Macrophage phagocytosis of human norovirus-infected cells in an ex vivo human enteroid-macrophage coculture model.

Human norovirus (HuNoV) causes acute gastroenteritis in immunocompetent hosts and chronic infection in immunocompromised individuals. Many recent studies of replication and innate immune responses following HuNoV infection have utilized epithelium-only human intestinal enteroids (HIEs), which lack immune cells. Here, we utilized an ex vivo enteroid-macrophage coculture model consisting of HIEs and different subtypes of human peripheral blood mononuclear cell-derived macrophages to better recapitulate in vivo gut biology and explore the role of macrophages in HuNoV replication and pathogenesis. We show that HuNoV infection in HIEs polarized on Transwells leads to bilateral release of the viral genome, with predominant apical virus release. Coculture with naïve M0, pro-inflammatory M1, or anti-inflammatory M2 macrophages does not change levels of HuNoV replication. We found that macrophages respond to HuNoV infection by phagocytosis of virus-infected cells with pro-inflammatory M1 macrophages exhibiting the greatest phagocytic activity. Apical release of chemokines (IP-10, MIP-1α, and RANTES), acute-phase inflammatory mediators (IL-1α, IL1-RA, IL-6, and TNF-α), and anti-inflammatory mediators IL-10 and IL-1RA are increased following HuNoV infection only in HIEs cocultured with activated macrophages. These findings underscore the importance of epithelial-macrophage crosstalk in chemokine production during the early stages of infection required for leukocyte recruitment and initiation of the host response.IMPORTANCEHuman noroviruses (HuNoVs) are important human enteric pathogens that cause outbreaks and sporadic gastroenteritis in people of all ages. HuNoV-induced illness can become chronic and debilitating in immunocompromised hosts. Previously, we observed an uptake of HuNoV-infected epithelial cells by macrophages in intestinal biopsies derived from chronically infected transplant patients. Based on this, we put forward the hypothesis that intestinal macrophages are important in contributing to an anti-HuNoV response during the early stages of infection. Here, we investigated the potential role of different macrophage subtypes in the induction of immune responses following HuNoV infection using HIE-macrophage coculture models. We found that pro-inflammatory macrophages showed the greatest capacity for phagocytosing virus-infected cells. This study highlights the importance of the interaction of the intestinal epithelium with activated macrophages in an anti-viral response that may be critical for enhancing viral clearance and reducing viral spread.