Abstract
SummaryHuman enteroids—epithelial spheroids derived from primary gastrointestinal tissue—are a promising model to study pathogen-epithelial interactions. However, accessing the apical enteroid surface is challenging because it is enclosed within the spheroid. We developed a technique to reverse enteroid polarity such that the apical surface everts to face the media. Apical-out enteroids maintain proper polarity and barrier function, differentiate into the major intestinal epithelial cell (IEC) types, and exhibit polarized absorption of nutrients. We used this model to study host-pathogen interactions and identified distinct polarity-specific patterns of infection by invasive enteropathogens. Salmonella enterica serovar Typhimurium targets IEC apical surfaces for invasion via cytoskeletal rearrangements, and Listeria monocytogenes, which binds to basolateral receptors, invade apical surfaces at sites of cell extrusion. Despite different modes of entry, both pathogens exit the epithelium within apically extruding enteroid cells. This model will enable further examination of IECs in health and disease.
Highlights
The intestinal epithelium is the barrier that mediates interactions between the intestinal lumen and the rest of the body
By manipulating extracellular matrix (ECM) proteins in the culture system, we successfully produced apical-out enteroids, which maintain their ability to differentiate to the various intestinal epithelial cell (IEC) lineages, maintain proper polarity and barrier function, and are able to absorb nutrients in a polarity-specific manner
We have previously reported in polarized Madin-Darby canine kidney (MDCK) monolayers, in rabbit intestinal loop infections (Pentecost et al, 2006), and in a murine model of oral infection (Pentecost et al, 2010) that L. monocytogenes entry at the apical epithelial surface is restricted to sites of cell extrusion where cell polarity is transiently disrupted
Summary
The intestinal epithelium is the barrier that mediates interactions between the intestinal lumen and the rest of the body. Proper intestinal function depends on the development and maintenance of organization of epithelial cells into two distinct compartments, the apical and basolateral epithelial regions. The apical surface of intestinal epithelial cells (IECs) faces the lumen and regulates interactions with lumenal contents. The apical surface mediates nutrient absorption, detects microbial products, and secretes molecules that protect the epithelium from potentially harmful agents in the lumen. The basolateral surface anchors epithelial cells to the underlying basement membrane, delivers nutrients from the lumen to the bloodstream, and communicates with nearby cells. In addition to proper polarity, intestinal epithelial function relies on differentiation into the various IEC types, which each carry out specific processes. Transformed cell lines have enabled investigation of epithelial cells in vitro; these cancerderived cultures do not differentiate into the various IEC types and often do not develop proper polarity. Animal models have provided valuable insights to intestinal function and disorders, these systems often do not recapitulate human-specific infections and diseases
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