Insulin Receptor Substrate-2-dependent Interleukin-4 Signaling in Macrophages Is Impaired in Two Models of Type 2 Diabetes Mellitus

Matthew E Hartman,Jason C O'Connor,Jonathan P Godbout,Kyle D Minor,Valerie R Mazzocco,Gregory G Freund

doi:10.1074/jbc.m404368200

Matthew E Hartman, Jason C O'Connor + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m404368200

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Abstract

We have shown previously that hyperinsulinemia inhibits interferon-alpha-dependent activation of phosphatidylinositol 3-kinase (PI3-kinase) through mammalian target of rapamycin (mTOR)-induced serine phosphorylation of insulin receptor substrate (IRS)-1. Here we report that chronic insulin and high glucose synergistically inhibit interleukin (IL)-4-dependent activation of PI3-kinase in macrophages via the mTOR pathway. Resident peritoneal macrophages (PerMPhis) from diabetic (db/db) mice showed a 44% reduction in IRS-2-associated PI3-kinase activity stimulated by IL-4 compared with PerMPhis from heterozygote (db/+) control mice. IRS-2 from db/db mouse PerMPhis also showed a 78% increase in Ser/Thr-Pro motif phosphorylation without a difference in IRS-2 mass. To investigate the mechanism of this PI3-kinase inhibition, 12-O-tetradecanoylphorbol-13-acetate-matured U937 cells were treated chronically with insulin (1 nm, 18 h) and high glucose (4.5 g/liter, 48 h). In these cells, IL-4-stimulated IRS-2-associated PI3-kinase activity was reduced by 37.5%. Importantly, chronic insulin or high glucose alone did not impact IL-4-activated IRS-2-associated PI3-kinase. Chronic insulin + high glucose did reduce IL-4-dependent IRS-2 tyrosine phosphorylation and p85 association by 54 and 37%, respectively, but did not effect IL-4-activated JAK/STAT signaling. When IRS-2 Ser/Thr-Pro motif phosphorylation was examined, chronic insulin + high glucose resulted in a 92% increase in IRS-2 Ser/Thr-Pro motif phosphorylation without a change in IRS-2 mass. Pretreatment of matured U937 cells with rapamycin blocked chronic insulin + high glucose-dependent IRS-2 Ser/Thr-Pro motif phosphorylation and restored IL-4-dependent IRS-2-associated PI3-kinase activity. Taken together these results indicate that IRS-2-dependent IL-4 signaling in macrophages is impaired in models of type 2 diabetes mellitus through a mechanism that relies on insulin/glucose-dependent Ser/Thr-Pro motif serine phosphorylation mediated by the mTOR pathway.

Highlights

The first member of the insulin receptor substrate (IRS)1 family, IRS-1, was initially discovered in Fao hepatoma cells as a tyrosine-phosphorylated substrate of the insulin receptor [1]
IL-4-activated IRS-2-associated PI3-Kinase Activity Is Impaired in peritoneal macrophages (PerM⌽s) from db/db Mice—We have shown previously that chronic insulin exposure-dependent serine phosphorylation of IRS-1 can block cytokine signaling in myeloma cells in vitro [13]
Shows that PerM⌽s from 8-week-old db/db mice treated with IL-4 (5.5 ng/ml, 15 min) had a 44% reduction in IRS-2-associated PI3kinase activity compared with treated PerM⌽s from db/ϩ control mice

Summary

Introduction

The first member of the insulin receptor substrate (IRS)1 family, IRS-1, was initially discovered in Fao hepatoma cells as a tyrosine-phosphorylated substrate of the insulin receptor [1]. Pretreatment of matured U937 cells with rapamycin blocked chronic insulin ؉ high glucose-dependent IRS-2 Ser/Thr-Pro motif phosphorylation and restored IL-4-dependent IRS-2-associated PI3-kinase activity.

Results

Conclusion