Publications by authors named "Eduardo Arilla-Ferreiro"

23 Publications

  • Page 1 of 1

Possible Role of IRS-4 in the Origin of Multifocal Hepatocellular Carcinoma.

Cancers (Basel) 2021 May 23;13(11). Epub 2021 May 23.

Ramón y Cajal Institute of Sanitary Research (IRYCIS), 28034 Madrid, Spain.

New evidence suggests that insulin receptor substrate 4 (IRS-4) may play an important role in the promotion of tumoral growth. In this investigation, we have evaluated the role of IRS-4 in a pilot study performed on patients with liver cancer. We used immunohistochemistry to examine IRS-4 expression in biopsies of tumoral tissue from a cohort of 31 patient suffering of hepatocellular carcinoma (HCC). We simultaneously analyzed the expression of the cancer biomarkers PCNA, Ki-67, and pH3 in the same tissue samples. The in vitro analysis was conducted by studying the behavior of HepG2 cells following IRS-4 overexpression/silencing. IRS-4 was expressed mainly in the nuclei of tumoral cells from HCC patients. In contrast, in healthy cells involved in portal triads, canaliculi, and parenchymal tissue, IRS-4 was observed in the cytosol and the membrane. Nuclear IRS-4 in the tumoral region was found in 69.9 ± 3.2%, whereas in the surrounding healthy hepatocytes, nuclear IRS-4 was rarely observed. The percentage of tumoral cells that exhibited nuclear PCNA and Ki-67 were 52.1 ± 7%, 6.1 ± 1.1% and 1.3 ± 0.2%, respectively. Furthermore, we observed a significant positive linear correlation between nuclear IRS-4 and PCNA (r = 0.989; < 0.001). However, when we correlated the nuclear expression of IRS-4 and Ki-67, we observed a significant positive curvilinear correlation (r = 0.758; < 0.010). This allowed us to define two populations, (IRS-4 + Ki-67 ≤ 69%) and (IRS-4 + Ki-67 > 70%). The population with lower levels of IRS-4 and Ki-67 had a higher risk of suffering from multifocal liver cancer (OR = 16.66; CI = 1.68-164.8 (95%); < 0.05). Immunoblot analyses showed that IRS-4 in normal human liver biopsies was lower than in HepG2, Huh7, and Chang cells. Treatment of HepG2 with IGF-1 and EGF induced IRS-4 translocation to the nucleus. Regulation of IRS-4 levels via HepG2 transfection experiments revealed the protein's role in proliferation, cell migration, and cell-. Nuclear IRS-4 is increased in the tumoral region of HCC. IRS-4 and Ki-67 levels are significantly correlated with the presence of multifocal HCC. Moreover, upregulation of IRS-4 in HepG2 cells induced proliferation by a β-catenin/Rb/cyclin D mechanism, whereas downregulation of IRS-4 caused a loss in cellular polarity and in its adherence to collagen as well as a gain in migratory and invasive capacities, probably via an integrin α2 and focal adhesion cascade (FAK) mechanism.
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http://dx.doi.org/10.3390/cancers13112560DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8197110PMC
May 2021

Leptin Modulates the Response of Brown Adipose Tissue to Negative Energy Balance: Implication of the GH/IGF-I Axis.

Int J Mol Sci 2021 Mar 11;22(6). Epub 2021 Mar 11.

Department of Endocrinology, Instituto de Investigación La Princesa, Hospital Infantil Universitario Niño Jesús, E-28009 Madrid, Spain.

The growth hormone (GH)/insulin-like growth factor I (IGF-I) axis is involved in metabolic control. Malnutrition reduces IGF-I and modifies the thermogenic capacity of brown adipose tissue (BAT). Leptin has effects on the GH/IGF-I axis and the function of BAT, but its interaction with IGF-I and the mechanisms involved in the regulation of thermogenesis remains unknown. We studied the GH/IGF-I axis and activation of IGF-I-related signaling and metabolism related to BAT thermogenesis in chronic central leptin infused (L), pair-fed (PF), and control rats. Hypothalamic somatostatin mRNA levels were increased in PF and decreased in L, while pituitary GH mRNA was reduced in PF. Serum GH and IGF-I concentrations were decreased only in PF. In BAT, the association between suppressor of cytokine signaling 3 and the IGF-I receptor was reduced, and phosphorylation of the IGF-I receptor increased in the L group. Phosphorylation of Akt and cyclic AMP response element binding protein and glucose transporter 4 mRNA levels were increased in L and mRNA levels of uncoupling protein-1 (UCP-1) and enzymes involved in lipid anabolism reduced in PF. These results suggest that modifications in UCP-1 in BAT and changes in the GH/IGF-I axis induced by negative energy balance are dependent upon leptin levels.
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http://dx.doi.org/10.3390/ijms22062827DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001882PMC
March 2021

Bisphenol A impaired cell adhesion by altering the expression of adhesion and cytoskeleton proteins on human podocytes.

Sci Rep 2020 10 6;10(1):16638. Epub 2020 Oct 6.

Laboratory of Renal Physiology and Experimental Nephrology, Department of System Biology/Physiology Unit, University of Alcalá, Alcalá de Henares, Spain.

