BUSCADOR RECOLECTA

PGC-1 alpha controls, to a large extent, the capacity of cells to respond to changing nutritional requirements and energetic demands. The key role of metabolic reprogramming in tumor development has highlighted the potential role of PGC-1 alpha in cancer. To investigate how loss of PGC-1 alpha activity in primary cells impacts the oncogenic characteristics of spontaneously immortalized cells, and the mechanisms involved, we used the classic 3T3 protocol to generate spontaneously immortalized mouse embryonic fibroblasts (iMEFs) from wild-type (WT) and PGC-1 alpha knockout (KO) mice and analyzed their oncogenic potential in vivo and in vitro. We found that PGC-1 alpha KO iMEFs formed larger and more proliferative primary tumors than WT counterparts, and fostered the formation of lung metastasis by B16 melanoma cells. These characteristics were associated with the reduced capacity of KO iMEFs to respond to cell contact inhibition, in addition to an increased ability to form colonies in soft agar, an enhanced migratory capacity, and a reduced growth factor dependence. The mechanistic basis of this phenotype is likely associated with the observed higher levels of nuclear beta-catenin and c-myc in KO iMEFs. Evaluation of the metabolic adaptations of the immortalized cell lines identified a decrease in oxidative metabolism and an increase in glycolytic flux in KO iMEFs, which were also more dependent on glutamine for their survival. Furthermore, glucose oxidation and tricarboxylic acid cycle forward flux were reduced in KO iMEF, resulting in the induction of compensatory anaplerotic pathways. Indeed, analysis of amino acid and lipid patterns supported the efficient use of tricarboxylic acid cycle intermediates to synthesize lipids and proteins to support elevated cell growth rates. All these characteristics have been observed in aggressive tumors and support a tumor suppressor role for PGC-1 alpha, restraining metabolic adaptations in cancer., This work was funded by grants from the Spanish "Ministerio de Ciencia, Innovacion y Universidades" (MICINN) and ERDF/FEDER funds, SAF2012-37693, SAF2015-63904-R, SAF2015-71521-REDC, RTI2018-093864-B-I00 to M.M., SAF2017-83043-R and B2017/BMD-3724 to S.C., PI15/00107 to A.M.R, the University of the Basque Country UPV/EHU grant GIU16/62) to J.l.R.S. and M.B.R.L., and the European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement 721236-TREATMENT to M.M.

Schizophrenia is a neuropsychiatric disorder that chronically affects 21 million people worldwide. Secondgeneration
antipsychotics (SGAs) are the cornerstone in the management of schizophrenia. However,
despite their efficacy in counteracting both positive and negative symptomatology of schizophrenia, recent clinical
observations have described an increase in the prevalence of metabolic disturbances in patients treated with
SGAs, including abnormal weight gain, hyperglycemia and dyslipidemia. While the molecular mechanisms responsible
for these side-effects remain poorly understood, increasing evidence points to a link between SGAs and
adipose tissue depots of white, brown and beige adipocytes. In this review, we survey the present knowledge in
this area, with a particular focus on the molecular aspects of adipocyte biology including differentiation, lipid metabolism, thermogenic function and the browning/beiging process, This work was funded by H2020 Marie Skłodowska-Curie
ActionsITN-TREATMENT (Grant Agreement 721236, European
Commission). We also acknowledge grants RTI2018-094052-B-100
(MICINN/FEDER, Spain), S2017/BMD-3684 MOIR2-CM (Comunidad
de Madrid, Spain) and CIBERdem (ISCIII, Spain).

Aims: Pupillography is a noninvasive and cost-effective method to determine autonomic nerve activity. Genetic variants in cytochrome P450 (CYP), dopamine receptor (DRD2, DRD3), serotonin receptor (HTR2A, HTR2C) and ATP-binding cassette subfamily B (ABCB1) genes, among others, were previously associated with the pharmacokinetics and pharmacodynamics of antipsychotic drugs. Our aim was to evaluate the effects of aripiprazole and olanzapine on pupillary light reflex related to pharmacogenetics. Methods: Twenty-four healthy volunteers receiving 5 oral doses of 10 mg aripiprazole and 5 mg olanzapine tablets were genotyped for 46 polymorphisms by quantitative polymerase chain reaction. Pupil examination was performed by automated pupillometry. Aripiprazole, dehydro-aripiprazole and olanzapine plasma concentrations were measured by high-performance liquid chromatography–tandem mass spectrometry. Results: Aripiprazole affected pupil contraction: it caused dilatation after the administration of the first dose, then caused constriction after each dosing. It induced changes in all pupillometric parameters (P '.05). Olanzapine only altered minimum pupil size (P =.046). Polymorphisms in CYP3A, HTR2A, UGT1A1, DRD2 and ABCB1 affected pupil size, the time of onset of constriction, pupil recovery and constriction velocity. Aripiprazole, dehydro-aripiprazole and olanzapine pharmacokinetics were significantly affected by polymorphisms in CYP2D6, CYP3A, CYP1A2, ABCB1 and UGT1A1 genes. Conclusions: In conclusion, aripiprazole and its main metabolite, dehydro-aripiprazole altered pupil contraction, but olanzapine did not have such an effect. Many polymorphisms may influence pupillometric parameters and several polymorphisms had an effect on aripiprazole, dehydro-aripiprazole and olanzapine pharmacokinetics. Pupillography could be a useful tool for the determination of autonomic nerve activity during antipsychotic treatment., Consejería de Educación, Juventud y Deporte, Comunidad de Madrid, Grant/Award Number: PEJD-2017-PRE/BMD-4164; H2020 Marie Skłodowska-Curie Actions, Grant/Award Number: 721236

Background and Objectives
Aripiprazole is an atypical antipsychotic drug that is metabolized by cytochrome P450 (CYP) 2D6 and CYP3A4, which mainly form its active metabolite dehydro-aripiprazole. Because of the genetic polymorphism of CYP2D6, plasma concentrations are highly variable between different phenotypes. In this study, phenotype-related physiologically based pharmacokinetic models were developed and evaluated to suggest phenotype-guided dose adjustments.

Methods
Physiologically based pharmacokinetic models for single dose (oral and orodispersible formulation), multiple dose, and steady-state condition were built using trial data from genotyped healthy volunteers. Based on evaluated models, dose adjustments were simulated to compensate for genetically caused differences.

Results
Physiologically based pharmacokinetic models were able to accurately predict the pharmacokinetics of aripiprazole and dehydro-aripiprazole according to CYP2D6 phenotypes, illustrated by a minimal bias and a good precision. For single-dose administration, 92.5% (oral formulation) and 79.3% (orodispersible formulation) of the plasma concentrations of aripiprazole were within the 1.25-fold error range. In addition, physiologically based pharmacokinetic steady-state simulations demonstrate that the daily dose for poor metabolizer should be adjusted, resulting in a maximum recommended dose of 10 mg, but no adjustment is necessary for intermediate and ultra-rapid metabolizers.

Conclusions
In clinical practice, CYP2D6 genotyping in combination with therapeutic drug monitoring should be considered to personalize aripiprazole dosing, especially in CYP2D6 poor metabolizers, to ensure therapy effectiveness and safety, Open Access funding enabled and organized by Projekt DEAL. Dora Koller and the multiple-dose phase I trial (EUDRA-CT: 2018-000744-26) were co-financed by the H2020 Marie Sklodowska-Curie Innovative Training Network 721236 grant

Aims/hypothesis: Second-generation antipsychotic (SGA) drugs have been associated with the development of type 2 diabetes and the metabolic syndrome in patients with schizophrenia. In this study, we aimed to investigate the effects of two different SGA drugs, olanzapine and aripiprazole, on metabolic state and islet function and plasticity. Methods: We analysed the functional adaptation of beta cells in 12-week-old B6;129 female mice fed an olanzapine- or aripiprazole-supplemented diet (5.5–6.0 mg kg−1 day−1) for 6 months. Glucose and insulin tolerance tests, in vivo glucose-stimulated insulin secretion and indirect calorimetry were performed at the end of the study. The effects of SGAs on beta cell plasticity and islet serotonin levels were assessed by transcriptomic analysis and immunofluorescence. Insulin secretion was assessed by static incubations and Ca2+ fluxes by imaging techniques. Results: Treatment of female mice with olanzapine or aripiprazole for 6 months induced weight gain (p<0.01 and p<0.05, respectively), glucose intolerance (p<0.01) and impaired insulin secretion (p<0.05) vs mice fed a control chow diet. Aripiprazole, but not olanzapine, induced serotonin production in beta cells vs controls, likely by increasing tryptophan hydroxylase 1 (TPH1) expression, and inhibited Ca2+ flux. Of note, aripiprazole increased beta cell size (p<0.05) and mass (p<0.01) vs mice fed a control chow diet, along with activation of mechanistic target of rapamycin complex 1 (mTORC1)/S6 signalling, without preventing beta cell dysfunction. Conclusions/interpretation: Both SGAs induced weight gain and beta cell dysfunction, leading to glucose intolerance; however, aripiprazole had a more potent effect in terms of metabolic alterations, which was likely a result of its ability to modulate the serotonergic system. The deleterious metabolic effects of SGAs on islet function should be considered while treating patients as these drugs may increase the risk for development of the metabolic syndrome and diabetes, Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. This work was funded by H2020
Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement 721236, European Commission). We also acknowledge grants
RTI2018-094052-B-100/ AEI/10.13039/501100011033 (Ministerio de Ciencia e Innovación y Fondo Europeo de Desarrollo Regional [FEDER]) and S2017/BMD-3684 (Comunidad de Madrid, Spain), and grants from Fundación Ramón Areces (Spain) and CIBERDEM (ISCIII, Spain)

PGC-1α controls, to a large extent, the capacity of cells to respond to changing nutritional requirements and energetic demands. The key role of metabolic reprogramming in tumor development has highlighted the potential role of PGC-1α in cancer. To investigate how loss of PGC-1α activity in primary cells impacts the oncogenic characteristics of spontaneously immortalized cells, and the mechanisms involved, we used the classic 3T3 protocol to generate spontaneously immortalized mouse embryonic fibroblasts (iMEFs) from wild-type (WT) and PGC-1α knockout (KO) mice and analyzed their oncogenic potential in vivo and in vitro. We found that PGC-1α KO iMEFs formed larger and more proliferative primary tumors than WT counterparts, and fostered the formation of lung metastasis by B16 melanoma cells. These characteristics were associated with the reduced capacity of KO iMEFs to respond to cell contact inhibition, in addition to an increased ability to form colonies in soft agar, an enhanced migratory capacity, and a reduced growth factor dependence. The mechanistic basis of this phenotype is likely associated with the observed higher levels of nuclear β-catenin and c-myc in KO iMEFs. Evaluation of the metabolic adaptations of the immortalized cell lines identified a decrease in oxidative metabolism and an increase in glycolytic flux in KO iMEFs, which were also more dependent on glutamine for their survival. Furthermore, glucose oxidation and tricarboxylic acid cycle forward flux were reduced in KO iMEF, resulting in the induction of compensatory anaplerotic pathways. Indeed, analysis of amino acid and lipid patterns supported the efficient use of tricarboxylic acid cycle intermediates to synthesize lipids and proteins to support elevated cell growth rates. All these characteristics have been observed in aggressive tumors and support a tumor suppressor role for PGC-1α, restraining metabolic adaptations in cancer., This work was funded by grants from the Spanish “Ministerio de Ciencia, Innovación y Universidades” (MICINN) and ERDF/FEDER funds, SAF2012-37693, SAF2015-63904-R, SAF2015-71521-REDC, RTI2018-093864-B-I00 to M.M., SAF2017-83043-R and B2017/BMD-3724 to S·C., PI15/00107 to A.M.R, the University of the Basque Country UPV/EHU grant GIU16/62) to J.l.R.S. and M.B.R.L., and the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement 721236-TREATMENT to M.M.

