RESUMEN
ABSTRACT Objective: To verify the physiological action of triiodothyronine T3 on the expression of transforming growth factor α (TGFA) mRNA in MCF7 cells by inhibition of RNA Polymerase II and the MAPK/ERK pathway Materials and methods: The cell line was treated with T3 at a physiological dose (10−9M) for 10 minutes, 1 and 4 hour (h) in the presence or absence of the inhibitors, α-amanitin (RNA polymerase II inhibitor) and PD98059 (MAPK/ERK pathway inhibitor). TGFA mRNA expression was analyzed by RT-PCR. For data analysis, we used ANOVA, complemented with the Tukey test and Student t-test, with a minimum significance of 5%. Results: T3 increases the expression of TGFA mRNA in MCF7 cells in 4 h of treatment. Inhibition of RNA polymerase II modulates the effect of T3 treatment on the expression of TGFA in MCF7 cells. Activation of the MAPK/ERK pathway is not required for T3 to affect the expression of TGFA mRNA. Conclusion: Treatment with a physiological concentration of T3 after RNA polymerase II inhibition altered the expression of TGFA. Inhibition of the MAPK/ERK pathway after T3 treatment does not interfere with the TGFA gene expression in a breast adenocarcinoma cell line.
Asunto(s)
Humanos , Femenino , Triyodotironina/genética , Neoplasias de la Mama/genética , Adenocarcinoma/genética , Regulación Neoplásica de la Expresión Génica/genética , Factor de Crecimiento Transformador alfa/genética , Sistema de Señalización de MAP Quinasas/genética , Triyodotironina/metabolismo , Triyodotironina/farmacología , Proto-Oncogenes/genética , Neoplasias de la Mama/metabolismo , ARN Mensajero/genética , Adenocarcinoma/metabolismo , Factor de Crecimiento Transformador alfa/efectos de los fármacos , Factor de Crecimiento Transformador alfa/metabolismo , Línea Celular Tumoral/metabolismo , Células MCF-7/metabolismoRESUMEN
OBJECTIVE: To verify the physiological action of triiodothyronine T3 on the expression of transforming growth factor α (TGFA) mRNA in MCF7 cells by inhibition of RNA Polymerase II and the MAPK/ERK pathway. MATERIALS AND METHODS: The cell line was treated with T3 at a physiological dose (10-9M) for 10 minutes, 1 and 4 hour (h) in the presence or absence of the inhibitors, α-amanitin (RNA polymerase II inhibitor) and PD98059 (MAPK/ERK pathway inhibitor). TGFA mRNA expression was analyzed by RT-PCR. For data analysis, we used ANOVA, complemented with the Tukey test and Student t-test, with a minimum significance of 5%. RESULTS: T3 increases the expression of TGFA mRNA in MCF7 cells in 4 h of treatment. Inhibition of RNA polymerase II modulates the effect of T3 treatment on the expression of TGFA in MCF7 cells. Activation of the MAPK/ERK pathway is not required for T3 to affect the expression of TGFA mRNA. CONCLUSION: Treatment with a physiological concentration of T3 after RNA polymerase II inhibition altered the expression of TGFA. Inhibition of the MAPK/ERK pathway after T3 treatment does not interfere with the TGFA gene expression in a breast adenocarcinoma cell line.
Asunto(s)
Adenocarcinoma/genética , Neoplasias de la Mama/genética , Regulación Neoplásica de la Expresión Génica/genética , Sistema de Señalización de MAP Quinasas/genética , Factor de Crecimiento Transformador alfa/genética , Triyodotironina/genética , Adenocarcinoma/metabolismo , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral/metabolismo , Femenino , Humanos , Células MCF-7/metabolismo , Proto-Oncogenes Mas , Proto-Oncogenes/genética , ARN Mensajero/genética , Factor de Crecimiento Transformador alfa/efectos de los fármacos , Factor de Crecimiento Transformador alfa/metabolismo , Triyodotironina/metabolismo , Triyodotironina/farmacologíaRESUMEN
O feocromocitoma é um tumor secretor de catecolaminas de difícil diagnóstico, pois as manifestações clínicas nem sempre estão presentes, já que a secreção de epinefrina e norepinefrina ocorre de maneira episódica. O objetivo deste trabalho é relatar o caso de um cão maltês macho, de nove anos de idade, assintomático, que apresentava uma massa adrenal identificada ao acaso durante um exame ultrassonográfico. Foi indicado o tratamento cirúrgico, que no entanto não foi realizado inicialmente. Após oito meses, o animal apresentou trombo neoplásico em veia cava caudal, sendo submetido a adrenalectomia e trombectomia. A análise histopatológica revelou um feocromocitoma maligno e trombo neoplásico. Ressaltase aqui a importância do diagnóstico e do tratamento cirúrgico dos tumores adrenais, mesmo quando identificados ao acaso em animais assintomáticos. Atualmente, cinco anos após o tratamento cirúrgico, o paciente encontra-se estável, saudável e sem recidiva ou evidências de metástase do tumor.