Bisphenol A (BPA), a chemical -xenoestrogen- used in food containers is present in the urine of almost the entire population. Recently, several extensive population studies have proven a significant association between urinary excretion of BPA and albuminuria. The alteration of glomerular podocytes or "podocytopathy" is a common event in chronic albuminuric conditions. Since many podocytes recovered from patients' urine are viable, we hypothesized that BPA could impair podocyte adhesion capabilities. Using an in vitro adhesion assay, we observed that BPA impaired podocyte adhesion, an effect that was abrogated by Tamoxifen (an estrogen receptor blocker). Genomic and proteomic analyses revealed that BPA affected the expression of several podocyte cytoskeleton and adhesion proteins. Western blot and immunocytochemistry confirmed the alteration in the protein expression of tubulin, vimentin, podocin, cofilin-1, vinculin, E-cadherin, nephrin, VCAM-1, tenascin-C, and β-catenin. Moreover, we also found that BPA, while decreased podocyte nitric oxide production, it lead to overproduction of ion superoxide. In conclusion, our data show that BPA induced a novel type of podocytopathy characterizes by an impairment of podocyte adhesion, by altering the expression of adhesion and cytoskeleton proteins. Moreover, BPA diminished production of podocyte nitric oxide and induced the overproduction of oxygen-free metabolites. These data provide a mechanism by which BPA could participate in the pathogenesis and progression of renal diseases.
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http://dx.doi.org/10.1038/s41598-020-73636-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7538920PMC
October 2020

A nitric oxide synthase inhibitor, L-NAME, prevents L-arginine-induced downregulation of the rat cortical somatostatinergic system.

Neuroreport 2020 01;31(2):87-91

Biochemistry and Molecular Biology Unit, Department of Systems Biology, Faculty of Medicine, Universidad de Alcalá, Alcalá de Henares.

Activation of NMDA receptors leads to nitric oxide (NO) synthesis by NO synthase (NOS) from L-arginine. Neuronal NOS colocalizes with somatostatinergic (SRIF) neurons and there is growing evidence of an interaction between NO and the cerebral SRIFergic system in several neurological diseases. Our aim was to study the effect of L-arginine on the regulation of the SRIFergic system in the frontoparietal cortex of male Sprague-Dawley rats. Intraperitoneal administration of L-arginine (150 mg/Kg), twice-daily during eight days, induced a decrease in SRIF receptor density, which was accompanied by a reduction in the capacity of SRIF to stimulate inositol-1,4,5-triphosphate (IP3) accumulation and SRIF-like immunoreactivity (SRIF-LI) levels. To determine if these changes were related to L-arginine-derived NO synthesis, a NOS inhibitor, Nω-nitro-L-arginine methyl ester was coadministered with L-arginine. Its coadministration prevented the reduction in the SRIF receptor density, accumulation of IP3 and SRIF-LI content. These findings indicate that L-arginine induces a deleterious effect on the cortical somatostatinergic system and that the inhibition of NOS could be helpful in some neurological disorders where this neurotransmitter system is affected.
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http://dx.doi.org/10.1097/WNR.0000000000001373DOI Listing
January 2020

Oxidative Stress and Lymphocyte Alterations in Chronic Relapsing Experimental Allergic Encephalomyelitis in the Rat Hippocampus and Protective Effects of an Ethanolamine Phosphate Salt.

Mol Neurobiol 2020 Feb 10;57(2):860-878. Epub 2019 Sep 10.

Grupo de Neurobioquímica, Unidad de Bioquímica y Biología Molecular, Departamento de Biología de Sistemas, Facultad de Medicina, Universidad de Alcalá, Alcalá de Henares, Madrid, Spain.

Chronic relapsing experimental allergic encephalomyelitis (CR-EAE) exhibits neuropathological and immunological dysfunctions similar to those found in multiple sclerosis (MS) and has been used as an animal model of MS. Inflammatory infiltrates and oxidative stress have been linked to the development of both diseases. Ethanolamine plasmalogen derivates have been shown to be powerful antioxidants and immunomodulators. Therefore, the objective of this study was to analyse inflammatory infiltrates, the state of the oxidative defences and the possible protective effects of calcium, magnesium and phosphate ethanolamine (EAP) in the CR-EAE rat hippocampus. To this aim, we evaluated, by immunohistochemistry, T cell infiltrates, Iba-1 (a marker of activated microglia) immunoreactivity and TUNEL (+) cells. We also measured the protein levels and activity of the antioxidant enzymes catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GP) and glutathione reductase (GR). In addition, reduced (GSH) and oxidized (GSSG) glutathione levels, lipid peroxidation and cholesterol as well as desmosterol content were determined. We found an increase in T cell infiltrates and Iba1 immunoreactivity, lipid peroxidation, SOD, GP and GR activities as well as enhanced cholesterol levels and a decrease in CAT activity, GSH and desmosterol levels in the first and second attack in the CR-EAE rat hippocampus. Pretreatment of CR-EAE rats with EAP led to a delay in the onset of the clinical signs of the disease as well as a decrease in inflammatory infiltrates and alterations of the antioxidant defences in the hippocampus. Altogether, the present results suggest a protective role of EAP in the CR-EAE rat hippocampus.
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http://dx.doi.org/10.1007/s12035-019-01774-8DOI Listing
February 2020

Improvement in inflammation is associated with the protective effect of Gly-Pro-Glu and cycloprolylglycine against Aβ-induced depletion of the hippocampal somatostatinergic system.

