RESUMO
p53 is a tumor suppressor with a pro-death role in many conditions. However, in some contexts, evidence supports a pro-survival function. p53 has been shown to be activated in acetaminophen (APAP) toxicity but the impact of this on toxicity is uncertain. In the present study, we have found that p53 plays a protective role in APAP-induced liver injury. We inhibited p53 using three different approaches in mice, pifithrin-α (PFTα), knockdown of p53 expression with antisense oligonucleotide, and p53 knockout. Mice were treated with APAP (300mg/kg) i.p. and after 24h in all three conditions, the liver injury was more severe as reflected in higher ALT levels and great area of necrosis in histology of the liver. Conversely, a p53 activator, nutlin-3a, decreased the liver injury induced by APAP. In the p53 inhibition models, enhanced sustained JNK activation was seen in the early time course, while the JNK was suppressed with the p53 activator. In conclusion, p53 plays a novel protective role in APAP induced liver injury through inhibiting the activation of JNK, a key mediator in APAP-induced oxidative stress.
Assuntos
Acetaminofen/toxicidade , Doença Hepática Induzida por Substâncias e Drogas/genética , Fígado/efeitos dos fármacos , MAP Quinase Quinase 4/genética , Proteína Supressora de Tumor p53/genética , Animais , Benzotiazóis/administração & dosagem , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/fisiopatologia , Técnicas de Silenciamento de Genes , Técnicas de Inativação de Genes , Hepatócitos/efeitos dos fármacos , Hepatócitos/patologia , Humanos , Imidazóis/administração & dosagem , Fígado/fisiopatologia , Camundongos , Oligonucleotídeos Antissenso/genética , Piperazinas/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Tolueno/administração & dosagem , Tolueno/análogos & derivados , Proteína Supressora de Tumor p53/antagonistas & inibidoresRESUMO
Oxidative stress is a persistent threat to the genome and is associated with major causes of human mortality, including cancer, atherosclerosis, and aging. Here we established a method to generate libraries of genomic DNA fragments containing oxidatively modified bases by using specific monoclonal antibodies to immunoprecipitate enzyme-digested genome DNA. We applied this technique to two different base modifications, 8-hydroxyguanine and 1,N6-propanoadenine (acrotein-Ade), in a ferric nitrilotriacetate-induced murine renal carcinogenesis model. Renal cortical genomic DNA derived from 10- to 12-week-old male C57BL/6 mice, of untreated control or 6 hours after intraperitoneal injection of 3 mg iron/kg ferric nitrilotriacetate, was enzyme digested, immunoprecipitated, cloned, and mapped to each chromosome. The results revealed that distribution of the two modified bases was not random but differed in terms of chromosomes, gene size, and expression, which could be partially explained by chromosomal territory. In the wild-type mice, low GC content areas were more likely to harbor the two modified bases. Knockout of OGG1, a repair enzyme for genomic 8-hydroxyguanine, increased the amounts of acrolein-Ade as determined by quantitative polymerase chain reaction analyses. This versatile technique would introduce a novel research area as a high-throughput screening method for critical genomic loci under oxidative stress.
Assuntos
Adenina/análogos & derivados , Transformação Celular Neoplásica/genética , Mapeamento Cromossômico/métodos , Genes Neoplásicos/genética , Guanina/análogos & derivados , Neoplasias Renais/genética , Estresse Oxidativo , Acroleína/química , Adenina/análise , Adenina/química , Animais , Anticorpos Monoclonais/imunologia , DNA/química , DNA/genética , DNA Glicosilases/genética , Expressão Gênica , Biblioteca Gênica , Genoma/genética , Guanina/análise , Guanina/imunologia , Imunoprecipitação , Rim/química , Masculino , Camundongos , Camundongos Knockout , OxirreduçãoRESUMO
Oxidative stress is a persistent threat to the genome and is associated with major causes of human mortality, including cancer, atherosclerosis, and aging. Here we established a method to generate libraries of genomic DNA fragments containing oxidatively modified bases by using specific monoclonal antibodies to immunoprecipitate enzyme-digested genome DNA. We applied this technique to two different base modifications, 8-hydroxyguanine and 1,N6-propanoadenine (acrotein-Ade), in a ferric nitrilotriacetate-induced murine renal carcinogenesis model. Renal cortical genomic DNA derived from 10- to 12-week-old male C57BL/6 mice, of untreated control or 6 hours after intraperitoneal injection of 3 mg iron/kg ferric nitrilotriacetate, was enzyme digested, immunoprecipitated, cloned, and mapped to each chromosome. The results revealed that distribution of the two modified bases was not random but differed in terms of chromosomes, gene size, and expression, which could be partially explained by chromosomal territory. In the wild-type mice, low GC content areas were more likely to harbor the two modified bases. Knockout of OGG1, a repair enzyme for genomic 8-hydroxyguanine, increased the amounts of acrolein-Ade as determined by quantitative polymerase chain reaction analyses. This versatile technique would introduce a novel research area as a high-throughput screening method for critical genomic loci under oxidative stress.
