Objectives: The aim of the study is to create assays able to detect the beginning of islet cell specific cellular immune response, the destructive capacity of this response and its natural development as well as the possible effect of intranasal insulin treatment on the direction of this response.
Basis: The proposal is based on the ongoing Finnish Diabetes Prediction and Prevention project identifying a large number of study subjects who are in various stages of prediabetes and have in part been recruited to a secondary prevention trial testing the effect of intranasal insulin treatment.
Rationale: It is known that a long prediabetic period exists during which antibodies to several diabetes-associated autoantigens can be detected. Insulin is the only well characterised antigens which is beta-cell specific and autoantibodies to insulin are usually the first to appear. In addition, there are several other autoantigens, GAD65 and IA-2 being the best characterised. Autoimmunity as such does not necessarily lead to beta-cell destruction and clinical disease, but the balance of immune response between cytotoxic/destructive Th1 and protective/tolerance promoting Th2/Th3 type of responsiveness is important as supported also by animal models. It is thus conceivable that assays able to discern the type of ongoing autoimmunity could be developed, and one might, by tolerazing treatment, affect the direction of the response.
Significance: Reliable assays able to show the ongoing beta-cell destruction in early phase would allow the targeting of preventive treatment to subjects who without treatment would in most cases develop the clinical disease. The developed assays might also be able to monitor the efficiency of tolerization treatment. This will be tested in an associated intranasal treatment trial.
Protocol Summary: The protocols tested have included stimulation of peripheral blood lymphocytes with insulin and detection of T-cell responses using production of intracellular and secreted cytokines representing Th1, Th2 and Th3 type of responsiveness. Flow cytometry and quantitative RT-PCR have been used for the quantitation of the responses. The precursor frequency of cells recognising dominant insulin and GAD65 epitopes have also been measured using tetramer technology. Microarray analysis of gene expression in peripheral blood leukocytes has been used to identify changes in expression levels associated with development of the clinical disease. Additional markers like autoantigen-specific class I tetramers are under consideration.