Development of an intestinal mucosa ex vivo co-culture model to study viral infections

Studying viral infections necessitates well-designed cell culture models to deepen our understanding of diseases and develop effectivetreatments. In this study, we present a readily available ex vivo 3D co-culture model replicating the human intestinal mucosa. The model combines fully differentiatedhuman intestinal epithelium (HIE) with human monocyte-derived macrophages (hMDMs) and faithfully mirrors the in vivo structural and organizational properties of intestinal mucosal tissues. Specifically,it mimics the lamina propria, basement membrane, and the air-exposed epithelial layer, enabling the pioneering observation of macrophage migration through the tissue to the site of viral infection. In this study, we applied the HIE-hMDMs model for the firsttime in viral infection studies, infecting the model with two globally significantviruses: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and human norovirus GII.4. The results demonstrate the model's capability to support the replication of both viruses and show the antiviral role of macrophages, determined by their migration to the infection site and subsequent direct contact with infected epithelial cells. In addition, we evaluated the production of cytokines and chemokines in the intestinal niche, observing an increased interleukin-8 production during infection. A parallel comparison using a classical in vitro cell line model comprising Caco-2 and THP-1 cells for SARS-CoV-2 experiments confirmedthe utility of the HIE-hMDMs model in viral infection studies. Our data show that the ex vivo tissue models hold important implications for advances in virology research.