Mouse Anti-Human INSR Hybridoma [EJJ 44.2]

The insulin receptor (INSR) gene was analyzed in four patients with severe insulin resistance, revealing five novel mutations and a deletion that removed exon 2. A patient with Donohue syndrome (DS) had a novel p.V657F mutation in the second fibronectin type III domain (FnIII-2), which contains the - cleavage site and part of the insulin-binding site. The mutant INSR was expressed in Chinese hamster ovary cells, revealing that it reduced insulin proreceptor processing and impaired activation of downstream signaling cascades. Using online databases, we analyzed 82 INSR missense mutations and demonstrated that mutations causing DS were more frequently located in the FnIII domains than those causing the milder type A insulin resistance (P = 0.016). In silico structural analysis revealed that missense mutations predicted to severely impair hydrophobic core formation and stability of the FnIII domains all caused DS, whereas those predicted to produce localized destabilization and to not affect folding of the FnIII domains all caused the less severe Rabson-Mendenhall syndrome. These results suggest the importance of the FnIII domains, provide insight into the molecular mechanism of severe insulin resistance, will aid early diagnosis, and will provide potential novel targets for treating extreme insulin resistance.

An adverse intrauterine environment may induce adult disease in offspring, but the mechanisms are not well understood. It is reported that fresh embryo transfer (ET) in assisted reproductive technology leads to high maternal estradiol (E-2), and prenatal high E-2 exposure increases the risk of organ disorders in later life. We found that male newborns and children of fresh ET showed elevated fasting insulin and homeostasis model of assessment for insulin resistance index (HOMA-IR) scores. Male mice with high prenatal estradiol exposure (HE) grew heavier than control mice and developed insulin resistance; they also showed increased food intake, with increased orexigenic hypothalamic neuropeptide Y (NPY) expression. The hypothalamic insulin receptor (INSR) was decreased in male HE mice, associated with elevated promoter methylation. Chronic food restriction (FR) in HE mice reversed insulin resistance and rescued hypothalamic INSR expression by correcting the elevated Insr promoter methylation. Our findings suggest that prenatal exposure to high E-2 may induce sex-specific metabolic disorders in later life through epigenetic programming of hypothalamic Insr promoter, and dietary intervention may reverse insulin resistance by remodeling its methylation pattern.