Presentation at the Kick-off meeting of H2020-funded Metabolic Dysfunctions associated with Pharmacological Treatment of Schizophrenia project (Treatment) held in Madrid at the CSIC Institute de Investigaciones Biomédicas Alberto Sols on June 15-16, 2017., No

ARS Open., [Aims]: Sirtuin 1 (SIRT1) is a key player in liver physiology and a therapeutic target against hepatic inflammation. We evaluated the role of SIRT1 in the proinflammatory context and oxidative stress during acetaminophen (APAP)-mediated hepatotoxicity., [Results]: SIRT1 protein levels decreased in human and mouse livers following APAP overdose. SIRT1-Tg mice maintained higher levels of SIRT1 on APAP injection than wild-type mice and were protected against hepatotoxicity by modulation of antioxidant systems and restrained inflammatory responses, with decreased oxidative stress, proinflammatory cytokine messenger RNA levels, nuclear factor kappa B (NFκB) signaling, and cell death. Mouse hepatocytes stimulated with conditioned medium of APAP-treated macrophages (APAP-CM) showed decreased SIRT1 levels; an effect mimicked by interleukin (IL)1β, an activator of NFκB. This negative modulation was abolished by neutralizing IL1β in APAP-CM or silencing p65-NFκB in hepatocytes. APAP-CM of macrophages from SIRT1-Tg mice failed to downregulate SIRT1 protein levels in hepatocytes. In vivo administration of the NFκB inhibitor BAY 11-7082 preserved SIRT1 levels and protected from APAP-mediated hepatotoxicity., [Inovation]: Our work evidenced the unique role of SIRT1 in APAP hepatoprotection by targeting oxidative stress and inflammation., [Conclusion]: SIRT1 protein levels are downregulated by IL1β/NFκB signaling in APAP hepatotoxicity, resulting in inflammation and oxidative stress. Thus, maintenance of SIRT1 during APAP overdose by inhibiting NFκB might be clinically relevant., [Rebound Track]: This work was rejected during standard peer review and rescued by Rebound Peer Review (Antioxid Redox Signal 16:293-296, 2012) with the following serving as open reviewers: Rafael de Cabo, Joaquim Ros, Kalervo Hiltunen, and Neil Kaplowitz. Antioxid. Redox Signal. 28, 1187-1208., This work was funded by SAF2015-65267-R (MINECO/FEDER), CIBERdem (ISCIII, Spain), and INFLAMES (ISCIII PIE14/00045, cofunded by ERDF, ‘‘Investing in your
future’’) to A.M.V.; IJCI-2014-19381 (MINECO/FEDER) to P.R. and A.M.V.; SAF2014-52009-R (MINECO/FEDER) to S.A.; SAF2015-63904-R (MINECO/FEDER) to M.M.; CP14/00181 (ISCIII/FEDER) to A.G.-R.; and PI13/00021 (ISCIII/FEDER) to J.M. CIBERehd (ISCIII) to A.G.-R. and J.M.; Grant 37/371 from Al Jouf University to M.A.M. We also acknowledge H2020 Marie Sklodowska-Curie ITNTREATMENT
(Grant Agreement No. 721236) (European Commission)., Peer Reviewed

Fentanyl is an agonist of the μ‐opioid receptor commonly used in the treatment of moderate‐severe pain. In order to study whether pharmacogenetics explains some of the variability in the response to fentanyl, several genes related to fentanyl receptors, transporters and metabolic enzymes have been analysed. Thirty‐five healthy volunteers (19 men and 16 women) receiving a single 300 μg oral dose of fentanyl were genotyped for 9 polymorphisms in cytochrome P450 (CYP) enzymes (CYP3A4 and CYP3A5), ATP‐binding cassette subfamily B member 1 (ABCB1), opioid receptor mu 1 (OPRM1), catechol‐O‐methyltransferase (COMT) and adrenoceptor beta 2 (ADRB2) by real‐time PCR. Fentanyl concentrations were measured by ultra‐performance liquid chromatography combined with tandem mass spectrometry (UPLC‐MS/MS). Fentanyl pharmacokinetics is affected by sex. Carriers of the CYP3A4*22 allele, which is known to reduce the mRNA expression, showed higher area under the concentration‐time curve (AUC) and lower clearance (Cl) values. Although this finding might be of importance, its validity needs to be confirmed in other similar settings. Furthermore, carriers of the ABCB1 C1236T T/T genotype presented a lower AUC and higher Cl, as well as lower half‐life (T1/2). As volunteers were blocked with naltrexone, the effect of fentanyl on pharmacodynamics might be biased; however, we could observe that fentanyl had a hypotensive effect. Moreover, ADRB2 C523A A allele carriers showed a tendency towards reducing systolic blood pressure. Likewise, OPRM1 and COMT minor allele variants were risk factors for the development of somnolence. CYP3A5*3, ABCB1 C3435T and ABCB1 G2677T/A were not associated with fentanyl's pharmacokinetics, pharmacodynamics and safety profile., F. Abad‐Santos and D. Ochoa have been consultants or investigators in clinical trials sponsored by the following pharmaceutical companies: Abbott, Alter, Chemo, Cinfa, FAES, Farmalíder, Ferrer, GlaxoSmithKline, Galenicum, Gilead, Janssen‐Cilag, Kern, Normon, Novartis, Servier, Silverpharma, Teva and Zambon. D. Koller is co‐financed by the H2020 Marie Sklodowska‐Curie Innovative Training Network 721236 grant. P. Zubiaur is co‐financed by Consejería de Educación, Juventud y Deporte from Comunidad de Madrid and Fondo Social Europeo., Peer reviewed

Taurine (TAU) is a sulfur-containing beta amino acid that is not involved in protein composition and anabolism, conditionally essential in mammals provided through diet. Growing evidence supports a protective role of TAU supply in osmoregulation, calcium flux, and reduction of inflammation and oxidant damage in renal diseases like diabetes. Endoplasmic reticulum (ER) stress, due to abnormal proteostasis, is a contributor to nephrotic syndrome and related renal damage. Here, we investigated the effect of dietary TAU (1.5% in drinking water for 15 days) in an established rat model that mimics human minimal change nephrosis, consisting of a single puromycin aminonucleoside (PAN) injection (intraperitoneally 15 mg/100 g body weight), with sacrifice after eight days. TAU limited proteinuria and podocytes foot processes effacement, and balanced slit diaphragm nephrin and glomerular claudin 1 expressions. In cortical proximal tubules, TAU improved lysosomal density, ER perimeter, restored proper ER-mitochondria tethering and mitochondrial cristae, and decreased inflammation. Remarkably, TAU downregulated glomerular ER stress markers (GRP78, GRP94), pro-apoptotic C/EBP homologous protein, activated caspase 3, tubular caspase1, and mitochondrial chaperone GRP75, but maintained anti-apoptotic HSP25. In conclusion, TAU, by targeting upstream ER stress separate from mitochondria dysfunctions at crucial renal sites, might be a promising dietary supplement in the treatment of the drug-resistant nephrotic syndrome., This study was supported by New Pet Food Italia S.r.l. (Italy) donation, ex 60% grants from the University of Brescia (Italy), FFARB 2017, SAF2015-63904-R grant from the Spanish Ministerio de Economía Industria y Competitividad (MINEICO), FEDER funds and MSCA-ITN-2016-721236 from the European Commission H2020-MSCA-ITN program., Peer Reviewed

Protein tyrosine phosphatase 1B (PTP1B) is widely expressed in mammalian tissues, in particular in immune cells, and plays a pleiotropic role in dephosphorylating many substrates. Moreover, PTP1B expression is enhanced in response to pro-inflammatory stimuli and to different cell stressors. Taking advantage of the use of mice deficient in PTP1B we have investigated the effect of γ-radiation in these animals and found enhanced lethality and decreased respiratory exchange ratio vs. the corresponding wild type animals. Using bone-marrow derived macrophages and mouse embryonic fibroblasts (MEFs) from wild-type and PTP1B-deficient mice, we observed a differential response to various cell stressors. PTP1B-deficient macrophages exhibited an enhanced response to γ-radiation, UV-light, LPS and S-nitroso-glutathione. Macrophages exposed to γ-radiation show DNA damage and fragmentation, increased ROS production, a lack in GSH elevation and enhanced acidic β-galactosidase activity. Interestingly, these differences were not observed in MEFs. Differential gene expression analysis of WT and KO macrophages revealed that the main pathways affected after irradiation were an up-regulation of protein secretion, TGF-β signaling and angiogenesis among other, and downregulation of Myc targets and Hedgehog signaling. These results demonstrate a key role for PTP1B in the protection against the cytotoxicity of irradiation in intact animal and in macrophages, which might be therapeutically relevant., This work was supported by grants SAF2017-82436R, SAF2016-75004R and SAF2015-65267 from MINIECO, S2017/BMD-3686 and BMD3684 from Comunidad de Madrid, Fundación Ramón Areces (2016/CIVP18A3864) and Cibercv, Ciberdem and Ciberehd (funded by the Instituto de Salud Carlos III and by Fondos FEDER). We also acknowledge H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement number 721236) (European Commission). P.R. was recipient a Juan de la Cierva postdoctoral contract from MINIECO (IJCI-2014-19381)., Peer Reviewed

