Expression of the microfold cells in three-dimensional coculture system for in vitro cultivation of human norovirus

Introduction: Human Norovirus (HuNoV), a food-borne virus is the leading cause for acute gastroenteritis. However, its inability to propagate in vitropersists as major challenge in understanding HuNoV biology.Objective: This study aims to determine an effective culture system for HuNoV.Methods: The Caco-2 cells were cocultured with Raji B cells on alginate hydrogel beads. Scanning electron microscopy (SEM) was performed to confirmthe three-dimensional (3D) cells morphology. Western blot (WB) analysis was performed to detect protein markers expressed by Microfold (M) cells.Results: Optimization of Caco-2 cells monoculture in the alginate hydrogel beads showed optimum number of cells of 1 × 106 cells/ml, indicatedby the intact structure of the beads. Result of SEM showed clear structure of monoculture in the alginate hydrogel beads indicated by the presenceof smooth and regular apical surface while the coculture showed reduced apical surface of M cells. The result of WB showed downregulation ofUlex europaeus antibody expression.Conclusion: It is evident that the expression of M cells grown in 3D alginate hydrogel beads was successful, indicated by the structural morphologyseen under SEM as well as expression of protein marker by M cells. This established in vitro system is highly potential for cultivation of HuNoV.