Engineering Flexible Electronics for Monitoring Epithelial Integrity In-Vitro and Ex-Vivo

Inflammatory bowel disease (IBD) is a widespread condition, with a forecasted prevalence of 1.1% by 2025 [1]. It presents with chronic symptoms such as abdominal pain and diarrhoea and is characterised by a leaky gut barrier, which has increased permeability due to changes in localisation of tight junction proteins [2]. However, a detailed understanding of the disease pathway is limited, and current methods of assessing gut tissue health<i> ex vivo</i> are impeding progress in the field. The techniques need re-designing in the modern era given advancements in technology. This work presents flexible bioelectronic devices that permit real-time recordings of changes in gut tissue. The ability to assess gut health in living tissue is important in establishing and monitoring phenotypes associated with gastrointestinal diseases.<br/><br/>All-planar conformable devices with multiple working electrode sizes (from 25 to 1000 µm) and an internal counter/reference electrode (CE/RE) were fabricated using previously developed techniques [3]. On the bottom, and between each of the layers, parylene C was used for insulation. Conductive tracks, outlines, and contact pads for the electrodes were formed using photolithography and gold deposition. Electrodes were coated with PEDOT:PSS, a conducting polymer used to increase volumetric capacitance. The electrodes were characterised by electrochemical impedance spectroscopy (EIS) and brightfield microscopy. <i>In-vitro</i> testing utilized a human colonic epithelial cell line, Caco-2. The cells were grown on Transwell® inserts and differentiated for 21 days. The ability of the device to measure barrier integrity, perturbations, and recovery in real-time was assessed via the transient addition of EGTA. The devices were tested e<i>x-vivo </i>on small intestine and colonic tissue from wild type-C57BI6 male mice. Cell viability was confirmed with confocal microscopy and staining. Extracted transepithelial resistance values, based on simple equivalent circuit models, were confirmed against the gold standard methods for <i>in-vitro</i> and <i>ex-vivo</i>, using the EVOM3™ and Ussing Chambers respectively.<br/><br/>This work highlights the first instance of an all-planar flexible electronic device that can monitor epithelial barrier function and perturbations at the air liquid interface. The mechanical flexibility of the device allows for direct and conformable contact with biological surfaces, to provide a quantifiable electrical readout under physiological conditions. The multi-electrode design established a minimum electrode radius (400 µm) needed to capture the epithelial properties, and chemical disruption, of colon cell lines and mouse colon tissue. The sensing area achieved is one order of magnitude smaller than previously reported, highlighting the increased sensing efficiency of the design. The device achieves comparable sensitivity with a planar CE/RE compared to an external CE/RE. As this device was the first of its kind, readouts were validated against gold standard TEER measurements, which showed comparable trends with a reduction in measurement error. We confirm the device is biocompatible and does not negatively affect the barrier properties under study. Future work could look to apply this technology for spatially mapping local and real-time changes in epithelial barrier properties for toxicology and drug screening applications.<br/><br/>[1]. Freeman, K. et al. BMC Gastroenterology 21, 1–7 (2021).<br/>[2]. Michielan, A. & D’Inc`a, R. Mediators of inflammation (2015).<br/>[3]. Khodagholy, D. et al. Advanced Materials 23, H268–H272 (2011)