Modelling of Live Bacteria or Ligand-Induced Acute and Chronic Inflammation in the Gut using In Vitro 3D-Primary Reconstructed Human Small Intestinal Tissues (EpiIntestinal)

Abstract: Intestinal epithelium is known to be involved in innate immune responses by recognizing potential pathogens through cellular pattern recognition receptors (PRRs). The purpose of this study was to investigate PRR responses following exposure of an in vitro reconstructed 3D human small intestinal (SMI) tissue to live bacteria or various Toll-like receptor (TLRs) and Node-like receptor (NOD) ligands. The SMI tissues are cultured using human intestinal fibroblasts and enterocytes and their 3-dimensional polarity and morphology mimics that of native in vivo tissues. Characterization of the SMI tissues included evaluation of structural features, barrier properties, and expression of drug transporters and drug metabolizing enzymes. Exposure of intestinal tissues to live bacterial (commensals and pathogens) or ligands to TLR4 (LPS) and NOD2 (Muramyl dipeptide; MDP) induced gene expression of pro-inflammatory cytokines such as IFN-β, IL-1β, IL-6, and RANTES. Prolonged exposure of intestinal tissues to IL-1β also resulted in reduced membrane integrity and induction of pro-inflammatory cytokines (IL-6 and CCL20) known to stimulate acquired immune cell responses by inducing cytokine release such as IL-17, TNF-α, and IFN-Ƴ or by initiating the migration of inflammatory cells. All these responses may be precursors to IBD-like disease. To simulate the effect of acquired immune cell responses on the intestinal epithelium, we also exposed the tissues to TNF-α and IFN- Ƴ, and IL-1 β, which compromised the barrier integrity and induced the release of pro-inflammatory cytokines. The effect of TNF-α and IFN- Ƴ on the intestinal epithelium was further exacerbated if antigen-presenting cells such as dendritic cells were incorporated into the 3D intestinal tissues.

In summary, our results suggest that the EpiIntestinal tissue is capable of modelling innate immune responses and can be an useful tool to study the complex interactions of human intestinal epithelium with microbiome in vitro.



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