Pheochromocytoma is a catecholamine secreting tumor that may be difficull to diagnose because clinical signs are not always present due to the episodic secretion of epinephrine and norepinephrine. We report a case of pheochromocytoma in an asymptomatic 9-year-old intact male maltese, The adrenal mass was an incidental finding during an abdominal ultrasonographic exam. Surgical treatment was declined by the owner. Eight months laler the animal was presented with neoplastic thrombus in the caudal vena cava, and was submitted to adrenalectomy with venotomy and thrombectomy. The histopathological exam confirmed the diagnosis malignant pheochromocytoma and neoplastic thrombus. The importance of diagnosis and surgical treatment of adrenal tumors, including cases of incidental finding in asymptomatic animais is emphasized here. Currently, five years after surgery, the dog is stable, healthy and without recurrence or evidences of tumor metastasis.
El feocromocitoma es un tumor secretor de catecolaminas de difícil diagnóstico debido a que pocas veces presenta signos clínicos, dado que la secreción de epinefrina y norepinefrina no son constantes. El objetivo de este trabajo fue relatar el caso de un perro Maltés macho de nueve años, asintomático, que presentaba una masa adrenal encontrada casualmente durante un examen ecográfico. Se indicó el tratamiento quirúrgico, pero no fue realizado en forma inmediata. Ocho meses después, el paciente presentó un trombo neoplásico en vena cava caudal, por lo que se realizó una adrenalectomía y trombectomía. En el examen histopatológico fue diagnosticado un feocromocitoma maligno y presencia de trombo neoplásico. Destacamos la importancia del diagnóstico y del tratamiento quirúrgico de los tumores adrenales, aún cuando se los identifica en forma casual en animales asintomáticos. Actualmente, cinco años después de la cirugía, el paciente se encuentra estable, sin alteraciones clínicas ni evidencias de metástasis del tumor.
Asunto(s)
Animales , Perros , Feocromocitoma/diagnóstico por imagen , Feocromocitoma/veterinaria , Neoplasias de las Glándulas Suprarrenales/diagnóstico por imagen , Neoplasias de las Glándulas Suprarrenales/veterinaria , Adrenalectomía/veterinaria , Trombectomía/veterinariaRESUMEN
O feocromocitoma é um tumor secretor de catecolaminas de difícil diagnóstico, pois as manifestações clínicas nem sempre estão presentes, já que a secreção de epinefrina e norepinefrina ocorre de maneira episódica. O objetivo deste trabalho é relatar o caso de um cão maltês macho, de nove anos de idade, assintomático, que apresentava uma massa adrenal identificada ao acaso durante um exame ultrassonográfico. Foi indicado o tratamento cirúrgico, que no entanto não foi realizado inicialmente. Após oito meses, o animal apresentou trombo neoplásico em veia cava caudal, sendo submetido a adrenalectomia e trombectomia. A análise histopatológica revelou um feocromocitoma maligno e trombo neoplásico. Ressaltase aqui a importância do diagnóstico e do tratamento cirúrgico dos tumores adrenais, mesmo quando identificados ao acaso em animais assintomáticos. Atualmente, cinco anos após o tratamento cirúrgico, o paciente encontra-se estável, saudável e sem recidiva ou evidências de metástase do tumor.(AU)
Pheochromocytoma is a catecholamine secreting tumor that may be difficull to diagnose because clinical signs are not always present due to the episodic secretion of epinephrine and norepinephrine. We report a case of pheochromocytoma in an asymptomatic 9-year-old intact male maltese, The adrenal mass was an incidental finding during an abdominal ultrasonographic exam. Surgical treatment was declined by the owner. Eight months laler the animal was presented with neoplastic thrombus in the caudal vena cava, and was submitted to adrenalectomy with venotomy and thrombectomy. The histopathological exam confirmed the diagnosis malignant pheochromocytoma and neoplastic thrombus. The importance of diagnosis and surgical treatment of adrenal tumors, including cases of incidental finding in asymptomatic animais is emphasized here. Currently, five years after surgery, the dog is stable, healthy and without recurrence or evidences of tumor metastasis.(AU)
El feocromocitoma es un tumor secretor de catecolaminas de difícil diagnóstico debido a que pocas veces presenta signos clínicos, dado que la secreción de epinefrina y norepinefrina no son constantes. El objetivo de este trabajo fue relatar el caso de un perro Maltés macho de nueve años, asintomático, que presentaba una masa adrenal encontrada casualmente durante un examen ecográfico. Se indicó el tratamiento quirúrgico, pero no fue realizado en forma inmediata. Ocho meses después, el paciente presentó un trombo neoplásico en vena cava caudal, por lo que se realizó una adrenalectomía y trombectomía. En el examen histopatológico fue diagnosticado un feocromocitoma maligno y presencia de trombo neoplásico. Destacamos la importancia del diagnóstico y del tratamiento quirúrgico de los tumores adrenales, aún cuando se los identifica en forma casual en animales asintomáticos. Actualmente, cinco años después de la cirugía, el paciente se encuentra estable, sin alteraciones clínicas ni evidencias de metástasis del tumor.(AU)
