The Regulation of Systemic Immune Responses by the Dietary Antigen Ovalbumin

Oral tolerance is the specific immunological unresponsiveness normally induced by feeding a soluble antigen. Though it is an obstacle to oral vaccination, it is probably the mechanism that prevents intestinal hypersensitivity reactions to food antigens and there is currently a great deal of interest in the manipulation of the phenomenon to provide a novel strategy for immunotherapy of autoimmune and inflammatory disorders. However, the mechanisms of oral tolerance and the major factors that influence them remain controversial and require to be clarified before practical application. The principal aim of my project was to assess the role for a number of the mechanisms that had been proposed to regulate peripheral immune responses to orally administered antigen. At the time I began my study, the effects of oral tolerance were known to be influenced by the dose of antigen and the dogma was that low feeding doses could invoke active regulatory mechanisms, while high doses could inactivate T cells directly. However, this theory was based on reports from different model systems where the findings were not consistent in every aspect and in general were restricted to comparisons of only a few antigen doses. As active regulation and direct T cell inactivation have different implications for the clinical application of oral tolerance, I attempted to clarify this issue by using the dietary antigen ovalbumin (OVA) to examine the systemic effects of feeding mice a wide range of doses (100mug-25mg). A single dose of 10-25mg fed OVA reduced every antigen-specific effector function examined after parenteral immunisation with OVA/CFA, including DTH, IgG, IgGl and IgG2a responses in vivo and PLN cell proliferation and production of IL3, IL5, ILIO and IFNgamma in vitro. That the activity associated with both Thl and Th2 cells was reduced suggests that neither T cell subset could have been involved in modulating the other thereby ruling out this mechanism of active regulation in high dose oral tolerance. Feeding OVA at doses below 10mg had different effects on individual effector functions, and although all responses were progressively reduced by increasing antigen doses, this appeared to follow a pattern generally consistent with individual T helper cell subsets. Th1-despendent DTH, IgG2a and IFNy responses were the most susceptible to inhibition and were reduced by feeding as little as 100mug OVA, while PLN cell proliferative responses became inhibited at doses ≥2mg fed OVA. The regulation of Th2-dependent responses was less clear cut. IL5 production was tolerised by as little as 100mug fed OVA, whereas IgGl and ILIO responses resisted the effects of tolerance induced by feeding 100mug, Img, 2mg and 5mg OVA, only becoming significantly inhibited at doses of 100mug fed OVA. These findings indicate that although the regulatory factors mediating IL5 production may differ from those controlling the other Th2-dependent responses, it is possible that low dose oral tolerance may be mediated by regulatory Th2 cells, particularly since IgGl and ILIO responses were preferentially activated by 100mug -1mg fed OVA. However, I was unable to detect evidence for the non-specific bystander suppression that is reported to accompany Th2-dependent crossregulation of Thl cells. Bystander suppression is induced in an antigen-specific manner but inhibits subsequent responses to unrelated antigen present in the same microenvironment as the onginal antigen. To examine for the presence of this phenomenon, I assessed the responses of cells restimulated with the antigen PPD, which had been present in the CFA used for parenteral immunisation with OVA. Under these circumstances, proliferation and cytokine production by orally tolerised cells was comparable with control responses and as this finding was irrespective of the dose of fed OVA, it suggests that bystander suppression may not have been operating in either low or high dose oral tolerance. Feeding very low doses of antigen is known to prime systemic immune responses. As this would be useful for the development of oral vaccines but potentially hazardous for the clinical application of fed antigens as immunotherapy, I also assessed this feature in my dose response study. My results showed that OVA-specific DTH, proliferation and IFNgamma responses were enhanced by 10-50?mug fed OVA, while upregulation of IgGl, IL3 and IL5 responses required feeding 50mug OVA and IgG2a antibody production was not augmented by either feeding dose. These findings suggest that individual effector responses vary in their susceptibility to upregulation by fed OVA. However, these findings were observed only in mice challenged with a suboptimal dose of OVA/CFA after feeding, indicating that the extent of oral priming may have been weak. The capacity of certain Th2-dependent functions to remain resistant to inhibition by low doses of fed antigen indicated that oral tolerance may be the result of different mechanisms depending on the dose fed antigen administered. (Abstract shortened by ProQuest.)