Dietary Insulin and the Gut Immune System in Type 1 Diabetes

.........................................................................................................................................................7 LIST OF ORIGINAL PUBLICATIONS ................................................................................................................9 ABBREVIATIONS .............................................................................................................................................10 INTRODUCTION...............................................................................................................................................12 REVIEW OF THE LITERATURE ......................................................................................................................13 TYPE 1 DIABETES ........................................................................................................................................13 Epidemiology...............................................................................................................................................13 Pathogenesis ................................................................................................................................................14 Genetic background.................................................................................................................................................. 14 Autoimmunity.......................................................................................................................................................... 18 Description of autoimmune diabetes ..................................................................................................................... 18 Cell-mediated immunity....................................................................................................................................... 18 Th1/Th2 paradigm................................................................................................................................................ 20 Autoantigens and autoantibodies........................................................................................................................... 22 ICA ................................................................................................................................................................. 22 Insulin ............................................................................................................................................................. 23 Glutamic acid decarboxylase (GAD)................................................................................................................. 24 Protein tyrosine phosphatase-like islet cell antigens IA-2 and IA-2β .................................................................. 26 Environmental factors............................................................................................................................................... 27 Dietary factors ..................................................................................................................................................... 27 Cow`s milk proteins ......................................................................................................................................... 27 Plant proteins ................................................................................................................................................... 30 Nitrates and nitrites .......................................................................................................................................... 30 Vitamin D deficiency ....................................................................................................................................... 30 Viral infections .................................................................................................................................................... 31 ADHESION MOLECULES.............................................................................................................................33 GUT IMMUNITY AND ORAL TOLERANCE ...............................................................................................37 Gut immune system......................................................................................................................................37 Structure and function .............................................................................................................................................. 37 Oral tolerance........................................................................................................................................................... 38 The gut immune system, oral tolerance, and type 1 diabetes..........................................................................41 The gut immune system and oral tolerance in other autoimmune diseases and in food allergy........................43 AIMS OF THE STUDY ......................................................................................................................................45 SUBJECTS AND METHODS .............................................................................................................................46 Subjects .......................................................................................................................................................46 Antigens ......................................................................................................................................................49 Assay for measurement of soluble ICAM-1 and L-selectin............................................................................50 T-cell proliferation assay..............................................................................................................................50 Depletion of α4β7-expressing lymphocyte population from PBMCs ............................................................51 Depletion of CD8 lymphocytes from the PBMC population ..........................................................................51 Flow-cytometry analysis of lymphocyte surface antigens ..............................................................................52 Immunoblotting of BI in study formulas .......................................................................................................52 Radioimmunoassay (RIA) for detection of insulin in CM products ................................................................52 Enzyme immunoassay (EIA) for IgG antibodies to BI and HI .......................................................................53 IAA radioligand assay..................................................................................................................................53 EIA for IgG to antibodies to BLG.................................................................................................................53 HLA typing..................................................................................................................................................54 Statistical analysis........................................................................................................................................54 RESULTS ...........................................................................................................................................................55 Soluble adhesion molecules in infants ..........................................................................................................55 Cellular response after depletion of α4β7-expressing PBMCs.......................................................................56 Cellular response to GAD65 after depletion of CD8+ lymphocytes ...............................................................58 Presence of insulin in study formulas............................................................................................................59 Insulin-binding antibodies to BI and HI ........................................................................................................59 Insulin-binding antibodies in children at risk for type 1 diabetes ................................................................................ 59 Insulin-binding antibodies in children at increased genetic risk for type 1 diabetes...................................................... 61 Insulin-binding antibodies in children at risk for allergy............................................................................................. 63 Cellular immune responses to insulin............................................................................................................65 Effect of maternal type 1 diabetes on immunization to dietary insulin ...........................................................68