RESUMEN
To study the effect of oleate on ATP binding cassette transporter A1 (ABCA1) expression and cholesterol efflux in THP-1 macrophage-derived foam cells, after exposure of the cultured THP-1 macrophage-derived foam cells to oleate for different time, cholesterol efflux was determined by FJ-2107P type liquid scintillator. ABCA1 mRNA and its protein level were determined by RT-PCR and Western blot, respectively. The mean ABCA1 fluorescence intensity of THP-1 macrophage-derived foam cells was detected by flow cytometry. The results showed that oleate markedly inhibited ABCA1-mediated cholesterol efflux from THP-1 macrophage-derived foam cells. This was accompanied by a reduction in the membrane content of ABCA1. Oleate did not alter ABCA1 mRNA abundance, indicating that decreased ABCA1 transcription, enhanced mRNA decay, or impaired translation efficiency did not account for these inhibitory effects. Oleate, however, increased ABCA1 turnover when protein synthesis was blocked by cycloheximide. Oleate reduces cholesterol efflux and the level of ABCA1 protein in THP-1 macrophage-derived foam cells.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Colesterol/metabolismo , Células Espumosas/efectos de los fármacos , Ácido Oléico/farmacología , Transportador 1 de Casete de Unión a ATP , Transportadoras de Casetes de Unión a ATP/metabolismo , Línea Celular , Cromatografía Líquida de Alta Presión , Citometría de Flujo , Células Espumosas/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Lipoproteínas LDL/farmacología , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de TiempoRESUMEN
OBJECTIVE: To investigate the relationship between aldose reductase like protein (ARL-1) gene overexpressed in HCC cells and drug-resistance of the cell to drugs containing carbonyl group. METHODS: To establish ARL-1 stable expression positive cell line, eukaryotic expression vectors containing ARL-1 gene cDNA were transfected into Hep cell mediated by lipofect AMINE. The positive monoclones were determined by PCR and RT-PCR, respectively. Then MTT assay was used to study the drug resistance ability of the cells to drugs containing carbonyl after incubating three days with those drugs. RESULTS: After ARL-1 gene transfection mediated by lipofect AMINE, one positive monoclonal cell overexpressing ARL-1 gene was selected. Compared with the control cell group, drug resistance ability of the positive cells to ADM and MMC which contain carbonyl group increased 2.3 and 3.17 fold, respectively (t=6.39, P=0.016 in ADM group and t=30.06, P=0.001 in MMC group). In the same time, drug resistance ability to 5-FU which has no carbonyl group had no statistical difference between positive monoclonal cell group and control cell group (t=0.684, P=0.531). CONCLUSIONS: The Hep ARL-1 positive cell line with stable expression of ARL-1 gene has been established successfully and the up-regulation of ARL-1 gene may plays an important role in drug resistance of the cells to anticancer drugs containing carbonyl group.