Max Planck Institute for Dynamics of Complex Technical Systems, Magdeburg, Germany.
Chair of Bioprocess Engineering, Institute of Process Engineering, Faculty of Process & Systems Engineering, Otto-von-Guericke University, Magdeburg, Germany.
PLoS Comput Biol. 2021 Sep 7;17(9):e1009357. doi: 10.1371/journal.pcbi.1009357. eCollection 2021 Sep.
Cell culture-derived defective interfering particles (DIPs) are considered for antiviral therapy due to their ability to inhibit influenza A virus (IAV) production. DIPs contain a large internal deletion in one of their eight viral RNAs (vRNAs) rendering them replication-incompetent. However, they can propagate alongside their homologous standard virus (STV) during infection in a competition for cellular and viral resources. So far, experimental and modeling studies for IAV have focused on either the intracellular or the cell population level when investigating the interaction of STVs and DIPs. To examine these levels simultaneously, we conducted a series of experiments using highly different multiplicities of infections for STVs and DIPs to characterize virus replication in Madin-Darby Canine Kidney suspension cells. At several time points post infection, we quantified virus titers, viable cell concentration, virus-induced apoptosis using imaging flow cytometry, and intracellular levels of vRNA and viral mRNA using real-time reverse transcription qPCR. Based on the obtained data, we developed a mathematical multiscale model of STV and DIP co-infection that describes dynamics closely for all scenarios with a single set of parameters. We show that applying high DIP concentrations can shut down STV propagation completely and prevent virus-induced apoptosis. Interestingly, the three observed viral mRNAs (full-length segment 1 and 5, defective interfering segment 1) accumulated to vastly different levels suggesting the interplay between an internal regulation mechanism and a growth advantage for shorter viral RNAs. Furthermore, model simulations predict that the concentration of DIPs should be at least 10000 times higher than that of STVs to prevent the spread of IAV. Ultimately, the model presented here supports a comprehensive understanding of the interactions between STVs and DIPs during co-infection providing an ideal platform for the prediction and optimization of vaccine manufacturing as well as DIP production for therapeutic use.
细胞培养衍生的缺陷干扰颗粒 (DIPs) 因其能够抑制甲型流感病毒 (IAV) 的产生而被考虑用于抗病毒治疗。DIPs 在其八个病毒 RNA (vRNAs) 之一中含有一个大的内部缺失，使其复制失活。然而，它们可以在感染期间与同源标准病毒 (STV) 一起传播，以争夺细胞和病毒资源。迄今为止，针对 IAV 的实验和建模研究在研究 STVs 和 DIPs 的相互作用时，要么侧重于细胞内水平，要么侧重于细胞群体水平。为了同时检查这些水平，我们使用 STV 和 DIP 的高度不同的感染复数进行了一系列实验，以研究 Madin-Darby 犬肾悬浮细胞中的病毒复制。在感染后的几个时间点，我们使用成像流式细胞术定量病毒滴度、活细胞浓度、病毒诱导的细胞凋亡，使用实时逆转录 qPCR 定量 vRNA 和病毒 mRNA 的细胞内水平。基于获得的数据，我们开发了一个描述 STV 和 DIP 共感染动力学的数学多尺度模型，该模型使用单个参数集可紧密描述所有情况下的动力学。我们表明，应用高浓度的 DIP 可以完全关闭 STV 的传播并防止病毒诱导的细胞凋亡。有趣的是，观察到的三种病毒 mRNA（全长片段 1 和 5、缺陷干扰片段 1）积累到差异极大的水平，这表明存在内部调节机制和较短病毒 RNA 的生长优势之间的相互作用。此外，模型模拟预测 DIP 的浓度应该至少比 STV 高 10000 倍才能阻止 IAV 的传播。最终，本文提出的模型支持了对 STV 和 DIP 共感染期间相互作用的全面理解，为疫苗制造的预测和优化以及用于治疗的 DIP 生产提供了理想的平台。
