Fe-MOF纳米探针在乳腺癌光动力增敏铁死亡及MR激活成像中实验研究

Experimental Study of Fe-MOF Nanoprobe in Photodynamically Sensitized Ferroptosis and MR Activation Imaging of Breast Cancer

目的:探讨肿瘤酸性微环境响应铁基金属有机框架(Iron-containing Metal Organic Frameworks,Fe-MOF)纳米探针对乳腺癌光动力增敏铁死亡及其体外MR T1激活效应。方法:制备Fe-MOF纳米探针,用透射电子显微镜(Transmission Electron Microscope,TEM)及原子力显微镜(Atomic Force Microscope,AFM)对其形态进行表征;采用3,3’,5,5’-四甲基联苯胺(TMB)及5,5’-二硫双(2-硝基苯甲酸)(DTNB)评价其在溶液水平ROS生成及GSH消耗能力;通过细胞毒性实验(MTT)测定纳米探针在暗毒性及光毒性条件下对4T1乳腺癌细胞的光动力增敏铁死亡的效能;将4T1细胞与Fe-MOF共孵育后,协同激光照射处理,使用荧光显微镜观察其活性氧(Reactive Oxygen Specis,ROS)、脂质过氧化物(Lipid PerOxide,LPO)的生成以及细胞活/死染色情况;观察纳米探针在不同的pH条件下T1加权成像的激活效应,并在细胞水平测量不同时间点的T1激活效能。结果:制备的Fe-MOF纳米探针呈针状结构,厚度约为44 nm;在体外溶液水平可有效促进ROS的生成及LPO的消耗;荧光显微镜结果显示Fe-MOF触发的铁死亡效应联合光动力治疗可有效促进细胞内ROS和LPO的生成,以及肿瘤细胞死亡率升高(P<0.001);MR成像结果显示,在酸性条件下纳米探针的T1信号可被特异性激活,pH 5.0条件下r1弛豫率为4.954 m M^(-1)s^(-1),具有较好pH响应性及细胞水平时间依赖性激活效能。结论:pH响应诊疗一体化的Fe-MOF纳米探针可实现乳腺癌的光动力增敏铁死亡效能以及MR微环境响应激活成像。

Objective:To synthesize tumor acidic microenvironment-responsive Fe-MOF nanoprobe and investigate its synergistic therapeutic effect of ferroptosis-photodynamic therapy for breast cancer and in vitro MR T1 activation effect.Methods:Fe-MOF nanoprobes were prepared and characterized by transmission electron microscope(TEM)and atomic force microscopy(AFM);Assess its reactive oxygen specis(ROS)generation capability and GSH depletion ability at the solution level using 3,3',5,5'-Tetramethylbenzidine(TMB)and 5,5'-Dithiobis(2-nitrobenzoic acid)(DTNB);The MTT assay was used to determine the cytotoxicity of the nanoprobe on 4T1 breast cancer cells under dark and light conditions,so as to evaluate synergistic therapeutic effect of ferroptosis-photodynamic therapy;After the 4T1 cells were co-incubated with Fe-MOF and treated with laser irradiation,the generation of ROS,lipid peroxide(LPO)and cell live/dead staining were observed using the fluorescence microscope;Observe the activation effect of Fe-MOF in T1-weighted imaging under different pH conditions,and measure the T1 activation efficiency at different time points at the cellular level.Results:The prepared Fe-MOF nanoprobes exhibit a needle-like structure,with a thickness of approximately 44 nm;Fe-MOF can effectively promote the generation of ROS and the consumption of LPO at the solution level;Fluorescence microscope results show that ferroptosis effect triggered by Fe-MOF combined with photodynamic therapy can effectively promote the generation of intracellular ROS and LPO and promote an increase in the death rate of tumor cells(P<0.001);MR imaging results show that the T1 signal of Fe-MOF can be specifically activated under acidic conditions and the r1 relaxation rate under pH 5.0 is 4.954 mM^(-1)s^(-1).It has good pH responsiveness at the solution level and time-dependent activation efficiency at the cellular level.Conclusion:The pH-responsive diagnostic and therapeutic integrated nanoprobe,Fe-MOF,is capable of achieving synergistic therapeutic efficacy through ferroptosis-photodynamic treatment and magnetic resonance T1 contrast effect.