Human colon-on-a-chip enables continuous in vitro analysis of colon mucus layer accumulation and physiology

Abstract Background & Aims The mucus layer in the human colon protects against commensal bacteria and pathogens, and defects in its unique bilayered structure contribute to intestinal disorders, such as ulcerative colitis. However, our understanding of colon physiology is limited by the lack of in vitro models that replicate human colonic mucus layer structure and function. Here, we investigated if combining organ-on-a-chip and organoid technologies can be leveraged to develop a human-relevant in vitro model of colon mucus physiology. Methods A human colon-on-a-chip (Colon Chip) microfluidic device lined by primary patient-derived colonic epithelial cells was used to recapitulate mucus bilayer formation, and to visualize mucus accumulation in living cultures non-invasively. Results The Colon Chip supports spontaneous goblet cell differentiation and accumulation of a mucus bilayer with impenetrable and penetrable layers, and a thickness similar to that observed in human colon, while maintaining a subpopulation of proliferative epithelial cells. Live imaging of the mucus layer formation on-chip revealed that stimulation of the colonic epithelium with prostaglandin E2, which is elevated during inflammation, causes rapid mucus volume expansion via an NKCC1 ion channel-dependent increase in its hydration state, but no increase in de novo mucus secretion. Conclusion This study is the first to demonstrate production of colonic mucus with a physiologically relevant bilayer structure in vitro, which can be analyzed in real-time non-invasively. The Colon Chip may offer a new preclinical tool to analyze the role of mucus in human intestinal homeostasis as well as diseases, such as ulcerative colitis and cancer.