Human organoid-derived Gut-on-Chip model for studying Virus–Host interactions

Organ-on-Chip (OoC) systems are an emerging cell culture technology that bridge the gap between conventional 2D culture and animal models or patients. These microfluidic systems typically consist of one or more compartments containing tissue-specific cells grown on scaffolds such as membranes or hydrogels. OoC models can recapitulate key organotypic functions, while integrated pumps provide continuous microfluidic perfusion to mimic blood flow, resulting in the removal of waste, providing nutrients and growth factors, thus overall better mimic essential in vivo conditions. We established an adult stem cell–derived human intestinal organoid gut-on-chip system using Dynamic42 biochips to study virus–host interactions. The model integrates human intestinal epithelial cells that self-organize into a 3D architecture featuring villus- and crypt-like structures. The biochip is cultured connected to a peristaltic pump that perfuses the culture medium. Each biochip contains two microfluidic chambers separated by a porous polyethylene terephthalate (PET) membrane, which serves both as a scaffold for cell attachment and as a semi-permeable barrier enabling cell–cell communication and migration. Each chamber is accessible via two Luer-lock ports. Upon perfusion, the cells form 3D structures that provide a physiologically relevant interface for colonization with live microorganisms. Virus infection can be performed under static or perfused conditions. Multiple readouts can be applied to both effluent and tissue layers, including assessment of viral replication kinetics (RT-qPCR), virus tropism (confocal microscopy), cell viability, metabolic activity, immune responses, barrier integrity, and single-cell RNA sequencing.