RESUMEN

AIM: To study recombinant plasmid pEGFP-AFP-TK delivered by nano-magnetic fluids targeting killed AFP positive HepG2 cells in vitro. METHODS: Constructed recombinant plasmid pEGFP-AFP-TK which was delivered by nano-magnetic fluids into AFP positive HepG2 cells and AFP negative SMMC-7721.The fluorescence was detected in order to evaluate the transfection rate, and RT-PCR and Western blot were used to detect the expression of TK gene after transfection. MTT assay was used to evaluate the effect of TK on the proliferation and bystander effect of HepG2 cells in ganciclovir (GCV). Flow cytometry was used to analysis the apoptosis of HepG2 cells. RESULTS: Nano-magnetic fluids delivered plasmid pEGFP-AFP-TK into HepG2 cells and TK gene was successfully expressed, and the transfection efficacy of nano-magnetic fluids was superior to that of Lipofectamine. RT-PCR and Western blot results demonstrated that TK gene was expressed in HepG2 cells after being transfected with nano-magnetic fluids/pEGFP-AFP-TK. MTT and flow cytometry results showed TK gene exerts cell-killing and bystander effect. CONCLUSION: Nano-magnetic fluids could successfully deliver pEGFP-AFP-TK into HepG2 cells and induce expression of TK gene, which is promising gene vector for liver cancer gene therapy. AFP enhancer can specifically enhance the expression of target TK gene within the AFP positive cell, and induce bystander effect and apoptosis in the AFP positive HepG2 cell with with GCV.

Asunto(s)

Ganciclovir/farmacología , Proteínas Recombinantes de Fusión/metabolismo , Timidina Quinasa/metabolismo , alfa-Fetoproteínas/metabolismo , Antivirales/farmacología , Apoptosis/efectos de los fármacos , Western Blotting , Efecto Espectador , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/terapia , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Elementos de Facilitación Genéticos/genética , Citometría de Flujo , Terapia Genética/métodos , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/terapia , Magnetismo , Microscopía Fluorescente , Nanotecnología/instrumentación , Nanotecnología/métodos , Plásmidos/genética , Plásmidos/metabolismo , Proteínas Recombinantes de Fusión/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Timidina Quinasa/genética , Transfección/instrumentación , Transfección/métodos , alfa-Fetoproteínas/genética