Neuropharmacology 2019 06 11;151:112-126. Epub 2019 Apr 11.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Instituto de Investigación Sanitaria La Princesa, Madrid, Spain; CIBER Fisiopatología Obesidad y Nutrición (CB06/03), Instituto de Salud Carlos III, Madrid, Spain. Electronic address:

Glycine-proline-glutamate (GPE) is a cleaved tripeptide of IGF-I that can be processed to cycloprolylglycine (cPG) in the brain. IGF-I protects the hippocampal somatostatinergic system from β-amyloid (Aβ) insult and although neither IGF-I-derived peptides bind to IGF-I receptors, they exert protective actions in several neurological disorders. As their effects on the hippocampal somatostatinergic system remain unknown, the objective of this study was to evaluate if cPG and/or GPE prevent the deleterious effects of Aβ infusion on this system and whether changes in intracellular-related signaling and interleukin (IL) content are involved in their protective effect. We also determined the effect of cPG or GPE co-administration with Aβ on IL secretion in glial cultures and the influence of these ILs on signaling activation and somatostatin synthesis in neuronal cultures. cPG or GPE co-administration reduced Aβ-induced cell death and pro-inflammatory ILs, increased IL-4 and partially avoided the reduction of components of the somatostatinergic system affected by Aβ. GPE increased activation of Akt and CREB and reduced GSK3β activation and astrogliosis, whereas cPG increased phosphorylation of extracellular signal-regulated kinases. Both peptides converged in the activation of mTOR and S6 kinase. Co-administration of these peptides with Aβ to glial cultures increased IL-4 and reduced IL-1β; this release of IL-4 could be responsible for activation of Akt and increased somatostatin in neuronal cultures. Our findings suggest that cPG and GPE exert protective effects against Aβ on the somatostatinergic system by a reduction of the inflammatory environment that may activate different pro-survival pathways in these neurons.
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http://dx.doi.org/10.1016/j.neuropharm.2019.04.008DOI Listing
June 2019

Ellagic acid protects from myelin-associated sphingolipid loss in experimental autoimmune encephalomyelitis.

Biochim Biophys Acta Mol Cell Biol Lipids 2018 09 21;1863(9):958-967. Epub 2018 May 21.

CIBER Fisiopatología de la Obesidad y Nutrición (CIBERobn), ISCIII, Spain; Servicio de Bioquímica Clínica, Hospital Universitario Ramón y Cajal (IRyCIS), Madrid, Spain. Electronic address:

Experimental autoimmune encephalomyelitis (EAE), the most common model for multiple sclerosis, is characterized by inflammatory cell infiltration into the central nervous system and demyelination. Previous studies have demonstrated that administration of some polyphenols may reduce the neurological alterations of EAE. In this work, we show that ellagic acid, a polyphenolic compound, is beneficial in EAE, most likely through stimulation of ceramide biosynthesis within the brain. EAE was induced in Lewis rats by injection of guinea-pig spinal cord tissue along with Freund's complete adjuvant containing Mycobacterium tuberculosis. Clinical signs first appeared at day 8 post-immunization and reached a peak within 3 days, coincident with reduction of myelin basic protein (MBP) in the cortex. Sphingolipids, the other major components of myelin, also decreased at the acute phase of EAE, both in the cerebral cortex and in the spinal cord. In rats receiving ellagic acid in the drinking water from 2 days before immunization, the onset of the disease was delayed and clinical signs were reduced. This amelioration of clinical signs was accompanied by sustained levels of both MBP and sphingolipid in the cortex, without apparent changes in infiltration of inflammatory CD3+ T-cells, microglial activation, or weight loss, which together suggest a neuroprotective effect of ellagic acid. Finally, in glioma and oligodendroglioma cells we demonstrate that urolithins, the ellagic acid metabolites that circulate in plasma, stimulate the synthesis of ceramide. Together these data suggest that ellagic acid consumption protects against demyelination in rats with induced EAE, likely by a mechanism involving sphingolipid synthesis.
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http://dx.doi.org/10.1016/j.bbalip.2018.05.009DOI Listing
September 2018

Improvement in glycemia after glucose or insulin overload in leptin-infused rats is associated with insulin-related activation of hepatic glucose metabolism.

Nutr Metab (Lond) 2016 1;13:19. Epub 2016 Mar 1.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Instituto de Investigación La Princesa, Avda. Menéndez Pelayo, 65, E-28009 Madrid, Spain ; Centro de Investigación Biomédica en Red de Fisiopatología de la Obesidad y Nutrición (CIBEROBN), Instituto de Salud Carlos III, Madrid, E-28009 Spain.

Background: Insulin regulates glucose homeostasis through direct effects on the liver, among other organs, with leptin modulating insulin's hepatic actions. Since central leptin may modify insulin signaling in the liver, we hypothesized that leptin infusion activates hepatic glycogen synthesis following peripheral administration of a bolus of glucose or insulin, thus regulating glycemia.

Findings: Oral glucose and intraperitoneal insulin tolerance tests were performed in control, intracerebroventricular leptin-treated and pair-fed rats during 14 days. An improvement in glycemia and an increase in hepatic free glucose and glycogen concentrations after glucose or insulin overload were observed in leptin-treated rats. In order to analyze whether the liver was involved in these changes, we studied activation of insulin signaling by Western blotting and multiplex bead immunoassay after leptin infusion. Our studies revealed an increase in phosphorylation of insulin receptor substrate-1 and Akt in leptin-treated rats. Examination of parameters related to glucose uptake and metabolism in the liver revealed an augment in glucose transporter 2 and a decrease in phosphoenolpyruvate carboxylase protein levels in this group.

Conclusions: These results indicate that central leptin increases hepatic insulin signaling, associated with increased glycogen concentrations after glucose or insulin overload, leading to an improvement in glycemia.
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http://dx.doi.org/10.1186/s12986-016-0079-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4774133PMC
March 2016

Reduction in Aβ-induced cell death in the hippocampus of 17β-estradiol-treated female rats is associated with an increase in IGF-I signaling and somatostatinergic tone.

