These data show that IGF-I levels are reduced in subjects with NAFLD and suggest that hepatic insulin resistance may affect IGF-I levels by modulating GH-stimulated synthesis of hepatic IGF-I.
Interleukin 6 (IL-6) is an independent predictor of type 2 diabetes and cardiovascular disease and is correlated with insulin resistance. Insulin stimulates nitric oxide (NO) production through the IRS-1/PI3-kinase/Akt/eNOS pathway (where IRS-1 is insulin receptor substrate 1, PI3-kinase is phosphatidylinositol 3-kinase, and eNOS is endothelial NO synthase). We asked if IL-6 affects insulin vasodilator action both in human umbilical vein endothelial cells (HUVEC) and in the aortas of C57BL/6J mice and whether this inhibitory effect was caused by increased Ser phosphorylation of IRS-1. We observed that IL-6 increased IRS-1 phosphorylation at Ser312 and Ser616; these effects were paralleled by increased Jun N-terminal protein kinase (JNK) and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and reversed by JNK and ERK1/2 inhibition. In addition, IL-6 treatment resulted in impaired IRS-1 phosphorylation at Tyr612, a site essential for engaging PI3-kinase. Furthermore, IL-6 treatment reduced insulin-stimulated phosphorylation of eNOS at the stimulatory Ser1177 site and impaired insulin-stimulated eNOS dephosphorylation at the inhibitory Thr495 site. Insulin-stimulated eNOS activation and NO production were also inhibited by IL-6; these effects were reversed by inhibition of JNK and ERK1/2. Treatment of C57BL/6J mice with IL-6 resulted in impaired insulin-dependent activation of the Akt/eNOS pathway in the aorta as a result of JNK and ERK1/2 activation. Our data suggest that IL-6 impairs the vasodilator effects of insulin that are mediated by the IRS-1/PI3-kinase/Akt/eNOS pathway through activation of JNK and ERK1/2.
Aims/hypothesis We determined the contribution to insulin resistance of the PH domain leucine-rich repeat protein phosphatase (PHLPP), which dephosphorylates Akt at Ser473, inhibiting its activity. We measured the abundance of PHLPP in fat and skeletal muscle from obese participants. To study the effect of PHLPP on insulin signalling, PHLPP (also known as PHLPP1) was overexpressed in HepG2 and L6 cells. Methods Subcutaneous fat samples were obtained from 82 morbidly obese and ten non-obese participants. Skeletal muscle samples were obtained from 12 obese and eight non-obese participants. Quantification of PHLPP-1 in human tissues was performed by immunoblotting. The functional consequences of recombinant PHLPP1 overexpression in hepatoma HepG2 cells and L6 myoblasts were investigated. Results Of the 82 obese participants, 31 had normal fasting glucose, 33 impaired fasting glucose and 18 type 2 diabetes. PHLPP-1 abundance was twofold higher in the three obese groups than in non-obese participants (p=0.004). No differences were observed between obese participants with normal fasting glucose, impaired fasting glucose or type 2 diabetes. PHLPP-1 abundance was correlated with basal Akt Ser473 phosphorylation (r=−0.48; p=0.001), BMI (r=0.44; p< 0.0001), insulin (r=0.35; p<0.0001) and HOMA (r=0.38; p<0.0001). PHLPP-1 abundance was twofold higher in the skeletal muscle of 12 obese participants than in that of eight non-obese participants (p<0.0001). Insulin treatment of HepG2 cells resulted in a dose-and time-dependent upregulation of PHLPP-1. Overexpression of PHLPP1 in HepG2 cells and L6 myoblasts resulted in impaired insulin signalling involving Akt/glycogen synthase kinase 3, glycogen synthesis and glucose transport. Conclusions/interpretation Increased abundance of PHLPP-1, production of which is regulated by insulin, may represent a new molecular defect in insulin-resistant states such as obesity.
