Characterization of cells with different mitochondrial membrane potential during apoptosis
Characterization of cells with different mitochondrial membrane potential during apoptosis
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DOI：
10.1002/cyto.a.20188
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发表时间：
2005-11-01
期刊：
影响因子：
3.7
通讯作者：
Cossarizza, A
中科院分区：
文献类型：
--
作者：
Lugli, E;Troiano, L;Cossarizza, A
背景：到目前为止，由于常用染料的光谱特性，不可能同时分析细胞凋亡过程中的几个参数，包括DNA含量和线粒体膜电位（Delta Psi）。利用多波长激光和紫外灯激发的多色流式细胞术，我们对细胞凋亡过程中δ Psi的变化进行了表征。方法：用类黄酮槲皮素（Qu）处理U937细胞，用JC-1染色检测AT，用碘化丙啶（PI）染色检测细胞活力，用Hoechst 33342染色检测DNA含量，用Annexin V与Alexa Fluor-647偶联检测磷脂二丝氨酸（PS）暴露，检测早期凋亡标志物，用Mitotracker Deep Red检测线粒体质量。结果：曲霉可引起三种不同δ Psi的细胞群发病：(1)健康细胞，δ Psi正常，DNA含量和物理参数正常，线粒体质量高，PI-和Annexin v阴性；(2) δ Psi中等，DNA含量正常，但具有典型凋亡细胞的物理参数，线粒体质量低；以PI+和Annexin V+居多；(3) Delta Psi塌陷的细胞，线粒体质量低，Annexin-V+, PI+；其中一半显示DNA含量减少。在其他细胞凋亡模型中也观察到类似的结果，即存在中等δ Psi的细胞。结论：在q诱导的细胞凋亡过程中，δ Psi的丧失、PS暴露和线粒体质量的减少是PI通透性和DNA丢失之前的早期事件。JC-1染色后流式细胞术显示，不同δ Psi的细胞群在与凋亡相关的其他参数上也存在差异。因此，通过多色流式细胞术同时分析几个参数，可以更好地识别细胞死亡的许多阶段，更一般地说，可以评估被研究群体对给定化合物的最终异质敏感性。(c) 2005年国际分析细胞学学会。
Background: Until now, the simultaneous analysis of several parameters during apoptosis, including DNA content and mitochondrial membrane potential (Delta Psi), has not been possible because of the spectral characteristics of the commonly used dyes. Using polychromatic flow cytometry based upon multiple laser and UV lamp excitation, we have characterized cells with different Delta Psi during apoptosis.Methods: U937 cells were treated with the flavonoid quercetin (Qu) and stained with JC-1 to detect AT, propidium iodide (PI) for cell viability, Hoechst 33342 for DNA content, Annexin V conjugated with Alexa Fluor-647 for detection of phosphatidilserine (PS) exposure, marker of early apoptosis, or Mitotracker Deep Red for the determination of mitochondrial mass.Results: Treatment with Qu provoked the onset of three cell populations with different Delta Psi: (1) healthy cells, with normal Delta Psi, DNA content and physical parameters, high mitochondrial mass, PI- and Annexin V-negative; (2) cells with intermediate Delta Psi and normal DNA content, but with physical parameters typical of apoptotic cells and low mitochondrial mass; most of them were PI+ and Annexin V+; (3) cells with collapsed Delta Psi that had low mitochondrial mass and were Annexin-V+, PI+; half of them showed diminished DNA content. Similar results, i.e. the presence of cells with intermediate Delta Psi, were observed in other models of apoptosis.Conclusions: During Qu-induced apoptosis, loss of Delta Psi, PS exposure, and decrease of mitochondrial mass are early events that precede permeability to PI and loss of DNA. Populations of cells with different Delta Psi, as revealed by flow cytometry after JC-1 staining, differed also for other parameters associated to apoptosis. Thus, the simultaneous analysis of several parameters by polychromatic flow cytometry permits a better identification of many stages of cell death, and, more in general, allows to evaluate the eventual heterogenic sensibility of the population under study to a given compound. (c) 2005 international Society for Analytical Cytology.