Zhang Zhuoxi, Bai Zhonghu, Liu Guangyin, Nie Jianqi, Yang Yankun
The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, Jiangsu, China.
National Engineering Research Center of Cereal Fermentation and Food Biomanufacturing, Jiangnan University, Wuxi 214122, Jiangsu, China.
Sheng Wu Gong Cheng Xue Bao. 2023 Aug 25;39(8):3364-3378. doi: 10.13345/j.cjb.230142.
With various diseases ravaging internationally, the demands for recombinant adenoviral vector (Adv) vaccines have increased dramatically. To meet the demand for Adv vaccine, development of a new cell culture process is an effective strategy. Applying hyperosmotic stress in cells before virus infection could increase the yield of Adv in batch culture mode. Emerging perfusion culture can significantly increase the yield of Adv as well. Therefore, combining the hyperosmotic stress process with perfusion culture is expected to improve the yield of Adv at high cell density. In this study, a shake flask combined with a semi-perfusion culture was used as a scaled-down model for bioreactor perfusion culture. Media with osmotic pressure ranging from 300 to 405 mOsm were used to study the effect of hyperosmotic stress on cell growth and Adv production. The results showed that using a perfusion culture process with a hyperosmotic pressure medium (370 mOsm) during the cell growth phase and an isosmotic pressure medium (300 mOsm) during the virus production phase effectively increased the yield of Adv. This might be due to the increased expression of HSP70 protein during the late phases of virus replication. The Adv titer in a bioreactor with such a process reached 3.2×10 IFU/mL, three times higher than that of the traditional perfusion culture process. More importantly, this is the first time that a strategy of combining the hyperosmotic stress process with perfusion culture is applied to the production of Adv in HEK 293 cells. It also reveals the reason why the hyperosmotic stress process increased the yield of Adv, which may facilitate the process optimization of for producing other Adv in HEK 293 cells.
随着各种疾病在全球肆虐，对重组腺病毒载体（Adv）疫苗的需求急剧增加。为满足Adv疫苗的需求，开发新的细胞培养工艺是一种有效的策略。在病毒感染前对细胞施加高渗应激可提高分批培养模式下Adv的产量。新兴的灌注培养也能显著提高Adv的产量。因此，将高渗应激过程与灌注培养相结合有望在高细胞密度下提高Adv的产量。在本研究中，将摇瓶与半灌注培养相结合用作生物反应器灌注培养的缩小模型。使用渗透压范围为300至405 mOsm的培养基研究高渗应激对细胞生长和Adv生产的影响。结果表明，在细胞生长阶段使用高渗压培养基（370 mOsm）的灌注培养过程，在病毒生产阶段使用等渗压培养基（300 mOsm），可有效提高Adv的产量。这可能是由于病毒复制后期HSP70蛋白表达增加所致。采用这种工艺的生物反应器中Adv滴度达到3.2×10⁹ IFU/mL，比传统灌注培养工艺高出三倍。更重要的是，这是首次将高渗应激过程与灌注培养相结合的策略应用于HEK 293细胞中Adv的生产。它还揭示了高渗应激过程提高Adv产量的原因，这可能有助于优化在HEK 293细胞中生产其他Adv的工艺。
