Inducible liver-specific knockdown of protein tyrosine phosphatase 1B improves glucose and lipid homeostasis in adult mice.

AIMS/HYPOTHESIS Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of insulin signalling. Hepatic PTP1B deficiency, using the Alb-Cre promoter to drive Ptp1b deletion from birth in mice, improves glucose homeostasis, insulin sensitivity and lipid metabolism. The aim of this study was to investigate the therapeutic potential of decreasing liver PTP1B levels in obese and insulin-resistant adult mice. METHODS Inducible Ptp1b liver-specific knockout mice were generated using SA-Cre-ER(T2) mice crossed with Ptp1b floxed (Ptp1b(fl/fl)) mice. Mice were fed a high-fat diet (HFD) for 12 weeks to induce obesity and insulin resistance. Tamoxifen was administered in the HFD to induce liver-specific deletion of Ptp1b (SA-Ptp1b(-/-) mice). Body weight, glucose homeostasis, lipid homeostasis, serum adipokines, insulin signalling and endoplasmic reticulum (ER) stress were examined. RESULTS Despite no significant change in body weight relative to HFD-fed Ptp1b(fl/fl) control mice, HFD-fed SA-Ptp1b(-/-) mice exhibited a reversal of glucose intolerance as determined by improved glucose and pyruvate tolerance tests, decreased fed and fasting blood glucose and insulin levels, lower HOMA of insulin resistance, circulating leptin, serum and liver triacylglycerols, serum NEFA and decreased HFD-induced ER stress. This was associated with decreased glycogen synthase, eukaryotic translation initiation factor-2α kinase 3, eukaryotic initiation factor 2α and c-Jun NH2-terminal kinase 2 phosphorylation, and decreased expression of Pepck. CONCLUSIONS/INTERPRETATION Inducible liver-specific PTP1B knockdown reverses glucose intolerance and improves lipid homeostasis in HFD-fed obese and insulin-resistant adult mice. This suggests that knockdown of liver PTP1B in individuals who are already obese/insulin resistant may have relatively rapid, beneficial therapeutic effects

A 1H NMR metabolic profiling to the assessment of protein tyrosine phosphatase 1B role in liver regeneration after partial hepatectomy

10.1016/j.biochi.2012.11.015Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of the tyrosine kinase growth factor signaling pathway, which is involved in major physiological mechanisms such as liver regeneration. We investigate early hepatic metabolic events produced by partial hepatectomy (PHx) for PTP1B deficient (PTP1B KO) and wild type (WT) mice using proton nuclear magnetic resonance spectroscopy. Metabolic response of the two genotypes produced 24 h upon PHx is compared using magic angle spinning high-resolution nuclear magnetic resonance (1H-HR-MAS-NMR) on intact liver tissues. In addition, genotype-associated metabolic profile changes were monitored during the first 48 h after PHx using high-resolution nuclear magnetic resonance (1H-HR-NMR) on liver extracts. A marked increase of lipid-related signals in regenerating livers was observed after 24 h PHx in either intact tissues or liver extracts studies. In spite of this common initial metabolic response, results obtained 48 h after PHx on liver extracts indicate a genotype-differential metabolic pattern. This metabolic pattern resulted in line with well known regenerative features such as more sustained cell proliferation, a better management of lipids as energy fuel and lessened liver injury for PTP1B KO mice as compared to WT. Taken together, these findings suggest the metabolic basis to the pivotal role of PTP1B in liver regeneration

IRS2 deficiency limits the effects of inhibition of protein tyrosine phosphatase 1B in IGF-IR-mediated signalling in the retina

