Studies On Size Dependent Localization And Penetration Of Gold Nanoparticles In Tumor-associated Macrophages

Tumor-associated macrophages are a critical component of tumor microenvironment and have complex and diverse effects on tumor growth,angiogenesis,immunosuppression,metastasis and chemical resistance.Especially polarization of macrophages showed completely opposite immune characteristics,significantly affected the clinical efficacy of cancer treatment.In addition,with the rapid development of kinase inhibitors and cytokines,tumor-associated macrophages have received increasing attention as a potential target with immunotherapeutic potential,targeting tumor-associated macrophages The study is also deepening.In the face of the above problems,this study selected inorganic nanoparticles with high physical and chemical properties,simple production and high controllability of inorganic nanoparticles.The results showed that gold nanoparticles could be used to investigate the effects of gold nanoparticles on tumor-related giant The particle size dependence of phage localization and osmotic capacity.So as to provide an important basis for screening nanoparticles with suitable particle size for targeting tumor-associated macrophages.Polyethylene glycol(PEG)is non-toxic,non-irritating,has good water solubility while both good biocompatibility,and because of its strong anti-protein adsorption capacity,is widely used reticular endothelium System stealth material.In this study,the gold nanoparticles were modified by PEG,and the PEGylation particle size-dependent localization and penetration of nanoparticles in tumor-associated macrophages were investigated.The linear range: 0.25 ~ 2.5?mol / L was used to adjust the HS-PEG-FITC fluorescence linear equation: Y = 191.95 X + 10.319 R2 = 0.9998,HS-PEG-NH2 and HS-PEG-FITC ratio,so that fluorescein isothiocyanate(FITC)can quantitatively labeled gold nanoparticles to meet the subsequent flow cytometry and confocal observation of intracellular localization of the experimental requirements.The macrophage cell line RAW264.7,the mouse hepatoma cell line Hepa1-6 and the human breast cancer cell MCF-7 were selected as the model cells.The co-culture model of tumor cells and macrophages was constructed by trans well,and the tumor microenvironment was simulated in vitro.The results showed that 5nm and 20 nm had stronger ability to penetrate gold nanoparticles than those with larger diameter,and the degree of macrophage internalization was significantly higher than that of macrophage cells Tumor cells.Under the co-culture condition,the uptake of macrophages did not change,but the amount of nanoparticles in tumor cells showed a slight decrease.In the migration experiment of macrophages in cell co-culture model,20 nm Au NPs showed stronger ability to induce macrophage aggregation than other nanoparticles.C57BL/6 male mice and BALB/c female mice were used to establish the in situ tumor model,and the tumor-associated macrophages in solid tumor tissue were extracted and purified by gradient density centrifugation and magnetic bead sorting.The macrophages were extracted from the healthy mice as a control,and TAMs were taken for cell uptake.The results showed that 20 nm Au NPs had the highest degree of intracellularization in TAMs,and the results were also confirmed by flow cytometry and laser confocal observation with Au-PEG-FITC.Besides,The results showed that 5nm Au NPs had better nucleus localization compared with other nanoparticles in larger particle diameter.The model of breast cancer was established and the in vivo uptake experiment was carried out.The Au NPs with 5nm and 20 nm particle size were selected as the object of study.The penetration of nanoparticles was investigated by in vivo local injection and in vitro flow detection.The results showed that the infiltration effect of 5 nm Au NPs was slightly better than that of 20 nm,but there was no significant difference.At the same time,TAMs reached the total uptake of tumor cells in less than 10% The amount of 50%,showing that the lower particle size of Au NPs on TAMs penetration efficiency is much higher than the tumor cells.In this study,gold nanoparticles with functional modification as a tool and the main experimental object,provides insights of particle size of 5 ~ 100 nm gold nanoparticles on TAMs in vitro and in vivo infiltration and positioning of particle size dependence.The results showed that low-diameter nanoparticles showed a unique advantage in the penetration of tumor-related cells in vivo and in vivo,and the degree of internalization was much higher than that of tumor cells.In contrast,5 nm Au NPs The localization effect is good,20 nm Au NPs on macrophages to recruit more ability.This study can be used as an important basis for targeting tumor-associated macrophage nanocarrier design and functional selection.