| Object:The research indicated the p53 gene has the high rate of mutation in the major tumor, this kind of mutuation change and the tumor cell multi-medicines drug resistant to have the close relation. This research plans to discuss that the wtp53 gene was transfected into cell line HepG2/5-Fu of the human liver cancer using magnetic nano particles t to analyses the apoptosis and drug resisitant induced by 5-Fu.Methods:(1) Analyze multi-medicine drug resistance the of human liver cancer cell line HepG2/5-Fu in order to examines the HepG2/5-Fu cell line to the different anti-tumor medicine sensitivity.(2) Analyze the to apoptosiss and the 5-Fu tolerance of HepG2/5-Fu cell induced by 5-Fu through FAS.(3) Analyzes apoptosiss correlation genes expression on the HepG2/5-Fu cell line such as p53, bcl-2, bcl-xl, bax et al.by RT-PCR.(4) Analyzes the p53 gene mutaion in the HepG2/5-Fu cell line through the SSCP method.(5) Prepares IONP-PLL nano particles to construct the oxidation ferromagnetism nanometer transportation system, and analyzes its biology effect.(6) The wtp53 gene was transfected into the HepG2/5-Fu by IONP-PLL nano particles as the gene transportation carrier, observes the lethal effect that the cell line HepG2/5-Fu was treated with 5-Fu.Result:(1) The HepG2/5-Fu cell has the varying degree drug resistance to 5-Fu and other several kind of commonly used chemotherapy medicine compared with HepG2 , among of total, the drug resistance to 5-Fu reach 107.0 times, to Paclitaxel reach 49.0 times compared with the drug resistanct of HepG2.(2) When treated the HepG2 with the concent ration of 5-Fu (0μmo/L , 30μmo/L or 150μmol/L), the AI was (6.5±1.0) %,(14.0±4.0) % and (20.0±5.0) %, inspectively.The apoptosis rateincreased 1.2 time and 2.1 time compared with not treated with 5-Fu, inspectively, which wrer significantly ( P< 0.05) .But treated the HepG2/5-Fu with the same concent ration of 5-Fu (30μmo/L or 150μmol/L), the AI was (6.7±0.7) %,(7.1±2.2)% and (6.5±2.0)(3) When treated with the same concent ration of 5-Fu (30μmo/L or 150μmol/L) , the proportion of apoptosis was significantly increased and G0/G1 phase was increased and S and G2/M phases were reduced in HepG2 cells , but the proportion of apoptosis and cell cycle was not changed in HepG2/5-Fu cells. (4) The expression of bcl-xl , bcl-xs , and bax mRNA were significantly increased and the expression of p53 and cpp32 mRNA were significantly decreased in resistant HepG2/5-Fu cells , as compared with HepG2 cells by RT-PCR.(5) IONP was prepared by alkaline precipitation of divalent and trivalent iron chloride in the presence of high concentration of dextran. In this system, iron oxide nanoparticles was equably dispersed in surfactant in suspend state.The diameter of IONP-PLLIDNA was 47nm±15nm. Negative staining showed DNA bourid on the surface of IONP-PLL. The DNA binding assay showed that at acid pH, IONP can bind DNA, but at pH 7 and 9, IONP has no the potential to bind DNA. The binding is achieved by interaction between the negatively charged DNA phosphate groups and positively charged gene vectors. When titrated to pH 3, the IONP showed an increase binding of plasmid DNA by this type of particle. The electronstatistic interaction can also protect DNA against the digestion of nuclease at acid pH.(6) Whether IONP-PLL and IONP-PLL/DNA complexes influenced cell viability was investigated in several cell lines by MTT assays. IONP-PLL and IONP-PLL/DNA complexes showed no obvious negative effect on cell viability.(7) The cell line HepG2/5-Fu tansfected wtp53 displayed the dosage dependence effect and the time dependence effect to 5-Fu. The cell line HepG2/5-Fu tansfected wtp53 appeared bviously apoptosis induced by 5-Fu. Conclusion:(1) wtp53 gene mutation was relative with cell cycle change and the drug resistance of liver cancer, which is one of multi-medicine drug resistance mechanisms of the liver cancer cell HepG2/5-Fu.(2)Poly-L-Lysine modified Iron Oxide Nanoparticles (IONP-PLL) system was optimized to enhance the gene delivery efficiency. IONP-PLL/DNA complexes showed no obvious negative effect on cell viability and safty.(3) Take the wtp53 gene as the target to treat later period liver cancer shall be hopefully one kind of new treatment strategy.
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