Hydrogel incorporated microfluidic device for in vitro culture and dynamic observation of human intestinal epithelial cells

Abstract

To complement limitations of the traditional methods for intestinal microbiota researches, a biomimetic in vitro model is demanded. Microfluidic models have been developed to meet such demand, but even they had to be sliced into thin sectional samples in order to observe microbiota environment such as epithelial monolayer, mucus and microorganisms. This study aims to develop a thin-section, microfluidic model allowing dynamic, simultaneous observation of microbiota environment. 20 μm thin collagen scaffold was constructed inside the microfluidic device as a barrier separating apical from basal flow and extracellular matrix (ECM) for epithelial culture. The collagen scaffold successfully allowed adherence of epithelial culture consisted of HT-29 intestinal cell line. Seeding density and cell incubation time suitable for covering the collagen scaffold with monolayer culture of epithelial cells were determined. HT-29 cells were found to secrete more mucus, which provides microbiota niche, when adhered to collagen. By quantifying mucus secretion of the HT-29 cells cultured inside the proposed device, the increased mucus secretion was found to be more dependent on cell-ECM adhesion than cell-cell adhesion. Dynamic, simultaneous observation of the collagen scaffold, the cell culture and green fluorescent protein (GFP)-expressing bacteria on single focal plane was demonstrated using the microfluidic device. During three days of cultivation, HT-29 cells displayed excessive proliferation, requiring inhibition of their tumorous phenotype. The proposed device offers a capability for live imaging of intestinal microbiota and its surroundings. Findings of this thesis study will be implemented to improve the device as truly biomimetic modeling of human intestinal microbiota.