Tiffany-Castiglioni E, Garcia D M, Wu J N, Zmudzki J, Bratton G R
Department of Veterinary Anatomy, Texas A&M University, College Station 77843.
J Toxicol Environ Health. 1988;23(2):267-79. doi: 10.1080/15287398809531112.
Cultured C6 rat glioma cells were exposed to lead (Pb) acetate (0, 1, 10, or 100 microM) for 3-4 d. Cells were analyzed for changes in viability and intracellular lead, iron, and copper concentrations after Pb treatment was discontinued. The results were compared with previous findings on astroglia and oligodendroglia in culture in order to evaluate C6 cultures as a model for Pb toxicity in glia. Viability was measured by three methods on the day Pb was removed from the cells (designated d 0), and 2 and 9 d after Pb treatment was discontinued (designated d 2 and 9). The methods used were trypan blue dye exclusion, total cell counts, and incorporation of [3H]-L-leucine into proteins. Small, dose-dependent reductions were observed on d 2 in the percentages of cells excluding dye. Decreased cell numbers were seen at all three Pb doses only on d 0. With respect to these two viability measurements, C6 cells responded like astroglia in culture to Pb, but not like oligodendroglia, which are more Pb-sensitive. We expected decreased amino acid incorporation to accompany the decreased viability of the cultures. Instead, increased amino acid incorporation, which indicates increased protein synthesis, was seen on d 0 and 2 at all three Pb doses, though total cellular protein did not increase. A similar response has been reported previously in oligodendroglial cultures. C6 cells treated for 3 with 1 or 100 microM Pb acetate were analyzed for intracellular metal content by atomic absorption aspectroscopy on d 4 and 11 after exposure to Pb was discontinued. The cells were found to take up large amounts of Pb intracellularly and store it for at least 11 d. Cells treated with FeCl2 instead of Pb took up Fe, but required a higher extracellular Fe concentration to achieve an intracellular Fe level comparable to that of Pb in Pb-treated cells. Pb uptake did not affect intracellular Fe or Cu concentrations. With respect to Pb and Fe uptake, C6 cells closely resembled immature astroglia in culture. Unlike C6 cells, however, astroglia showed elevations of intracellular Fe and Cu after Pb treatment. Thus, Pb effects on C6 cells resembled those on cultured oligodendroglia and astroglia in some respects but not in others. C6 cells appear to be an adequate model for selected events in glial toxicosis, such as Pb-stimulated protein synthesis in oligodendroglia and Pb uptake in astroglia, but not Pb-induced alterations of intracellular Cu and Fe in astroglia. Their use as a model for glial progenitor cells in Pb toxicity studies remains to be determined.
将培养的C6大鼠胶质瘤细胞暴露于醋酸铅（0、1、10或100微摩尔）中3 - 4天。在停止铅处理后，分析细胞的活力变化以及细胞内铅、铁和铜的浓度。将结果与之前关于培养的星形胶质细胞和少突胶质细胞的研究结果进行比较，以评估C6细胞培养物作为胶质细胞铅毒性模型的可行性。在从细胞中去除铅的当天（标记为第0天）、停止铅处理后2天和9天（分别标记为第2天和第9天），通过三种方法测量细胞活力。所使用的方法为台盼蓝染料排斥法、细胞总数计数法以及[3H]-L-亮氨酸掺入蛋白质的方法。在第2天观察到，排斥染料的细胞百分比出现了小幅度的剂量依赖性降低。仅在第0天，在所有三个铅剂量组中细胞数量均减少。就这两种活力测量方法而言，C6细胞在培养中对铅的反应与星形胶质细胞相似，但与对铅更敏感的少突胶质细胞不同。我们预期氨基酸掺入量会随着培养物活力的降低而减少。然而，在第0天和第2天，在所有三个铅剂量组中均观察到氨基酸掺入量增加，这表明蛋白质合成增加，尽管细胞总蛋白并未增加。之前在少突胶质细胞培养物中也报道过类似的反应。在停止铅暴露后第4天和第11天，通过原子吸收光谱法分析用1或100微摩尔醋酸铅处理3天的C6细胞的细胞内金属含量。发现细胞在细胞内摄取大量铅并将其储存至少11天。用氯化铁代替铅处理的细胞摄取铁，但需要更高的细胞外铁浓度才能达到与铅处理细胞中铅相当的细胞内铁水平。铅的摄取并不影响细胞内铁或铜的浓度。就铅和铁的摄取而言，C6细胞与培养中的未成熟星形胶质细胞非常相似。然而，与C6细胞不同的是，星形胶质细胞在铅处理后细胞内铁和铜含量升高。因此，铅对C6细胞的影响在某些方面与对培养的少突胶质细胞和星形胶质细胞的影响相似，但在其他方面则不同。C6细胞似乎是胶质细胞中毒某些特定事件的合适模型，例如铅刺激少突胶质细胞中的蛋白质合成以及星形胶质细胞摄取铅，但不是铅诱导星形胶质细胞内铜和铁的改变。它们作为铅毒性研究中胶质祖细胞模型的用途仍有待确定。
