ROS响应性不饱和脂肪酸脂质体促进肿瘤细胞铁死亡的研究

Study of reactive oxygen species responsive unsaturated fatty acid liposomes promoting ferroptosis in tumor cells

目的制备一种花生四烯酸脂质体(liposomes containing arachidonic acid,Lipo-DAPE),研究该脂质体的活性氧(reactive oxygen species,ROS)响应性和毒性情况,考察其促进肿瘤细胞发生铁死亡的能力。方法以粒径、Zeta电位为评价指标优化Lipo-DAPE的处方;通过透射电子显微镜和动态光散射表征脂质体的形貌、稳定性等理化性质,考察其ROS响应性及药物释放速率;通过激光扫描共聚焦显微镜观察4T1细胞对脂质体的摄取行为;通过MTT实验考察Lipo-DAPE的抗肿瘤活性;以细胞内ROS、脂质过氧化物及谷胱甘肽水平考察Lipo-DAPE促进肿瘤细胞铁死亡的能力。结果Lipo-DAPE粒径为(74.4±1.0)nm,Zeta电位为(29.4±1.2)mV,可在48 h内维持稳定;在模拟肿瘤微环境ROS条件下脂质体骨架崩解,促进药物释放;Lipo-DAPE能够被4T1细胞摄取并抑制细胞活力;此外,Lipo-DAPE促进了细胞中ROS及脂质过氧化物含量,降低了GSH水平。结论Lipo-DAPE具有促进肿瘤细胞发生铁死亡的效果,为新型全活性纳米药物递送载体的设计提供了新思路与理论依据。

Objective To prepare an arachidonic acid modified liposome(Lipo-DAPE)and to investigate the reactive oxygen species(ROS)responsiveness and cytotoxicity of Lipo-DAPE,as well as its ability to promote ferroptosis in tumor cells.Methods The formulation of Lipo-DAPE was optimized using hydrodynamic size and Zeta potential as evaluation indexes.The morphology,stability and other physicochemical properties along with the ROS responsiveness and drug release rate of liposomes were thoroughly characterized and investigated by transmission electron microscope and dynamic light scattering techniques.The cellular uptake of liposomes by 4T1 cells was observed with the aid of confocal laser scanning microscope.The cytotoxicity of Lipo-DAPE was evaluated by MTT assay.The variation of intracellular ROS,lipid peroxide and glutathione(GSH)level in tumor cells were detected to reflect the effect of Lipo-DAPE on promoting ferroptosis.Results The hydrodynamic diameter of Lipo-DAPE was(74.4±1.0)nm and the Zeta potential was(29.4±1.2)mV.This kind of liposome could maintain stability within 48 h and enable rapid cargo release upon ROS triggering in tumor microenvironment.Lipo-DAPE could be successfully internalized by 4T1 tumor cells and showed cytotoxicity due to lipid peroxidation.The increased amounts of ROS and lipid peroxides,and the decreased level of GSH after Lipo-DAPE treatment were significant.Conclusion This study provides a facile approach to enhance the efficacy of ferroptosis by using unsaturated fatty acid liposomes.It also provides a novel strategy and theoretical basis for the design of all-active drug delivery systems.