Asunto(s)
Animales , Perros , Feocromocitoma/diagnóstico por imagen , Feocromocitoma/veterinaria , Neoplasias de las Glándulas Suprarrenales/diagnóstico por imagen , Neoplasias de las Glándulas Suprarrenales/veterinaria , Adrenalectomía/veterinaria , Trombectomía/veterinariaRESUMEN
Human adipose tissue-derived stem cells (hASCs) have been subjected to extensive investigation because of their self-renewal properties and potential to restore damaged tissues. In the literature, there are several protocols for differentiating hASCs into osteoblasts, but there is no report on the control of cell viability during this process. In this study, we used osteoblasts derived from hASCs of patients undergoing abdominoplasty. The cells were observed at the beginning and end of bone matrix formation, and the expression of proteins involved in this process, including alkaline phosphatase and osteocalcin, was assessed. RANKL, Osterix, Runx2, Collagen3A1, Osteopontin and BSP expression levels were analyzed using real-time PCR, in addition to a quantitative assessment of protein levels of the markers CD45, CD105, STRO-1, and Nanog, using immunofluorescence. Rhodamine (Rho123), cytochrome-c, caspase-3, P-27, cyclin D1, and autophagy cell markers were analyzed by flow cytometry to demonstrate potential cellular activity and the absence of apoptotic and tumor cell processes before and after cell differentiation. The formation of bone matrix, along with calcium nodules, was observed after 16 days of osteoinduction. The gene expression levels of RANKL, Osterix, Runx2, Collagen3A1, Osteopontin, BSP and alkaline phosphatase activity were also elevated after 16 days of osteoinduction, whereas the level of osteocalcin was higher after 21 days of osteoinduction. Our data also showed that the cells had a high mitochondrial membrane potential and a low expression of apoptotic and tumor markers, both before and after differentiation. Cells were viable after the different phases of differentiation. This proposed methodology, using markers to evaluate cell viability, is therefore successful in assessing different phases of stem cell isolation and differentiation.
Asunto(s)
Tejido Adiposo/citología , Supervivencia Celular , Osteoblastos/citología , Células Madre/citología , Fosfatasa Alcalina/metabolismo , Biomarcadores/metabolismo , Diferenciación Celular , Células Cultivadas , Colágeno Tipo III/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Regulación de la Expresión Génica , Humanos , Potencial de la Membrana Mitocondrial , Microscopía Fluorescente , Modelos Biológicos , Osteoblastos/metabolismo , Osteopontina/metabolismo , Ligando RANK/metabolismo , Vimentina/metabolismoRESUMEN
Human adipose tissue-derived stem cells (hASCs) have been subjected to extensive investigation because of their self-renewal properties and potential to restore damaged tissues. In the literature, there are several protocols for differentiating hASCs into osteoblasts, but there is no report on the control of cell viability during this process. In this study, we used osteoblasts derived from hASCs of patients undergoing abdominoplasty. The cells were observed at the beginning and end of bone matrix formation, and the expression of proteins involved in this process, including alkaline phosphatase and osteocalcin, was assessed. RANKL, Osterix, Runx2, Collagen3A1, Osteopontin and BSP expression levels were analyzed using real-time PCR, in addition to a quantitative assessment of protein levels of the markers CD45, CD105, STRO-1, and Nanog, using immunofluorescence. Rhodamine (Rho123), cytochrome-c, caspase-3, P-27, cyclin D1, and autophagy cell markers were analyzed by flow cytometry to demonstrate potential cellular activity and the absence of apoptotic and tumor cell processes before and after cell differentiation. The formation of bone matrix, along with calcium nodules, was observed after 16 days of osteoinduction. The gene expression levels of RANKL, Osterix, Runx2, Collagen3A1, Osteopontin, BSP and alkaline phosphatase activity were also elevated after 16 days of osteoinduction, whereas the level of osteocalcin was higher after 21 days of osteoinduction. Our data also showed that the cells had a high mitochondrial membrane potential and a low expression of apoptotic and tumor markers, both before and after differentiation. Cells were viable after the different phases of differentiation. This proposed methodology, using markers to evaluate cell viability, is therefore successful in assessing different phases of stem cell isolation and differentiation.