J Neurochem 2015 Dec 6;135(6):1257-71. Epub 2015 Nov 6.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Instituto de Investigación La Princesa, Madrid, Spain.

Several studies indicate that 17β-estradiol (E2) protects against amyloid β-peptide (Aβ)-induced cell death and activates factors associated with learning and memory, a function involving the hippocampal somatostatinergic system. As alterations in somatostatin have been demonstrated in Alzheimer's disease, we examined whether E2 prevents changes in the hippocampal somatostatinergic system induced by Aβ25-35 and cell death, as well as the possible involvement of leptin and insulin-like growth factor (IGF)-I signaling. We also measured the levels of Aβ proteases neprilysin and insulin-degrading-enzyme. Co-administration of E2 with Aβ25-35 reduced both its levels and cell death, in addition to preventing the Aβ-induced depletion of some somatostatinergic parameters. Activation of leptin and IGF-I pathways increased after E2 co-administration, and this correlated with changes in the somatostatinergic system. Changes in some components of this system were inversely related with Aβ levels and cell death. Moreover, neprilysin levels were increased only in Aβ plus E2-treated rats and E2 prevented the Aβ-induced insulin-degrading-enzyme reduction. Our results suggest that the E2-induced reduction in cell death is related to lower Aβ levels, probably because of IGF-I and somatostatin modulation of Aβ proteases. We asked how 17β-estradiol (E2) protects against β-amyloid (Aβ)-induced cell death. E2 co-administration prevents Aβ-produced depletion of hippocampal somatostatin (SRIF) by an IGF-I-mediated mechanism, being related this protective effect with an increase in Aβ proteases. Our results suggest that the E2-induced reduction in cell death is related to lower Aβ levels, probably because of SRIF modulation of Aβ proteases. CREB, cAMP response element-binding protein; IGF-I, insulin-like growth factor-I; STAT3, signal transducer and activator of transcription-3.
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http://dx.doi.org/10.1111/jnc.13381DOI Listing
December 2015

Peptides and food intake.

Front Endocrinol (Lausanne) 2014 24;5:58. Epub 2014 Apr 24.

Biochemistry and Molecular Biology Unit, Department of Systems Biology, Faculty of Medicine, University of Alcalá , Alcalá de Henares , Spain.

The mechanisms for controlling food intake involve mainly an interplay between gut, brain, and adipose tissue (AT), among the major organs. Parasympathetic, sympathetic, and other systems are required for communication between the brain satiety center, gut, and AT. These neuronal circuits include a variety of peptides and hormones, being ghrelin the only orexigenic molecule known, whereas the plethora of other factors are inhibitors of appetite, suggesting its physiological relevance in the regulation of food intake and energy homeostasis. Nutrients generated by food digestion have been proposed to activate G-protein-coupled receptors on the luminal side of enteroendocrine cells, e.g., the L-cells. This stimulates the release of gut hormones into the circulation such as glucagon-like peptide-1 (GLP-1), oxyntomodulin, pancreatic polypeptides, peptide tyrosine tyrosine, and cholecystokinin, which inhibit appetite. Ghrelin is a peptide secreted from the stomach and, in contrast to other gut hormones, plasma levels decrease after a meal and potently stimulate food intake. Other circulating factors such as insulin and leptin relay information regarding long-term energy stores. Both hormones circulate at proportional levels to body fat content, enter the CNS proportionally to their plasma levels, and reduce food intake. Circulating hormones can influence the activity of the arcuate nucleus (ARC) neurons of the hypothalamus, after passing across the median eminence. Circulating factors such as gut hormones may also influence the nucleus of the tractus solitarius (NTS) through the adjacent circumventricular organ. On the other hand, gastrointestinal vagal afferents converge in the NTS of the brainstem. Neural projections from the NTS, in turn, carry signals to the hypothalamus. The ARC acts as an integrative center, with two major subpopulations of neurons influencing appetite, one of them coexpressing neuropeptide Y and agouti-related protein (AgRP) that increases food intake, whereas the other subpopulation coexpresses pro-opiomelanocortin (POMC) and cocaine and amphetamine-regulated transcript that inhibits food intake. AgRP antagonizes the effects of the POMC product, α-melanocyte-stimulating hormone (α-MSH). Both populations project to areas important in the regulation of food intake, including the hypothalamic paraventricular nucleus, which also receives important inputs from other hypothalamic nuclei.
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http://dx.doi.org/10.3389/fendo.2014.00058DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4005944PMC
June 2014

Leptin-induced downregulation of the rat hippocampal somatostatinergic system may potentiate its anorexigenic effects.

Neurochem Int 2012 Dec 13;61(8):1385-96. Epub 2012 Oct 13.

Neurobiochemistry Unit, Department of Biochemistry and Molecular Biology, Facultad de Medicina, Universidad de Alcalá, Alcalá de Henares, E-28871 Madrid, Spain.

