RNA interference-mediated silencing of cytochrome P450 3A4 gene in CHL-3A4 cells / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of small hairpin interfering RNA (shRNA) in suppressing cytochrome P450 3A4 (CYP3A4) gene expression in CHL-3A4 cells.</p><p><b>METHODS</b>Three shRNA expression vectors targeting CYP3A4 gene (CYP3A4 I, C YP3A4 II, and CYP3A4 III, respectively) were designed, synthesized and transfected into CHL-3A4 cells via liposomes. The inhibitory effect of shRNA on CYP 3A4 gene expression was detected by Western blotting and RT-PCR, and the effect of shRNA transfection in suppressing cyclophosphamide-induced cytotoxicity was measured using MTT assay.</p><p><b>RESULTS</b>The vector carrying CYP3A4 III shRNA significantly reduced the expression of CYP3A4 gene at both the mRNA (75%) and protein levels (80%) in CHL3A4 cells. The cytotoxicity of cyclophosphamide was markedly inhibited by CYP3A4 III-mediated suppression of CYP3A4 gene expression by 75% in CHL-3A4 cells.</p><p><b>CONCLUSION</b>The vector-mediated RNA interference can suppress CYP3A4 gene expression in CHL-3A4 cells, and RNA interference technique provides a new means for studying cytochrome P450 gene function in mammalian cells.</p>