A novel human enteroid-anaerobe co-culture system to study microbial-host interaction under physiological hypoxia

Abstract

Mechanistic investigations of host-microbe interactions in the human gut are limited by current coculture model systems. The intestinal epithelium requires oxygen for viability, while gut bacteria are facultative or obligate anaerobes. The ability to model host-commensal interactions under dynamic oxygen conditions is critical to understanding host-pathogen interactions in the human gut. Here, we demonstrate a simple, cost-effective method for co-culturing obligate anaerobic bacteria with human intestinal enteroid monolayers under variable oxygen conditions. The Enteroid-Anaerobe Co-Culture (EACC) system is able to recapitulate the steep oxygen gradient seen in vivo and induce expression of hypoxia-associated phenotypes such as increased barrier integrity and expression of antimicrobial peptide genes. Using clinical strains of the commensal anaerobes Bacteroides thetaiotaomicron and Blautia sp. on established patient-derived intestinal enteroid cell lines under physiological hypoxia, the EACC system can sustain host-anaerobe interactions for at least 24 hours. Following co-culture with anaerobic bacteria, we demonstrate patient-specific differences in epithelial response, reinforcing the potential to develop a personalized medicine approach to bacteriotherapy and host-microbe interaction investigations. Our innovative EACC system provides a robust model for investigating host-microbe interactions in complex, patient-derived intestinal tissues, that facilitates study of mechanisms underlying the role of the microbiome in health and disease.

In VitroModels of the Small Intestine: Engineering Challenges and Engineering Solutions

“…Other devices have also established coculture of anaerobic bacteria and epithelial monolayers, including a microfluidic device 143 and a system that establishes an oxygen gradient across each Transwell in an array. 144 These devices share this design element of two chambers separated by a porous membrane, because most intestinal device development has been focused on transport of nutrients, drugs, and infectious agents between the apical and basolateral sides of the intestinal epithelium. Further design innovations to recapitulate physiological microstructure, signaling gradients, and mechanical stimuli could advance research in other…”

“…Larger devices also make it easier to co-culture intestinal cells with other cell types and bacteria and to establish oxygen gradients. 144 Moreover, in cases where the component of interest is produced in small quantities, having a larger device that contains more cells could be a useful strategy.…”

Stem cell‐derived enteroid cultures as a tool for dissecting host‐parasite interactions in the small intestinal epithelium

“…These conflicting studies demonstrate that different isolates of commensal bacteria can have opposing actions against Cryptosporidium infection, and that the specific effects of different bacterial isolates deserves further study. In this respect, recently described enteroid‐anaerobe co‐culture models, which maintain a physiologically relevant oxygen gradient across the epithelial layer, would prove particularly useful 97 …”

Organoid-derived adult human colonic epithelium responds to co-culture with a probiotic strain ofBifidobacterium longum

“…Organoid-derived human colonic epithelia retain the genetic and epigenetic state of the tissue from which it was derived (9). This model was recently extended to allow for co-culture of the human epithelium with anaerobic microbes typically found within the human gut (10). To establish the physiological effect of a common anaerobe in the human gut, we co-cultured a probiotic strain of Bifidobacterium longum (35624) with organoid-derived adult human colonic epithelium in asymmetric gas conditions (anoxic apically, 5% oxygen basolaterally) and compared its behavior to axenic ('germ-free') epithelium.…”