Modeling influenza virus replication in primary human lung cells

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DESCRIPTION (provided by applicant): Description Concern about a human pandemic of highly pathogenic avian (H5N1) influenza has focused attention on several related problems: understanding the pathogenesis and transmission of influenza A infections, pre-pandemic recognition of potential pandemic strains, and pre-pandemic vaccine development. Although molecular biology has provided a detailed understanding of the replication cycle in immortalized cells, influenza replication at the intact tissue level among phenotypically diverse epithelial cells of the human respiratory tract remains poorly understood. We are missing the quantitative kinetics accounting in the human airway and how one strain, but not a closely related strain, can initiate person-to-person transmission. We propose to model early influenza virion productivity in human airway epithelial cells. The in vitro model uses primary human differentiated tracheobronchial and bronchiolo-alveolar epithelial cells grown in air-liquid interface (ALI) culture to document the kinetics of virion productivity of human pathogens, both H3N2 strains and H5N1 avian influenza strains. The agent-based computer model is a two-dimensional cellular automata style simulation, designed and implemented by one of us (CB). Kinetic data collected from the in vitro model will be used validate the details and parameter values of the computer model, and then the computer model will be used to study unmeasurable variables and outcomes in the in vitro model. The goal of the project is to use experimentally derived kinetic data of influenza virus replication in primary human lung epithelial cells to calculate metrics of viral growth, particularly the Ro (reproductive rate, average number of secondary infected cells produced by one primary cell), in order to obtain a human tissue-relevant indicator of virulence and transmissibility, capable of distinguishing strains without expensive in vivo models. Using data from the in vitro model, we will determine the following parameters in the computer model: unit rate of viral production per cell, virion diffusion rate, and the unit rate of virion absorption by epithelial cells. The kinetics of strains of H3N2 human pathogenic influenza A will be compared to those of selected human pathogenic H5N1 avian influenza strains, both experimentally and in the computer model. We will manipulate the environment of the ALI culture experimentally to augment or diminish components of the innate host response including mucins and surfactant collectins. Analogous manipulations will be implemented in the computer model and results compared between the two models for all the different strains. We will then use the agent-based model to systematically study the relative contribution of each innate mucosal defense element, both in terms of individual and collaborative impacts on the R0 for each influenza strain. This project develops a test designed to distinguish threatening bird flu strains from non-threatening strains. The test measures influenza virus growth in cultured mature human lung airway cells and uses mathematical analysis to calculate the spread of the virus through the culture. Bird flu is considered by many to be potentially the next greatest threat to public health in the world.