Hung A C, Huang H M, Tsay H J, Lin T N, Kuo J S, Sun S H
Institute of Neuroscience, National Yang Ming University, Taipei, Taiwan, ROC.
J Cell Biochem. 2000 Mar;77(2):323-32. doi: 10.1002/(sici)1097-4644(20000501)77:2<323::aid-jcb14>3.0.co;2-w.
The stimulus-transcriptional coupling during ischemia/hypoxia was examined for ATP-stimulated expression of immediate early genes (IEGs; c-fos, zif268, c-myc and nur77) in a rat brain-derived type 2 astrocyte cell line, RBA-2. Incubation of cells with 1 mM of extracellular ATP stimulated time-dependent expression of c-fos and zif268. ATP induced the largest increases in zif268 mRNA and a lesser one in c-fos mRNA. ATP also induced a slight increase in nur77 mRNA but was ineffective in inducing c-myc expression in these cells. Brief exposure of cells to potassium cyanide to simulate chemical hypoxia induced 9-fold and 7-fold transient increases in c-fos and zif268 expression, respectively, but did not affect c-myc or nur77 expression. When cyanide and ATP were added together, the expression of c-fos and zif268 expression was inhibited, and the effect was mimicked by simulating chemical hypoxia with sodium azide. To elucidate the mechanism involved, the effect of cyanide on ATP-stimulated increases in intracellular Ca(2+) concentrations, Ca(2+), and phospholipase D (PLD) activities were measured. Cyanide induced an increase in Ca(2&plus); and further enhanced the ATP-stimulated increases in Ca(2+) and PLD activities. Nevertheless, metabolic inhibitor, iodoacetate, blocked the ATP-induced c-fos and partially inhibited zif268 expression, and deprivation of cells with glucose also inhibited the ATP-induced c-fos expression. Taken together, these results demonstrate that both extracellular ATP and chemical hypoxia induce c-fos and zif268 expression in RBA-2 type 2 astrocytes. The chemical hypoxia inhibited ATP-stimulated c-fos and zif268 expression is not due to alterations in Ca(2+) and PLD signaling, and is at least partially related to metabolic disturbance in these cells.
在大鼠脑源性2型星形胶质细胞系RBA - 2中，研究了缺血/缺氧期间的刺激-转录偶联，以观察ATP刺激的即刻早期基因（IEGs；c - fos、zif268、c - myc和nur77）的表达。用1 mM细胞外ATP孵育细胞可刺激c - fos和zif268的时间依赖性表达。ATP诱导zif268 mRNA的增加幅度最大，而c - fos mRNA的增加幅度较小。ATP还诱导nur77 mRNA略有增加，但对这些细胞中c - myc的表达无诱导作用。短暂暴露细胞于氰化钾以模拟化学性缺氧分别诱导c - fos和zif268表达瞬时增加9倍和7倍，但不影响c - myc或nur77的表达。当氰化钾和ATP一起添加时，c - fos和zif268的表达受到抑制，用叠氮化钠模拟化学性缺氧可模拟该效应。为阐明其中涉及的机制，测量了氰化钾对ATP刺激的细胞内Ca(2+)浓度[Ca(2+)]i增加及磷脂酶D（PLD）活性的影响。氰化钾诱导[Ca(2+)]i增加，并进一步增强ATP刺激的[Ca(2+)]i和PLD活性增加。然而，代谢抑制剂碘乙酸可阻断ATP诱导的c - fos表达并部分抑制zif268表达，用葡萄糖剥夺细胞也可抑制ATP诱导的c - fos表达。综上所述，这些结果表明细胞外ATP和化学性缺氧均可诱导RBA - 2型2星形胶质细胞中c - fos和zif268的表达。化学性缺氧抑制ATP刺激的c - fos和zif268表达并非由于Ca(2+)和PLD信号改变，且至少部分与这些细胞中的代谢紊乱有关。
