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Insulin antibodies and T-cell lymphocyte cytotoxic reactivity against insulin and its related peptides were studied longitudinally in 3 groups of patients with type I diabetes mellitus (DM). Group 1 patients were those in whom the diagnosis was made within 1 week of the initiation diagnosis. They were subdivided into those receiving MC porcine (A) or MC bovine (B) insulin. Group 2 patients were those with a duration of DM for 2-6 years who were receiving either MC porcine (A) or MC bovine (B) insulins. Group 3 subjects were those who had been on conventional recrystallized insulin and then switched to MC porcine (A) or MC bovine (B) insulins for 2 weeks before the start of the study. The incidence of cytotoxic reactions and insulin antibodies were approximately 40-50% for group 1 (either 1A or 1B) at the initiation of the study. At 3-month follow up all patients in group 1B developed insulin antibodies (p less than 0.02) and a significant increase in the frequency of cytotoxic reactions (p less than 0.01). By contrast there was a decline in the frequency of cytotoxic reactions in group 1A (p less than 0.01 at 1 year) and the increase in insulin antibodies was non-significant. Group 2B had higher frequency in cytotoxic reactions (p less than 0.005) and of insulin antibodies (p less than 0.05) than group 2A. A significant decrease (p less than 0.01) in cytotoxic reactions was observed at 3 months following the switch of patients from conventional bovine insulin preparations to 'A' but not to 'B'. However in both subgroups insulin antibodies persisted for at least 12 months. Cross-reactivity between antibodies to human, porcine and bovine insulins was evident in all groups. The early cellular and humoral immune phenomena were positively correlated in both group 1A and 1B suggesting their common involvement in the pathogenesis of DM.

Using a cellular cytotoxicity assay, we have investigated the antigenicity of pharmaceutical insulins and of highly purified insulin constituents (unformulated monocomponent (MC) insulin, insulin B-component and C-peptide) in control, insulin naive, subjects and in three groups of type I diabetic patients. These were: Group 1, newly diagnosed patients receiving either porcine or bovine MC insulins for less than one week; Group 2, established patients receiving porcine or bovine MC insulin for 2-6 years; Group 3, established patients receiving conventional bovine insulin for 2-6 years, and tested within two weeks of switching to either porcine or bovine MC insulins. In control subjects, there was a higher incidence of cytotoxic reactions with beef (72%) than with porcine (33%) pharmaceutical preparations (p less than 0.01) but there was a similar incidence of reactions with the purified beef and pork constituents. All patients, except those Group 3 patients receiving bovine MC insulins, had a significantly increased incidence of aggregate reactions to the spectrum of antigens. In porcine treated patients, there was an increase in the incidence of reactions to pharmaceutical preparations in Group 1 (p less than 0.05) and Group 3 (p less than 0.001), but this was absent from each of the bovine treated groups. All patient groups showed significant increases in the incidence of reactions against insulin constituents of their therapeutic analog. In Groups 1 and 2, but not 3, there was significant analog cross reactivity. We deduce that the reactivity seen in control subjects is principally directed against unidentified formulation constituents of bovine pharmaceutical preparations. Patients on both acute and chronic therapy with insulin show T-cell sensitization to MC insulin and its components. Chronic therapy with conventional bovine insulin induces tolerogenesis, but this is reversed on exposure to a fresh insulin analog.