| objective: Preparation and characterization of a novel nanosystem FA-QDS-FUA with safe,sustained release and good anti-tumor effect.Methods: 1.Preparation of FA-QDs-FUA nanosystem: FA-QDs-FUA nanosystem was prepared by amide condensation reaction.FA-QDs-FUA nanocarrier system was finally obtained through amide condensation of the amino groups on the surface of QDs with the carboxyl groups of folate and 5-fluorouracil by the double cross-linker method.2.Characterization of FA-QDs-FUA: The optical properties of as-prepared FA-QDs-FUA were characterized by UV-Vis spectra and fluorescence spectra.The conjugation of FUA to the FA-PEG-QDs matrix was confirmed using FT-IR spectrometer.The surface morphology of the FA-QDs-FUA was observed using scanning electron microscopy(TEM).Particle size and zeta potential of FA-QDs-FUA was determined using Nano ZS90 Zeta sizer.The concentration of drug from FA-QDs-FUA and the in vitro drug release was determined in triplicate by UV.Hemolytic test was used to investigate the biocompatibility of FA-QDs-FUA.3.In vitro cell viability and proliferation assay: In vitro cytotoxicity of the PEG-QDs and FA-PEG-QDs nanoparticles in the normal hepatocyte line L02 was examined by MTT assay.The antiproliferative activity of conjugates FA-QDs-FUA was also tested in three different cell lines(L02,Hep G2 and SMMC-7721)in vitro and compared with the activity of free 5-FU.The ability of FA-QDs-FUA to inhibit the invasion and metastasis of tumor cell was investigated by wound-healing assay and transwell assay.4.In vitro targeting specificity: The normal human hepatocytes L02,human hepatocellular carcinoma cells Hep G2 and SMMC-7721 were selected as experimental models in vitro and were imaged under an inverted fluorescence microscope.5.In vivo antineoplastic efficacy study: The tumor-bearing liver cancer in nude mice model was establish,and it was randomized intraperitoneal injection,record the changes of survival rate,body weight,tumor volume.After 21 days,the tumor tissue was taken for tissue section.After 7 days of administration,blood samples were obtained from the SD rat model for evaluation of the major organ toxicities and side effects by automatic analyzer 7600 series.Results: The optimum conditions for the synthesis of FA-QDs-FUA nanosystem were as follows: the best ratio of FA-QDs-FUA was FA: PEG-QDs: FUA = 10: 1: 100,the optimal temperature was 37 ℃ and the reaction time was 2 h.The safe concentration of PEG-QDs was 40 n M.Under the optimized conditions,the drug loading capacity of the FA-QDs-FUA was 36.85%(n = 3).The average particle size was 220.28 nm and the zeta potential was-13.3 m V.The TEM showed that the particle size distribution of the system was uniform.And the fluorescence spectra of FA-QDs-FUA and FA-QDs-FUA showed that the FA-QDs-FUA was successfully synthesized without changing the fluorescence characteristics of QDs-FUA.The percent hemolysis ratio of FA-QDs-FUA nanosystem was below 5%,which showed very little hemolysis and were considered safe for intravenous administration.The results of MTT assay showed that FA-QDs-FUA had a good inhibition effect on tumor cells.Wound-healing assay and transwell showed that FA-QDs-FUA could effectively inhibit the invasion and migration of tumor.The results of animal experiments showed that FA-QDs-FUA can inhibit tumor proliferation and metastasis in vivo.Conclusion: FA-PEG-QDs not only exhibited a satisfactory loading capacity but also increased drug uptake toward FA-positive tumor cells via a specific FA receptor-mediated endocytosis mechanism.The successful synthesis of FA-QDs-FUA was characterized by FTIR,SEM,and zeta potential measurement.The results of this study provide a promising nanoplatform for the targeting of hepatocellular carcinoma. |
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