Retinal degenerative diseases are a group of heterogeneous diseases that include age-related macular degeneration (AMD), retinitis pigmentosa (RP), and diabetic retinopathy (DR). The progressive degeneration of the retinal neurons results in a severe deterioration of the visual function. Neuroinflammation is an early hallmark of many neurodegenerative disorders of the retina including AMD, RP and DR. Microglial cells, key components of the retinal immune defense system, are activated in retinal degenerative diseases. In the microglia the interplay between the proinflammatory/classically activated or antiinflammatory/alternatively activated phenotypes is a complex dynamic process that occurs during the course of disease due to the different environmental signals related to pathophysiological conditions. In this regard, an adequate transition from the proinflammatory to the anti-inflammatory response is necessary to counteract retinal neurodegeneration and its subsequent damage that leads to the loss of visual function. Insulin like-growth factor-1 (IGF-1) has been considered as a pleiotropic factor in the retina under health or disease conditions and several effects of IGF-1 in retinal immune modulation have been described. In this review, we provide recent insights of inflammation as a common feature of retinal diseases (AMD, RP and RD) highlighting the role of microglia, exosomes and IGF-1 in this process., The work of AIA and ÁMV has been funded by grants from European Union (project H2020-MSCA-ITN TREATMENT Grant Agreement number: 721236 and project EUROCONDOR FP7 Grant Agreement number 278040), grant from the Spanish Ministry of Economy and Competitiveness: SAF2015-65267-R (MINECO/FEDER) and
grants from the Spanish ISCIII (CIBERdem) and INFLAMES (ISCIII PIE14/00045, co-funded by ERDF, Investing in your future)., Peer Reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: Second-generation Antipsychotics (SGAs) have become the treatment of choice over the typical antipsychotics as they provide excellent efficacy and fewer extrapyramidal symptoms. However, the compliance of the patients to SGAs is negatively affected by their ability to induce or aggravate metabolic syndrome, namely, weight gain, insulin resistance, and Type 2 Diabetes. The exact underlying mechanism of metabolic effects of SGAs is not fully elucidated and it is assumed to
be at least partially due to their effect on central nervous system. However, whether SGAs have a direct effect on insulin action in the tissues is still to be elucidated. The effect of SGAs on body metabolism varies and we have chosen two drugs, Olanzapine and Aripiprazole, which are associated with high and low risk of metabolic side-effects, respectively. Our research is focused on studying the effect of both SGAs on insulin resistance in human adipose tissue. Aside from lipid storage function, adipose tissue has been recognised, as an endocrine organ, producing hormones, such as adiponectin and leptin, indispensable for energy homeostasis. The set of experiments performed as a part of this study includes measuring the effect of Olanzapine and Aripiprazole on the lipolysis in human isolated adipocytes., [Methods]: Biopsies of subcutaneous adipose tissue (SAT) were collected from 6 patients (3 men, 3 women; age: 20-76 years; BMI: 20.9-34.5 kg/m2). Subjects were free of antidepressants or antipsychotics treatment. At the moment, only the effect of Olanzapine has been tested and measured, the experiments with Aripiprazole are in progress. A 6% adipocyte suspension was incubated with olanzapine (0.004, 0.04, 0.1, 0.2, 2 and 20 μM) or aripiprazole (0.02, 0.2, 0.5, 1, 10 and 100 μM). This was followed by 10 minutes incubation with 4 concentrations of insulin (0.1 μU; 1.0 μU; 10 μU; 100
μU) and then incubated with 0.5 μM ß-adrenergic receptor agonist, Isopretenerol, for 1h 50 min. ß-adrenergic stimulation activates hormone-sensitive lipase (HSL) enzyme via cAMP-dependent pathway. HSL, in turn, hydrolyses tritriacylglycerol (TAG), diacylglycerol (DAG) or monoacylglycerol (MAG) molecules producing free fatty acids and glycerol. The supernatant was then collected and used for glycerol measurement., [Results]: Short incubations of adipocytes with therapeutic concentrations of Olanzapine show no effect in lipolysis. The highest concentration of the drug hints at a reduced rate of lipolysis in adipocytes by more than 50% for each insulin concentration (p<0.0001) and in control conditions (p<0.01)., [Conclusions]: Therefore, it seems that short-term incubation of adipocytes with 20 μM Olanzapine reduces the rate of lipolysis, while the therapeutic concentrations do not seem to alter lipolysis in adipocytes., This work is being supported by Marie Skłodowska Curie Actions (H2020-MSCA-ITN-2016)., Peer reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: Second-generation Antipsychotics (SGAs) are preferable pharmacological treatment for patients with Schizophrenia, mainly due to their efficacy and reduced risk of extrapyramidal effects when compared with first generation antipsychotics. However, there are metabolic side effects associated with the administration of SGAs, namely weight gain, dyslipidaemia and impaired glucose metabolism. Literature reports that even in the absence of antipsychotic treatment patients with Schizophrenia have a propensity to develop metabolic changes that can lead to cardiovascular diseases, insulin resistance, obesity and Type 2 Diabetes. Olanzapine and aripiprazole belong to SGAs and have been reported as drugs with the highest and the lowest risk of inducing metabolic changes, respectively. However, the pharmacological mechanisms underlying their metabolic side effects remain unclear and they will be the main focus of our study. Adipose tissue is not only specialized in storing lipids but it is also an endocrine organ that produces and secretes numerous
biological active compounds that regulate metabolic homeostasis. Our aim is to evaluate the effect of olanzapine and aripiprazole in the glucose uptake on human isolated adipocytes., [Methods]: Biopsies of subcutaneous adipose tissue were collected from 16 healthy volunteers (4 men, 12 women; age: 20-76 years; BMI: 20.9-34.5 kg/m2). Subjects taking antidepressants or antipsychotics were not included. The effect of short-term incubation (30 min) with different concentrations of olanzapine (0.004, 0.04, 0.1, 0.2, 2 and 20 μM) or aripiprazole (0.02, 0.2, 0.5, 1, 10 and 100 μM) on basal and insulin-stimulated (25 and 1000 μU/ml) D-[U-14C]-glucose uptake of isolated adipocytes
was measured and compared with control., [Results]: Short incubation of adipocytes with olanzapine or aripiprazole showed no effect on basal or insulin-stimulated glucose uptake, with the exception of supra-physiological concentrations of aripiprazole (10 and 100 μM) where we see a systematic decrease of basal and insulin-stimulated glucose uptake; 10 μM by ͌20-25% (p<0.05) and 100 μM by ͌60-70% (p<0.01)., [Conclusions]: Short-term treatment of isolated adipocytes with therapeutic doses of olanzapine or aripiprazole did not affect glucose uptake, suggesting no acute alteration in insulin activity. These data suggest that the plasma glucose increase seen in patients taking Olanzapine cannot be justified by acute alteration in insulin-signalling in adipocytes. Incubation with aripiprazole at 10 and 100 μM decreased the adipocyte glucose uptake but this might be justified by cell death, which will be explored in future experiments., This work is being supported by Marie Skłodowska Curie Actions (H2020-MSCA-ITN-2016)., Peer reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: The liver is the main organ in charge of drug catabolism and also the major site susceptible to drug injury. Sirtuin 1 (SIRT1), a NAD+-dependent histone deacetylase, is a key player in liver physiology and a therapeutic target against hepatic inflammation. In this study, we evaluated the role of SIRT1 in the pro-inflammatory context and oxidative stress during acetaminophen (APAP)-mediated hepatotoxicity., [Material and Methods]: SIRT1 expression was analyzed in APAP-induced liver failure in humans and mice. Hepatotoxicity was assessed in wild-type and transgenic mice overexpressing SIRT1 (SIRT1 Tg) poisoned with APAP (300 mg/kg). Raw 264.7 and peritoneal macrophages were treated with APAP and conditioned medium (CM) was added to mouse hepatocytes. siRNA was used to reduce inflammatory mediators in hepatocytes., [Results]: SIRT1 protein levels decreased in human and mouse livers following APAP overdose. SIRT1-Tg mice maintained higher levels of SIRT1 upon APAP injection than wild-type mice and were protected against hepatotoxicity by modulation of antioxidant systems and restrained inflammatory responses, with decreased oxidative stress, pro-inflammatory cytokine mRNA levels, nuclear factor kappa B (NFκB) signaling, and cell death. Mouse hepatocytes stimulated with conditioned medium of APAP-treated macrophages (APAP-CM) showed decreased SIRT1 levels; an effect mimicked by interleukin 1β (IL1β), an activator of NFκB. This negative modulation was abolished by neutralizing IL1β in APAP-CM or silencing p65-NFκB in hepatocytes. APAP-CM of macrophages from SIRT1-Tg mice failed to downregulate SIRT1 protein levels in hepatocytes. In vivo administration of the NFκB inhibitor BAY 11-7082 preserved SIRT1 levels and protected from APAP-mediated hepatotoxicity., [Conclusion]: SIRT1 protein levels are downregulated by IL1β/NFκB signaling in APAP hepatotoxicity, resulting in inflammation and oxidative stress. Thus, maintenance of SIRT1 during APAP overdose by inhibiting NFκB might be clinically relevant. Our work evidenced the unique role of SIRT1 in APAP hepatoprotection by targeting oxidative stress and inflammation., This work was funded by SAF2015-65267-R (MINECO/FEDER), CIBERdem (ISCIII, Spain), INFLAMES (ISCIII PIE14/00045, co-funded by ERDF, “Investing in your future”) to A.M.V; IJCI-2014-19381 (MINECO/FEDER) to P.R and A.M.V; SAF2014-52009-R (MINECO/FEDER) to S.A; SAF2015-63904-R (MINECO/FEDER) to M.M.; CP14/00181 (ISCIII/FEDER) to A. G-R; PI13/00021 (ISCIII/FEDER) to J.M. CIBERehd (ISCIII, Spain) to A. G-R and J.M; Grant 37/371 from Al Jouf University (to M.A.M). We also acknowledge H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement number 721236) (European Commission)., Peer reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: Schizophrenia (SCZ) is a chronic brain disorder with symptoms of hallucinations, delusions, disturbed behavior and emotional and cognitive impairment. These symptoms are managed by administration of antipsychotic drugs (APDs), either as a single APD or a combination of different APDs depending on the individual. It is always recommended to take a maintenance dose of APD even after the reduction of SCZ symptoms which makes it a lifelong treatment. APD treatment is currently known to be the only effective treatment against SCZ. However, they are observed to trigger metabolic
dysfunctions such as insulin resistance, obesity, coronary heart diseases and dyslipedemia in SCZ patients at the later stage during APD treatment. The main aim of the project is to identify a biomarker signature that could be detected in blood to diagnose the metabolic dysfunction due to APD administration at an early stage. The talk will provide an insight into omic approaches for biomarker discovery and a worked example of these approaches using relevant existing data sets from public
repositories., [Materials and Methods]: The public repository Array Express (AE) (https://www.ebi.ac.uk/arrayexpress/) was used to shortlist the datasets from transcriptomic studies of primary human hepatocytes (PHH) that has been treated with any antipsychotic drug. R studio was used to process the data and shortlist genes that had a minimum fold change of 2. Bioinfominer online tool (https://bioinfominer.com/) was used for Gene ontology enrichment analysis., [Results and conclusion]: We found a microarray dataset of PHH that has been treated with Chlorpromazine (APD) from AE (AE id: E-MTAB-1747). The deregulated gene list from PHH that were treated with CPZ was significantly grouped under inflammatory and immune response from the gene ontology term enrichment analysis. Since the gene products of the inflammatory/immune response are mostly secreted, they are likely to be detected in blood which makes them potential biomarker candidates. However, these candidates were identified from PHH isolated from a single donor, so more work would be required to confirm them as valid biomarkers. Despite that caveat, the gene list can be considered as a useful reference list for future experiments., H2020-MSCA-ETN-2016-72123., Peer reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: The P-glycoprotein (P-gp) is an efflux pump widely expressed in the organism that exports xenobiotic compounds out of the tissue where it is expressed. It plays a central role in the Blood-Brain Barrier permeability, being responsible of Central Nervous System side-effects or ineffectiveness of many drugs. Several single-nucleotide polymorphisms (SNPs) in ABCB1 (the gene
encoding for P-gp) have been identified. The most relevant ones, C3435T, C2677T/A and C1236T have been associated with variable pharmacokinetic parameters in healthy volunteers that received single oral doses of antidepressants and antipsychotics. There is no consensus regarding the in vivo effect they have. Here, we compile relevant information in order to simplify the understanding of
materials, methods and cell lines classically used to assess polymorphic P-gp activity in in vitro cell culture models., [Methods]: A comprehensive research of the studies performed in this regard has been accomplished. More than 389 articles have been reviewed, corresponding to the topics “ABCB1 polymorphisms”, “assessment of P-gp function” and “P-gp expression in cell cultures” published in PubMed Search Engine., [Results]: Twenty-four articles have been summarised and classified. Site-directed mutagenesis has been acknowledged as a convenient approach to obtain cell lines expressing mutant P-gp. Ten different techniques have been identified as key in the assessment of P-gp function: Transfection; Western Blot; Flow Cytometry; Transcriptional Analysis; TEER measurements; Calcein-AM, Rhodamine-123 or Radioactivity based accumulation and transport assays; Transwell® inserts; MTT viability assays., [Conclusion]: Site-directed mutagenesis performed in plasmids that contain wild-type ABCB1, followed by transfection of the plasmid into cells (HeLa, Caco-2 cells) may lead to cell lines expressing P-gp with the SNPs of interest (C3435T, C2677T/A and C1236T). Assessment of P-gp function may be accomplished by Calcein-AM or Rhodamine-123 accumulation assays. The actual effect of these SNPs in P-gp on antidepressants or antipsychotics efflux through membranes could be assessed by Transwell® insert transport assays., P. Zubiaur is co-financed by Consejería de Educación, Juventud y Deporte from Comunidad de Madrid and Fondo Social Europeo. D. Koller is co-financed by the H2020 Marie Sklodowska-Curie Innovative Training Network 721236 grant., Peer reviewed

