Induction of high and low dose oral tolerance in isolated small bowel loops with and without peyers patches

Abstract

Oral Tolerance (OT) is defined as the systemic unresponsiveness to an antigen following administration by the oral route. While many of the effector mechanisms involved in the development of OT has been characterized, the cells and microenvironment responsible for the induction of oral tolerance remains elusive. We have previously developed a murine model consisting of loops of small bowel with or without Peyers Patches (PP). Results from this model indicate that intestinal epithehal cells may play a direct role in the induction of 02. In the present study we used OVA-peptide 323-339 to investigate the induction of high and low dose tolerance in loops with and without PP. Tolerance was induced by administration of OVA-peptide into the loops for 5 days, followed by immunization with OVA-peptide in Complete Freunds Adjuvant and subsequent restimulation of spleen and lymph node cells in vitro. Administration of 0.5 mg OVA-peptide into the ligated loops resulted in the induction of high dose tolerance, characterized by reduced T-cell proliferation as well as reduced IFN-gamma, IL-2, IL-4 and IL-10 production. Furthermore, at this dose, tolerance could not be transferred from fed mice to naive mice, indicating that regulatory cells were not generated. There was no significant difference in the degree of tolerance induced in loops containing PP compared to loops without PP. This demonstrates that induction of htgh dose tolerance is independent of PP, We then attempted to induce low dose tolerance in loops containing PP using 0.01 mg OVA peptide. This protocol resulted in both reduced proliferation and reduced production of IFN-gamma, IL-2, IL-4 and ILl0. However, in this ease spleen cells from fed mice were both able to transfer tolerance to naive mice and to downregulate the proliferation of cells from nonfed mice in vitro, indicating the generation of regulatory cells. In conclusion, this study demonstrates that both high and low dose tolerance can be induced in small bowel loops. High dose tolerance is independent of PP, indicating that intestinal epithelial cells may play a role in the induction of this type of tolerance in vivo,