RESUMO
Genomic methylation patterns of Dunning R-3327 cell lines anaplastic tumor-1 (AT-1), anaplastic tumor-3 (AT-3), metastasis-lymph and lung (Mat-LyLu) and metastasis-lung (Mat-Lu), and Mat-LyLu cells treated with difluoromethylornithine (DFMO), and retinoic acid (RA) have been analyzed. Each cell line was digested with HpaII and MspI restriction endonuclease enzymes to characterize methylation patterns, at the interior cytosine of the sequence CmCGG. Identical molecular weight banding patterns were found for both HpaII and MspI digests in normal dorsal prostate (NDP) used as a control. Both the treated and non-treated Dunning R-3327 cells digested with HpaII and MspI, displayed similar banding profiles from those seen in NDP solid tissues, indicative of a progressive loss of methylation at CCGG sites.
Assuntos
Adenocarcinoma/genética , Adenocarcinoma/metabolismo , DNA de Neoplasias/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Antineoplásicos/farmacologia , DNA de Neoplasias/genética , Eflornitina/farmacologia , Humanos , Masculino , Metilação/efeitos dos fármacos , Próstata/metabolismo , Valores de Referência , Tretinoína/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacosRESUMO
Detection of point mutations in the Dunning System with the Ki-ras2 gene in the first and second positions of codon 12 exon I, has been performed using allele-specific oligonucleotide (ASO) primers. PCR generated 94bp templates and genomic DNA extracted from the Dunning cell lines AT-1, AT-3, Mat-Lu, and Mat-Lylu were tested. Our investigation revealed that the first and second positions of codon 12 have undergone either transitions or transversions from the wild type sequence (GG). Dorsal prostate (DP) solid tissues used as controls revealed the wild type configuration as well as transversions at both positions, in addition to a transition in the first position for Mat-Lu. The most aberrant of the Dunning lines (AT-3 and Mat-LyLu) collectively displayed sequence changes (transitions and transversions) with no evidence of the wild type configuration. These findings are supportive of biometric studies (area, shape factor, and motility) along with adhesion and invasion assays done in our laboratory in correlating genotypic and phenotypic properties to metastatic potential.