Study on Using Magnetic Iron Oxide Nanoparticles as HIF-1α shRNA Gene Carrier to Reverse Cisplatin Resistance of A549/CDDP Cell Lines*

Abstract

To evaluate the feasibility of using magnetic iron oxide nanoparticles (pll-DCIONP) as HIF-1α shRNA gene carrier for transfection in vitro and in vivo,and the effect of HIF-1α targeted RNA interference for reversing cisplatin resistance in human lung adenocarcinoma A549/CDDP.The HIF-1α shRNA was constructed and transfected into A549/CDDP and its xenograft animal model by pll-DCIONP and lipo2000,respectively.Fluorescent microscopy was employed to compare the transfection efficiency in vitro.The expression levels of HIF-1α,MRP1 and LRP after transfection were detected by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry analysis.Immunohistochemical analysis was performed to compare the levels of HIF-1α,MRP1 and LRP in transplanted tumors among different groups.In orderto calculate the cisplatin resistant,MTT assay was performed to detect the cell half-maximum inhibitory concentration (IC50).The growth index of transplanted tumors after transfection were detected among groups.Additionally,HE staining of liver,kidney and brain tissues was used aftermagnetic iron oxide nanoparticles transfection.The method of pll-DCIONP was more efficient on transferring plasmid into cells than the lipids examined in vitro (P0.01).The mRNA and protein levels of HIF-1α,MRP1 and LRP of A549/CDDP were decreased after transfection with HIF-1α shRNA,and the resistance to cisplatin of A549/CDDP was reversed by 82%.The protein levels of HIF-1α,MRP1 and LRP in A549/CDDP transplanted tumors were decreased after transfection with HIF-1α shRNA;also,the growth of A549/CDDP transplanted tumors were inhibited by HIF-1α shRNA,cooperating with the synergistic effect of cisplatin.No necrosis of liver,kidney and brain tissue were observed after magnetic iron oxide nanoparticles transfection.The pll-DCIONP could be used as one of the ideal gene carriers for HIF-1α shRNA gene delivery in vitro and in vivo.HIF-1α can be an effective target forreversing cisplatin resistance in lung cancer,the mechanism underlying may be related to the decreased expression levels of HIF-1α,MRP1 and LRP after transfection with HIF-1α shRNA,and magnetic nanoparticle-mediated HIF-1α shRNA transfection has biological safety to some extent.