阿尔茨海默病转线粒体DNA细胞模型胞质钙稳态的改变

目的观察阿尔茨海默病(Alzheimer's disease,AD)转线粒体DNA细胞模型胞质游离钙水平,探讨线粒体DNA(mitochondrial DNA,mtDNA)缺陷在AD发病中的作用。方法 将正常青年人、正常老年人和AD患者的血小板分别与无mtDNA细胞融合,建立转mtDNA细胞模型。采用微测量法测定细胞色素C氧化酶(cytochrome C oxidase,COX)的活性;用荧光探针Fluo-3标记胞质内游离钙离子,激光共聚焦显微镜和流式细胞仪观察细胞胞质钙离子荧光强度。结果 AD转mtDNA细胞COX活性与正常老年对照相比差异有统计学意义(P<0.05);静息状态胞质钙离子浓度升高,氧化磷酸化解耦联剂CCCP刺激后胞质钙离子的调节能力明显降低。结论 AD患者线粒体存在COX缺陷,使线粒体钙库功能下降,导致细胞内胞质钙稳态失衡。

Alteration of Cytosolic Calcium in mtDNA-transferred Cells of Alzheimer's Disease

Objective To investigate the level of cytosolic free calcium in mitochondria DNA-transferred cells of Alzheimer’s disease and the influence of mitochondrial DNA(mtDNA) deficiency on calcium regulation capacity of cells.
Methods Platelets from AD patients and aged control persons were fused with mtDNA-depleted cells to develop mtDNA transferred cell model. Cytochrome C oxidase(COX) activity was determined by microplate assay; cytosolic calcium was labeled with Fluo-3 and fluorescence density was detected with laser scan confocal microscope(LSCM) and flow cytometer(FCM).
Results COX activity was reduced in AD mtDNA-transferred cells when compared with aged control and young control. The resting cytosolic calcium was elevated in AD mtDNA-transferred cells compared with the controls and decreased when stimulated by carbonyl cyanide m-chlorophenyl hydrazone(CCCP).
Conclusion The transfer of AD mtDNA was sufficient to produce pathological changes in cytosolic calcium regulation.