RESUMO
Biological functions of metallothionein (MT) proteins which are encoded by 10 functional MT isoforms, include cell proliferation, differentiation and apoptosis. The aim of this study was to compare the relative expression levels of functional MT mRNA isoforms in three nasopharyngeal cancer (NPC) cell lines with laryngeal carcinoma and embryonic lung cell lines by quantitative real-time RT-PCR. All the NPC lines exhibited expression of the MT-2A transcript, whereas the MT-1E isoform was expressed in well differentiated HK1 and moderately differentiated TW01 but not in poorly differentiated CNE2 cells. Interestingly, TW01 and HEp-2 laryngeal cancer cells exhibited similar expression profiles with both MT-1E and MT-2A isoforms being detected at levels below those of MRC-5 embryonic lung fibroblasts. Functional studies of the MT-2A isoform by down-regulating expression of this gene with MT-2A antisense oligonucleotide in CNE2 cells, showed a reduction in cell viability and proliferation. These findings may provide valuable information in the search for novel therapeutic strategies against NPC.
Assuntos
Antineoplásicos/farmacologia , Inibidores do Crescimento/farmacologia , Metalotioneína/antagonistas & inibidores , Neoplasias Nasofaríngeas/metabolismo , Oligodesoxirribonucleotídeos Antissenso/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Regulação para Baixo , Inibidores do Crescimento/genética , Humanos , Metalotioneína/genética , Metalotioneína/metabolismo , Neoplasias Nasofaríngeas/genética , Oligodesoxirribonucleotídeos Antissenso/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Células Tumorais CultivadasRESUMO
Metallothionein (MT), a low-molecular weight protein with pleiotropic functions, is believed to play an important role in tumorigenesis. The aim of this study was to compare the expression of functional MT-1 and MT-2 mRNA isoforms in five breast cancer cell lines ranging from noninvasive MCF7 breast cancer cells to highly aggressive MDA-MB-231 breast cancer cells together with breast myoepithelial cells in vitro by conventional semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and quantitative real-time RT-PCR. The MT-2A isoform was observed to be differentially upregulated in the invasive phenotype. The MT-1E isoform was found to be present in estrogen receptor-negative breast cancer cell lines (MDA-MB-231 and Hs578T) but not detectable in the estrogen receptor-positive cell lines (T47D, MCF7, and ZR75-1 cells). Only the myoepithelial cells exhibited the presence of the MT-1G transcript. Direct sequencing of the RT-PCR products revealed the occurrence of a variant MT-1H isoform with changes in amino acid residues in the protein sequence and notable differences in the predicted secondary protein structure. The observations in this study are relevant to the development of novel approaches to metastatic breast cancer disease, and may herald the search for novel MT mutants and the elucidation of their biological roles.