Reactive Oxygen Species Limit Intestinal Mucosa-Bacteria Homeostasis in Vitro
Abstract The gut microbiome and its interactions with epithelial and immune cells have wide-ranging effects on many aspects of human health. Thus, in vitro models that enable highly controlled studies of these interactions are of value, yet critical parameters enabling long term homeostasis between bacteria and mammalian cultures have not been established. In this study, we developed a model incorporating epithelial and immune cells as well as different bacterial species (B. fragilis, E. coli, L. rhamnosus, or R. gnavus) over a 50 hour culture period. Interestingly, both obligate and facultative anaerobes grew to similar extents in aerobic culture environments in co-culture with epithelial and immune cells, potentially due to measured microaerobic oxygen levels near the epithelial apical surface. It was demonstrated that bacteria elicited reactive oxygen species (ROS) production, and that these species heavily contribute to observed epithelial barrier damage in these static cultures. Introduction of a ROS scavenger significantly mitigated ROS-mediated damage, improving cell monolayer integrity and reducing lipid peroxidation, although not to control (bacteria-free culture) levels. These results indicate that monitoring and mitigating ROS concentrations can enable longer term bacteria-intestinal epithelial cultures, but also highlight the significance of additional factors that impact homeostasis in mammalian cell-bacteria systems.