Resumen del póster presentado al 48th European Association for the Study of Diabetes (EASD) Annual Meeting, celebrado en Berlin (Alemania) del 1 al 5 de octubre de 2012.[Background and aims]: Mice with complete deletion of insulin receptor substrate (IRS) 2 develop type 2 diabetes. In the retina, IRS2 deficiency induced photoreceptor degeneration and apoptosis that is not rescued by normalization of glucose levels. On the other hand, protein tyrosine phosphatase 1B (PTP1B) inhibition by genetic deletion in IRS2-/- mice (double mutant IRS2-/-PTP1B-/-) restored peripheral insulin resistance and normalized glucose homeostasis. Since IGF-IR promotes survival of photoreceptors and also is a substrate of PTP1B, we aimed to investigate IGF-IR-mediated survival signalling and visual function in PTP1B-/-and IRS2-/-/PTP1B-/-in comparison to wild-type and IRS2-/-mice. [Materials and methods]: IGF-IR tyrosine phosphorylation and Akt serine 473 phosphorylation were analyzed by western blot in organotypic retinal explants stimulated with IGF-I. Immunohistochemistry was used to evaluate retinal structure preservation in mice at 10-12 weeks and programmed cell death was assessed by TUNEL. Visual function was evaluated by Electroretinographic (ERG) recording in mice at 5 and 10 weeks. [Results]: IGF-IR tyrosine phosphorylation and Akt serine 473 phosphorylation increased in retinal explants stimulated for 15 min with IGF-I in a dose-dependent manner. In PTP1B-/- retinal explants, these responses increased by twofold in IGF-IR phosphorylation and by threefold in Akt phosphorylation compared to the wild-type control (p=0.037 and p=0.04, respectively). Conversely, in IRS2-/- mice the response to IGF-I in Akt phosphorylation decreased compared to the wild-type control (p=0.04). Moreover, in IRS2-/- mice PTP1B deletion (double mutant IRS2-/- /PTP1B-/-) also enhanced IGF-IR tyrosine phosphorylation (p=0.01) but, unexpectedly, the response to IGF-I in the activation of Akt remained decreased as observed in the IRS2-/- mice. Histological evaluation revealed a significant thickness in whole retina in both IRS2-/- and double mutant IRS2-/-/PTP1B-/- mice, specifically in the outer nuclear layer (ONL) and retinal outer segments (ROS) (p<0.001). ERG analysis showed that PTP1B deficiency did not restored normal visual function in IRS2-/- mice. [Conclusion]: Although PTP1B deficiency significantly enhanced IGF-IR tyrosine phosphorylation in retinal explants of IRS2-/- mice, it was unable to restore Akt phosphorlyation and, therefore, the structural and functional visual of these mice defects were not improved.Supported by: SAF-2009 and NEURORED-DIAB.Peer Reviewe

Loss of protein tyrosine phosphatase 1B increases IGF-I receptor tyrosine phosphorylation but does not rescue retinal defects in IRS2-deficient mice

et al.[Purpose]: Mice with deletion of insulin receptor substrate (IRS) 2 develop type 2 diabetes and photoreceptor degeneration. Loss of protein tyrosine phosphatase 1B (PTP1B) in diabetic IRS2-/- mice restores insulin sensitivity and normalizes glucose homeostasis. Since insulinlike growth factor (IGF)-IR promotes survival of photoreceptors and is a substrate of PTP1B, we investigated IGF-IR-mediated survival signaling and visual function in PTP1B-/- and double mutant IRS2-/-/PTP1B-/- mice. [Methods]: IGF-IR-mediated Akt signaling was evaluated in IGF-I-stimulated retinal explants. Histologic and electroretinogram analysis was performed in wild-type (WT), IRS2-/-, PTP1B-/-, and the double mutant IRS2-/-/PTP1B/ mice. [Results]: IGF-I stimulated the tyrosine phosphorylation of its receptor and Akt activation in retinal explants of WT mice. In PTP1B-/- retinal explants, these responses were enhanced. Conversely, in retinas from IRS2-/- mice, expression of PTP1B was increased, coincident with decreased IGF-I-mediated Akt serine 473 phosphorylation. PTP1B deletion in IRS2 -/-mice also enhanced IGF-IR tyrosine phosphorylation but, unexpectedly, did not rescue Akt activation in response to IGF-I. One potential explanation is that PTEN was increased in retinas of IRS2-/- and IRS2-/-/PTP1B-/- mice. Histologic evaluation revealed alterations in various structures of the retina in IRS2-/- and IRS2-/-/PTP1B-/- mice, specifically in the outer nuclear layer (ONL) and retinal outer segments (ROS). Electroretinogram (ERG) analysis confirmed that PTP1B deficiency did not restore visual function in IRS2-/- mice. [Conclusions]: Although loss of PTP1B enhances tyrosine phosphorylation of the IGF-IR in retinal explants of IRS2-/- mice, Akt activation remains defective owing to elevated PTEN levels and, thus, structural and functional visual defects persist in this model. © 2013 The Association for Research in Vision and Ophthalmology, Inc.Supported by Grants SAF2012-33283, SAF2010-21879-C02-01 (MINECO, Spain), NEURORET-DIAB from CIBERDEM (Centro de Investigacion Biomedica en Red de Diabetes y Enfermedades Metabolicas Asociadas, Instituto Salud Carlos III, Spain), Comunidad de Madrid S2010/BMD-2423, and The European Consortium for the Early Treatment of Diabetic Retinopathy (278040 PCOLSMALL), a project funded by the FP7 Program of the European Community. AIA, JR-C, and AG-R hold postdoctoral contracts from CIBERDEM. ThPeer Reviewe