[Objectives]: Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. Protein tyrosine phosphatase 1B (PTP1B), a negative modulator of insulin and cytokine signaling, is a therapeutic target for type 2 diabetes and obesity. We investigated the impact of PTP1B deficiency during NAFLD, particularly in non-alcoholic steatohepatitis (NASH). [Methods]: NASH features were evaluated in livers from wild-type (PTP1BWT) and PTP1B-deficient (PTP1BKO) mice fed methionine/choline-deficient diet (MCD) for 8 weeks. A recovery model was established by replacing MCD to chow diet (CHD) for 2–7 days. Non-parenchymal liver cells (NPCs) were analyzed by flow cytometry. Oval cells markers were measured in human and mouse livers with NASH, and in oval cells from PTP1BWT and PTP1BKO mice. [Results]: PTP1BWT mice fed MCD for 8 weeks exhibited NASH, NPCs infiltration, and elevated Fgf21, Il6 and Il1b mRNAs. These parameters decreased after switching to CHD. PTP1B deficiency accelerated MCD-induced NASH. Conversely, after switching to CHD, PTP1BKO mice rapidly reverted NASH compared to PTP1BWT mice in parallel to the normalization of serum triglycerides (TG) levels. Among NPCs, a drop in cytotoxic natural killer T (NKT) subpopulation was detected in PTP1BKO livers during recovery, and in these conditions M2 macrophage markers were up-regulated. Oval cells markers (EpCAM and cytokeratin 19) significantly increased during NASH only in PTP1B-deficient livers. HGF-mediated signaling and proliferative capacity were enhanced in PTP1BKO oval cells. In NASH patients, oval cells markers were also elevated. [Conclusions]: PTP1B elicits a dual role in NASH progression and reversion. Additionally, our results support a new role for PTP1B in oval cell proliferation during NAFLD., This work was funded by SAF2015-65267-R (MINECO/FEDER), CIBERdem (ISCIII, Spain), INFLAMES (ISCIII PIE14/00045, co-funded by ERDF, “Investing in your future”) to A.M.V; IJCI-2014-19381 (MINECO/FEDER) to P.R and A.M.V; CP14/00181 and PI16/00823 (ISCIII/FEDER) and CIBERehd (ISCIII, Spain) to A.G-R; BFU2015-65937-R (MINECO/FEDER), 67-2015 (Department of Health, Navarra Government) and CIBERobn (ISCIII, Spain) to M.J.M; SAF2015-69145-R (MINECO/FEDER) to A.S. A.A. was recipient of a Marie Curie ESR contract from IT-LIVER (Marie Curie Action of the FP7-2012, grant agreement 316549). We also acknowledge the European Union, project H2020-MSCA-ITN TREATMENT Grant Agreement number: 721236., Peer Reviewed

[Background]: Agomelatine is an agonist of the melatoninergic receptors used for the treatment of depression. Our aim was to evaluate the effect of genetic polymorphisms in metabolising enzymes and the P-glycoprotein transporter on agomelatine pharmacokinetics and pharmacodynamics., [Methods]: Twenty-eight healthy volunteers receiving a single 25 mg oral dose of agomelatine, were genotyped for nine polymorphisms in cytochrome P450 enzymes (CYP1A2, CYP2C9 and CYP2C19) and adenosine triphosphate-binding cassette subfamily B member 1 (ABCB1), by real-time polymerase chain reaction. Agomelatine concentrations were measured by high performance liquid chromatography coupled to a tandem mass spectrometry detector., [Results]: We calculated a CYP1A2 activity score that was directly correlated with agomelatine pharmacokinetics. Individuals with a decreased enzyme activity (*1C carriers) had a lower clearance and accumulated higher concentrations of agomelatine. In contrast, individuals with a high CYP1A2 inducibility (*1F or *1B carriers) showed an extensive clearance and lower agomelatine concentrations. The apparently marked differences between races were due to the different CYP1A2 genotype distribution. CYP2C9 intermediate/poor metabolisers showed a higher area under the concentration-time curve and maximum concentration. ABCB1 G2677T/A polymorphism affected the time to reach maximum concentration, as subjects carrying A/A+A/T genotypes showed higher values. No association was found for CYP2C19 phenotype. Agomelatine did not produce any change in blood pressure, heart rate or QT interval., [Conclusions]: CYP1A2 polymorphisms affect agomelatine pharmacokinetics. CYP1A2 phenotype inferred from the genotyping of CYP1A2*1C, *1F and *1B alleles might be a potential predictor of agomelatine exposure. ABCB1 G2677T/A could affect agomelatine absorption, as subjects with A/A+A/T genotypes had lower agomelatine concentration and they take more time to reach the maximum concentration., D. Koller is co-financed by the H2020 Marie Sklodowska-Curie Innovative Training Network 721236 grant. P. Zubiaur is co-financed by Consejería de Educación, Juventud y Deporte from Comunidad de Madrid and Fondo Social Europeo, Peer reviewed

Inflammation is typically associated with the development of fibrosis, cirrhosis and hepatocellular carcinoma. The key role of protein tyrosine phosphatase 1B (PTP1B) in inflammatory responses has focused this study in understanding its implication in liver fibrosis. Here we show that hepatic PTP1B mRNA expression increased after bile duct ligation (BDL), while BDL-induced liver fibrosis was markedly reduced in mice lacking Ptpn1 (PTP1B−/−) as assessed by decreased collagen deposition and α-smooth muscle actin (α-SMA) expression. PTP1B−/− mice also showed a significant increase in mRNA levels of key markers of monocytes recruitment (Cd68, Adgre1 and Ccl2) compared to their wild-type (PTP1B+/+) littermates at early stages of injury after BDL. Interestingly, the lack of PTP1B strongly increased the NADPH oxidase (NOX) subunits Nox1/Nox4 ratio and downregulated Cybb expression after BDL, revealing a pro-survival pattern of NADPH oxidase induction in response to liver injury. Chimeric mice generated by transplantation of PTP1B−/− bone marrow (BM) into irradiated PTP1B+/+ mice revealed similar hepatic expression profile of NOX subunits than PTP1B−/− mice while these animals did not show differences in infiltration of myeloid cells at 7 days post-BDL, suggesting that PTP1B deletion in other liver cells is necessary for boosting the early inflammatory response to the BDL. PTP1B−/− BM transplantation into PTP1B+/+ mice also led to a blockade of TGF-β and α-SMA induction after BDL. In vitro experiments demonstrated that deficiency of PTP1B in hepatocytes protects against bile acid-induced apoptosis and abrogates hepatic stellate cells (HSC) activation, an effect ameliorated by NOX1 inhibition. In conclusion, our results have revealed that the lack of PTP1B switches NOX expression pattern in response to liver injury after BDL and reduces HSC activation and liver fibrosis., This work was funded by grants SAF2015-65267-R and RTI2018-094052-B-100 (MICINN/FEDER, Spain), S2017/BMD-3684 MOIR2-CM (Comunidad de Madrid, Spain) and CIBERdem (ISCIII, Spain) to A.M.V; IJCI-2014-19381 (MINECO/FEDER, Spain) to P.R and A.M.V. We also acknowledge H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement 721236, European Commission)., Peer reviewed

Insulin receptor substrate 2 (IRS2) is a key downstream mediator of insulin and insulin-like growth factor 1 (IGF1) signalling pathways and plays a major role in liver metabolism. The aim of this study was to investigate whether IRS2 had an impact on the hepatic fibrotic process associated with cholestatic injury. Bile duct ligation (BDL) was performed in wild-type (WT) and Irs2-deficient (IRS2KO) female mice. Histological and biochemical analyses, together with fibrogenic and inflammatory responses were evaluated in livers from mice at 3, 7 and 28 days following BDL. We also explored whether activation of human hepatic stellate cells (HSCs) induced by IGF1 was modulated by IRS2. IRS2KO mice displayed reduced disruption of liver histology, such hepatocyte damage and excess deposition of extracellular matrix components, compared with WT mice at 3 and 7 days post-BDL. However, no histological differences between genotypes were found at 28 days post-BDL. The less pro-inflammatory profile of bile acids accumulated in the gallbladder of IRS2KO mice after BDL corresponded with the reduced expression of pro-inflammatory markers in these mice. Stable silencing of IRS2 or inhibition of ERK1/2 reduced the activation of human LX2 cells and also reduced induction of MMP9 upon IGF1 stimulation. Furthermore, hepatic MMP9 expression was strongly induced after BDL in WT mice, but only a slight increase was found in mice lacking IRS2. Our results have unravelled the signalling pathway mediated by IGF1R-IRS2-ERK1/2-MMP9 as a key axis in regulating HSC activation, which might be therapeutically relevant for targeting liver fibrosis., This work was supported by the Instituto de Salud Carlos III (ISCIII/FEDER) (PI17/00535 and CIBEREHD) to C.G.-M.; Ministerio de Ciencia e Innovación (MICINN/FEDER) (RTI2018-094052-B-100), Comunidad de Madrid (S2017/BMD-3684 MOIR2-CM), Instituto de Salud Carlos III (ISCIII/FEDER) (CIBERDEM) and H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant agreement number 721236) to A.M.V.; Instituto de Salud Carlos III (ISCIII/FEDER) (CP14/00181 and PI16/00823) and Beca Eduardo Gallego 2016 (Fundación Francisco Cobos) to A.G.-R., Peer reviewed

This article belongs to the Special Issue Cellular Stress Response in Health and Disease., The interplay of mitochondria with the endoplasmic reticulum and their connections, called mitochondria-ER contacts (MERCs) or mitochondria-associated ER membranes (MAMs), are crucial hubs in cellular stress. These sites are essential for the passage of calcium ions, reactive oxygen species delivery, the sorting of lipids in whole-body metabolism. In this perspective article, we focus on microscopic evidences of the pivotal role of MERCs/MAMs and their changes in metabolic diseases, like obesity, diabetes, and neurodegeneration., This research was funded by FFARB 2017 of A.S., ex 60% grants of University of Brescia (Italy) of A.S. and R.R. and Spanish “Ministerio de Economía Industria y Competitividad” (MINEICO) that includes FEDER funds (SAF2015-63904-R) and from the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie agreement 721236-TREATMENT of M.M., Peer reviewed

