Fallopian Tube Organoid-Integrated Microfluidic Platform for Real-Time Imaging and Outflow Analysis

Abstract

The fallopian tube is a dynamic, hormonally regulated epithelial conduit whose luminal microenvironment governs oocyte transport, fertilisation, early embryo development, and the initiation of gynaecological diseases including ectopic pregnancy, hydrosalpinx, and serous tubal intraepithelial carcinoma (STIC), yet existing models such as 2D monolayers, intact organoids, and static Transwell systems fail to recapitulate a polarised epithelium lining a perfused lumen under physiological fluid shear while preserving native organoid secretory function. We present a robust and accessible workflow integrating mechanically opened fallopian tube organoids into a custom-designed polydimethylsiloxane (PDMS) microfluidic device, in which organoid fragments form epithelial sheets and re-establish luminal-like architecture under controlled perfusion. The focus of this study is organoid integration and the characterisation of the resulting organoid-derived epithelium, rather than establishing chip cultures from individual cell lines. As this epithelium is built from expandable organoid stock rather than primary tissue, which typically survives only a few days in culture, the approach removes a major bottleneck to establishing organ-on-chip models, and we maintain viable cultures for up to 10 days. Epithelial sheets reproducibly covered more than 90% of the channel area by day 6 (n = 5 devices) and formed continuous ZO-1-positive tight junctions. The platform supports longitudinal brightfield and fluorescence microscopy at subcellular resolution, continuous collection of conditioned flow-through for ELISA, and quantitative analysis of cell coverage and health using CellProfiler software. We characterise the organoid-derived epithelium by benchmarking against tissue culture (TC) formats, comparing nuclear morphology on-chip with conventional well culture, and showing that the damage-associated cytokine interleukin-1 alpha (IL1 alpha) was undetectable in chip outflow, as in intact organoid secretions, indicating that cell health on-chip is comparable to that of intact organoids. Computational flow modelling characterises perfusion through the device and predicts how factors secreted by healthy epithelium and by tumour cells are distributed along the channel. The feasibility of tumour epithelium co culture was explored using OVCAR3 ovarian cancer spheroids as a proof of concept model of early tumour interaction with the tubal epithelium. Together, these results establish the technical feasibility of the platform and support its further development as an accessible system for studying reproductive health conditions affecting the fallopian tube, including tubal damage from infection, endometriosis, STIC lesions, and occlusion.