RESUMO
Introduction: Life on Earth depends on oxygen; human tissues require oxygen signaling, whereas many microorganisms, including bacteria, thrive in anoxic environments. Despite these differences, human tissues and bacteria coexist in close proximity to each other such as in the intestine. How oxygen governs intestinal-bacterial interactions remains poorly understood. Methods: To address to this gap, we created a dual-oxygen environment in a microfluidic device to study the role of oxygen in regulating the regulation of intestinal enzymes and proteins by gut bacteria. Two-layer microfluidic devices were designed using a fluid transport model and fabricated using soft lithography. An oxygen-sensitive material was integrated to determine the oxygen levels. The intestinal cells were cultured in the upper chamber of the device. The cells were differentiated, upon which bacterial strains, a facultative anaerobe, Escherichia coli Nissle 1917, and an obligate anaerobe, Bifidobacterium Adolescentis, were cultured with the intestinal cells. Results: The microfluidic device successfully established a dual-oxygen environment. Of particular importance in our findings was that both strains significantly upregulated mucin proteins and modulated several intestinal transporters and transcription factors but only under the anoxic-oxic oxygen gradient, thus providing evidence of the role of oxygen on bacterial-epithelial signaling. Conclusions: Our work that integrates cell and molecular biology with bioengineering presents a novel strategy to engineer an accessible experimental system to provide tailored oxygenated environments. The work could provide new avenues to study intestine-microbiome signaling and intestinal tissue engineering, as well as a novel perspective on the indirect effects of gut bacteria on tissues including tumors. Supplementary Information: The online version contains supplementary material available at 10.1007/s12195-022-00735-x.
RESUMO
The intestinal environment is unique because it supports the intestinal epithelial cells under a normal oxygen environment and the microbiota under an anoxic environment. Due to importance of understanding the interactions between the epithelium and the microbiota, there is a strong need for developing representative and simple experimental models. Current approaches do not capture the partitioned oxygen environment, require external anaerobic chambers, or are complex. Another major limitation is that with the solutions that can mimic this oxygen environment, the oxygenation level of the epithelial cells is not known, raising the question whether the cells are hypoxic or not. We report standalone microfluidic devices that form a partitioned oxygen environment without the use of an external anaerobic chamber or oxygen scavengers to coculture intestinal epithelial and bacterial cells. By changing the thickness of the device cover, the oxygen tension in the chamber was modulated. We verified the oxygen levels using several tests: microscale oxygen sensitive sensors which were integrated within the devices, immunostaining of Caco-2 cells to determine hypoxia levels, and genetically encoded bacteria to visualize the growth. Collectively, these methods monitored oxygen concentrations in the devices more comprehensively than previous reports and allowed for control of oxygen tension to match the requirements of both intestinal cells and anaerobic bacteria. Our experimental model is supported by the mathematical model that considered diffusion of oxygen into the top chamber. This allowed us to experimentally determine the oxygen consumption rate of the intestinal epithelial cells under perfusion.
