Development of microphysiological system of small intestine mucosa to investigate effects of microbiota on transepithelial transport and host immune response

Small Intestinal Bacterial Overgrowth (SIBO) is a medical condition characterized by an excessive growth of bacteria in the small intestine. The prevalence of SIBO is high in patients with concurrent gastrointestinal complications, such as irritable bowel syndrome (~35.5%), inflammatory bowel disease (~22.3%), and non-alcoholic liver disease (~35%). Surprisingly, studies report a high (up to 13%) prevalence of SIBO in healthy control groups. Currently, the culture of small intestine aspirate represents the only option to identify bacteria causing SIBO, but it is highly invasive for the patient and therefore only used in emergency cases. The main aim of this project is to establish an in vitro model that can efficiently discover species-specific SIBO-associated biomarkers, which would support the development of more convenient SIBO diagnostics in the future. We will study the transport of metabolites from the luminal to the vascular side of an in vitro epithelium integrated into a microphysiological system (MPS). This model will be challenged with metabolites as well as cell material from bacterial species known to frequently cause SIBO. Untargeted metabolomics will provide data on which compounds transition over the epithelium, and more importantly, which are species-specific and predictive of the SIBO causative agent. The major aim of this project is to develop an experimental platform that integrates advanced 3D bioprinting approaches with microphysiological systems (MPS) to engineer novel in vitro models of the small intestine mucosa. This integration will allow the manufacturing of bioscaffolds with precisely defined composition and geometrical features as well as a high degree of control over culturing conditions to mimic the native physiological environment and steer tissue development in the desired direction. With steadily increasing complexity and an improving simulation of GIT function, MPS models are creating new opportunities to study the cellular and molecular levels of its physiology in health and disease. The advantages of the proposed experimental platform over existing in vitro models of the small intestine mucosa system come from the specific integration of the 3D bioprinted scaffold and the MPS design, which will align the intestinal epithelium and the underlying vascularized tissue within a clear optical pathway. Consequently, this will allow continuous imaging and monitoring of molecular transport from the simulated intestinal lumen through the epithelium and into the vasculature. The project will fulfill three specific objectives, each realized in its respective work package, namely: O1 – Development of the experimental platform and engineering of the in vitro SIM (MPS development and integration into the culturing set-up to allow bright-field imaging and sample collection at the luminal and vascular circuits. O2 – Isolation, characterization, and cultivation of SIBO-associated bacterial species to obtain a representative collection of strains that will be further used for biomarker discovery in the in vitro SIM. O3 – In vitro transport studies of metabolites and cell wall components of bacterial origin in healthy and challenged in vitro SIM MPS with untargeted metabolomics. The interdisciplinary skills and experience required for successful implementation will be ensured by a balanced team of experienced and early-career researchers from a range of backgrounds and the state-of-the-art infrastructure available at the Faculty of Medicine, University of Maribor, and the National Laboratory of Health, Environment, and Foodstaffs.