RESUMEN
The insulin receptor substrates (IRS) are adapter proteins mediating insulin's and IGF1's intracellular effects. Recent data suggest that IRS2 in the central nervous system (CNS) is involved in regulating fuel metabolism as well as memory formation. The present study aims to specifically define the role of chronically increased IRS2-mediated signal transduction in the CNS. We generated transgenic mice overexpressing IRS2 specifically in neurons (nIRS2 (tg)) and analyzed these in respect to energy metabolism, learning, and memory. Western blot (WB) analysis of nIRS2 (tg) brain lysates revealed increased IRS2 downstream signaling. Histopathological investigation of nIRS2 (tg) mice proved unaltered brain development and structure. Interestingly, nIRS2 (tg) mice showed decreased voluntary locomotoric activity during dark phase accompanied with decreased energy expenditure (EE) leading to increased fat mass. Accordingly, nIRS2 (tg) mice develop insulin resistance and glucose intolerance during aging. Exploratory behavior, motor function as well as food and water intake were unchanged in nIRS2 (tg) mice. Surprisingly, increased IRS2-mediated signals did not change spatial working memory in the T-maze task. Since FoxO1 is a key mediator of IRS2-transmitted signals, we additionally generated mice expressing a dominant negative mutant of FoxO1 (FoxO1DN) specifically in neurons. This mutant mimics the effect of increased IRS2 signaling on FoxO-mediated transcription. Interestingly, the phenotype observed in nIRS2 (tg) mice was not present in FoxO1DN mice. Therefore, increased neuronal IRS2 signaling causes decreased locomotoric activity in the presence of unaltered exploratory behavior and motor coordination that might lead to increased fat mass, insulin resistance, and glucose intolerance during aging independent of FoxO1-mediated transcription.
Asunto(s)
Envejecimiento/metabolismo , Encéfalo/metabolismo , ADN/genética , Regulación del Desarrollo de la Expresión Génica , Intolerancia a la Glucosa/genética , Proteínas Sustrato del Receptor de Insulina/genética , Resistencia a la Insulina , Animales , Southern Blotting , Encéfalo/fisiopatología , Calorimetría Indirecta , Células Cultivadas , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Fibroblastos/metabolismo , Genotipo , Intolerancia a la Glucosa/metabolismo , Intolerancia a la Glucosa/fisiopatología , Inmunohistoquímica , Proteínas Sustrato del Receptor de Insulina/biosíntesis , Locomoción/fisiología , RatonesRESUMEN
Recent studies have discovered changes in the insulin-/IGF1 signaling affecting glucose metabolism and the molecular pathogenesis of human hepatocellular cancer. Insulin/IGF1 receptor mediates its intracellular effects by recruitment of one out of the four different insulin receptor substrates (IRS). To investigate mechanisms of IRS2 expression, we analyzed transcriptional regulation of IRS2 in human HepG2 cells. We identified a region 688 bp upstream of the translation start codon responsible for approximately 90% of basal human IRS2 promoter activity in HepG2 cells, and confirmed binding of specificity protein 1 (also called Sp1 transcription factor, SP1) and nuclear factor 1 (NFI) in this region. Mutation of both SP1 and NFI binding sites or inhibition of extracellular signal regulated kinase (ERK) suppressed IRS2 promoter activity almost completely, revealing a major role of MAP kinases (MAPK) for IRS2 transcription. Activating this cascade with oxidative stress increased IRS2 promoter activity and endogenous IRS2 expression substantially. IRS2 promoter activity rose even more after additional inhibition of p38MAPK indicating an inhibitory effect of p38MAPK on ERK mediated IRS2 transcription. Activation of the MAPK pathway using interleukin 1, beta (IL1B) increased IRS2 promoter activity similar to oxidative stress. In contrast IL1B decreases and inhibition of the MAPK pathway increases IRS1 promoter activity revealing opposed effects of IL1B and ERK on the expression of different IRS proteins. In conclusion we discovered a specific region (-688 to -611 bp) in the IRS2 promoter essential for basal promoter activity and oxidative stress induced transcription depending on ERK activation and SP1 and NFI binding in human hepatocytes.
Asunto(s)
Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Proteínas Sustrato del Receptor de Insulina/genética , Factores de Transcripción NFI/metabolismo , Regiones Promotoras Genéticas/genética , Factor de Transcripción Sp1/metabolismo , Estrés Fisiológico/genética , Transcripción Genética , Activación Enzimática/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Regulación de la Expresión Génica/efectos de los fármacos , Glucosa/farmacología , Células Hep G2 , Humanos , Interleucina-1beta/farmacología , Modelos Genéticos , Especificidad de Órganos/efectos de los fármacos , Especificidad de Órganos/genética , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , PPAR gamma/metabolismo , Unión Proteica/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Estrés Fisiológico/efectos de los fármacos , Transcripción Genética/efectos de los fármacosRESUMEN
Recent studies suggest that the perinatal period is a sensitive part in human development with respect to the pathogenesis of metabolic diseases in adulthood. Neonates, who are either small or large for gestational age (SGA or LGA) have a greater risk of developing obesity and insulin resistance in later life. The term "perinatal priming" is used to describe this phenomenon. Therefore, in the present study we first aimed to investigate if birth weight influences fetal adiponectin and RBP-4 metabolism. Umbilical cord blood was obtained form 40 neonates born on term+/-4 weeks and the adipokine concentrations in the serum were measured. In this analysis adiponectin but not RBP-4 levels showed a positive significant correlation to birth weight. Since maternal preconceptional obesity is associated with an increased birth weight and the risk for LGA neonates, we further aimed to investigate, if the maternal nutritional state influences fetal adiponectin and RBP-4. Therefore umbilical cord blood levels of the adipokines were correlated to maternal preconceptional BMI. In this analysis, neither adiponectin nor RBP-4 levels showed a significant correlation. Taken together, in the present study for the first time we directly compare fetal adiponectin and RBP-4 levels in respect to birth weight and maternal preconceptional BMI. Our data suggest that (1) adiponectin is more likely to have a role in perinatal priming of obesity and insulin resistance than RBP-4 and (2) that birth weight has a greater impact on fetal adipokine serum levels than maternal preconceptional obesity.