To define the mechanisms involved in the evolution of diabetes in the Zucker diabetic fatty (ZDF) rat, beta-cell mass and replication rates were determined by immunochemistry, point-counting morphometry, and 6-h 5-bromo-2'-deoxyuridine (BrdU) incorporation. The beta-cell mass in 5- to 7-week-old prediabetic ZDF rats (4.3 +/- 0.06 mg) was similar to age-matched insulin-resistant Zucker fatty (ZF) rats (3.7 +/- 0.05 mg) and greater than that in Zucker lean control (ZLC) rats (1.9 +/- 0.3, P < 0.05). At 12 weeks (after diabetes onset), beta-cell mass in the ZDF rats (8.1 +/- 1.7 mg) was significantly lower than the ZF rats (15.7 +/- 1.8 mg). The mass in the ZF rats was significantly greater than in the ZLC rats (4.3 +/- 0.8 mg, P < 0.05). The beta-cell proliferation rate (mean of both time points) was significantly greater in the ZDF rats (0.88 +/- 0.1%) compared with the ZF and ZLC rats (0.53 +/- 0.07%, 0.62 +/- 0.07%, respectively, P < 0.05), yet ZDF rats have a lower beta-cell mass than the ZF rats despite a higher proliferative rate. Morphological evidence of neogenesis and apoptosis is evident in the ZF and ZDF rats. In addition, even at 5-7 weeks a modest defect in insulin secretion per beta-cell unit was found by pancreas perfusion. These studies provide evidence that the expansion of beta-cell mass in response to insulin resistance and insulin secretory defects in diabetic ZDF rats is inadequate. This failure of beta-cell mass expansion in the ZDF rat does not appear to be from a reduction in the rate of beta-cell proliferation or neogenesis, suggesting an increased rate of cell death by apoptosis.

Mutations in the gene for the transcription factor hepatocyte nuclear factor (HNF) 1 ␣ cause maturity-onset diabetes of the young (MODY) 3, a form of diabetes that results from defects in insulin secretion. Since the nature of these defects has not been defined, we compared insulin secretory function in heterozygous

Besides the genetic framework, there are two critical requirements for the development of tissue-specific autoimmune diseases. First, autoreactive T cells need to escape thymic negative selection. Second, they need to find suitable conditions for autoantigen presentation and activation in the target tissue. We show here that these two conditions are fulfilled in diabetic NOD mice. A set of autoreactive CD4+ T cells specific for an insulin peptide, with the noteworthy feature of not recognizing the insulin protein when processed by the antigen presenting cells (APC) escape thymic control, participate in diabetes and can cause disease. We also find that APCs situated in close contact with the beta cells in the islets of Langerhans bear vesicles with the antigenic insulin peptides and activate the peptide-specific T cells. These findings may be relevant for other cases of endocrine autoimmunity.

To determine whether loss of beta-cell mass and function in the NOD mouse occurs gradually, beginning after the onset of insulitis, or abruptly, just before the onset of overt diabetes, beta-cell mass and rates of beta-cell proliferation and insulin secretory responses from the perfused pancreas were measured in NOD and control NOD/Scid mice at 8-9, 13, and 18 weeks of age. Of the NOD mice, 11 and 70% had diabetes (fasting blood glucose >8.3 mmol/l) at 13 and 18 weeks of age, respectively. Beta-cell mass in 8-week-old NOD mice was 69% of control mice (P>0.05), but the rate of 5-bromo-2-deoxyuridine uptake was greater, suggesting a compensatory proliferative response to ongoing autoimmune beta-cell destruction. Despite an increase in the rate of beta-cell proliferation, beta-cell mass was significantly reduced by 42% in 13-week-old nondiabetic NOD mice and by 73% in 18-week-old diabetic NOD mice. Insulin secretory responses to glucose and arginine demonstrated reductions of similar magnitude. In 18-week-old diabetic NOD mice, insulin secretion was reduced to a greater degree than beta-cell mass, suggesting the presence of beta-cell dysfunction in addition to reduced mass. These results suggest that in the NOD mouse, beta-cell destruction begins soon after the onset of insulitis. Despite a compensatory beta-cell proliferative response, beta-cell mass progressively falls and is significantly reduced by 13 weeks despite normal blood glucose concentrations. Diabetes may be present when residual beta-cell mass represents 30% of control levels.