types of brackets manufactured by GAC/DENTSPLY® were evaluated: ceramic (n=30) and polycarbonate (n=30). The brackets were divided into 6 groups. Two control groups (n=6) were immersed in absolute ethanol: GI - ceramic brackets and GII - polycarbonate brackets. Four experimental groups (n=12) were immersed in different plaque disclosing solutions: GIII (ceramic brackets) and GIV (polycarbonate brackets) were immersed in Replak®; GV (ceramic brackets) and GVI (polycarbonate brackets) were immersed in Replasul S®. Relative quantitative analysis of the influence of plaque disclosing tablets on bracket staining was performed using reflectance spectrophotometry of stain deposition. Exploratory analysis of the data was performed using Analysis of Variance (ANOVA) in a 2x2 factorial setup (bracket x immersion) with additional treatments (controls). Results: The results demonstrated that the ceramic brackets presented the highest amount of staining when Replasul S® was used (pd0.05). However, when Replak® was used, no statistically significant difference was found in comparison with the control group (p>0.05). For polycarbonate brackets, staining was detected for both disclosing solutions (p>0.05). Conclusions: The disclosing solutions caused stain formation on polycarbonate brackets and, under the tested conditions, use of Replak® on ceramic brackets did not cause staining

Cellular senescence and autophagy of myoepithelial cells are involved in the progression of in situ areas of carcinoma Ex-pleomorphic adenoma to invasive carcinoma. An in vitro model

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)During tumor invasion, benign myoepithelial cells of carcinoma ex-pleomorphic adenoma (CXPA) surround malignant epithelial cells and disappear. The mechanisms involved in the death and disappearance of these myoepithelial cells were investigated via analysis of the expression of regulatory proteins for apoptosis, autophagy and cellular senescence in an in situ in vitro model. Protein expression relating to apoptosis (Bax, Bcl-2, Survivin), autophagy (Beclin-1, LC3B) and cellular senescence (p21, p16) was evaluated using indirect immunofluorescence. beta-galactosidase expression was assessed via histochemistry. Biopsies of CXPA (ex vivo) allowed immunhistochemical evaluation of p21 and p16, whilst LC3B, p21 and p16 protein expression was analyzed by western blotting. In the in vitro model, the myoepithelial cells were positive for LC3B (cytoplasm) and p21 (nucleus), whilst in vivo positivity for p21 and p16 was observed. In vitro, beta-galactosidase activity increased in the myoepithelial cells over time. Western blotting analysis revealed an increased LC3B, p16 and p21 expression in the myoepithelial cells with previous contact with the malignant cells when compared with those without contact. The investigation of behavior of benign myoepithelial cells in ductal areas of CXAP revealed that the myoepithelial cells are involved in the autophagy-senescence phenotype that subsequently leads to their disappearance.During tumor invasion, benign myoepithelial cells of carcinoma ex-pleomorphic adenoma (CXPA) surround malignant epithelial cells and disappear. The mechanisms involved in the death and disappearance of these myoepithelial cells were investigated via analy93255265FAPESP - FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULOCNPQ - CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2011/21157-0]CNPq [473939/2011-8]2011/21157-0473939/2011-