Resumen del póster presentado al 6th Symposium on Biomedical Research: Advances and Perspectives in Molecular Endocrinology "In Homage to Gabriella Morreale", celebrado en el Instituto de Investigaciones Biomédicas Alberto Sols (IIBM-CSIC) el 31 de mayo de 2019., Schizophrenia is a chronic mental disorder that affects approximately 50 million people worldwide. Also, it is associated with psychotic experiences that not allow patients to have a normal life. Fortunately, treatments used can suppress the symptomatology and lead to a productive life and integration in the society. According to the current guidelines, the first line in schizophrenia’s therapy are the second generation antipsychotic drugs (SGA). Besides being D2 receptors antagonists/partial agonists, the SGAs have the ability to interact also with serotonergic, histaminergic and other receptors; minimizing
the symptomatology and not resulting in neurological side effects (contrarily to first generation antipsychotic drugs). Nevertheless, recent clinical observations show a variety of metabolic dysfunctions in patients under SGAs treatment, such as abnormal gain weight, hyperglycemia and dyslipidemia. However, the molecular mechanisms behind these alterations are still very poorly understood. Having this in consideration, the objective of this work was to study the effect of Olanzapine administration in wild-type (WT) and PTP1B-deficient (KO) mice on glucose homeostasis and the mechanisms involved. Of interest, PTP1B is the main tyrosine phosphatase of the insulin receptor, and PTP1B KO mice are protected against insulin resistance and development of type 2 diabetes mellitus. After administration of 10 mg/kg/day of Olanzapine for 8 weeks to WT and PTP1B-KO mice, the glucose homeostasis tests suggest that Olanzapine induced insulin resistance only in WT mice. Also, primary hepatocytes from WT mice treated with Olanzapine showed lower glucose uptake than those from non-treated WT mice. In contrast, Olanzapine treatment of PTP1B KO mice significantly increased hepatocyte glucose uptake, suggesting that Olanzapine-induced insulin resistance is at least in part dependent on PTP1B. These results support the relevance of PTP1B in
Olanzapine-induced insulin resistance in hepatocytes., European Union's EU Framework Programme for Research and Innovation Horizon 2020. ITN-TREATMENT under Grant Agreement No GA 721236., Peer reviewed

Melatonin, an indole produced by pineal and extrapineal tissues, but also taken with a vegetarian diet, has strong anti-oxidant, anti-inflammatory and anti-obesogenic potentials. Non-alcoholic fatty liver disease (NAFLD) is the hepatic side of the metabolic syndrome. NAFLD is a still reversible phase but may evolve into steatohepatitis (NASH), cirrhosis and carcinoma. Currently, an effective therapy for blocking NAFLD staging is lacking. Silent information regulator 1 (SIRT1), a NAD+ dependent histone deacetylase, modulates the energetic metabolism in the liver. Micro-RNA-34a-5p, a direct inhibitor of SIRT1, is an emerging indicator of NAFLD grading. Thus, here we analyzed the effects of oral melatonin against NAFLD and underlying molecular mechanisms, focusing on steatosis, ER stress, mitochondrial shape and autophagy. Male C57BL/6J (WT) and SIRT1 heterozygous (HET) mice were placed either on a high-fat diet (58.4% energy from lard) (HFD) or on a standard maintenance diet (8.4% energy from lipids) for 16 weeks, drinking melatonin (10 mg/kg) or not. Indirect calorimetry, glucose tolerance, steatosis, inflammation, ER stress, mitochondrial changes, autophagy and microRNA-34a-5p expression were estimated. Melatonin improved hepatic metabolism and steatosis, influenced ER stress and mitochondrial shape, and promoted autophagy in WT HFD mice. Conversely, melatonin was ineffective in HET HFD mice, maintaining NASH changes. Indeed, autophagy was inconsistent in HET HFD or starved mice, as indicated by LC3II/LC3I ratio, p62/SQSTM1 and autophagosomes estimation. The beneficial role of melatonin in dietary induced NAFLD/NASH in mice was related to reduced expression of microRNA-34a-5p and sterol regulatory element-binding protein (SREBP1) but only in the presence of full SIRT1 availability., This research was funded by FFABR 2017, 60% from grants of University of Brescia (Italy) and grants by the Spanish “Ministerio de Economía Industria y Competitividad” (MINEICO) including FEDER funds (SAF2015-63904-R) and the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie agreement 721236-TREATMENT to M.M., Peer reviewed

Las alteraciones identificables tanto en estructura como en marcadores moleculares mediante análisis histológico de muestras de tejido tiroideo son esenciales tanto para su clasificación como para la estadificación del riesgo., Este trabajo ha sido realizado con fondos aportados por proyectos de investigación del Ministerio de Economía Industria y Competitividad (MINEICO) y fondos FEDER [proyectos con referencias
SAF2012-37693, SAF2015-63904-R, SAF2015-71521-REDC] así como por el proyecto europeo 721236-TREATMENT del programa marco Horizon 2020., Peer reviewed

This article belongs to the Special Issue Oxidative stress and Applied Biology., High levels of reactive oxygen species (ROS) can lead to impairment of cell structure, biomolecules' loss of function and cell death and are associated with liver diseases. Cells that survive increased ROS often undergo malignant transformation. Many cancer cells tolerate high levels of ROS. Here we report a transiently increased production of H2O2 and concomitant upregulation of antioxidative enzymes triggered by hepatocyte isolation; the H2O2 levels revert in about two days in culture. Three-day survival rate of the isolated cells in the presence of 2.5-fold increase of H2O2 is almost 80%. Apoptosis activation through the mitochondrial pathway is meanwhile reduced by inhibition of caspase-9 triggering. This reduction depends on the amount of H2O2 production, as decreased production of H2O2 in the presence of an antioxidant results in increased apoptosis triggering. These stress adaptations do not influence urea production, which is unchanged throughout the normal and stress adapted phases. We conclude that hepatocytes' stress adaptation is mediated by increased ROS production. In this case, high ROS improve cell survival., This research was funded by the Slovenian Research Agency (research core funding No. P3-0019).
This work was also partially supported by the H2020-MSCA-ITN:721236 TREATMENT project., Peer reviewed

Schizophrenia is a mental disease that results in decreased life expectancy and wellbeing, by promoting obesity and sedentary lifestyles. Schizophrenia is treated by antipsychotic drugs. While the second generation of antipsychotics (SGA), Olanzapine and Aripiprazole are more effective in treating schizophrenia, they display a higher risk of metabolic side effects, mostly by development of diabetes and insulin resistance, weight gain as well as dyslipidemia. Endoplasmic reticulum (ER) stress is induced when ER homeostasis of lipid biosynthesis and protein folding is impaired. This leads to the activation of the unfolded protein response (UPR), a signaling cascade that aims to restore ER homeostasis or initiate cell death. Chronic conditions of ER stress in the liver are associated with diabetes and perturbed lipid metabolism. These metabolic dysfunctions resemble the pharmacological side effects of SGAs. We, therefore, investigated whether SGAs promote the UPR in human and mouse hepatocytes. We observed full-fledged activation of ER stress by Aripiprazole, not by Olanzapine. This occurred at low micromolar concentrations and to variable intensities in different cell types, such as hepatocellular carcinoma, melanoma and glioblastoma. Mechanistically, Aripiprazole caused depletion of ER calcium, leading to activation of IRE1 and PERK, two major transducers of the UPR. Cells underwent apoptosis upon Aripiprazole treatment, which coincided with UPR induction, and this effect was reduced by adding glutathione without affecting UPR itself. Deletion of IRE1 from HepG2 cells protected cells from Aripiprazole toxicity. Our study reveals for the first time a cytotoxic effect of Aripiprazole that involves the induction of ER stress., FF, PD and VF are Marie Curie fellows [Treatment H2020-MSCA-ITN721236]. AMV, PR and IGM were supported by grants [RTI2018-094052-B-100] (MCIU/AEI/FEDER, UE)) and [S2017/BMD-3684] MOIR2-CM (Comunidad de Madrid, Spain) and CIBERdem (ISCIII, Spain)., Peer reviewed

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease around the world estimated to affect up to one-third of the adult population and is expected to continue rising in the coming years. Nonalcoholic fatty liver disease is considered as the hepatic manifestation of the metabolic syndrome because it is strongly associated with obesity, insulin resistance, type 2 diabetes mellitus, and cardiovascular complications. Despite its high prevalence, factors leading to NAFLD progression from simple steatosis to nonalcoholic steatohepatitis, cirrhosis, and, ultimately hepatocellular carcinoma remain poorly understood. To date, no treatment has proven efficacy, and also no reliable method is currently available for diagnosis or staging of NAFLD beyond the highly invasive liver biopsy. Recently, extracellular vesicles (EVs) have emerged as potential candidate biomarkers for the diagnosis of NAFLD. Extracellular vesicles are circulating, cell-derived vesicles containing proteins and nucleic acids, among other components, that interact with and trigger a plethora of responses in neighbor or distant target cells. Several mechanisms implicated in NAFLD progression, such as inflammation, fibrosis, and angiogenesis, all related to metabolic syndrome–associated lipotoxicity, trigger EV production and release by liver cells. As hepatocytes represent ~80% of the liver volume, in this review we will focus on hepatocyte-derived EVs as drivers of the interactome between different liver cell types in NAFLD pathogenesis, as well as in their role as noninvasive biomarkers for NAFLD diagnosis and progression. Based on that, we will highlight the research that is currently available on EVs in this topic, the current limitations, and future directions for implementation in a clinical setting as biomarkers or targets of liver disease., We acknowledge grant RTI2018-094052-B-100 (MCIU/AEI/FEDER, UE), Fundación Ramón Areces (Spain), S2017/BMD-3684 (Comunidad de Madrid, Spain), and H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement 721236, European Commission)., Peer reviewed

MicroRNAs (miRNAs) regulate gene expression posttranscriptionally and control biological processes (BPs), including fibrogenesis. Kidney fibrosis remains a clinical challenge and miRNAs may represent a valid therapeutic avenue. We show that miR‐9‐5p protected from renal fibrosis in the mouse model of unilateral ureteral obstruction (UUO). This was reflected in reduced expression of pro‐fibrotic markers, decreased number of infiltrating monocytes/macrophages, and diminished tubular epithelial cell injury and transforming growth factor‐beta 1 (TGF‐β1)‐dependent de‐differentiation in human kidney proximal tubular (HKC‐8) cells. RNA‐sequencing (RNA‐Seq) studies in the UUO model revealed that treatment with miR‐9‐5p prevented the downregulation of genes related to key metabolic pathways, including mitochondrial function, oxidative phosphorylation (OXPHOS), fatty acid oxidation (FAO), and glycolysis. Studies in human tubular epithelial cells demonstrated that miR‐9‐5p impeded TGF‐β1‐induced bioenergetics derangement. The expression of the FAO‐related axis peroxisome proliferator‐activated receptor gamma coactivator 1 alpha (PGC‐1α)‐peroxisome proliferator‐activated receptor alpha (PPARα) was reduced by UUO, although preserved by the administration of miR‐9‐5p. We found that in mice null for the mitochondrial master regulator PGC‐1α, miR‐9‐5p was unable to promote a protective effect in the UUO model. We propose that miR‐9‐5p elicits a protective response to chronic kidney injury and renal fibrosis by inducing reprogramming of the metabolic derangement and mitochondrial dysfunction affecting tubular epithelial cells., This work was supported by Grants from the Ministerio de Economía y Competitividad (MINECO) SAF 2012‐31388 (SL) and SAF2015‐66107‐R (SL), both cofunded by the European Regional Development Fund, Instituto de Salud Carlos III REDinREN RD12/0021/0009 and RD16/0009/0016 (SL and MRO), PI17/00119 (MRO), SAF2015‐63904‐R (MM), cofunded by the European Regional Development Fund, European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska‐Curie grant agreement 721236‐TREATMENT (MM), Comunidad de Madrid “NOVELREN” B2017/BMD­3751 (SL and MRO), and Fundación Renal “Iñigo Alvarez de Toledo” (SL), all from Spain. The CBMSO receives institutional support from Fundación “Ramón Areces”. VM and CNT were supported by predoctoral fellowships of the FPI Program (BES‐2013‐065986 and BES‐2014‐068929) from MINECO., Peer reviewed