The learning and memory mechanisms in the hippocampus translate hormonal signals of energy balance into behavioral outcomes involved in the regulation of food intake. As leptin and its receptors are expressed in the hippocampus and somatostatin (SRIF), an orexigenic neuropeptide, may inhibit leptin-mediated suppression of food intake in other brain areas, we asked whether chronic leptin infusion induces changes in the hippocampal somatostatinergic system and whether these modifications are involved in leptin-mediated effects. We studied 18 male Wistar rats divided into three groups: controls (C), treated intracerebroventricularly (icv) with leptin (12 μg/day) for 14 days (L) and a pair-fed group (PF) that received the same amount of food consumed by the L group. Food restriction increased whereas leptin decreased the hippocampal SRIF receptor density, due to changes in SRIF receptor 2 protein levels. These changes in the PF group were concurrent with an increase of hippocampal G protein-coupled receptor kinase 2 protein levels and activation of Akt and cyclic AMP response element binding protein. The inhibitory effect of SRIF on adenylyl cyclase (AC) activity, however, was decreased in L rats, coincident with lower G inhibitory α3 and higher AC-I levels as well as signal transducer and activator of transcription factor 3 activation. In addition, 20 male Wistar rats were included to analyze whether the leptin antagonist L39A/D40A/F41A and the SRIF receptor agonist SMS 201-995 modify SRIF signaling and food intake, respectively. Administration of L39A/D40A/F41A reversed changes in SRIF signaling, whereas SMS 201-995 ameliorated food consumption in L. Altogether, these results suggest that increased somatostatinergic tone in PF rats may be a mechanism to improve the hippocampal orexigenic effects in a situation of metabolic demand, whereas down-regulation of this system in L rats may represent a mechanism to enhance the anorexigenic effects of leptin.
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http://dx.doi.org/10.1016/j.neuint.2012.09.019DOI Listing
December 2012

Adipose tissue promotes a serum cytokine profile related to lower insulin sensitivity after chronic central leptin infusion.

PLoS One 2012 2;7(10):e46893. Epub 2012 Oct 2.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Universidad Autónoma de Madrid, Madrid, Spain.

Obesity is an inflammatory state characterized by an augment in circulating inflammatory factors. Leptin may modulate the synthesis of these factors by white adipose tissue decreasing insulin sensitivity. We have examined the effect of chronic central administration of leptin on circulating levels of cytokines and the possible relationship with cytokine expression and protein content as well as with leptin and insulin signaling in subcutaneous and visceral adipose tissues. In addition, we analyzed the possible correlation between circulating levels of cytokines and peripheral insulin resistance. We studied 18 male Wistar rats divided into controls (C), those treated icv for 14 days with a daily dose of 12 μg of leptin (L) and a pair-fed group (PF) that received the same food amount consumed by the leptin group. Serum leptin and insulin were measured by ELISA, mRNA levels of interferon-γ (IFN-γ), interleukin-2 (IL-2), IL-4, IL-6, IL-10 and tumor necrosis factor-α (TNF-α) by real time PCR and serum and adipose tissue levels of these cytokines by multiplexed bead immunoassay. Serum leptin, IL-2, IL-4, IFN-γ and HOMA-IR were increased in L and TNF-α was decreased in PF and L. Serum leptin and IL-2 levels correlate positively with HOMA-IR index and negatively with serum glucose levels during an ip insulin tolerance test. In L, an increase in mRNA levels of IL-2 was found in both adipose depots and IFN-γ only in visceral tissue. Activation of leptin signaling was increased and insulin signaling decreased in subcutaneous fat of L. In conclusion, leptin mediates the production of inflammatory cytokines by adipose tissue independent of its effects on food intake, decreasing insulin sensitivity.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0046893PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3462753PMC
May 2013

Vitamin E deficiency impairs the somatostatinergic receptor-effector system and leads to phosphotyrosine phosphatase overactivation and cell death in the rat hippocampus.

J Nutr Biochem 2013 May 16;24(5):848-58. Epub 2012 Aug 16.

Biochemical and Molecular Biology Department, Neuro-Biochemical Group, Faculty of Medicine, Universidad de Alcalá de Henares, Madrid, Spain.

Vitamin E plays an essential role in maintaining the structure and function of the nervous system, and its deficiency, commonly associated with fat malabsorption diseases, may reduce neuronal survival. We previously demonstrated that the somatostatinergic system, implicated in neuronal survival control, can be modulated by α-tocopherol in the rat dentate gyrus, increasing cyclic adenosine monophosphate response element binding protein phosphorylation. To gain a better understanding of the molecular actions of tocopherols and examine the link among vitamin E, somatostatin and neuronal survival, we have investigated the effects of a deficiency and subsequent administration of tocopherol on the somatostatin signaling pathway and neuronal survival in the rat hippocampus. No changes in somatostatin expression were detected in vitamin-E-deficient rats. These rats, however, showed a significant increase in the somatostatin receptor density and dissociation constant, which correlated with a significant increase in the protein levels of somatostatin receptors. Nevertheless, vitamin E deficiency impaired the ability of the somatostatin receptors to couple to the effectors adenylyl cyclase and phosphotyrosine phosphatase by diminishing Gi protein functionality. Furthermore, vitamin E deficiency significantly increased phosphotyrosine phosphatase activity and PTPη expression, as well as PKCδ activation, and decreased extracellular-signal-regulated kinase phosphorylation. All these changes were accompanied by an increase in neuronal cell death. Subsequent α-tocopherol administration partially or completely reversed all these values to control levels. Altogether, our results prove the importance of vitamin E homeostasis in the somatostatin receptor-effector system and suggest a possible mechanism by which this vitamin may regulate the neuronal cell survival in the adult hippocampus.
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http://dx.doi.org/10.1016/j.jnutbio.2012.05.005DOI Listing
May 2013

Chronic central leptin infusion modifies the response to acute central insulin injection by reducing the interaction of the insulin receptor with IRS2 and increasing its association with SOCS3.

J Neurochem 2011 Apr 9;117(1):175-85. Epub 2011 Feb 9.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Instituto Investigación Sanitaria Princesa, Madrid, Spain.

