Engineering Advanced in Vitro Infection Models via Microfluidics and Transwell-Based Organoid Systems
EMBARGOED UNTIL 2026-12-04
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Date
2025
Publication Type
Doctoral Thesis
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EMBARGOED UNTIL 2026-12-04
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Abstract
Antimicrobial resistance (AMR) is one of the most urgent global health challenges, expected to cause a significant rise in infection-related morbidity and mortality over the coming decades. Although the prevalence of multi-drug-resistant pathogens continues to increase, the development of new antibiotics has stagnated, which is largely due to their high attrition rates during the drug screening process and limitations of current drug screening assays. These challenges call for innovative approaches to both deepen our understanding of resistance mechanisms and to establish more efficient methods for evaluating potential antimicrobial candidates. Conducted within the framework of the NCCR AntiResist, this thesis contributes to such efforts as it includes developing physiologically relevant in vitro infection models and establishing associated experimental systems.
In this thesis, I present the design and fabrication of poly(methylmethacrylate) (PMMA)-based microfluidic platforms for infection studies using transwells with human organoid models. The system overcomes key limitations of conventional static transwell cultures by introducing physiologically relevant fluid perfusion, while ensuring system compatibility with high-resolution, real-time imaging. These features enable direct real-time visualization of host-pathogen interactions and support mechanistic studies of infection onset under physiologically relevant conditions. By bridging the gap between traditional in vitro models and complex in vivo systems, the platform provides a versatile tool for organ-specific infection research and predictive antimicrobial assessment.
The methodology compiled in this thesis details the process of growing the organoids, their integration into the microfluidic platform, and the establishment of infection assays under physiologically relevant conditions. Furthermore, the models were successfully applied to drug testing, encompassing both traditional and non-traditional antimicrobial therapies.
The thesis work focused on two organ-specific infection models:
1. Human airway epithelium model:
A system for Pseudomonas aeruginosa infection of the airway epithelium that supports dynamic transitions between air-liquid and liquid-liquid interfaces. This system enabled detailed monitoring of lung tissue infection and its gradual destruction over time at high spatial and temporal resolution.
2. Human bladder epithelium model:
A system for culturing stratified bladder epithelium under urine perfusion, which simultaneously enabled continuous monitoring of Uropathogenic Eschericia coli (UPEC) infections and colonization. This system was also employed to evaluate antimicrobial interventions, including D-mannose and bacteriophage therapies in a physiologically relevant context.
In addition, the microfluidic platform developed in this study was augmented with two additional key features:
1. Real-time impedance profiling of the tissue barrier function:
The integration of a custom-designed four-electrode sensor into the microfluidic platform enabled non-invasive, real-time assessment of transepithelial electrical resistance (TEER), performed simultaneously with high-resolution live imaging.
2. Higher-throughput pharmacokinetic analysis of antimicrobial compounds:
To increase the assay throughput from 4 to 24 transwell inserts per experimental run under varying antimicrobial concentrations, the microfluidic platform and associated fluidic circuits were re-designed and characterized. Initial tests for antimicrobial compound permeability were conducted.
Overall, this work demonstrates how the combination of microfluidics with transwell-based organoid cultures can significantly advance our ability to study bacterial infections and evaluate therapeutic strategies under representative physiological conditions.
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Contributors
Examiner: Hierlemann, Andreas
Examiner : Jenal, Urs
Examiner : Dehio, Christoph
Examiner : Boos, Julia A.
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Pages / Article No.
Publisher
ETH Zurich
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Subject
Microfluidics; Organoids; Transwell insert; Barrier model; Infection modelling; High-resolution imaging; Antimicrobial resistance;
Organisational unit
03684 - Hierlemann, Andreas / Hierlemann, Andreas