Hepatic fat-specific protein 27 [cell death-inducing DNA fragmentation effector protein C (Cidec)/Fsp27] mRNA levels have been associated with hepatic lipid droplet extent under certain circumstances. To address its hepatic expression under different dietary conditions and in both sexes, apolipoprotein E (Apoe)-deficient mice were subjected to different experimental conditions for 11 wk to test the influence of cholesterol, Western diet, squalene, oleanolic acid, sex, and surgical castration on Cidec/Fsp27 mRNA expression. Dietary cholesterol increased hepatic Cidec/Fsp27β expression, an effect that was suppressed when cholesterol was combined with saturated fat as represented by Western diet feeding. Using the latter diet, neither oleanolic acid nor squalene modified its expression. Females showed lower levels of hepatic Cidec/Fsp27β expression than males when they were fed Western diets, a result that was translated into a lesser amount of CIDEC/FSP27 protein in lipid droplets and microsomes. This was also confirmed in low-density lipoprotein receptor (Ldlr)-deficient mice. Incubation with estradiol resulted in decreased Cidec/Fsp27β expression in AML12 cells. Whereas male surgical castration did not modify the expression, ovariectomized females did show increased levels compared with control females. Females also showed increased expression of peroxisome proliferator-activated receptor-γ coactivator 1-α (Pgc1a), suppressed by ovariectomy, and the values were significantly and inversely associated with those of Cidec/Fsp27β. When Pgc1a-deficient mice were used, the sex differences in Cidec/Fsp27β expression disappeared. Therefore, hepatic Cidec/Fsp27β expression has a complex regulation influenced by diet and sex hormonal milieu. The mRNA sex differences are controlled by Pgc1a., This research was supported by Ministerio de Economía y CompetitividadFondo Europeo de Desarrollo Regional (Grants SAF2015-63904-R, SAF2016-75441-R, and RTI2018-093864-B-I00), Fondo Social Europeo-Gobierno de Aragón (Grant B16_17R), and the European Union’s Horizon 2020 research and innovation program under Marie Skłodowska-Curie Grant Agreement
721236-TREATMENT to M. Monsalve. Centro de Investigación Biomédica en Red Fisiopatología de la Obesidad y Nutrición (CB06/03/1012) is an initiative of Instituto de Salud Carlos III. L. V. Herrera-Marcos and S. Sancho-Knapik were recipients of Fondo Social Europeo-Gobierno de Aragón and Fundación Cuenca-Villoro (BE 203/2009) fellowships, respectively.

Comunicación presentada en el 6th Symposium on Biomedical Research Advances and Perspectives in Molecular Endocrinology, celebrado en Madrid (España), el 31 de mayo de 2019., Schizophrenia is a chronic mental disorder that affects approximately 50 million people worldwide. Also, it is associated with psychotic experiences that not allow patients to have a normal life. Fortunately, treatments used can suppress the
symptomatology and lead to a productive life and integration in the society. According to the current guidelines, the first line in schizophrenia’s therapy are the second generation antipsychotic drugs (SGA). Besides being D2 receptors antagonists/ partial agonists, the SGAs have the ability to interact also with serotonergic, histaminergic and other receptors; minimizing the symptomatology and not resulting in neurological side effects (contrarily to first generation antipsychotic drugs). Nevertheless, recent clinical observations show a variety of metabolic dysfunctions in patients under SGAs treatment, such as abnormal gain weight, hyperglycemia and dyslipidemia. However, the molecular mechanisms behind these alterations are still very poorly understood. Having this in consideration, the objective of this work was to study the effect of Olanzapine administration in wild-type (WT) and PTP1B-deficient (KO) mice on glucose homeostasis and the mechanisms involved. Of interest, PTP1B is the main tyrosine phosphatase of the insulin receptor, and PTP1B KO mice are protected against insulin resistance and development of type 2 diabetes mellitus. After administration of 10 mg/kg/day of Olanzapine for 8 weeks to WT and PTP1B-KO mice, the glucose homeostasis tests suggest that Olanzapine induced insulin resistance only in WT mice. Also, primary hepatocytes from WT mice treated with Olanzapine showed lower glucose uptake than those from non-treated WT mice. In contrast, Olanzapine treatment of PTP1B KO mice significantly increased hepatocyte glucose uptake, suggesting that Olanzapine-induced insulin resistance is at least in part dependent on PTP1B. These results support the relevance of PTP1B in Olanzapine-induced insulin resistance in hepatocytes., European Union's EU Framework Programme for Research and Innovation Horizon 202. ITN-TREATMENT under Grant Agreement No GA 721236

© 2020 by the authors., Cancer is one of the highest prevalent diseases in humans. The chances of surviving cancer and its prognosis are very dependent on the affected tissue, body location, and stage at which the disease is diagnosed. Researchers and pharmaceutical companies worldwide are pursuing many attempts to look for compounds to treat this malignancy. Most of the current strategies to fight cancer implicate the use of compounds acting on DNA damage checkpoints, non-receptor tyrosine kinases activities, regulators of the hedgehog signaling pathways, and metabolic adaptations placed in cancer. In the last decade, the finding of a lipid peroxidation increase linked to 15-lipoxygenases isoform 1 (15-LOX-1) activity stimulation has been found in specific successful treatments against cancer. This discovery contrasts with the production of other lipid oxidation signatures generated by stimulation of other lipoxygenases such as 5-LOX and 12-LOX, and cyclooxygenase (COX-2) activities, which have been suggested as cancer biomarkers and which inhibitors present anti-tumoral and antiproliferative activities. These findings support the previously proposed role of lipid hydroperoxides and their metabolites as cancer cell mediators. Depletion or promotion of lipid peroxidation is generally related to a specific production source associated with a cancer stage or tissue in which cancer originates. This review highlights the potential therapeutical use of chemical derivatives to stimulate or block specific cellular routes to generate lipid hydroperoxides to treat this disease., This research was funded by Ministerio de Ciencia e Innovación grant number RTI2018-093864-B-I00 and the European Union’s Horizon 2020 Research and Innovation Programme MSCA GA 721236-TREATMENT.

© The Author(s) 2020., Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease worldwide. NAFLD stages range from simple steatosis (NAFL) to non-alcoholic steatohepatitis (NASH) which can progress to cirrhosis and hepatocellular carcinoma. One of the crucial events clearly involved in NAFLD progression is the lipotoxicity resulting from an excessive fatty acid (FFA) influx to hepatocytes. Hepatic lipotoxicity occurs when the capacity of the hepatocyte to manage and export FFAs as triglycerides (TGs) is overwhelmed. This review provides succinct insights into the molecular mechanisms responsible for lipotoxicity in NAFLD, including ER and oxidative stress, autophagy, lipoapotosis and inflammation. In addition, we highlight the role of CD36/FAT fatty acid translocase in NAFLD pathogenesis. Up-to-date, it is well known that CD36 increases FFA uptake and, in the liver, it drives hepatosteatosis onset and might contribute to its progression to NASH. Clinical studies have reinforced the significance of CD36 by showing increased content in the liver of NAFLD patients. Interestingly, circulating levels of a soluble form of CD36 (sCD36) are abnormally elevated in NAFLD patients and positively correlate with the histological grade of hepatic steatosis. In fact, the induction of CD36 translocation to the plasma membrane of the hepatocytes may be a determining factor in the physiopathology of hepatic steatosis in NAFLD patients. Given all these data, targeting the fatty acid translocase CD36 or some of its functional regulators may be a promising therapeutic approach for the prevention and treatment of NAFLD., We acknowledge grants RTI2018-094052-B-100 (MCIU/AEI/FEDER, UE), Fundación Ramón Areces (Spain), S2017/BMD-3684 (Comunidad de Madrid, Spain) and H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement 721236, European Commission) to AMV; grant PI17/00535 and CIBEREHD from Instituto de Salud Carlos III (ISCIII/FEDER, Spain) to CGM; grants PI16/00823, CPII19/0032 and PI19/00123 (ISCIII/FEDER, Spain), and Beca Eduardo Gallego 2016 (Fundación Francisco Cobos, Spain) to AGR. We thankfully acknowledge Esther Rey for helpful assistance.

DATA EXPLANATION:
[Folder 1]. Analysis of Drug concentration in plasma and mitochondria (1h and 24 h). The xls file shows the statistics analysis of the detected concentration in plasma and mitochondrial samples after 1 and 24 hours after adding the treatment by aripripazole (Ari), dehydroaripripazole (Dehydroari) or olanzapine (Ola). The .pzfx file can be opened with GraphPad Prism 8 XML Project and shows the decreased in the concentrations across the time in plasma samples.
[Folder 2]. Seahorse Data Healthy volunteer study. The two xls files show (1) the raw data obtained from the measurement of oxygen flux, oxygen consumption rate [OCR] and proton flux, extracellular acidification rate [ECAR], in the medium immediately surrounding the cells in a microplate in a Seahorse analyzer and (2) the linear regression between these data. And, the .pzfx files can be opened with GraphPad Prism 8 XML Project and show the relationships between some of the parameters showed in xls files., Funding agency and Grant Agreetment Number : European Comission ITN Treatment H2020-MSCA-ITN-721236, Peer reviewed

[Purpose] The pentose phosphate pathway (PPP) is an important component of hepatic intermediary metabolism. Jin et al developed an elegant 13C-NMR method for measuring hepatic PPP flux by quantifying the distribution of glucose 13C-isotopomers formed from [U-13C]glycerol. We demonstrate that this approach can be extended to exogenous [U-13C]fructose and [U-13C]glucose precursors by 13C-NMR analysis of glycogen., [Methods] Twelve male C57BL/6 mice fed standard chow were provided a 55/45 mixture of fructose and glucose at 30% w/v in the drinking water for 18 wk. On the evening before sacrifice, the fructose component was enriched with 20% [U-13C]fructose for 6 mice, while the glucose component was enriched with 20% [U-13C]glucose for the remaining 6 mice. Mice were allowed to feed and drink naturally overnight, and then, euthanized. Livers were freeze-clamped and glycogen was extracted and derivatized for 13C NMR spectroscopy. Flux of each sugar into the PPP relative to its incorporation into glycogen was quantified from selected 13C glycogen isotopomer ratios., [Results] Both [U-13C]fructose and [U-13C]glucose precursors yielded glycogen 13C-isotopomer distributions that were characteristic of PPP activity. The fraction of [U-13C]glucose utilized by the PPP relative to its conversion to glycogen via the direct pathway was 14 ± 1%, while that from [U-13C]fructose relative to its conversion to glycogen via the indirect pathway was significantly lower (10 ± 1%, P = .00032)., [Conclusions] Hepatic PPP fluxes from both [U-13C]glucose and [U-13C]fructose precursors were assessed by 13C NMR analysis of glycogen 13C-isotopomers. Glucose-6-phosphate generated via glucokinase and the direct pathway is preferentially utilized by the PPP., This work was financed by the European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme under project FCT-FEDER-02/SAICT/2017/028147 and through the COMPETE 2020 - Operational Programme for Competitiveness and Internationalisation and Portuguese national funds via FCT – Fundação para a Ciência e a Tecnologia, under project UIDB/04539/2020. The UC-NMR facility is supported in part by FEDER – European Regional Development Fund through the COMPETE Programme and the Portuguese Foundation for Science and Technology through grants POCI-01-0145-FEDER-007440; REEQ/481/QUI/2006, RECI/QEQ-QFI/0168/2012, CENTRO-07-CT62-FEDER-002012, and Rede Nacional de Ressonância Magnética Nuclear. GDB is supported by the European Union’s Horizon 2020 Research and Innovation programme under the Marie Skłodowska-Curie Grant Agreement No. 722619 (Project FOIE GRAS). GDN and ANT are supported by Marie Skłodowska-Curie Grant Agreement No. 721236 (Project TREATMENT)., Peer reviewed

[Objective] To assess adverse events (AEs) and safety of aripiprazole (ARI) and olanzapine (OLA) treatment.

