乙酸胁迫酵母细胞凋亡过程中单个线粒体损伤的拉曼光谱分析

Raman Spectra Analysis for Single Mitochondria after Apoptosis Process of Yeast Cells Stressed by Acetic Acid

应用激光光镊拉曼光谱技术(LTRS)作为一种非入侵分析线粒体的工具,结合氧电极法、紫外分光光度计法,研究乙酸胁迫酵母细胞后提取的线粒体,以及直接胁迫正常酵母细胞的离体线粒体的拉曼光谱。结果显示,乙酸胁迫酵母细胞后提取的线粒体的拉曼光谱,归属于核酸的谱峰(1081和1301 cm!1)、蛋白质的谱峰(872,1445,1604和1657 cm!1)、脂类的谱峰(1125,1301,1445和1657 cm!1)、Cyt c的特征峰(750和1125 cm!1)、线粒体呼吸特征峰(1604 cm!1),都随着诱导程度的加深而降低,与常规法测定的指标呼吸率,磷氧比,细胞色素C含量的降低,MDA含量的升高的变化趋势相似;乙酸直接胁迫线粒体的拉曼光谱,归属于核酸的谱峰(1081和1301 cm!1),蛋白质的谱峰(872,1445,1604和1657 cm!1),脂类的谱峰(1125,1301,1445和1657 cm!1),Cyt c的特征峰(750和1125 cm!1),线粒体呼吸特征峰(1604 cm!1),都随着诱导程度的加深而降低,与常规法测定的指标呼吸率、磷氧比、细胞色素C含量的降低,MDA含量的升高的变化趋势相似。结果表明,乙酸胁迫细胞过程中,乙酸进到细胞内可能直接作用于线粒体,引起线粒体内物质的释放,导致依赖于线粒体途径的细胞凋亡。

Laser tweezers Raman spectroscopy （LTRS） as a non-invasive tool for mitochondria analysis, combined with oxygen electrode and ultraviolet spectrophotometric method, was used to investigate the Raman spectra of the mitochondria of yeast cells in vitro and in vivo after induced with acetic acid. The results showed that when the mitochondfia of yeast cells in vivo was induced by acetic acid, the spectral peaks of the nucleic acid （1081, 1301 cm-1）, proteins （872, 1604, 1445, 1657 cm-l）, lipids （1125, 1301, 1445, 1657 cm-1）, cytochrome c （750, 1125 cm-1 ） and mitochondria respiration （ 1604 cm-1 ） were significantly decreased as a function of the duration of the acetic acid stress, and the obtained physiological and biochemical indexes of respiration rate, p/o ration and cytochrome c content were similar to that of by the conventional method. Besides, when the mitochondria of yeast cells in vitro was induced by acetic acid, the spectral peaks of the nucleic acid （ 1081, 1301 cm-1 ）, proteins （ 872, 1604, 1445, 1657 cm-1 ）, lipids （1125, 1301, 1445, 1657 cm-1 ）, cytochrome c （750, 1125 cm-1 ） and mitochondria respiration （1604 cm-1 ） were significantly decreased as the function of the duration of the acetic acid stress, and the obtained physiological and biochemical indexes of respiration rate, p/o ration and cytochrome c content were similar to that of by conventional method. The results indicated that acetic acid could penetrate into the cell interior and directly impacted the mitochondria possibly, resulted in the release of inclusions from the mitochondria, subsequently, caused the apoptosis of yeast cells which relied on the mitochondria pathways.