Leptin and insulin have overlapping intracellular signaling mechanisms and exert anorexigenic actions in the hypothalamus. We aimed to determine how chronic exposure to increased leptin affects the hypothalamic response to a rise in insulin. We analyzed the activation and interactions of components of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in the hypothalamus of rats treated icv for 14 days with leptin followed by a central injection of insulin and killed 15 min later. Insulin increased glycemia and chronic leptin reduced this insulin induced rise in glucose. Leptin decreased the association between the insulin receptor beta chain (IRβ) and insulin receptor substrate 2 (IRS2), augmented the association between Janus kinase 2 and IRS2, increased levels of the catalytic subunit of PI3K and pAkt-Ser473 and decreased forkhead box O number 1 levels. Insulin reduced the association between suppressor of the cytokine signaling 3 and IRβ, increased IRβ-IRS2 association and pAkt-Thr308 levels, with chronic leptin exposure blunting these effects. In conclusion, chronic exposure to leptin decreases the central response to insulin by increasing suppressor of the cytokine signaling 3 association to IR, which inhibits insulin signaling at the level of interaction of its receptor with IRS2 and activates PI3K by promoting Janus kinase 2-IRS2 association. Thus, these results suggest that this mechanism could be a target for the treatment of insulin resistance.
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http://dx.doi.org/10.1111/j.1471-4159.2011.07191.xDOI Listing
April 2011

Effects of the antipsychotic drug haloperidol on the somastostatinergic system in SH-SY5Y neuroblastoma cells.

J Neurochem 2009 Jul 11;110(2):631-40. Epub 2009 May 11.

Hospital Ramón y Cajal, Madrid, Spain.

Antipsychotics are established drugs in schizophrenia treatment which, however, are not free of side effects. Lipid rafts are critical for normal brain function. Several G protein-coupled receptors, such as somatostatin (SRIF) receptors, have been shown to localize to lipid rafts. The aim of this study was to investigate whether haloperidol treatment affects the composition and functionality of lipid rafts in SH-SY5Y neuroblastoma cells. Haloperidol inhibited cholesterol biosynthesis, leading to a marked reduction in cell cholesterol content and to an accumulation of sterol intermediates, particularly cholesta-8,14-dien-3beta-ol. These changes were accompanied by a loss of flotillin-1 and Fyn from the lipid rafts. We next studied the functionality of the SRIF receptor. Treatment with haloperidol reduced the inhibitory effect of SRIF on adenylyl cyclase (AC) activity. On the other side, haloperidol decreased basal AC activity but increased forskolin-stimulated AC activity. Addition of free cholesterol to the culture medium abrogated the effects of haloperidol on lipid raft composition and SRIF signaling whereas the AC response to forskolin remained elevated. The results show that haloperidol, by affecting cholesterol homeostasis, ultimately alters SRIF signaling and AC activity, which might have physiological consequences.
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http://dx.doi.org/10.1111/j.1471-4159.2009.06159.xDOI Listing
July 2009

The N-terminal tripeptide of insulin-like growth factor-I protects against beta-amyloid-induced somatostatin depletion by calcium and glycogen synthase kinase 3 beta modulation.

J Neurochem 2009 Apr 11;109(2):360-70. Epub 2009 Feb 11.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Madrid, Spain.

The protective effects of insulin-like growth factor I on the somatostatin (SRIF) system in the temporal cortex after beta-amyloid (Abeta) injury may be mediated through its N-terminal tripeptide glycine-proline-glutamate (GPE). GPE is cleaved to cyclo[Pro-Gly] (cPG), a metabolite suggested to mediate in neuroprotective actions. We evaluated the effects of GPE and cPG in the temporal cortex of Abeta25-35-treated rats on SRIF and SRIF receptor protein and mRNA levels, adenylyl cyclase activity, cell death, Abeta25-35 accumulation, cytosolic calcium levels ([Ca(2+)](c)) and the intracellular signaling mechanisms involved. GPE and cPG did not change Abeta25-35 levels, but GPE partially restored SRIF and SRIF receptor 2 protein content and mRNA levels and protected against cell death after Abeta25-35 insult, which was coincident with Akt activation and glycogen synthase kinase 3beta inhibition. In addition, GPE displaced glutamate from NMDA receptors and blocked the glutamate induced rise in cytosolic calcium in isolated rat neurons and moderately increased Ca(2+) influx per se. Our findings suggest that GPE, but not its metabolite, mimics insulin-like growth factor I effects on the SRIF system through a mechanism independent of Abeta clearance that involves modulation of calcium and glycogen synthase kinase 3beta signaling.
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http://dx.doi.org/10.1111/j.1471-4159.2009.05980.xDOI Listing
April 2009

17Beta-estradiol protects depletion of rat temporal cortex somatostatinergic system by beta-amyloid.

Neurobiol Aging 2007 Sep 14;28(9):1396-409. Epub 2006 Jul 14.

Department of Endocrinology, Hospital Infantil Universitario Niño Jesús, Universidad Autónoma de Madrid, Avda. Menéndez Pelayo, Madrid, Spain.

Estradiol prevents amyloid-beta peptide (Abeta)-induced cell death through estrogen receptors (ERs) and modulates somatostatin (SRIF) responsiveness in the rat brain. As intracerebroventricular (ICV) Abeta25-35 administration reduces SRIFergic tone in the temporal cortex of ovariectomized (Ovx) rats, we asked whether 17beta-estradiol (E2) treatment can restore the Abeta25-35 induced changes in SRIF content, SRIF receptor density and adenylyl cyclase (AC) activity, as well as if these effects are mediated by ERs. E2 treatment did not change Abeta25-35 levels in the temporal cortex, but partially restored the SRIFergic parameters affected by Abeta insult and decreased cell death, which was correlated with Akt activation. The ER antagonist ICI 182,780 prevented the protective effect of E2 on sst2 levels, but did not modify SRIF levels. Furthermore, ICI 182,780 treatment further decreased sst2 protein and mRNA levels when administered alone to Abeta25-35-treated rats, suggesting that it may block the effects of endogenous estrogens. These findings indicate that E2 protects the temporal cortical SRIFergic system from Abeta-induced depletion independently of Abeta accumulation.
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http://dx.doi.org/10.1016/j.neurobiolaging.2006.06.009DOI Listing
September 2007

Differential effects of ethanol ingestion on somatostatin content, somatostatin receptors and adenylyl cyclase activity in the frontoparietal cortex of virgin and parturient rats.