[Methods] Twenty-four healthy volunteers receiving five daily oral doses of 10 mg ARI and 5 mg OLA in a crossover clinical trial were genotyped for 46 polymorphisms in 14 genes by qPCR. Drug plasma concentrations were measured by high-performance liquid chromatography tandem mass spectrometry. Blood pressure (BP) and 12-lead electrocardiogram were measured in supine position. AEs were also recorded.

[Results] ARI decreased diastolic BP on the first day and decreased QTc on the third and fifth day. OLA had a systolic and diastolic BP, heart rate and QTc lowering effect on the first day. Polymorphisms in ADRA2A, COMT, DRD3 and HTR2A genes were significantly associated to these changes. The most frequent adverse drug reactions (ADRs) to ARI were somnolence, headache, insomnia, dizziness, restlessness, palpitations, akathisia and nausea while were somnolence, dizziness, asthenia, constipation, dry mouth, headache and nausea to OLA. Additionally, HTR2A, HTR2C, DRD2, DRD3, OPRM1, UGT1A1 and CYP1A2 polymorphisms had a role in the development of ADRs.

[Conclusions] OLA induced more cardiovascular changes; however, more ADRs were registered to ARI. In addition, some polymorphisms may explain the difference in the incidence of these effects among subjects., D. Koller is co-financed by the H2020 Marie Sklodowska-Curie Innovative Training Network 721236 grant. M. Navares is co-financed by the “Consejería de Educación, Juventud y Deporte” PEJ-2018-TL/BMD-11080 grant from “Comunidad de Madrid” and “Fondo Social Europeo”., Peer reviewed

File 1. This xls files shows the data related to the main charateristics used for the development and validation of the new method as long as the measures for letting assure the repeatability and intermediate precision and accuracy values. The results obtained for the pharmacokinetic parameters are also included in the file
File 2 This jpg file shows the product ion spectra and chemical structures of the antipsychotics and their metabolites obtained by collision-induced dissociation (CID) of the indicated precursor ions [M+H]+. The fragmentation patterns of all analytes are indicated by an arrow on their chemical structure of each analyte. The results are presented as the percentage of counts versus Mass-to-Charge (m/z). All mass peaks have been normalized to the most abundant, A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated in human plasma for the simultaneous determination of aripiprazole (ARI) and its metabolite dehydro-aripiprazole (DARI); olanzapine (OLA), risperidone (RIS), paliperidone (PAL), quetiapine (QUE), clozapine (CLO) and caffeine (CAF). CAF is included to the method because it can have an influence on drug metabolism due to competitive inhibition. The above mentioned compounds and their isotope-labeled internal standards were extracted from 200 µL human plasma samples by both, effective phospholipids-eliminating three-step microelution-solid-phase extraction (µ-SPE) and protein precipitation (PPT) for comparison. A combination of formic acid (0.2%)-acetonitrile (pH 3.0; 65:35, v/v) was used as mobile phase and the chromatogram was run under gradient conditions at a flow rate of 0.6 mL/min. Run time lasted 6 min, followed by a re-equilibration time of 3 min. All analytes were monitored by mass spectrometric detection operating in multiple reaction monitoring mode and the method was validated covering the corresponding therapeutic ranges: 0.18-120 ng/mL for ARI, 0.25-80 ng/mL for DARI, 1.00-100 ng/mL for OLA, 0.70-60 ng/mL for RIS, 0.20-30 ng/mL for PAL, 0.50-160 ng/mL for QUE, 0.50-1000 ng/mL for CLO, and finally 1200-3700 ng/mL for CAF. The method was validated based on the recommendations of regulatory agencies through tests of precision, accuracy, extraction recovery, identity confirmation, trueness, matrix effect, process efficiency, stability, selectivity, linearity and carry-over effect fulfilling the guideline requirements. Our µ-SPE method results in the elimination of more than 99% of early eluting and more than 92% of late-eluting phospholipids compared to PPT. Additionally, the method was successfully applied for quantifying ARI and OLA plasma concentrations from healthy volunteers., European Comission ITN Treatment H2020-MSCA-ITN-721236, Peer reviewed

Aripiprazole treatment in schizophrenic patients was previously associated with lower or normalized prolactin levels. Genetic variants in cytochrome P450 (CYP) (CYP2D6), dopamine receptor (DRD2, DRD3) and serotonin receptor (HTR2A, HTR2C) genes were previously associated with antipsychotic-induced hyperprolactinaemia. Our aim was to evaluate whether aripiprazole affects prolactin secretion and its relationship with pharmacogenetics. Thirty-one healthy volunteers receiving a 10-mg single oral dose of aripiprazole were genotyped for 12 polymorphisms in CYP2D6, DRD2, DRD3, HTR2A and HTR2C genes by qPCR. Aripiprazole and dehydro-aripiprazole plasma concentrations were measured by HPLC-MS/MS. Prolactin concentrations of the 31 volunteers taking aripiprazole and 12 volunteers receiving ibuprofen were determined by ELISA. Prolactin concentrations after ibuprofen intake were considered as control, since it is known to cause no effect. Prolactin concentrations were slightly higher in the aripiprazole group compared to the ibuprofen group. All prolactin pharmacokinetic parameters were higher in females than in males. CYP2D6 poor and intermediate metabolizers had notably higher prolactin Cmax and AUC0-12 than normal and ultrarapid metabolizers. The DRD3 rs6280 polymorphism affected prolactin levels: volunteers carrying Ser/Ser genotype had significantly lower prolactin levels than volunteers carrying the Gly allele. Furthermore, HTR2C rs3813929 C/C homozygotes had significantly lower prolactin levels than T allele carriers. Nevertheless, aripiprazole did increase prolactin levels compared to ibuprofen., D. Koller is co-financed by the H2020 Marie Sklodowska-Curie Innovative Training Network 721236 grant. P. Zubiaur is co-financed by Consejería de Educación, Juventud y Deporte from Comunidad de Madrid and Fondo Social Europeo. F. Abad-Santos and D. Ochoa have been consultants or investigators in clinical trials sponsored by the following pharmaceutical companies: Abbott, Alter, Aptatargets, Chemo, Cinfa, FAES, Farmalíder, Ferrer, Galenicum, GlaxoSmithKline, Gilead, Italfarmaco, Janssen-Cilag, Kern, Normon, Novartis, Servier, Silverpharma, Teva and Zambon., Peer reviewed

File 1. Demographic characteristics of the volunteers from the aripiprazole study.
File 2. Prolactin pharmacokinetic parameters after aripiprazole and ibuprofen administration. This xls file shows the modifications in the prolactin concentrations after the administration of aripiprazole or ibuprofen A It is been shown the prolactin concentrations versus time in women and men treated with aripiprazole compared to ibuprofen
File 3 Genotype Matrix. The xls file show the detected genetic polymorphism associated with the effects of aripiprazole
File 4 Aripiprazole and dehydro-aripiprazole pharmacokinetic parameters.
File 5 The influence of polymorphisms on prolactin concentrations., Aripiprazole treatment in schizophrenic patients was previously associated with lower or normalized prolactin levels. Genetic variants in cytochrome P450 (CYP) (CYP2D6), dopamine receptor (DRD2, DRD3) and serotonin receptor (HTR2A, HTR2C) genes were previously associated with antipsychotic-induced hyperprolactinaemia. Our aim was to evaluate whether aripiprazole affects prolactin secretion and its relationship with pharmacogenetics. Thirty-one healthy volunteers receiving a 10-mg single oral dose of aripiprazole were genotyped for 12 polymorphisms in CYP2D6, DRD2, DRD3, HTR2A and HTR2C genes by qPCR. Aripiprazole and dehydro-aripiprazole plasma concentrations were measured by HPLC-MS/MS. Prolactin concentrations of the 31 volunteers taking aripiprazole and 12 volunteers receiving ibuprofen were determined by ELISA. Prolactin concentrations after ibuprofen intake were considered as control, since it is known to cause no effect. Prolactin concentrations were slightly higher in the aripiprazole group compared to the ibuprofen group. All prolactin pharmacokinetic parameters were higher in females than in males. CYP2D6 poor and intermediate metabolizers had notably higher prolactin Cmax and AUC0-12 than normal and ultrarapid metabolizers. The DRD3 rs6280 polymorphism affected prolactin levels: volunteers carrying Ser/Ser genotype had significantly lower prolactin levels than volunteers carrying the Gly allele. Furthermore, HTR2C rs3813929 C/C homozygotes had significantly lower prolactin levels than T allele carriers. Nevertheless, aripiprazole did increase prolactin levels compared to ibuprofen., European Comission ITN Treatment H2020-MSCA-ITN-721236, Peer reviewed

Folder 1. Analysis of in vivo chronic dietary treatment with both olanzapine and aripiprazole (5.5-6 mg/kg/d) for 6 months in WT females to assess disturbances in glucose metabolism as well as beta-cell dysfunction. Metabolic tests have been done before sacrifice: glucose tolerance test (GTT), insulin tolerance test (ITT), and glucose-stimulated insulin secretion (GSIS). The xls file shows the statistics analysis and raw data analysis of the parameters measured. The immunofluorescence and immunohistochemistry analysis are also included in xls file. The pzfx files can be opened with GraphPad Prism 8 XML Project show the relationships between some of the parameters showed in xls files.
Folder 2. In vitro data in INS-1 cells. The xls files showed the quantification of the short-term impact of olanzapine (1-6 µM) and the long-term impact of both olanzapine and aripiprazole (1-6 µM) in glucose-stimulated insulin secretion (GSIS) and the western blot quantification of the molecular machinery associated with ER stress activation (mainly PERK/eIF2a and IRE1-alpha) in INS-1 cells after olanzapine/aripiprazole treatment with different concentrations (1-6 µM) for 4h. The pzfx files can be opened with GraphPad Prism 8 XML Project and show the relationships between some of the parameters showed in xls files., The incidence of Type 2 Diabetes Mellitus (T2DM) is reaching epidemic proportions. Recent investigations have demonstrated that long-term treatment with Second Generation Antipsychotics (SGAs), the main-line treatment for schizophrenia, can induce T2DM. Beta cell dysfunction is proposed as a plausible mechanism by which SGAs cause T2DM, but the process remains largely unknown. In this Thesis, we have investigated whether two unrelated SGAs, olanzapine, a common prescribed SGA with diabetogenic properties, and aripiprazole, a more recently developed SGA with less explored metabolic-side effects, can impact on beta cells. We analyzed beta cell functionality and pancreatic islet plasticity in two in vivo studies: female mice treated with olanzapine for 6 weeks via intraperitoneal and female mice fed an olanzapine- or aripiprazole-supplemented diet for 6 months. Additionally, we conducted gene expression analysis in islets of mice receiving the medicated diet and in vitro studies to evaluate beta cell functionality and the molecular mechanisms associated to the treatments. Our results evidenced that long-term treatment with olanzapine or aripiprazole induced weight gain, glucose intolerance and beta cell dysfunction, but the mechanisms behind these alterations are specific for each SGA. Whereas olanzapine effects in the pancreas in female mice seem to be dependent on an obesogenic-like phenotype, it activated endoplasmic reticulum (ER) stress in both INS-1 cells and pancreatic islets. Alleviation of olanzapine-induced ER stress with Tauroursodeoxycholic acid (TUDCA) recovered insulin secretion, suggesting that inhibition of insulin secretion by olanzapine is dependent on ER stress activation. On the other hand, aripiprazole treatment during 6 months induced serotonin production through tryptophan hydroxylase 1 (TPH1) activation in pancreatic islets. Moreover, beta cell hypertrophy and higher beta cell mass were found in aripiprazole-treated mice concomitantly to the activation of mTORC1/S6. Additionally, ex vivo experiments using pancreatic islets revealed that aripiprazole inhibition of insulin secretion was due to a reduction in calcium entry into the beta cell. Until now, regulation of the serotonergic system in islets has been associated to beta cell compensation in pregnancy and postnatal growth. Thus, in this Thesis, we have described for the first time the modulation of the serotonergic system in pancreatic islets by pharmacological treatment with aripiprazole, showing that serotonin production induced by this SGA plays a critical role in intra-islet functionality and beta cell mass and it might also explain other metabolic disturbances associated with aripiprazole treatment, European Comission ITN Treatment H2020-MSCA-ITN-721236, Peer reviewed

