Role And Mechanisms Of Toll-like Receptors In Self-tolerance

Toll-like receptors (TLRs), the best-characterized receptors among the pattern recognition receptors (PRRs), recognize a diverse range of pathogen associated molecular patterns and play a critical role in antimicrobial host defense. Moreover, increasing evidences suggest the involvement of TLRs in self-tolerance and autoimmune diseases. On one hand, appropriate TLR signaling can break down the self-tolerance and promote the initiation or progression of autoimmune diseases, such as autoimmune diabetes, inflammatory bowel diseases, multiple sclerosis or systemic lupus erythematosus. On the other hand, TLR signaling has also been proposed in induction of self-tolerance and prevention of autoimmune diabetes. However, the mechanisms of how TLR signaling breaks down or promotes tolerance remain unclear. The present study aims to further investigate the role of TLR signaling in self-tolerance, especially to investigate the underlying mechanisms. The major results are shown as follow:1. TLR3 signaling induces severe small intestinal injury in mice. The mechanisms by which TLRs influence mucosal homeostasis are obscure and the objective of this part is to establish a mice model which mimics the autoimmune injury and then to investigate the underlying mechanisms. The present work reports that only poly I:C, a ligand for TLR3, induces severe small intestinal injury, and other TLR ligands have little effect. The effect of poly I:C is also confirmed by genomic double-stranded RNA (dsRNA) from rotavirus. The further experiments show that poly I.C can induce IECs to produce large amounts of IL-15 and pretreatment with anti-IL-15Rαneutralizing antibody can prevent the poly I:C-induced small intestinal injury. Moreover, IEC-derived IL-15 can enhance the cytotoxicity of IELs and induce IELs to express NK1.1. Depletion of NK1.1⁺ IELs with anti-NK.1.1 antibody can prevent the small intestinal injury, while depletion of NK cells with anti-AsGM1 antibody can not. Also, the CD3⁺NK1.1⁺ IELs are proved as CD8αα⁺ IELs and can be induced from CD3⁺NK1.1^- IELs by IEC-derived IL-15. In addition, the intestinal injury induced by poly I.C is impaired in TLR3^-/- mice. In this study, we have established a mice model which mimics the TLR signaling-induced autoimmune injury and provide the first evidence of pathogenic effects mediated by CD8αα⁺ IELs.2. NKG2D recognition mediates TLR3 signaling-induced breakdown of epithelial homeostasis in small intestine. TLRs and NK receptors are the two most important receptor families in innate immunity. Although it has been observed that TLR signaling can induce or up-regulate the expression of the ligands for stimulatory NK-receptors on monocytes or muscle cells, there is not yet a report indicating whether TLR signaling can break down self-tolerance through NK-receptors. The present work reports that TLR3 signaling by poly I:C can stimulate IECs to express retinoic acid early inducible-1 (Rael, a ligand for NKG2D), and also can induce NKG2D expression on CD8αα⁺IELs by interleukin-15 derived from TLR3-activated IECs. The blockade of interaction between NKG2D and Rael inhibits the cytotoxicity of IELs aganst IECs in a cell-cell contact dependent manner and therefore alleviates poly I:C-induced epithelial destruction and acute mucosal injury of small intestine. These results, for the first time, demonstrate that TLR signaling induces tissue injury through NKG2D pathway, suggesting that TLR signaling may break down self-tolerance via induction of abnormal expression of ligands for stimulatory NK-receptors.3. Long-term TLR3 activation promotes self-tolerance and prevents the onset of diabetes in NOD mice. TLRs play a critical role in breakdown of self-tolerance; however, increasing evidences suggest the role of TLR signaling in the induction of self-tolerance and in the prevention of onset of autoimmune diseases. The objective of this part is to investigate the mechanisms involved in the induction of self-tolerance by TLR signaling in NOD mice. The present work reports that the prevention of diabetes by poly I:C is associated with the formation of Th2-enriched environment in spleen and pancreas. The further experiments show that the prevention of diabetes and the formation of Th2-enriched environment depend on the presence of NK cells. Moreover, long-term poly I:C-treated NK cells exhibit a "NK3-like" phenotype, and are involved in the induction of Th2 bias of spleen cells in response to islet autoantigens via TGF-β dependent manner. These findings indicate that long-term TLR3 stimulation can induce NK cells to exhibit "NK3-like" phenotype and then inhibit the excessive Th1 responses.In conclusion, the present study not only provides a mice model which mimics the autoimmune injury, but also proposes detailed mechanisms through which TLR signaling breaks down or promotes the self-tolerance. These findings will contribute to our understanding of mechanisms for autoimmune diseases and provide the theoretical basis for clinical treatment.