Significant lethality following liver resection in A20 heterozygous knockout mice uncovers a key role for A20 in liver regeneration

Hepatic expression of A20, including in hepatocytes, increases in response to injury, inflammation and resection. This increase likely serves a hepatoprotective purpose. The characteristic unfettered liver inflammation and necrosis in A20 knockout mice established physiologic upregulation of A20 as integral to the anti-inflammatory and anti-apoptotic armamentarium of hepatocytes. However, the implication of physiologic upregulation of A20 in modulating hepatocytes' proliferative responses following liver resection remains controversial. To resolve the impact of A20 on hepatocyte proliferation and the liver's regenerative capacity, we examined whether decreased A20 expression, as in A20 heterozygous knockout mice, affects outcome following two-third partial hepatectomy. A20 heterozygous mice do not demonstrate a striking liver phenotype, indicating that their A20 expression levels are still sufficient to contain inflammation and cell death at baseline. However, usually benign partial hepatectomy provoked a staggering lethality (>40%) in these mice, uncovering an unsuspected phenotype. Heightened lethality in A20 heterozygous mice following partial hepatectomy resulted from impaired hepatocyte proliferation due to heightened levels of cyclin-dependent kinase inhibitor, p21, and deficient upregulation of cyclins D1, E and A, in the context of worsened liver steatosis. A20 heterozygous knockout minimally affected baseline liver transcriptome, mostly circadian rhythm genes. Nevertheless, this caused differential expression of >1000 genes post hepatectomy, hindering lipid metabolism, bile acid biosynthesis, insulin signaling and cell cycle, all critical cellular processes for liver regeneration. These results demonstrate that mere reduction of A20 levels causes worse outcome post hepatectomy than full knockout of bona fide liver pro-regenerative players such as IL-6, clearly ascertaining A20's primordial role in enabling liver regeneration. Clinical implications of these data are of utmost importance as they caution safety of extensive hepatectomy for donation or tumor in carriers of A20/TNFAIP3 single nucleotide polymorphisms alleles that decrease A20 expression or function, and prompt the development of A20-based liver pro-regenerative therapies

Significant lethality following liver resection in A20 heterozygous knockout mice uncovers a key role for A20 in liver regeneration

Hepatic expression of A20, including in hepatocytes, increases in response to injury, inflammation and resection. This increase likely serves a hepatoprotective purpose. The characteristic unfettered liver inflammation and necrosis in A20 knockout mice established physiologic upregulation of A20 as integral to the anti-inflammatory and anti-apoptotic armamentarium of hepatocytes. However, the implication of physiologic upregulation of A20 in modulating hepatocytes' proliferative responses following liver resection remains controversial. To resolve the impact of A20 on hepatocyte proliferation and the liver's regenerative capacity, we examined whether decreased A20 expression, as in A20 heterozygous knockout mice, affects outcome following two-third partial hepatectomy. A20 heterozygous mice do not demonstrate a striking liver phenotype, indicating that their A20 expression levels are still sufficient to contain inflammation and cell death at baseline. However, usually benign partial hepatectomy provoked a staggering lethality (>40%) in these mice, uncovering an unsuspected phenotype. Heightened lethality in A20 heterozygous mice following partial hepatectomy resulted from impaired hepatocyte proliferation due to heightened levels of cyclin-dependent kinase inhibitor, p21, and deficient upregulation of cyclins D1, E and A, in the context of worsened liver steatosis. A20 heterozygous knockout minimally affected baseline liver transcriptome, mostly circadian rhythm genes. Nevertheless, this caused differential expression of >1000 genes post hepatectomy, hindering lipid metabolism, bile acid biosynthesis, insulin signaling and cell cycle, all critical cellular processes for liver regeneration. These results demonstrate that mere reduction of A20 levels causes worse outcome post hepatectomy than full knockout of bona fide liver pro-regenerative players such as IL-6, clearly ascertaining A20's primordial role in enabling liver regeneration. Clinical implications of these data are of utmost importance as they caution safety of extensive hepatectomy for donation or tumor in carriers of A20/TNFAIP3 single nucleotide polymorphisms alleles that decrease A20 expression or function, and prompt the development of A20-based liver pro-regenerative therapies