(64e) In Vitro Reconstitution of Natural Mucins Captures pH and Ion-Dependent Collective Dynamic Mucus Barrier Complexity

A mucus lining along the gastrointestinal tract is the outermost and largest interface in the body that protects the epithelium, regulates the selective transport of nutrients, and maintains a sterile relationship with microbes. Despite its indisputable importance, a mucus barrier has been understudied due to the lack of relevant experimental models. Here, we introduce a functional and analytical mucus barrier model that was enabled by the development of tissue-inspired simple and scalable mucin extraction method. Retrieved mucins preserved intrinsic biochemical and biophysical characteristics. Reconstituted mucus barriers recapitulated pH and Ca²⁺ dependent mucus barrier properties and associated molecular transport and bacterial motility. At neutral pH, the thickness of a mucus barrier linearly increases as a function of Ca²⁺, but the increased thickness does not necessarily decrease mucus transport. Intriguingly, there exists an optimal Ca²⁺ concentration that maximally delays transport. Our model revealed that the mucus fluid forms a coacervate-like phase; densely packed small aggregates of mucin-ionic complexes minimize the interstitial space for percolation. This observation explains how secreted mucins collectively and autonomously regulate barrier properties in response to dynamically evolving luminal contents. We envision that the established methods facilitate various aspects of mucus barrier research and mucosal barrier tissue modeling with higher physiological complexity.