Life Sci 2005 Jul;77(10):1094-105

Research Laboratory, Universidad Autónoma, Hospital Infantil Universitario Niño Jesús, Avda. Menéndez Pelayo, 65; 28009 Madrid, Spain.

Chronic ethanol ingestion decreases the number of somatostatin (SRIF) receptors in the rat frontoparietal cortex and female sex hormones modulate the effects of ethanol in the brain. Therefore, we investigated the differential effects of ethanol consumption on the SRIFergic system in the frontoparietal cortex of virgin and parturient rats given ethanol in their drinking water before and during gestation. In parturient rats, ethanol consumption decreased the density of SRIF receptors (25%, p<0.01 vs control parturient group) whereas the SRIF-like immunoreactivity (SRIF-LI) content was increased (140%, p<0.01). In virgin rats, ethanol ingestion decreased the density of SRIF receptors (42%, p<0.01) more than in alcoholic parturient rats. SRIF-LI levels were unaffected. The inhibitory effect of SRIF on basal and forskolin-stimulated adenylyl cyclase was significantly lower in alcoholic virgin rats as compared to alcoholic parturient rats. No differences in the levels of the G inhibitory (Gi) alpha1 and Gialpha2 proteins were observed among the experimental groups. These results suggest that gestation may confer partial resistance to the ethanol-induced effect on the SRIFergic system.
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http://dx.doi.org/10.1016/j.lfs.2005.01.019DOI Listing
July 2005

Gly-Pro-Glu protects beta-amyloid-induced somatostatin depletion in the rat cortex.

Neuroreport 2004 Aug;15(12):1979-82

Department of Endocrinology and Research Laboratory Hospital Infantil Universitario Niño Jesús, Universidad Autónoma, E-28009 Madrid, Spain.

The effect of Gly-Pro-Glu (GPE) on the somatostatinergic system of the temporal cortex in amyloid beta-peptide (Abeta) treated rats was investigated. Intracerebroventricular Abeta25-35 administration for 14 days (300 pmol/day) to ovariectomized rats produced a marked reduction in somatostatin (SRIF) content, SRIF receptor density and reduced the inhibitory effect of SRIF on adenylyl cyclase activity. I.p. injection of three doses (300 microg) of GPE on days 0, 6 and 12 resulted in a partial recovery of the parameters affected by Abeta25-35 administration. These results indicate that GPE may have an in vivo effect protecting the temporal cortical somatostatinergic system from Abeta insult.
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http://dx.doi.org/10.1097/00001756-200408260-00029DOI Listing
August 2004

Activity of the hippocampal somatostatinergic system following daily administration of melatonin.

Brain Res Mol Brain Res 2004 Jul;126(2):107-13

Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Grupo de Neurobioquímica, Ctra. Madrid-Barcelona Km. 33,600, Universidad de Alcalá, E-28871 Alcalá de Henares, Madrid, Spain.

If melatonin or its analogs are to be used therapeutically in humans, their chronic effects on responsiveness of melatonin target cells need to be assessed. We have previously demonstrated that acute melatonin treatment regulates the somatostatinergic system in the rat hippocampus. In the present study, we have investigated the effects of subchronic and chronic daily treatment with melatonin on the somatostatinergic system in the rat hippocampus. Male Wistar rats (200-250 g) were injected with melatonin (25 microg/kg body weight, subcutaneously) daily for 4, 7 or 14 days and sacrificed 24 h after the last injection. Melatonin administration for 4 days induced a decrease in the hippocampal somatostatin (SRIF)-like immunoreactivity content as well as a decrease in the number of SRIF receptors and an increase in their apparent affinity. The decreased number of SRIF receptors in the melatonin (4 days)-treated rats was associated with a decreased capacity of SRIF to inhibit both basal and forskolin-stimulated adenylyl cyclase activity. These melatonin-induced effects reversed to control values after 7 or 14 days of treatment. Hippocampal membranes from control and melatonin-treated rats showed similar Gi and Gs activities. Melatonin treatment altered neither the functional Gi activity nor the Gialpha 1 or Gialpha 2 levels at any of the time periods studied. The present results suggest that chronic exposure to melatonin results in a tolerance of the hippocampus to this hormone.
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http://dx.doi.org/10.1016/j.molbrainres.2004.03.010DOI Listing
July 2004

Acutely administered melatonin decreases somatostatin-binding sites and the inhibitory effect of somatostatin on adenylyl cyclase activity in the rat hippocampus.

J Pineal Res 2004 Mar;36(2):87-94

Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Grupo de Neurobioquímica, Universidad de Alcalá, Madrid, Spain.

