RESUMEN
BACKGROUND: Cancer might be envisaged as the result of a genetic process causing the unregulated proliferation of a given cell as well as its inability to undergo differentiation and/or apoptosis. Alterations of genes regulating cell division cycle appear to play a key role in the development of human cancer. OBJECTIVE: On the bases of the above considerations, we decided to establish new cell lines from human melanoma specimens, in order to analyse the molecular alterations in primary preparations of malignant cells. RESULTS: The present paper describes two new established cell lines and their genetic and biochemical features. Both the melanoma cell lines show inactivation of the cyclin-dependent kinase inhibitor gene, CDKN2A/p16INK4A, thus demostrating that this alteration occurs in primary human melanomas. No other alterations were observable when we investigated several different cell cycle genes including those encoding cyclins, cyclin-dependent kinases and cyclin-dependent kinase inhibitors. Analyses at protein level by means of immunoblotting confirmed the results obtained at the genetic level. Moreover, the inducibility of a pivotal cyclin-dependent kinase inhibitor gene, namely p21CIP1 gene, was obtained by treating the cells with histone deacetylase inhibitors, namely butyrate and phenylbutyrate. CONCLUSIONS: Our results suggest a primary role of cyclin-dependent kinase inhibitor genes inactivation in the origin of human melanoma and allow the proposal of new therapeutic strategies based on the transcriptional activation of p21CIP1 gene.
Asunto(s)
Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Melanoma/genética , Neoplasias Cutáneas/genética , División Celular , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/genética , Inhibidores Enzimáticos , Genes p16 , Humanos , Immunoblotting , Melanoma/patología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Neoplasias Cutáneas/patología , Células Tumorales CultivadasRESUMEN
A wealth of evidence correlates the chemopreventive activity of a fiber-rich diet with the production of butyrate. In order to identify the genes transcriptionally modulated by the molecule, we analyzed the expression profile of butyrate-treated colon cancer cells by means of cDNA expression arrays. Moreover, the effect of trichostatin A, a specific histone deacetylase inhibitor, was studied. A superimposable group of 23 genes out of 588 investigated is modulated by both butyrate and trichostatin A. Among them, a major target was tob-1, a gene involved in the control of cell cycle. tob-1 is also up-regulated by butyrate in a neuroblastoma-derived cell line, and its overexpression in the colon cells caused growth arrest. Our findings represent an extensive analysis of genes modulated by butyrate and identify completely new effectors of its biological activities.
Asunto(s)
Butiratos/farmacología , Proteínas de Unión al ADN/genética , Expresión Génica/efectos de los fármacos , Histonas/metabolismo , Factores de Transcripción/genética , Acetilación , Neoplasias del Colon , Cicloheximida/farmacología , Proteínas de Unión al ADN/metabolismo , Inhibidores Enzimáticos/farmacología , Factor de Transcripción GATA2 , Perfilación de la Expresión Génica , Células HT29 , Inhibidores de Histona Desacetilasas , Histona Desacetilasas/metabolismo , Histonas/fisiología , Humanos , Ácidos Hidroxámicos/farmacología , Análisis de Secuencia por Matrices de Oligonucleótidos , Inhibidores de la Síntesis de la Proteína/farmacología , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Factores de Transcripción/metabolismo , Células Tumorales CultivadasRESUMEN
OBJECTIVES: Previous histochemical observations have suggested a possible involvement of the bcl-2 family genes in the acquisition of neoplastic phenotype of the endometrium. Since knowledge of the type and function of genes controlling the transformed cell may result in new diagnostic, prognostic, and therapeutic approaches, we have investigated at the molecular level the biological role of bcl-2 family genes in endometrial neoplastic cells. METHODS: To investigate the relationship between the sensitivity to apoptosis and the expression of the bcl-2 family genes, we set up a model system consisting of four human endometrial carcinoma cell lines. This system constitutes an array of two cell pairs presenting, respectively, endometrioid and adenosquamous phenotypes. G2 and G3 gradings are represented within each pair; in addition, each set contains one cell line that is apoptosis-sensitive and one that is resistant. Transfection of bcl-2 and bcl-XL into apoptosis-sensitive cells was used to monitor the biological function of protective genes. RESULTS: A differential pattern of expression of bcl-2 family genes was observed in apoptosis-sensitive versus resistant cells, independent from the histological subtype. Resistant lines exhibited high amounts of Bcl-XL and low amounts of Bcl-2. Bax expression clearly correlates with cellular susceptibility to apoptosis. Transfection of bcl-XL resulted in a dose-dependent enhancement in resistance toward apoptosis. In contrast, the main effect of bcl-2 constitutive overexpression was to drastically abate the proliferative potential of transfected cells. CONCLUSIONS: These data demonstrate, at the molecular level, that bcl-XL is selected as an apoptosis-protective gene in place of bcl-2 while bax retains its dominant proapototic role.