Effects of oxygen tension on the establishment and lactate dehydrogenase activity of murine embryonic stem cells
Effects of oxygen tension on the establishment and lactate dehydrogenase activity of murine embryonic stem cells
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DOI：
10.1089/clo.2006.8.117
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发表时间：
2006-06-01
期刊：
影响因子：
--
通讯作者：
Gardner, D. K.
中科院分区：
文献类型：
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作者：
Gibbons, J.;Hewitt, E.;Gardner, D. K.
胚胎干细胞系建立缺乏标准化的培养环境，阻碍了细胞的有序分化和技术的应用。氧气浓度对许多细胞类型的增殖和分化具有深远的影响。本研究测试了以下假设：建立动态、乳酸脱氢酶 (LDH) 同工型和 mRNA 表达模式会受到培养环境中氧张力的影响。回收的（第 4 天）小鼠囊胚在 6% CO2 和 20% 或 5% O-2（平衡补充 N-2）的气体环境中培养。与高 (57.1 +/- 0.1%) O-2 组相比，低 (79.3 +/- 0.1%) 组中产生更多 (p < 0.05) 胚泡，并且相对于高 02 组，低 02 组 (14/15; 93.3 +/- 0.1%) 中碱性磷酸酶染色呈阳性的集落更多 (p < 0.05) （9/15；60.6 +/- 0.1%）。氧气处理对氧化还原酶乳酸脱氢酶的活性没有影响。有趣的是，两种处理中的干细胞系都显示出多种 LDH 亚型（III、IV 和 V），而生长物则显示亚型 I 和 V。相比之下，双细胞胚胎和囊胚仅显示亚型 I，而成纤维细胞则显示亚型 IV 和 V。在生长物或建立的干细胞中，LDH α mRNA 表达没有处理差异。 LDH 从心脏 (1) 到肌肉 (V) 亚型的转变表明糖酵解活性增加，与孵化期/植入期一致。减少的 O-2 环境对小鼠干细胞的建立和维持具有显着的积极影响，支持了这一假设，而 LDH 同工酶转变在处理之间是一致的。
The lack of a standardized culture environment for establishment of embryonic stem cell lines has hindered the orchestrated differentiation of cells and the application of this technology. Oxygen concentration has a profound effect on proliferation and differentiation of many cell types. This study tested the hypothesis that establishment dynamics, lactate dehydrogenase (LDH) isoforms, and mRNA expression patterns would be affected by the oxygen tension in the culture environment. Recovered (day 4) murine blastocysts were cultured in a gas environment of 6% CO2 and either 20% or 5% O-2 (balance supplemented with N-2). More (p < 0.05) blastocysts produced outgrowths in the low (79.3 +/- 0.1%) compared to the high (57.1 +/- 0.1%) O-2 groups, and more (p < 0.05) colonies in the low 02 group (14/15; 93.3 +/- 0.1%) stained positive for alkaline phosphatase relative to the high 02 group (9/15; 60.6 +/- 0.1%). Oxygen treatment had no effect on the activity of the oxioreductase lactate dehydrogenase. Interestingly, the stem cell lines in both treatments displayed multiple isoforms (III, IV, and V) of LDH, whereas the outgrowths displayed isoforms I and V. In contrast, two-cell embryos and blastocysts displayed only isoform I, and fibroblasts displayed isoforms IV and V. There were no treatment differences in mRNA expression of LDH alpha in the outgrowths, or established stem cells. LDH transition from the heart (1) to the muscle (V) isoform indicated an increase in glycolytic activity, consistent with the peri-hatching/implantation time period. Reduced O-2 environment had significant positive effects on the establishment and maintenance of murine stem cells, supporting the hypothesis, whereas the LDH isozyme transition was consistent among treatments.