Analysis of the intracellular localization of amiodarone using live single-cell mass spectrometry
Analysis of the intracellular localization of amiodarone using live single-cell mass spectrometry
复制标题
使用活体单细胞质谱分析胺碘酮的细胞内定位
DOI：
10.1016/j.jpba.2021.114318
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发表时间：
2021
影响因子：
3.4
通讯作者：
Todoroki Kenichiro
中科院分区：
文献类型：
--
作者：
Yahata Kenji;Mizuno Hajime;Sugiyama Eiji;Todoroki Kenichiro
胺碘酮是一种众所周知的抗糖尿病药物，其副作用包括磷脂质沉积症。然而，尚不清楚胺碘酮及其代谢产物如何定位于细胞中。在本研究中，胺碘酮在单个HepG2人肝细胞癌细胞的胞液，空泡和脂滴中的定位直接使用活单细胞质谱法测定。在使用荧光探针可视化脂滴后，在荧光显微镜下使用纳米喷雾头分别捕获用胺碘酮处理的单个HepG2细胞的胞质溶胶、空泡和脂滴。该测定法在测定胺碘酮水平时呈线性，细胞裂解物和血清中的R2值分别为0.9996和0.9998。胺碘酮及其代谢产物在脂滴和空泡中的峰强度显著高于胞浆中的峰强度，脂滴中的峰强度高于空泡中的峰强度。在脂滴和细胞溶质中均检测到胺碘酮代谢产物。活单细胞质谱结合荧光成像显示胺碘酮及其代谢产物在脂滴中的清晰定位与空泡分开。该分析系统与荧光成像相结合，可用于研究各种药物及其代谢物的细胞内定位。
Amiodarone is a well-known antiarrhythmic drug with side effects including phospholipidosis. However, it is not clear how amiodarone and its metabolites are localized in the cell. In the present study, the localization of amiodarone in the cytosol, vacuoles, and lipid droplets of a single HepG2 human hepatocellular carcinoma cell was determined directly using live single-cell mass spectrometry. The cytosol, vacuoles, and lipid droplets of a single HepG2 cell treated with amiodarone were separately captured using a nano-spray tip under a fluorescence microscope after visualizing the lipid droplets using a fluorescent probe. This assay showed a linearity in the measurement of amiodarone levels withR2values of 0.9996 and 0.9998 in the cell lysates and serum, respectively. The peak intensities of amiodarone and its metabolites in lipid droplets and vacuoles were significantly higher than those in the cytosol, while those in lipid droplets were higher than those in vacuoles. Amiodarone metabolites were detected in both lipid droplets and the cytosol. Live single-cell mass spectrometry combined with fluorescence imaging demonstrated clear localization of amiodarone and its metabolites in lipid droplets separately from the vacuole. This assay system combined with fluorescence imaging could be useful for investigating the intracellular localization of various drugs and their metabolites.