Our current work, based on preliminary animal studies and human genetic data, is driven by the hypothesis that insulin-degrading enzyme (IDE) regulates insulin levels, and defects in IDE can lead to a diabetic phenotype.

Several studies support the idea that IDE may play a role in insulin metabolism and the pathogenesis of the diabetic phenotype in vivo. For example, IDE has been shown to degrade receptor-bound insulin in endosomes and may, through this action, be involved in terminating insulin-receptor signaling. In one of our preliminary studies, we performed ¹²⁵I-insulin trichloroacetic acid (TCA) precipitation degradation assays on both Na₂CO₃-washed membrane fractions and cytosolic fractions of liver from IDE KO and WT animals. Both IDE KO liver membrane and cytosolic fractions showed highly significant decreases in insulin degradation compared to WT (p<0.0001 for each fraction). To determine whether the insulin degradation deficit elevated endogenous insulin levels, we quantified fasting insulin levels from 17-20 week old mice (n=8 IDE KO and 8 WT). A ~2.8-fold increase in basal insulin levels (p<0.01) was detected in the IDE KO mice. Furthermore, an intraperitoneal glucose tolerance test revealed significant hyperglycemia at 60 (p<0.01), 90 (p<0.01), and 120 (p<0.05) minutes after glucose injection in the IDE KO mice (n=6 IDE KO and 4 WT males at 28-34 weeks). Thus, the preliminary data suggest that at least at one time point (~5-7 months), the IDE KO mice show basal hyperinsulinemia and impaired glucose tolerance, both hallmarks of type 2 diabetes mellitus.

The aim of our current study is to precisely define the metabolic phenotype of the IDE KO mice, and, in a second phase, determine the mechanisms underlying the metabolic phenotype resulting form IDE hypofunction.

All experimental animals were generated by synchronous crossing of a cohort of mice heterozygous for the IDE KO allele, which eliminated differential effects of maternal genotype on offspring metabolic phenotype. In a first cohort of mice, male and female KO and WT littermates were randomly have been fed standard chow (6% fat). In a second cohort, a high-fat diet (HFD) has been administered to female IDE KO, het or WT mice. Both groups undergo the same experimental procedures.

To precisely define the time of onset and natural history of hyperinsulinemia and insulin resistance, we have been collecting fasting blood serum at 4-week intervals starting at 4 weeks of. Additionally, we have been measuring blood glucose levels at 4 week- intervals, as well as body weight and food intake at weekly intervals.

Glucose tolerance tests (GTT), performed on the chow-diet mice at 5 months of age, revealed glucose intolerance in both sexes (p < 0.005 by repeated measures ANOVA for both males and females). 

Insulin tolerance tests (ITT) were performed on the chow-diet animals at 4.5 months of age.  Both the male and female IDE KO animals showed an interesting pattern.  There was an exaggerated drop in blood glucose at 30 and 60 minutes after the insulin injections, then at 120 minutes, the IDE KO mice showed a dramatic increase in blood glucose relative to the WT controls.

Next, we plan to measure insulin, glucagon, amylin, adiponectin, resistin, TNFa in the blood sera we collected from our mice.

Future directions for this study are more mechanistic studies to discover the underlying cause of the metabolic phenotype in our mice. The nature of these studies will be determined by the abnormal phenotypes identified in the longitudinal studies. For example, if, as suggested by the preliminary evidence, the IDE KO mice are hyperinsulinemic, we will measure serum C-peptide in the serial blood samples in order to evaluate the contribution of insulin production versus insulin clearance in generating the hyperinsulinemia. If the hyperinsulinemia is secondary to decreased clearance, we will determine whether it is entirely attributable to decreased peripheral degradation, or results of decreased intracellular degradation of newly generated insulin in the pancreatic cells, leading to higher insulin secretion. Similarly, if the preliminary evidence suggests that the IDE KO animals are glucose intolerant, we will investigate different possible mechanisms for insulin resistance and glucose intolerance, e.g. compensatory down-regulation of insulin receptors or factors in the insulin pathway.