Overexpression of inducible heat shock protein 70 and its mutants in astrocytes is associated with maintenance of mitochondrial physiology during glucose deprivation stress
Overexpression of inducible heat shock protein 70 and its mutants in astrocytes is associated with maintenance of mitochondrial physiology during glucose deprivation stress
复制标题
DOI：
10.1379/csc-182r.1
复制
发表时间：
2006-06-01
影响因子：
3.8
通讯作者：
Giffard, Rona G.
中科院分区：
文献类型：
--
作者：
Ouyang, Yi-Bing;Xu, Li-Jun;Giffard, Rona G.
野生型诱导型Hsp 70（WT）和2种折叠缺陷突变体在体内可保护脑免于局灶性脑缺血，在体外可保护脑细胞免于氧-葡萄糖剥夺（OGD），但其保护机制尚不清楚。线粒体对正常的生理功能和细胞死亡的调节都是至关重要的。我们测试了过表达Hsp 70和2个突变体，Hsp 70-K71 E，一个腺苷三磷酸酶（ATP酶）缺陷的点突变体，和Hsp 70 -381-640，一个缺失突变体缺乏ATP酶结构域对原代培养的星形胶质细胞在葡萄糖剥夺（GD）应激下线粒体生理学的影响。线粒体膜电位进行了评估，使用电位荧光染料四甲基罗丹明乙酯（TMRE）。到GD 5小时，LXSN对照转染的星形胶质细胞中的线粒体具有显著降低的膜电位。然而，在Hsp 70-WT、-K71 E和-381-640组中，在GD的5小时期间TMRE信号没有明显变化。测量耗氧量以评估氧化呼吸。Hsp 70-K71 E和-381-640的过表达防止了在5小时时观察到的状态III呼吸的降低，并且所有3种都防止了GD 5小时后在LXSN对照中发现的状态IV呼吸的增加。活性氧（ROS）的生产进行了评估与hydroethidine。热休克蛋白70及其突变体都显着减少增加的ROS积累在GD的5小时。结果表明，热休克蛋白70和全长分子的羧基末端的一半的保护作用与更好地维持线粒体膜电位，更好地维持状态IV呼吸，并减少ROS的产生在GD。
Wild-type inducible Hsp70 (WT) and 2 folding deficient mutants protect the brain against focal cerebral ischemia in vivo and brain cells from oxygen-glucose deprivation (OGD) in vitro, but the protective mechanisms remain unclear. Mitochondria are central to both normal physiological function and the regulation of cell death. We tested the effect of overexpressing Hsp70 and 2 mutants, Hsp70-K71E, an adenosine triphosphatase (ATPase)-deficient point mutant, and Hsp70-381-640, a deletion mutant lacking the ATPase domain on mitochondrial physiology under glucose deprivation (GD) stress in primary cultured astrocytes. Mitochondrial membrane potential was assessed using a potentiometric fluorescent dye tetramethylrhodamine ethyl ester (TMRE). By 5 hours of GD, the mitochondria in the LXSN control transfected astrocytes had markedly reduced membrane potential. However, in the Hsp70-WT, -K71E, and -381-640 groups, there was no apparent change in TMRE signal during 5 hours of GD. Oxygen consumption was measured to assess oxidative respiration. Overexpression of Hsp70-K71E and -381-640 prevented the decrease in state III respiration observed at 5 hours, and all 3 prevented the increase in state IV respiration found in LXSN controls after 5 hours of GD. Reactive oxygen species (ROS) production was assessed with hydroethidine. Hsp70 and its mutants all significantly reduced the increases in ROS accumulation during 5 hours of GD. The results demonstrate that the protective effect of the carboxyl-terminal half of Hsp70 and of the full-length molecule is associated with better maintained mitochondrial membrane potential, better maintained state IV respiration, and reduced ROS generation during GD.