Abe K, Saito H
Department of Chemical Pharmacology, Faculty of Pharmaceutical Sciences, The University of Tokyo, Japan.
Neurosci Res. 1998 Aug;31(4):295-305. doi: 10.1016/s0168-0102(98)00055-8.
Alzheimer's disease amyloid beta protein (Abeta) inhibits cellular reduction of the dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Kaneko et al. have previously hypothesized that Abeta works by suppressing mitochondrial succinate dehydrogenase (SDH), but Liu and Schubert have recently demonstrated that Abeta decreases cellular MTT reduction by accelerating the exocytosis of MTT formazan in neuronal cells. To ask which is the case in astrocytes, we compared the effects of Abeta and 3-nitropropionic acid (3-NP), a specific SDH inhibitor, on MTT reduction in cultured rat cortical astrocytes. Treatment with 3-NP (10 mM) decreased cellular activity of MTT reduction, regardless of the time of incubation with MTT. On the other hand. Abeta-induced inhibition of cellular MTT reduction was dependent on the time of incubation with MTT. The cells treated with Abeta (0.1-1000 nM) exhibited normal capacity for MTT reduction at an early stage of incubation ( < 30 min), but ceased to reduce MTT at the late stage (> 1 h). Microscopic examination revealed that Abeta treatment accelerated the appearance of needle-like MTT formazan crystals at the cell surface. These observations support that Abeta accelerates the exocytosis of MTT formazan in astrocytes. In addition to inhibition of MTT reduction, Abeta is known to induce morphological changes in astrocytes. Following addition of Abeta (20 microM), polygonal astrocytes changed into process-bearing stellate cells. To explore a possible linkage between these two effects of Abeta, we tested if astrocyte stellation is induced by agents that mimic the effect of Abeta on MTT reduction. Cholesterol (5 5000 nM) and lysophosphatidic acid (0.2-20 microg/ml) were found to accelerate the exocytosis of MTT formazan in a similar manner to Abeta, but failed to induce astrocyte stellation. Therefore, Abeta-induced inhibition of MTT reduction is unlikely to be directly linked to its effect on astrocyte morphology.
阿尔茨海默病β淀粉样蛋白（Aβ）可抑制细胞对染料3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）的还原作用。金子等人此前曾推测，Aβ通过抑制线粒体琥珀酸脱氢酶（SDH）发挥作用，但刘和舒伯特最近证明，Aβ通过加速神经元细胞中MTT甲臜的胞吐作用来降低细胞对MTT的还原。为了探究星形胶质细胞中情况如何，我们比较了Aβ和3-硝基丙酸（3-NP，一种特异性SDH抑制剂）对培养的大鼠皮质星形胶质细胞中MTT还原的影响。用3-NP（10 mM）处理会降低细胞对MTT还原的活性，且与MTT孵育时间无关。另一方面，Aβ诱导的细胞对MTT还原的抑制作用取决于与MTT的孵育时间。用Aβ（0.1 - 1000 nM）处理的细胞在孵育早期（< 30分钟）表现出正常的MTT还原能力，但在后期（> 1小时）则停止还原MTT。显微镜检查显示，Aβ处理加速了细胞表面针状MTT甲臜晶体的出现。这些观察结果支持Aβ加速星形胶质细胞中MTT甲臜的胞吐作用。除了抑制MTT还原外，已知Aβ还会诱导星形胶质细胞的形态变化。加入Aβ（20 μM）后，多角形星形胶质细胞会转变为有突起的星形细胞。为了探究Aβ的这两种作用之间可能的联系，我们测试了模拟Aβ对MTT还原作用的试剂是否会诱导星形胶质细胞形成星状。发现胆固醇（5 - 5000 nM）和溶血磷脂酸（0.2 - 20 μg/ml）以与Aβ类似的方式加速MTT甲臜的胞吐作用，但未能诱导星形胶质细胞形成星状。因此，Aβ诱导的对MTT还原的抑制不太可能与其对星形胶质细胞形态的影响直接相关。
