斑蝥素固体脂质纳米粒的制备及促肝癌细胞凋亡的研究

Preparation of Cantharidin Solid Lipid Nanoparticles and Its Effect on Promoting Apoptosis of Hepatocellular Carcinoma Cells

目的制备斑蝥素固体脂质纳米粒,并对其进行质量评价及体内外抗肝癌研究。方法采用薄膜超声分散法制备斑蝥素固体脂质纳米粒,以粒径、PDI(Polydispersity Index)、Zeta电位和包封率等为指标,对制备工艺进行优化,通过透射电镜观察纳米粒的形态,并考察其体外释放度,以荧光实验考察肝癌细胞摄取固体脂质纳米粒,以CCK8法研究固体脂质纳米粒使斑蝥素增效减毒的作用、以流式细胞术研究斑蝥素固体脂质纳米粒促细胞凋亡及阻滞细胞周期的情况,以体内裸鼠实验验证斑蝥素固体脂质纳米粒的药效。结果斑蝥素固体脂质纳米粒呈粒径大小均一,形状规整的类球实体形,具有较好的稳定性和良好的药物缓释作用,细胞摄取实验结果显示与游离斑蝥素相比,固体脂质纳米粒更能有效递送药物到细胞。细胞毒性实验结果显示CTD-SLN(cantharidin solid lipid nanoparticles)和CTD(cantharidin)对HepG2细胞均具有显著浓度依赖性的毒作用,且CTD-SLN能够起到增效减毒的作用。流式细胞术的结果发现CTD-SLN能够促进细胞凋亡且使细胞停滞在S期。结论制备得到的斑蝥素固体脂质纳米粒能增强药物促肝肿瘤细胞凋亡效果,可用于进一步研究。

Objective To prepare cantharidin solid lipid nanoparticles(CTD-SLN)and to evaluate their quality and anti-liver cancer actions in vitro and in vivo.Methods Cantharidin solid lipid nanoparticles(CTD-SLN)were prepared by thin film ultrasonic dispersion method.The preparation process was optimized by particle size,polydispersity index(PDI),Zeta potential and encapsulation efficiency,and the morphology of the nanoparticles was observed by transmission electron microscopy.Moreover,their release in vitro was investigated,the uptake of solid lipid nanoparticles by hepatic cancer cells was observed with fluorescence test,CCK8 assay was used to study the efficacy-enhancing and toxicity-attenuating actions of solid lipid nanoparticles on cantharidin,the effect of CTD-SLN on cell apoptosis and cell cycle was investigated by flow cytometry,and the efficacy of CTD-SLN in nude mice was verified in vivo.Results CTD-SLN was in spheroid solid shape with the even particle size and regular shape,and they had good stability and good drug sustained release effect.The cellular uptake experiment showed that compared with free cantharidin,solid lipid nanoparticles were effective on the delivery of drugs to cells.The results of cytotoxicity experiments showed that both CTD-SLN and CTD had significant concentrationdependent toxic effects on HepG 2 cells,and CTD-SLN presented efficacy-enhancing and toxicity-attenuating actions.Results of flow cytometry showed that CTD-SLN promoted the cell apoptosis and arrested cells at S phase.Conclusion CTD-SLN can enhance the apoptosis of liver tumor cells promoted by the drugs and can be used for further study.