Reversal of multidrug resistance by MDR1 shRNA expression vector in human leukemia K562/A02 cells / 中华肿瘤杂志
Chinese Journal of Oncology
; (12): 422-425, 2006.
Artigo
em Chinês
| WPRIM (Pacífico Ocidental)
| ID: wpr-236926
Biblioteca responsável:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To construct a short hairpin RNA (shRNA) eukaryotic expression vector specific to MDR1 gene in multidrug resistance (MDR) human leukemia cell line K562/A02 to observe its silencing effect on MDR1 and P-glycprotein (P-gp) expression.</p><p><b>METHODS</b>The shRNA expression vector was constructed by gene recombination, then transfected into the cultured K562/A02 cells. The transcription of MDR1 gene was detected by semi-quantitative RT-PCR and the expression level of P-gp was determined by Western blot. 50% inhibition concentration (IC50) of ADM in K562/A02 cells was determined by MTT method. The intracellular doxorubicin (ADM) concentration was determined by HPLC.</p><p><b>RESULTS</b>The introduction of pEGFP-C1/U6/MDR1-A or pEGFP-C1/U6/MDR1-B expression vector was shown to efficiently and specifically inhibit the expression of P-gp according to results of Western blot, with an inhibitory rate of 50.67%. Semi-quantitative RT-PCR showed that mRNA transcription of MDR1 gene was reduced by (48.2 +/- 2.5)%. On the contrast, the control plasmid did not exhibit inhibitory effect on the protein expression and mRNA transcription of MDR1. The relative efficiency of K562/A02 to ADM was 40.8% or 62.4%, respectively, and the intracellular accumulation of ADM increased after shRNA treatment.</p><p><b>CONCLUSION</b>The shRNA expression vector targeting MDR1 gene showed dramatic inhibition on RNA transcription and protein expression. It could effectively restore the sensitivity of K562/A02 cells to conventional chemotherapeutic agents.</p>
Texto completo:
Disponível
Base de dados:
WPRIM (Pacífico Ocidental)
Assunto principal:
Farmacologia
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RNA Mensageiro
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Transfecção
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Doxorrubicina
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Sobrevivência Celular
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Western Blotting
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Membro 1 da Subfamília B de Cassetes de Ligação de ATP
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Resistência a Múltiplos Medicamentos
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Resistencia a Medicamentos Antineoplásicos
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Subfamília B de Transportador de Cassetes de Ligação de ATP
Limite:
Humanos
Idioma:
Chinês
Revista:
Chinese Journal of Oncology
Ano de publicação:
2006
Tipo de documento:
Artigo