The Role Of Small Intestinal Intraepithelial Lymphocytes (iIELs) In Resistance To Salmonella Typhimurium Infection

ObjectThe intestinal immune system is crucial for the maintenance of mucosal homeostasis and has evolved under the dual pressure of protecting the host from pathogenic infection and co-existence with the dense and diverse commensal organisms in the lumen. Intestinal intraepithelial lymphocytes (iIELs) are T cells that reside in the intestinal epithelium, forming a highly specialized lymphoid compartment. These cells are considered to play an important role in the regulation of mucosal immune responses and are available to respond to oral infection by pathogens.This study aimed to explore the role and possible mechanism of the intestinal immune system in surveillance of pathogenic infection and maintenance of tolerance to normal commensal flora. A model of Salmonella infection of the intestine by oral infection with the virulent S. typhimurium strain was established for observation of changes in the frequency of small intestinal IEL subpopulations, associated NK-like cytotoxicity and the expression of effector molecules. Immune responses following administration of E.coli, a component of normal commensal flora, were also examined. This study was designed to identify the subsets of iIELs important in defense against pathogenic infection. Such information is beneficial in gaining an understanding of how immune responses and immunopathologies develop during intestinal infection.Methods1. C57BL/6 mice were orally infected with S. typhimurium or E. coli2. The cytolytic activity of iIELs against YAC-1 cells, MCA-38 cells and IECs was assessed by the MTT assay.3. Changes in receptor-related gene expression in iIELs were detected by RT-PCR and real-time PCR was used to investigate the effects of S. typhimurium infection on the gene expression of Qa-1 and all known NKG2D ligands (Rael, H60 and MULT1)byIECs.4. Small intestinal bacterial loads were detected after blockade of NKG2D in vivo.5. Flow cytometry was used to examine the changes in iIELs subpopulations and IECs during S. typhimurium infection.6. CD8+TCRγδ+T cells were enriched (>90%purity) from the iIELs by MACS. The cytolytic activity of purified CD8+TCRγδ+ iIELs against IECs after NKG2D or Qa-1b blockade was assessed by the MTT assay.7. Small intestinal and systemic bacterial loads were detected after anti-γδTCR Ab treatment.Results1. S. typhimurium infection induces mouse enteritis.2. S. typhimurium infection augments the proportion of CD8+TCRγδ+ iIELs and enhances the NK-like cytotoxicity of iIELs.3. NKG2D expression on CD8+TCRγδ+ iIELs is upregulated after S. typhimurium infection.4. Expression of MULT1 on IECs is upregulated by S. typhimurium infection.5. Blockade of NKG2D in vivo accelerates bacterial growth in small intestine following oral infection with S.typhimurium.6. Anti-γδTCR mAb treated mice display accelerated bacterial growth following oral infection with S. typhimurium.7. Expression of Qa-1b on IECs is downregulated by S. typhimurium infection.8. Mouse IELs maintain tolerance to commensal microbes.9. Nod1 may participate in the recognition of S. typhimurium by IECs.Conclusion1. iIELs in the small intestine, particularly CD8+TCRγδ+ iIELs, are critical for the detection of pathogenic bacteria during S. typhimurium infection.2. Infection by S. typhimurium alters the homeostatic balance regulating activation and cytolytic activity of iIELs, particularly CD8+TCRγδ+iIELs. The data presented here suggest that this occurs by induction of MULT1 expression and inhibition of Qa-lb on IECs, in addition to enhanced expression of NKG2D on iIELs.3. NKG2D recognition is involved in the cytotoxicity of CD8+TCRγδ+iIELs against S. typhimurium-infected IECs.4. The high expression of Qa-lb on IECs may be involved in maintaining tolerance to normal IECs by iIELs.