Mucosally induced immunological tolerance is an attractive strategy for preventing or treating illnesses resulting from untoward inf lammatory immune reactions against self-or non-self-antigens. Oral administration of relevant autoantigens and allergens has been reported to delay or suppress onset of clinical disease in a number of experimental autoimmune and allergic disorders. However, the approach often requires repeated feeding of large amounts of tolerogens over long periods and is only partly effective in animals already systemically sensitized to the ingested antigen such as in animals already harboring autoreactive T cells, and thus presumably also in humans with an autoimmune disease. We have recently shown that oral administration of microgram amounts of antigen coupled to cholera toxin B subunit (CTB), can effectively suppress systemic T cell reactivity in naive as well as in immune animals. We now report that feeding small amounts (2-20 g) of human insulin conjugated to CTB can effectively suppress beta cell destruction and clinical diabetes in adult nonobese diabetic (NOD) mice. The protective effect could be transferred by T cells from CTBinsulin-treated animals and was associated with reduced lesions of insulitis. Furthermore, adoptive co-transfer experiments involving injection of Thy-1,2 recipients with diabetogenic T cells from syngeneic mice and T cells from congenic Thy-1,1 mice fed with CTB-insulin demonstrated a selective recruitment of Thy-1,1 donor cells in the peripancreatic lymph nodes concomitant with reduced islet cell infiltration. These results suggest that protection against autoimmune diabetes can be achieved by feeding minute amounts of a pancreas islet cell autoantigen linked to CTB and appears to involve the selective migration and retention of protective T cells into lymphoid tissues draining the site of organ injury.

Regulatory T cells (Treg) are crucial for the maintenance of tolerance to auto-antigens and harmless exogenous antigens. Here, we studied the role of the commensal microbiota for the development and function of Treg. CD4 + CD25 + T cells were obtained from peripheral lymph nodes (PLN) and mesenteric lymph nodes (MLN) of germ-free (GF) and conventional (conv) NMRI mice and tested for phenotype and functional suppressive capacity. CD4 + CD25 + T cells from GF mice showed a lower relative gene expression of fork head box p3 gene (Foxp3) and were not as potent suppressors in vitro as CD4 + CD25 + T cells from conv animals. Intracellular staining for Foxp3 and CTLA-4 revealed proportional and regional differences in putative Treg subsets between conv and GF mice. Fewer of the CD4 + CD25 + T cells in GF MLN expressed Foxp3 and CTLA-4, while the expression of these markers was similar amongst the CD4 + CD25 + T cells in PLN of conv and GF mice. The largest difference between conv and GF Treg was observed in the liver draining celiac lymph node, where GF mice had fewer putative Treg as compared to conv mice. We propose that the presence of a microbial flora favors the development of a fully functional Treg population. IntroductionRegulatory T cells down-regulate unwanted and exaggerated immune reactions to auto-antigens as well as innocuous environmental antigens. Different subsets of Treg have been defined. Natural CD4 + CD25 + Treg are positively selected for tissue-specific self-Ag in the thymus and exert their suppressive effect by cell-cell contact-dependent mechanisms [1]. Deficiency in the Treg population or neonatal thymectomy result in multiorgan autoimmune diseases [2][3][4][5][6][7][8]. Treg are characterized by a dense surface expression of CD25 (the a-chain of the IL-2-receptor), glucocorticoidinduced TNF receptor (GITR) and intracellular CTLA-4, however, these markers may be present on activated T cells. CD4 + CD25 + Tregs appear to be unique in transcribing the fork head box p3 gene (Foxp3) transcription factor [6,9], which enable their identification either with PCR are or newly developed mAb to the Foxp3 protein. Lack of a functional Foxp3 gene result in the severe autoimmune syndrome IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome), characterized by organ-specific autoimmunity, colitis and high IgE levels [10,11]. Mice with spontaneous or induced mutations in the Foxp3 gene that impair Foxp3 expression fail to develop CD4 + CD25 + Treg and exhibit a syndrome similar to IPEX [5,6].Other subsets of Treg are induced in the periphery and are referred to as Th3 [12][13][14] and Tr1 cells [15][16][17][18][19] can also generate Treg. This implies that the state of the APC and the local cytokine milieu, e.g. in the normal gut where both IL-10 and TGF-b are abundant, can determine the outcome of an immune reaction, and that Treg are induced under these circumstances.The aim of this study was to investigate the phenotype and functionality of CD4 + CD25 + Treg from germ-free (GF) ...

Oral administration