RESUMEN
5-Fluorouracil (5-FU) is a widely used anticancer drug for the treatment of colorectal cancer (CRC). However, resistance to 5-FU often prevents the success of chemotherapy. Nuclear factor-erythroid 2-related factor 2 (Nrf2) is a transcriptional regulator and a possible target to overcome 5-FU resistance. The present study examined epigenetic changes associated with Nrf2 induction in a human CRC cell line (SNUC5) resistant to 5-FU (SNUC5/5-FUR). Nrf2 expression, nuclear translocation, and binding to promoter were higher in SNUC5/5-FUR cells than in SNUC5 cells. The activated Nrf2 in SNUC5/5-FUR cells led to an increase in the protein expression and activity of heme oxygenase-1 (HO-1), an Nrf2-regulated gene. SNUC5/5-FUR cells produced a larger amount of reactive oxygen species (ROS) than SNUC5 cells. The siRNA- or shRNA-mediated knockdown of Nrf2 or HO-1 significantly suppressed cancer cell viability and tumor growth in vitro and in vivo, resulting in enhanced 5-FU sensitivity. Methylation-specific (MS) or real-time quantitative MS-PCR data showed hypomethylation of the Nrf2 promoter CpG islands in SNUC5/5-FUR cells compared with SNUC5 cells. Expression of the DNA demethylase ten-eleven translocation (TET) was upregulated in SNUC5/5-FUR cells. ROS generated by 5-FU upregulated TET1 expression and function, whereas antioxidant had the opposite effect. These results suggested that the mechanism underlying the acquisition of 5-FU resistance in CRC involves the upregulation of Nrf2 and HO-1 expression via epigenetic modifications of DNA demethylation.
Asunto(s)
Neoplasias del Colon/genética , Metilación de ADN/genética , Proteínas de Unión al ADN/metabolismo , Resistencia a Antineoplásicos/genética , Epigénesis Genética/efectos de los fármacos , Fluorouracilo/farmacología , Factor 2 Relacionado con NF-E2/genética , Proteínas Proto-Oncogénicas/metabolismo , Animales , Biocatálisis/efectos de los fármacos , Línea Celular Tumoral , Neoplasias del Colon/patología , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Hemo-Oxigenasa 1/metabolismo , Humanos , Espacio Intracelular/metabolismo , Ratones Desnudos , Oxigenasas de Función Mixta , Factor 2 Relacionado con NF-E2/metabolismo , Regiones Promotoras Genéticas/genética , Especies Reactivas de Oxígeno/metabolismoRESUMEN
The purpose of this study was to assess the protective effects of an ethanol extract derived from the red alga Gracilaria bursa-pastoris (Gmelin) Silva (GBE) on ultraviolet B (UVB)-irradiated human HaCaT keratinocytes. GBE exhibited scavenging activity against intracellular reactive oxygen species that were induced by either hydrogen peroxide or UVB radiation. In addition, both the superoxide anion and the hydroxyl radical were scavenged by GBE in cell-free systems. GBE absorbed light in the UVB range (280-320 nm) of the electromagnetic spectrum and lessened the extent of UVB-induced oxidative damage to cellular lipids, proteins, and DNA. Finally, GBE-treated keratinocytes showed a reduction in UVB-induced apoptosis, as exemplified by fewer apoptotic bodies. These results suggest that GBE exerts cytoprotective actions against UVB-stimulated oxidative stress by scavenging ROS and absorbing UVB rays, thereby attenuating injury to cellular constituents and preventing cell death.