...............................................................................................................................................................................II Dansk resumé......................................................................................................................................................................III Preface................................................................................................................................................................................ IV Abbreviation.........................................................................................................................................................................V CHAPTER 1: INTRODUCTION...............................................................................................................................................1 The immune system.........................................................................................................................................................1 Dendritic cells ..................................................................................................................................................................1 DC and innate immunity..................................................................................................................................................2 Toll-like receptors .......................................................................................................................................................3 C-type lectin receptors ................................................................................................................................................3 Interaction of DCs with other cells ..................................................................................................................................4 Interaction with T cells................................................................................................................................................4 Interaction through costimulatory molecules ..............................................................................................................7 Interaction with B cells ...............................................................................................................................................8 CHAPTER 2:DIETARY FIBERS AS IMMUNOREGULATORY COMPOUNDS IN HEALTH AND DISEASE..................................10 Introduction....................................................................................................................................................................11 Specificity of carbohydrate-binding receptors on macrophages and DCs.....................................................................12 Bioactive NSPs – Structure/function relationship .........................................................................................................13 In vitro effects of NSP extracts on macrophages and DCs............................................................................................16 Macrophages and NSP effects...................................................................................................................................17 NSPs and immune modulation in DCs......................................................................................................................19 Immunostimulatory contaminants in NSP preparations............................................................................................19 Absorption of NSPs .......................................................................................................................................................20 Effects of NSPs in animal models .................................................................................................................................22 Anti-infectious activities of NSPs .............................................................................................................................23 Anti-tumor effects of NSPs .......................................................................................................................................24 Side effects of NSPs..................................................................................................................................................25 Conclusion .....................................................................................................................................................................26 THIS THESIS.......................................................................................................................................................................27 CHAPTER 3: COMPARATIVE ANALYSIS OF A LARGE PANEL OF NON-STARCH POLYSACCHARIDES REVEALS STRUCTURES WITH SELECTIVE REGULATORY PROPERTIES IN DENDRITIC CELLS .........................................................28 Introduction....................................................................................................................................................................29 Materials and methods...................................................................................................................................................30 NSP preparations.......................................................................................................................................................30 NSP analysis..............................................................................................................................................................31 Determination of molecular weight...........................................................................................................................31 Generation and stimulation of bone marrow-derived DC.........................................................................................32 Cytokine quantification in culture supernatants........................................................................................................32 Analysis of surface molecule expression on DC.......................................................................................................33 vii Endotoxin test............................................................................................................................................................33 Statistics ....................................................................................................................................................................34 Results............................................................................................................................................................................34 The influence of NSP structure and origin on modulation of LPS-induced cytokine production ............................34 Effects of NSP per se on cytokine production in DC................................................................................................37 Up-regulation of surface molecule display on DC by specific NSP .........................................................................38 Effect of source and molecular weight of cereal β-glucans on modulatory capacity in DC.....................................40 Discussion......................................................................................................................................................................41 CHAPTER 4: β-GLUCAN-INDUCED PHENOTYPIC PROFILE OF TOLL-LIKE RECEPTOR-TRIGGERED DENDRITIC CELLS.46 Introduction....................................................................................................................................................................47 Materials and methods...................................................................................................................................................48 Cell isolation and culture...........................................................................................................................................48 Stimulation of DC .....................................................................................................................................................49 Cytokine quantification in culture supernatants........................................................................................................49 Immunostaining and flow cytometry ........................................................................................................................49 Statistical analysis .....................................................................................................................................................50 Results............................................................................................................................................................................50 Modulation of Dectin-1, DC-SIGN and Mannose receptor expression by diverse β-glucans..................................51 Distinct patterns of cytokine and chemokine modulation in TLR-triggered DC by β-glucans ................................53 Diverse regulation of CD86 and CD40 expression by β-glucans in TLR-triggered DC ..........................................56 β-glucan-induced phenotypic profile in TLR-triggered DC .....................................................................................57 Discussion......................................................................................................................................................................58 CHAPTER 5: MODIFICATION OF IN VIVO ANTIGEN-SPECIFIC IMMUNE RESPONSES BY A RECOMBINANT FUSION VARIANT OF AN ANTIGEN AND A β-GLUCAN BINDING MODULE: ADJUVANT PROPERTIES OF LICHENAN.......................62 Introduction....................................................