RESUMO
Chronic inflammation in the gastrointestinal tract increases the risk for development of cancer by an incompletely understood pathway, which may involve microsatellite instability (MSI). Low frequency of MSI referred to as "MSI-L" occurs frequently in chronically inflamed nonneoplastic tissue. In this work, we have tested the hypothesis that oxidative stress may induce the accumulation of frameshift mutations in human microsatellite DNA. Mismatch repair (MMR)-proficient HCT116+chr3 and MMR-deficient HCT116 cells were transfected with pCMV-(CA)13-EGFP, a plasmid that contains a (CA)13 dinucleotide repeat, which disrupts the reading frame of the downstream enhanced green fluorescent protein gene. A dose-dependent increase in frameshift mutations restoring the enhanced green fluorescent protein reading frame was detected in HCT116 by flow cytometry. At 1 mM H2O2, the mutant fraction was 9-fold higher than that in mock-treated control cells. Although demonstrating stability at lower H2O2 concentrations, MMR-proficient HCT116+chr3 cells accumulated mutations at the 1 mM H2O2 level (4.1-fold above mock-treated control). No significant mutations were detected when HCT116 cells were transfected with the pCMV-(N)26-EGFP construct that contains 26 nucleotides in a random sequence. These data indicate that oxidative stress is a potential mutagen leading to accumulation of frameshift mutations and may contribute to MSI in the setting of chronic inflammation.
Assuntos
Neoplasias Colorretais/genética , Mutação da Fase de Leitura , Repetições de Microssatélites/genética , Estresse Oxidativo/genética , Pareamento Incorreto de Bases , Sobrevivência Celular/fisiologia , Neoplasias Colorretais/metabolismo , Reparo do DNA , Relação Dose-Resposta a Droga , Citometria de Fluxo , Humanos , Peróxido de Hidrogênio/farmacologia , Repetições de Microssatélites/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
Myeloperoxidase (MPO) is a myeloid-specific enzyme that generates hypochlorous acid and other reactive oxygen species. MPO is present at high levels in circulating neutrophils and monocytes but is not detectable in microglia, brain-specific macrophages, in normal brain tissue. However, an earlier study indicated that MPO is present in macrophage-microglia at multiple sclerosis lesions, suggesting that reactivation of MPO gene expression may play a role in neurodegenerative diseases involving macrophage-microglia. In the present study, MPO is shown to colocalize with amyloid beta (Abeta) in senile plaques in cerebral cortex sections from Alzheimer's disease (AD) brain tissue. Microglia costaining for MPO and CD68 are closely associated with plaques, suggesting that plaque components induce MPO expression in microglia. In support of this interpretation, treatment of rodent microglia with aggregated Abeta(1-42) was shown to induce MPO mRNA expression. Also, the ApoE4 allele, the major AD risk factor associated with increased Abeta deposition, was shown to correlate with increased MPO deposition in plaques (P = 0.01, ANOVA). Finally, a genetic polymorphism links MPO expression to Alzheimer's risk, in that a higher expressing SpSp MPO genotype was associated with increased incidence of AD in females, and decreased incidence in males (P = 0.006). These findings suggest that the MPO polymorphism is a gender-specific risk factor for Alzheimer's disease.