Dept. of Biological Sciences and Graduate School of Nanoscience and Technology (WCU), KAIST, Yuseong-gu, Daejeon 305-701, Republic of Korea.
Biotechnol Prog. 2012 Mar-Apr;28(2):349-57. doi: 10.1002/btpr.1512. Epub 2012 Jan 4.
Sodium butyrate (NaBu), which is widely used in recombinant Chinese hamster ovary cell (rCHO) cultures for high-level expression of therapeutic proteins, is known to induce apoptosis in a dose-dependent manner. Lately, the significance of autophagy has increased in the field of CHO cell culture due to the fact that autophagy is related to the programmed cell death mechanism. To determine the effect of NaBu on autophagy as well as apoptosis of rCHO cells, rCHO cells producing erythropoietin were subjected to NaBu treatment. NaBu treatment up to 5 mM increased cleaved forms of PARP, caspase-3, and Annexin V positive population, confirming the previous results that NaBu induces apoptosis. Concurrently, NaBu treatment increased the level of accumulation of the autophagic marker, LC3-II, independently of nutrient depletion, suggesting that NaBu induces autophagy. To elucidate the potential role of autophagy induced by NaBu, a representative autophagy inducer (rapamycin) or an inhibitor (bafilomycin A1) was added to cultures together with NaBu. It was found that autophagy had the potential role of a positive cell survival mechanism under NaBu treatment. Furthermore, gradual reduction in mitochondrial membrane potential/mass and recruitment of a mitophagy protein, Parkin, to the mitochondria were observed under NaBu treatment, suggesting that this positive function of autophagy might be mediated by the autophagic removal of damaged mitochondria. Taken together, autophagy was observed in rCHO cell culture under NaBu treatments and the results obtained here support the positive effects of autophagy induced by NaBu treatments.
丁酸钠（NaBu）广泛用于重组中国仓鼠卵巢细胞（rCHO）培养物中，以实现治疗性蛋白的高水平表达，已知其以剂量依赖的方式诱导细胞凋亡。最近，由于自噬与程序性细胞死亡机制有关，自噬在 CHO 细胞培养领域的重要性增加。为了确定 NaBu 对 rCHO 细胞自噬和凋亡的影响，用 NaBu 处理产生红细胞生成素的 rCHO 细胞。NaBu 处理高达 5 mM 会增加 PARP、caspase-3 和 Annexin V 阳性细胞的裂解形式，证实了先前的结果，即 NaBu 诱导细胞凋亡。同时，NaBu 处理会增加自噬标记物 LC3-II 的积累水平，与营养物质耗尽无关，表明 NaBu 诱导自噬。为了阐明 NaBu 诱导的自噬的潜在作用，将代表性的自噬诱导剂（雷帕霉素）或抑制剂（巴弗洛霉素 A1）与 NaBu 一起添加到培养物中。结果发现，自噬在 NaBu 处理下具有正向细胞存活机制的潜在作用。此外，在 NaBu 处理下观察到线粒体膜电位/质量逐渐降低，以及 Parkin 等线粒体自噬蛋白募集到线粒体，表明自噬的这种正向作用可能是通过自噬清除受损线粒体介导的。综上所述，在 rCHO 细胞培养物中观察到自噬，并且这里获得的结果支持 NaBu 处理诱导的自噬的积极作用。
