Dissecting the role of endogenous epithelial IL-10 using a small intestinal organoid model

Abstract

Abstract IL-10 is widely appreciated as a potent anti-inflammatory cytokine, acting on leukocytes in mucosal immunity. Far less attention has been paid to the impact of IL-10 on epithelial cells, which make up the crucial barrier interface between the host mucosa and the external environment. Furthermore, most studies look into the effects of exogenous IL-10, disregarding the possible presence and function of endogenous IL-10 in the epithelium. Using ex vivo organoids we aimed to dissect any role for endogenous epithelial IL-10. We discovered that cells in small intestinal organoids (enteroids) derived from C57BL/6 mice produced IL-10 constitutively throughout development. IL-10 mRNA increased considerably on day 3, the time when enteroids begin “budding” to create crypt-like structures. In addition, a second IL-10 increase was detected at day 6 when enteroids are typically “mature”. Immunofluorescent staining for IL-10 on day 4 enteroids was localized to Paneth cells. As gut epithelial cells are constantly exposed to bacteria, we added LPS at the start of enteroid cultures and observed that IL-10 staining was significantly increased. To further investigate the role of endogenous IL-10, we grew enteroids from IL-10 gene knockout mice. IL-10 deficient enteroids develop to morphologically resemble wild-type enteroids. We are proceeding to examine any difference in permeability, cell lineages and apoptosis between the two enteroid genotypes. In conclusion, IL-10 is present in the developing small intestinal epithelium and while development is IL-10-independent, whether IL-10 impacts subcellular components is under investigation.