| Objective: In recent years,various nanocarriers are receiving attention for medical purposes,and are increasingly used in anti-tumor therapy.However,nano drug delivery system is facing great challenges in maintaining the stability of the nano drug delivery system,reducing drug leakage during drug delivery,and controlling drug release with effectively respond to the stimulation of the tumor microenvironment.The purpose of this study is to provide a new theoretical framework and research method for the material selection,structure design and performance regulation of nano drug delivery system in tumor chemotherapy by constructing a model of MTX-SS-PGA NPs in response to reducing stimulation in tumor cells.Methods: In this study,methotrexate(MTX),a hydrophobic anticancer drug,was conjugated with polyglutamic acid(PGA)based on the amide bond through cystine dihydrochloride(disulfide bond),and a glutathione-responsive targeting micellar system for osteosarcoma was designed.The response of MTX-SS-PGA NPs to glutathione and the release rate of methotrexate were determined by drug release assay in vitro;the effects of different concentrations of MTX-SS-PGA NPs and MTX on the proliferation and viability of 143 B cells was detected by MTT assay;the uptake of MTX-SS-PGA NPs in tumor and the localization of MTX-SS-PGA NPs were determined by subcellular localization assay;the apoptotic morphology of 143 B cells was observed by AO-EB staining.Finally,the anti-tumor effect of MTX-SS-PGA NPs on mice bearing 143 B tumor was determined by in vivo anti-tumor experiment.The damage degree of MTX-SS-PGA NPs and methotrexate on the tumors was determined by H&E and TUNEL staining analysis.Results: The nanoparticles were characterized by laser particle size analyzer(DLS)and transmission electron microscope(TEM): the size of nanoparticles was 100 nm and the surface was negatively charged.Through in vitro drug release experiments,the responsiveness of MTX-SS-PGA NPs to glutathione was proved;the drug concentration of the nanoparticles was determined by UV spectrophotometer;the results of subcellular localization assay showed that MTX-SS-PGA NPs can be effectively internalized by 143 B cells,and the high glutathione concentration in 143 B cells would disintegrate the structure of MTX-SS-PGA NPs and release MTX;the MTT test showed that MTX-SS-PGA NPs can effectively inhibit the proliferation of 143 B cells,and had little effect on HUVEC cell proliferation.In vivo administration to tumor-bearing mice,through in vivo fluorescence imaging experiments and tumor size changes further proved that: MTX-SS-PGA NPs have tumor targeting to tumor-bearing mice and have certain inhibition on tumor growth effect.Conclusion: In this study,a new type of MTX-SS-PGA NPs with uniform size,regular morphology,and glutathione responsiveness was designed.It exhibits high-efficiency uptake and anti-tumor activity in osteosarcoma cells.In tumor-bearing mice,MTX-SS-PGA NPs have specific tumor targeting properties and can inhibit the growth of tumor tissues.A new type of methotrexate drug delivery system designed in this experiment shows that it has a good anti-osteosarcoma effect and provides a new idea and new strategy for the treatment of osteosarcoma. |
