Abstract:
This study demonstrates the capability of Raman microscopy for detecting structural differences in Giardia cells exposed to different drugs and incubation times. While metronidazole (MTZ) visibly affects the cells by inducing extracellular vesicle releases of toxic iron intermediates and modified triple-bond moieties, oseltamivir (OSM) alters the phenylalanine and lipid structures. Modifications in the heme protein environment and the transformation of iron from ferric to ferrous observed for both drug treatments are more notable for MTZ. Different contents and amounts of vesicle excretion are detected for 24 h or 48 h with MTZ incubation. At a shorter drug exposure, releases of altered proteins, glycogen, and phospholipids dominate. Agglomerates of transformed iron complexes from heme proteins and multiple-bond moieties prevail at 48 h of treatment. No such vesicle releases are present in the case of OSM usage. Drug incorporations into the cells and their impact on the plasma membrane and the dynamics of lipid raft confirmed by confocal fluorescence microscopy reveal a more destructive extent by OSM, corroborating the Raman results. Raman microscopy provides a broader understanding of the multifaceted factors and mechanisms responsible for giardiasis treatment or drug resistance by enabling a label-free, simultaneous monitoring of structural changes at the cellular and molecular levels.
摘要:
这项研究证明了拉曼显微镜检测暴露于不同药物和孵育时间的贾第虫细胞结构差异的能力。虽然甲硝唑(MTZ)通过诱导细胞外囊泡释放有毒的铁中间体和修饰的三键部分明显影响细胞,奥司他韦(OSM)改变苯丙氨酸和脂质结构。在两种药物治疗中观察到的血红素蛋白环境的改变和铁从三价铁到亚铁的转化对于MTZ更为显著。用MTZ孵育24小时或48小时检测不同的囊泡排泄物含量和数量。在较短的药物暴露时间内,释放改变的蛋白质,糖原,磷脂占主导地位。来自血红素蛋白和多键部分的转化铁络合物的团聚体在处理48小时时占优势。在使用OSM的情况下不存在这样的囊泡释放。通过共聚焦荧光显微镜证实的药物掺入细胞及其对质膜和脂筏动力学的影响揭示了OSM更具破坏性的程度,证实了拉曼的结果。拉曼显微镜通过实现无标记,提供了对负责贾第鞭毛虫病治疗或耐药性的多方面因素和机制的更广泛的理解,同时监测细胞和分子水平的结构变化。