The AMP-activated protein kinase (AMPK) lies upstream of Akt in the pathway leading to endothelial NO synthase (eNOS) activation. Whether leptin promotes eNOS activation via AMPK-dependent activation of Akt, and which of the two AMPKalpha catalytic subunits is involved, remains unknown. Leptin resistance may be partly attributed to interaction between leptin and C-reactive protein (CRP). We hypothesized that leptin effect on eNOS activation in human aortic endothelial cells might be blunted by direct interaction with human recombinant CRP. Small interfering RNAs (siRNAs) were used to knock down expression of alpha1- or alpha2-AMPK in transient transfection assay to evaluate which is involved in this pathway and whether leptin effect on eNOS activation in human aortic endothelial cells might be blunted by direct interaction with human CRP. siRNA-mediated down-regulation of AMPKalpha1, but not AMPKalpha2, abolished leptin-induced Akt-Ser(473) phosphorylation, eNOS-Ser(1177) phosphorylation, eNOS activation, and cGMP accumulation. By contrast, siRNA-mediated knockdown of Akt1 did not affect AMPKalpha1 phosphorylation, but it abolished leptin-induced phosphorylation of Akt-Ser(473) and eNOS-Ser(1177), suggesting that Akt functions downstream of AMPKalpha1. Preincubation of leptin with human recombinant CRP impaired leptin-induced AMPK activation, eNOS-Ser(1177) phosphorylation, eNOS activity, and intracellular cGMP accumulation. The data are consistent with a model implicating an AMPKalpha1-->Akt-->eNOS pathway leading to NO production in response to leptin supporting the idea that interaction between leptin and CRP may have a role in impairing leptin effect on eNOS activation, suggesting a link between leptin resistance, low-grade inflammation, and endothelial dysfunction.
OBJECTIVE -The C-174G promoter polymorphism of the interleukin (IL)-6 gene was found to influence transcriptional activity and plasma IL-6 levels in humans. We addressed the question of whether the C-174G IL-6 polymorphism contributes to variation of insulin sensitivity.RESEARCH DESIGN AND METHODS -Two cohorts of subjects were genotyped. Cohort 1 includes 275 nondiabetic subjects who underwent a euglycemic-hyperinsulinemic clamp. Cohort 2 includes 77 patients with morbid obesity who underwent laparoscopic adjustable gastric banding (LAGB).RESULTS -The genotypes were consistent with Hardy-Weinberg equilibrium proportions. In cohort 1, insulin sensitivity was reduced in carriers of the Ϫ174G/G genotype as compared with subjects carrying the C allele (P ϭ 0.004). Carriers of Ϫ174G/G displayed significantly higher plasma IL-6 levels in comparison with carriers of the C allele. In a stepwise linear regression analysis, the C-174G polymorphism was independently associated with insulin sensitivity; however, after inclusion of plasma IL-6 concentrations, the polymorphism was excluded from the model explaining insulin sensitivity variability, thus suggesting that the polymorphism was affecting insulin sensitivity by regulating IL-6 plasma levels. IL-6 mRNA levels were measured by real-time RT-PCR in subcutaneous fat obtained from obese patients of cohort 2 during LAGB. Carriers of Ϫ174G/G showed increased IL-6 expression compared with subjects carrying the C allele (P ϭ 0.04). There was a significant correlation between adipose IL-6 mRNA expression and insulin resistance assessed by homeostasis model assessment ( ϭ 0.28, P ϭ 0.014).CONCLUSIONS -These results indicate that the Ϫ174G/G genotype of the IL-6 gene may contribute to variations in insulin sensitivity. Diabetes Care 28:2007-2012, 2005T here is increasing evidence suggesting the concept that chronic lowgrade activation of the immune system may play a role in the pathogenesis of insulin resistance and type 2 diabetes (1-3). Cross-sectional and prospective studies have shown that increased levels of systemic inflammatory markers such as interleukin (IL)-6, a major proinflammatory cytokine expressed in several tissues including leukocytes, adipocytes, and endothelial cells, are associated with type 2 diabetes and glucose disorders (1-3). Both circulating levels of IL-6 and adipose tissue IL-6 content have been correlated with insulin resistance (4,5). Moreover, high circulating IL-6 concentrations have been found to predict the development of type 2 diabetes (6).It has been reported that the common C-174G polymorphism in the promoter of the human IL-6 gene regulates its transcription in vitro with the G allele, showing increased transcriptional activity both under basal condition and in response to inflammatory stimuli such as lipopolysaccharides or IL-1 (7). However, data on the effects of this polymorphism on IL-6 levels in vivo have led to conflicting results. Some studies have shown that individuals with the G/G genotype have higher circulating IL-6 levels, as w...
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