Anti-Insulin Receptor Autoantibodies Are Not Required for Type 2 Diabetes Pathogenesis in NZL/Lt Mice, a New Zealand Obese (NZO)-Derived Mouse Strain

The New Zealand obese (NZO) mouse strain shares with the related New Zealand black (NZB) strain a number of immunophenotypic traits. Among these is a high proportion of B-1 B lymphocytes, a subset associated with autoantibody production. Approximately 50% of NZO/HlLt males develop a chronic insulin-resistant type 2 diabetes syndrome associated with 2 unusual features: the presence of B lymphocyte–enriched peri-insular infiltrates and the development of anti-insulin receptor autoantibodies (AIRAs). To establish the potential pathogenic contributions ofBlymphocytes and AIRAs in this model, a disrupted immunoglobulin heavy chain gene (Igh-6) congenic on the NZB/BlJ background was backcrossed 4 generations into the NZO/HlLt background and was then intercrossed to produce mice that initially segregated for wild-type versus the mutant Igh-6 allele and thus permitted comparison of syndrome development. A new flow cytometric assay (AIRA binding to transfected Chinese hamster ovary cells stably expressing mouse insulin receptor) showed IgM and IgG subclass AIRAs in serum from Igh-6 intact males, but not in Igh6null male serum. However, the absence of B lymphocytes and antibodies distinguishing mutant from wild-type males failed to significantly affect diabetes-free survival. The Igh6nullmales gained weight less rapidly than wild-type males, probably accounting for a retardation, but not prevention, of hyperglycemia. Thus, AIRA and the Blymphocyte component of the peri-insulitis in chronic diabetics were not essential either to development of insulin resistance or to eventual pancreatic beta cell failure and loss. A new substrain, designated NZL, was generated by inbreeding Igh-6 wild-type segregants. Currently at the F10 generation, NZL mice exhibit the same juvenile-onset obesity as NZO/HlLt males, but develop type 2 diabetes at a higher frequency (> 80%). Also, unlike NZO/HlLt mice that are difficult to breed, the NZL/Lt strain breeds well and thus offers clear advantages to obesity/diabetes researchers

Ubiquitin–proteasome degradation of serum- and glucocorticoid-regulated kinase-1 (SGK-1) is mediated by the chaperone-dependent E3 ligase CHIP

SGK-1 (serum- and glucocorticoid-regulated kinase-1) is a stress-induced serine/threonine kinase that is phosphorylated and activated downstream of PI3K (phosphoinositide 3-kinase). SGK-1 plays a critical role in insulin signalling, cation transport and cell survival. SGK-1 mRNA expression is transiently induced following cellular stress, and SGK-1 protein levels are tightly regulated by rapid proteasomal degradation. In the present study we report that SGK-1 forms a complex with the stress-associated E3 ligase CHIP [C-terminus of Hsc (heat-shock cognate protein) 70-interacting protein]; CHIP is required for both the ubiquitin modification and rapid proteasomal degradation of SGK-1. We also show that CHIP co-localizes with SGK-1 at or near the endoplasmic reticulum. CHIP-mediated regulation of SGK-1 steady-state levels alters SGK-1 kinase activity. These data suggest a model that integrates CHIP function with regulation of the PI3K/SGK-1 pathway in the stress response

Glucocorticoid (GC)-Mediated Down-Regulation of Urokinase Plasminogen Activator Expression via the Serum and GC Regulated Kinase-1/Forkhead Box O3a Pathway

The glucocorticoid receptor (GR) and its ligand, cortisol, play a central role in human physiology. The exact mechanisms by which GR activation regulates these processes are the subject of intensive investigation. We and others have shown that GR activation can indirectly down-regulate specific genes via serum and glucocorticoid (GC) regulated kinase-1-mediated inhibition of forkhead box O3a (FOXO3a) transcriptional activity. We previously used gene expression microarrays, together with bioinformatic analyses, to identify putative FOXO3a target genes in breast epithelial cells. In this paper we refine our analysis through the use of FOXO3a chromatin immunoprecipitation (ChIP) microarrays. ChIP microarray results reveal urokinase plasminogen activator (uPA) as a putative novel target of FOXO3a in breast epithelial and breast cancer cell lines. We further show that uPA down-regulation after GC treatment requires serum and GC regulated kinase-1-mediated inactivation of FOXO3a activity. ChIP and luciferase assays confirm that FOXO3a can both occupy and transactivate the uPA promoter. Our data suggest that inactivation of FOXO3a after GR activation is an important mechanism contributing to GC-mediated repression of uPA gene expression in breast epithelial and cancer cells