RESUMO
Bisphenol A (BPA) is a compound released from plastics and other consumer products used in everyday life. BPA exposure early in fetal development is proposed to contribute to programming of chronic diseases like obesity and diabetes, by affecting DNA methylation levels. Previously, we showed that in utero and lactational exposure of C57BL/6JxFVB hybrid mice via maternal feed using a dose range of 0-3000µg/kg body weight/day resulted in a sex-dependent altered metabolic phenotype in offspring at 23 weeks of age. The most univocal effects were observed in females, with reduced body weights and related metabolic effects associated with perinatal BPA exposure. To identify whether the effects of BPA in females are associated with changes in DNA methylation, this was analyzed in liver, which is important in energy homeostasis. Measurement of global DNA methylation did not show any changes. Genome-wide DNA methylation analysis at specific CpG sites in control and 3000µg/kg body weight/day females with the digital restriction enzyme analysis of methylation (DREAM) assay revealed potential differences, that could, however, not be confirmed by bisulfite pyrosequencing. Overall, we demonstrated that the observed altered metabolic phenotype in female offspring after maternal exposure to BPA was not detectably associated with liver DNA methylation changes. Still, other tissues may be more informative.
Assuntos
Compostos Benzidrílicos/toxicidade , Metilação de DNA/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Poluentes Ambientais/toxicidade , Fígado/efeitos dos fármacos , Fenóis/toxicidade , Fatores Etários , Animais , Cromatografia Líquida de Alta Pressão , Biologia Computacional , Ilhas de CpG , Bases de Dados Genéticas , Metabolismo Energético/genética , Feminino , Idade Gestacional , Fígado/metabolismo , Exposição Materna , Camundongos Endogâmicos C57BL , Fenótipo , Reação em Cadeia da Polimerase , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Fatores SexuaisRESUMO
Head and neck cancer remains a morbid and often fatal disease and at the present time few effective molecular markers have been identified. The purpose of the present work was to identify new molecular markers for head and neck squamous cell carcinoma (HNSCC). We applied methylation-sensitive arbitrarily primed PCR (MS/AP-PCR) to isolate sequences differentially methylated in HNSCC. The most frequently hypermethylated fragment we found maps close to a cytosine guanine dinucleotide (CpG) island on chromosome 9q33.2, and hypermethylation of this CpG island was associated with transcriptional silencing of an alternative transcript of the LHX6 gene. Using combined bisulfite restriction analysis (COBRA), hypermethylation of this fragment was detected in 13 of 14 (92.8%) HNSCC cell lines studied and 21 of 32 (65.6%) primary tumors, whereas little or no methylation was seen in 10 normal oral mucosa samples. We extended this investigation to other cancer cell lines and methylation was found in those derived from colon, breast, leukemia and lung, and methylation was also found in 12/14 primary colon tumors. These findings suggest that differentially methylated (DIME)-6 hypermethylation is a good cancer marker in HNSCC as well as in other kinds of neoplasias and confirm the importance of searching for markers of epigenetic dysregulation in cancer.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/metabolismo , Metilação de DNA , Neoplasias de Cabeça e Pescoço/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Sequência de Bases , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Cromossomos Humanos Par 9 , Primers do DNA , DNA de Neoplasias/sangue , Neoplasias de Cabeça e Pescoço/genética , Humanos , Proteínas com Homeodomínio LIM , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TranscriçãoRESUMO
Hypoxia is a key factor contributing to the progression of human neoplasias and to the development of resistance to chemotherapy. BNIP3 is a proapoptotic member of the Bcl-2 protein family involved in hypoxia-induced cell death. We evaluated the expression and methylation status of BNIP3 gene to better understand the role of epigenetic alteration of its expression in haematopoietic tumours. Methylation of the region around the BNIP3 transcription start site was detected in four acute lymphocytic leukaemia, one multiple myeloma and one Burkitt lymphoma cell lines, and was closely associated with silencing the gene. That expression of BNIP3 was restored by treatment with 5-aza2'-deoxycytidine (5-aza-dC), a methyltransferase inhibitor, which confirmed the gene to be epigenetically inactivated by methylation. Notably, re-expression of BNIP3 using 5-aza2-dC also restored hypoxia-mediated cell death in methylated cell lines. Acetylation of histone H3 in the 5' region of the gene, which was assessed using chromatin immunoprecipitation assays, correlated directly with gene expression and inversely with DNA methylation. Among primary tumours, methylation of BNIP3 was detected in five of 34 (15%) acute lymphocytic leukaemias, six of 35 (17%) acute myelogenous leukaemias and three of 14 (21%) multiple myelomas. These results suggest that aberrant DNA methylation of the 5' CpG island and histone deacetylation play key roles in silencing BNIP3 expression in haematopoietic tumours.
