Ritter Joachim B, Wahl Aljoscha S, Freund Susann, Genzel Yvonne, Reichl Udo
Delft University of Technology, Bioprocess Technology, Julianalaan 67, 2628 BC Delft, the Netherlands.
BMC Syst Biol. 2010 May 13;4:61. doi: 10.1186/1752-0509-4-61.
Many details in cell culture-derived influenza vaccine production are still poorly understood and approaches for process optimization mainly remain empirical. More insights on mammalian cell metabolism after a viral infection could give hints on limitations and cell-specific virus production capacities. A detailed metabolic characterization of an influenza infected adherent cell line (MDCK) was carried out based on extracellular and intracellular measurements of metabolite concentrations.
For most metabolites the comparison of infected (human influenza A/PR/8/34) and mock-infected cells showed a very similar behavior during the first 10-12 h post infection (pi). Significant changes were observed after about 12 h pi: (1) uptake of extracellular glucose and lactate release into the cell culture supernatant were clearly increased in infected cells compared to mock-infected cells. At the same time (12 h pi) intracellular metabolite concentrations of the upper part of glycolysis were significantly increased. On the contrary, nucleoside triphosphate concentrations of infected cells dropped clearly after 12 h pi. This behaviour was observed for two different human influenza A/PR/8/34 strains at slightly different time points.
Comparing these results with literature values for the time course of infection with same influenza strains, underline the hypothesis that influenza infection only represents a minor additional burden for host cell metabolism. The metabolic changes observed after 12 h pi are most probably caused by the onset of apoptosis in infected cells. The comparison of experimental data from two variants of the A/PR/8/34 virus strain (RKI versus NIBSC) with different productivities and infection dynamics showed comparable metabolic patterns but a clearly different timely behavior. Thus, infection dynamics are obviously reflected in host cell metabolism.
细胞培养衍生流感疫苗生产中的许多细节仍了解不足，工艺优化方法主要仍基于经验。对病毒感染后哺乳动物细胞代谢的更多了解可为细胞特异性病毒生产能力的限制提供线索。基于细胞外和细胞内代谢物浓度测量，对流感感染的贴壁细胞系（MDCK）进行了详细的代谢特征分析。
对于大多数代谢物，感染（人甲型流感病毒A/PR/8/34）细胞与模拟感染细胞的比较显示，在感染后（pi）的前10 - 12小时内行为非常相似。在pi约12小时后观察到显著变化：（1）与模拟感染细胞相比，感染细胞中细胞外葡萄糖的摄取和乳酸释放到细胞培养上清液中的量明显增加。同时（pi 12小时），糖酵解上游的细胞内代谢物浓度显著增加。相反，感染细胞的核苷三磷酸浓度在pi 12小时后明显下降。在两个不同的人甲型流感病毒A/PR/8/34毒株中，在略有不同的时间点观察到了这种行为。
将这些结果与相同流感毒株感染时间进程的文献值进行比较，支持了流感感染仅对宿主细胞代谢构成轻微额外负担这一假设。pi 12小时后观察到的代谢变化很可能是由感染细胞中凋亡的开始引起的。对具有不同生产力和感染动态的A/PR/8/34病毒株的两个变体（德国罗伯特·科赫研究所（RKI）与英国国家生物制品检定所（NIBSC））的实验数据比较显示，代谢模式具有可比性，但时间行为明显不同。因此，感染动态明显反映在宿主细胞代谢中。