Melatonin is known to increase neuronal activity in the hippocampus, an effect contrary to that of somatostatin (somatotropin release-inhibiting factor, SRIF). Thus, the aim of this study was to investigate whether the somatostatinergic system is implicated in the mechanism of action of melatonin in the rat hippocampus. One group of rats was injected a single dose of melatonin [25 microg/kg subcutaneously (s.c.)] or saline containing ethanol (0.5%, s.c.) and killed 5 hr later. Melatonin significantly decreased the SRIF-like immunoreactivity levels and induced a significant decrease in the density of SRIF receptors as well as in the dissociation constant (Kd). SRIF-mediated inhibition of basal and forskolin-stimulated adenylyl cyclase activity was markedly decreased in hippocampal membranes from melatonin-treated rats. The functional activity of Gi proteins was similar in hippocampal membranes from melatonin-treated and control rats. Western blot analyses revealed that melatonin administration did not alter Gialpha1 or Gialpha2 levels. To determine if the changes observed were related to melatonin-induced activation of central melatonin receptors, a melatonin receptor antagonist, luzindole, was administered prior to melatonin injection. Pretreatment with luzindole (10 mg/kg, s.c.) did not alter the melatonin-induced effects on the above-mentioned parameters and luzindole, alone, had no observable effect. The present results demonstrate that melatonin decreases the activity of the SRIF receptor-effector system in the rat hippocampus, an effect which is apparently not mediated by melatonin receptors. As SRIF exerts an opposite effect to that of melatonin on hippocampal neuronal activity, it is possible that the SRIFergic system could be implicated in the mechanism of action of melatonin in the rat.
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http://dx.doi.org/10.1046/j.1600-079x.2003.00100.xDOI Listing
March 2004

Modulation of somatostatin receptors, somatostatin content and Gi proteins by substance P in the rat frontoparietal cortex and hippocampus.

J Neurochem 2003 Jan;84(1):145-56

Grupo de Neurobioquímica, Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad de Alcalá, Alcalá de Henares, Madrid, Spain.

Substance P (SP) and somatostatin (SRIF) are widely spread throughout the CNS where they play a role as neurotransmitters and/or neuromodulators. A colocalization of both neuropeptides has been demonstrated in several rat brain areas and SP receptors have been detected in rat cortical and hippocampal somatostatinergic cells. The present study was thus undertaken to determine whether SP could modulate SRIF signaling pathways in the rat frontoparietal cortex and hippocampus. A single intraperitoneal injection of SP (50, 250 or 500 micro g/kg) induced an increase in the density of SRIF receptors in membranes from the rat frontoparietal cortex at 24 h of its administration, with no change in the hippocampus. The functionality of the SRIF receptors was next investigated. Western blot analysis of Gi proteins demonstrated a significant decrease in Gialpha1 levels in frontoparietal cortical membranes from rats treated acutely (24 h) with 250 micro g/kg of SP, which correlated with a decrease in functional Gi activity, as assessed by use of the non-hydrolyzable GTP analog 5'-guanylylimidodiphosphate. SRIF-mediated inhibition of basal or forskolin-stimulated adenylyl cyclase activity was also significantly lower in the frontoparietal cortex of the SP-treated group, with no alterations in the catalytic subunit of the enzyme. SRIF-like immunoreactivity content was increased in the frontoparietal cortex after acute (24 h) SP administration (250 or 500 micro g/kg) as well as in the hippocampus in response to 7 days of SP (250 micro g/kg) administration. All these SP-mediated effects were prevented by pretreatment with the NK1 receptor antagonist RP-67580. Although the physiologic significance of these results are unknown, the increase in SRIF receptor density together with the desensitization of the SRIF inhibitory signaling pathway might be a mechanism to potentiate the stimulatory pathway of SRIF, inducing a preferential coupling of the receptors to PLC.
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http://dx.doi.org/10.1046/j.1471-4159.2003.01510.xDOI Listing
January 2003

Effects of subchronic and chronic melatonin treatment on somatostatin binding and its effects on adenylyl cyclase activity in the rat frontoparietal cortex.

J Pineal Res 2002 Nov;33(4):189-97

Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad de Alcalá, E-28871 Alcalá de Henares, Madrid, Spain.

Melatonin and somatostatin are known to exert similar effects on locomotor activity. We have previously demonstrated that acute melatonin treatment regulates somatostatin receptor function in the rat frontoparietal cortex. However, the effects of subchronic and chronic melatonin treatment on the somatostatin receptor-G protein-adenylyl cyclase system in the rat frontoparietal cortex are unknown. Melatonin was administered subcutaneously at a daily dose of 25 microg/kg for 4 days, 1 wk or 2 wk. Twenty-four hours after the last injection, the animals were sacrificed. Melatonin did not alter the somatostatin-like immunoreactivity content in the frontoparietal cortex from control and melatonin-treated rats during any of the previously indicated periods. Four days of melatonin administration induced both an increase in the number of [(125)I]-Tyr11-somatostatin receptors and a decrease in the affinity of somatostatin for its receptors in frontoparietal cortical membranes. The increased number of somatostatin receptors in the melatonin-treated rats was associated with an increased capacity of somatostatin to inhibit basal and forskolin-stimulated adenylyl cyclase activity. Melatonin administration for 4 days induced a higher adenylyl cyclase activity both under basal conditions and after direct stimulation of the enzyme with forskolin. No significant differences were observed in the function of Gi proteins in the 4-day melatonin-treated rats. Western blot analyses showed that the 4-day melatonin treatment reduced Gialpha(2) levels, without altering the amount of Gialpha(1). These melatonin-induced changes reverted to control values after 7 or 14 days of treatment. Altogether, the present findings suggest that subchronic melatonin treatment modulates the somatostatin receptor/effector system in the rat frontoparietal cortex.
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http://dx.doi.org/10.1034/j.1600-079x.2002.02906.xDOI Listing
November 2002
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