NF-kappa-B activation unveils the presence of inflammatory hotspots in human gut xenografts

Abstract

The single-epithelial cell layer of the gut mucosa serves as an essential barrier between the host and luminal microflora and plays a major role in innate immunity against invading pathogens. Nuclear factor kB (NF-κB), a central component of the cellular signaling machinery, regulates immune response and inflammation. NF-κB proteins are activated by signaling pathways downstream to microbial recognition receptors and cytokines receptors. Highly regulated NF-κB activity in intestinal epithelial cells (IEC) is essential for normal gut homeostasis; dysregulated activity has been linked to a number of disease states, including inflammatory bowel diseases (IBD) such as Crohn’s Disease (CD). Our aim was to visualize and quantify spatial and temporal dynamics of NF-κB activity in steady state and inflamed human gut. Lentivirus technology was used to transduce the IEC of human gut xenografts in SCID mice with a NF-κB luminescence reporter system. NF-κB signaling was visualized and quantified using low resolution, intravital imaging of the whole body and high resolution, immunofluorescence microscopic imaging of the tissues. We show that NF-κB is activated in select subset of IEC with low “leaky” NF-κB activity. These unique inflammatory epithelial cells are clustered in the gut into discrete hotspots of NF-κB activity that are visible in steady state and selectively activated by systemic LPS and human TNFα or luminal bacteria. The presence of inflammatory hotspots in the normal and inflamed gut might explain the patchy mucosal lesions characterizing CD and thus could have important implications for diagnosis and therapy.

Hyperinflammation evokes different antiviral TMPRSS2 and ADAM17 expression responses in human gut xenograft versus host mouse gut although overall genomic responses are similar

“…We have utilized an experimental platform, first reported by Winter et al in 1991 (13) and refined in our laboratories for study of enteric nervous system (14,15), human-specific pathogens (16)(17)(18) and IBD (19)(20)(21)(22). Fetal gut is obtained from pregnancy terminations performed legally at 12-18 weeks gestational age and transplanted subcutaneously in mature SCID mice, where it grows and can be experimentally manipulated over the course of the subsequent several months.…”