Reversal of multidrug resistance by magnetic chitosan-Fe₃O₄ nanoparticle-encapsulated MDR1 siRNA in glioblastoma cell line

Abstract

Objective: To investigate the reversal effects of MDR1 gene on multidrug resistance in the glioblastoma cell line BT325 by magnetic chitosan-Fe₃O₄ nanoparticle-encapsulated MDR1 siRNA.

Methods: The shRNA expression vector was constructed and the recombinant plasmids were cloned. Magnetic chitosan-Fe₃O₄ nanoparticles were prepared and the encapsulation rate was determined. After transfection, the BT325 cells were cultured to assay the transfection efficiency. The changing of MDR1 mRNA level and P-gp protein was evaluated. And the sensitivity to different chemotherapeutic drugs was assessed in BT325-siRNA transfected cell and untransfected cell by IC₅₀ values.

Results: The MDR1 RNAi plasmid was successfully designed and preparation. The encapsulation efficiency of the magnetic chitosan-Fe₃O₄ nanoparticle was 98–99%. The transfection efficiency of the siRNA-nanoparticles in BT325 cells was 70–80%. And the MDR1 mRNA levels were downregulated by reverse transcription (RT)-PCR assay. Furthermore, the results of P-gp protein expression decreased on immunocytochemical assay, Western blot and flow cytometry compared with control group. The IC₅₀ values of DOX and VCR were decreased between the transfected cell and normal BT325 cell.

Conclusion: After targeted transfection of the glioblastoma cell line with magnetic chitosan-Fe₃O₄ nanoparticle-encapsulated MDR1 siRNA, the expression of MDR1 at both the mRNA and protein level decreased, which increased sensitivity to chemotherapy in vitro. It might provide a basis for investigation of the mechanism involved in multidrug resistance in glioma.

Keywords:

• Gene therapy
• Glioma
• Nanoparticle
• RNA interference

This study was supported by the National Natural Science Fund of China (No. 30900342) and Beijing Nova program (2010B030).