Resumen del trabajo presentado al 5th Symposium on Biomedical Research: "Advances and Perspectives In Pharmacology, Drug Toxicity and Pharmacogenetics", celebrado en Madrid del 15 al 16 de marzo de 2018., [Introduction]: The liver is the main organ in charge of drug catabolism and also the major site susceptible to drug injury. Sirtuin 1 (SIRT1), a NAD+-dependent histone deacetylase, is a key player in liver physiology and a therapeutic target against hepatic inflammation. In this study, we evaluated the role of SIRT1 in the pro-inflammatory context and oxidative stress during acetaminophen (APAP)-mediated hepatotoxicity., [Material and Methods]: SIRT1 expression was analyzed in APAP-induced liver failure in humans and mice. Hepatotoxicity was assessed in wild-type and transgenic mice overexpressing SIRT1 (SIRT1 Tg) poisoned with APAP (300 mg/kg). Raw 264.7 and peritoneal macrophages were treated with APAP and conditioned medium (CM) was added to mouse hepatocytes. siRNA was used to reduce inflammatory mediators in hepatocytes., [Results]: SIRT1 protein levels decreased in human and mouse livers following APAP overdose. SIRT1-Tg mice maintained higher levels of SIRT1 upon APAP injection than wild-type mice and were protected against hepatotoxicity by modulation of antioxidant systems and restrained inflammatory responses, with decreased oxidative stress, pro-inflammatory cytokine mRNA levels, nuclear factor kappa B (NFκB) signaling, and cell death. Mouse hepatocytes stimulated with conditioned medium of APAP-treated macrophages (APAP-CM) showed decreased SIRT1 levels; an effect mimicked by interleukin 1β (IL1β), an activator of NFκB. This negative modulation was abolished by neutralizing IL1β in APAP-CM or silencing p65-NFκB in hepatocytes. APAP-CM of macrophages from SIRT1-Tg mice failed to downregulate SIRT1 protein levels in hepatocytes. In vivo administration of the NFκB inhibitor BAY 11-7082 preserved SIRT1 levels and protected from APAP-mediated hepatotoxicity., [Conclusion]: SIRT1 protein levels are downregulated by IL1β/NFκB signaling in APAP hepatotoxicity, resulting in inflammation and oxidative stress. Thus, maintenance of SIRT1 during APAP overdose by inhibiting NFκB might be clinically relevant. Our work evidenced the unique role of SIRT1 in APAP hepatoprotection by targeting oxidative stress and inflammation., This work was funded by SAF2015-65267-R (MINECO/FEDER), CIBERdem (ISCIII, Spain), INFLAMES (ISCIII PIE14/00045, co-funded by ERDF, “Investing in your future”) to A.M.V; IJCI-2014-19381 (MINECO/FEDER) to P.R and A.M.V; SAF2014-52009-R (MINECO/FEDER) to S.A; SAF2015-63904-R (MINECO/FEDER) to M.M.; CP14/00181 (ISCIII/FEDER) to A. G-R; PI13/00021 (ISCIII/FEDER) to J.M. CIBERehd (ISCIII, Spain) to A. G-R and J.M; Grant 37/371 from Al Jouf University (to M.A.M). We also acknowledge H2020 Marie Sklodowska-Curie ITN-TREATMENT (Grant Agreement number 721236) (European Commission).

Antipsychotics used to treat schizophrenia can cause drug-induced liver injury (DILI), according to the Roussel Uclaf Causality Assessment Method. The role of oxidative stress in triggering injury in these DILI cases is unknown. We repeatedly administrated two second-generation antipsychotics, aripiprazole and olanzapine, at laboratory alert levels to study underlying mechanisms in stress prevention upon acute oxidative stress. The drugs were administered continuously for up to 8 weeks. For this, hepatoma Fao cells, which are suitable for metabolic studies, were used, as the primary hepatocytes survive in the culture only for about 1 week. Four stress responses—the oxidative stress response, the DNA damage response and the unfolded protein responses in the endoplasmic reticulum and mitochondria—were examined in H2O2-treated cells by antioxidant enzyme activity measurements, gene expression and protein quantification. Oxidant conditions increased the activity of antioxidant enzymes and upregulated genes and proteins associated with oxidative stress response in aripiprazole-treated cells. While the genes associated with DNA damage response, Gadd45 and p21, were upregulated in both aripiprazole- and olanzapine-treated cells, only aripiprazole treatment was associated with upregulation in response to even more H2O2, which also coincided with better survival. Endoplasmic reticulum stress-induced Chop was also upregulated; however, neither endoplasmic reticulum nor mitochondrial unfolded protein response was activated. We conclude that only aripiprazole, but not olanzapine, protects liver cells against oxidative stress. This finding could be relevant for schizophrenia patients with high-oxidative-stress-risk lifestyles and needs to be validated in vivo., The authors acknowledge the financial support from European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement 721236- TREATMENT and from the Slovenian Research Agency (research core funding No. P3-0019). BK acknowledges the financial support of the Word Federation of Scientists within the framework of the Slovenian Science Foundation.

Second-generation antipsychotics (SGAs), used as the cornerstone treatment for schizophrenia and other mental disorders, can cause adverse metabolic effects (e.g. obesity and type 2 diabetes). We investigated the effects of SGAs on adipocyte differentiation and metabolism. The presence of therapeutic concentrations of aripiprazole (ARI) or its active metabolite dehydroaripiprazole (DARI) during human adipocyte differentiation impaired adipocyte glucose uptake while the expression of gene markers of fatty acid oxidation were increased. Additionally, the use of a supra-therapeutic concentration of ARI inhibited adipocyte differentiation. Furthermore, olanzapine (OLA), a highly obesogenic SGA, directly increased leptin gene expression but did not affect adipocyte differentiation and metabolism. These molecular insights are novel, and suggest that ARI, but not OLA, may directly act via alterations in adipocyte differentiation and potentially by causing a switch from glucose to lipid utilization in human adipocytes. Additionally, SGAs may effect crosstalk with other organs, such as the brain, to exert their adverse metabolic effects., This work was supported by research grants from the European Commission via the Marie Sklodowska Curie Innovative Training Network TREATMENT (H2020-MSCA-ITN-721236), Swedish Diabetes Foundation, the Excellence of Diabetes Research in Sweden (EXODIAB), Family Ernfors Foundation, the P.O. Zetterling Foundation, Swedish Society for Medical Research, Novo Nordisk Foundation, Agnes and Mac Rudbergs Foundation, and the Uppsala University Hospital ALF grants.

Prolonged use of atypical antipsychotics (AAPs) is commonly associated with increased cardiovascular disease risk. While weight gain and related health issues are generally considered the primary contributors to this risk, direct interference with mitochondrial bioenergetics, particularly in the liver where these drugs are metabolized, is emerging as an additional contributing factor. Here, we compared the effects of two AAPs with disparate metabolic profiles on the response of Fao hepatoma cells to oxidative stress: olanzapine (OLA), which is obesogenic, and aripiprazole (ARI), which is not. Results showed that cells treated with ARI exhibited resistance to H2O2-induced oxidative stress, while OLA treatment had the opposite effect. Despite enhanced survival, ARI-treated cells exhibited higher apoptotic rates than OLA-treated cells when exposed to H2O2. Gene expression analysis of pro- and anti-apoptotic factors revealed that ARI-treated cells had a generally blunted response to H2O2, contrasting with a heightened response in OLA-treated cells. This was further supported by the reduced activation of MAPKs and STAT3 in ARI-treated cells in response to H2O2, whereas OLA pre-treatment enhanced their activation. The loss of stress response in ARI-treated cells was consistent with the observed increase in the mitochondrial production of O2•-, a known desensitizing factor. The physiological relevance of O2•- in ARI-treated cells was demonstrated by the increase in mitophagy flux, likely related to mitochondrial damage. Notably, OLA treatment protected proteasome activity in Fao cells exposed to H2O2, possibly due to the better preservation of stress signaling and mitochondrial function. In conclusion, this study highlights the underlying changes in cell physiology and mitochondrial function by AAPs. ARI de-sensitizes Fao cells to stress signaling, while OLA has the opposite effect. These findings contribute to our understanding of the metabolic risks associated with prolonged AAP use and may inform future therapeutic strategies., This research was funded by the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement 721236-TREATMENT, by the Slovenian Research and Innovation Agency (ARIS), research core funding No. P3-0019, by the Spanish Ministry of Science Innovation and Universities (MCIU), Agencia Estatal de Investigación (AEI) grant PID2021-122765OB-I00, and by the Horizon Europe Program under the EIC-2023-PATFINDERCHALLENGES-01-3 action GA-101162517 (DiBaN)., Peer reviewed

Cellular energy metabolism varies depending on tissue and cell type, as well as the availability of energy substrates and energy demands. We recently investigated the variations in cellular metabolism and antioxidant responses in primary bovine vascular endothelial cells (BAECs) under different energetic substrate conditions in vitro, specifically glucose or galactose. In this context, pharmacological agents may affect cells differently depending on their energy metabolism status. In this study, we aimed to characterize the effects of diphenyl diselenide ((PhSe)2), a redox-active molecule known for its prominent cardiovascular effects, on redox-bioenergetic cellular pathways under glycolytic or oxidative conditions in BAECs. Under glucose conditions, (PhSe)2 positively impacted mitochondrial oxidative capacity, as assessed by respirometry, and was associated with changes in mitochondrial cellular dynamics. However, these changes were not observed in cells cultured with galactose. Although (PhSe)2 induced the nuclear translocation of the redox sensitive nuclear factor erythroid 2-related factor 2 (Nrf2) in both glucose and galactose media, Nrf2 remained in the nuclei of cells cultured in galactose for a longer duration. Additionally, activation of another redox sensitive transcription factor, forkhead O3 (FOXO3a) was only detected in galactose media. Notably, (PhSe)2 induced the expression of genes controlling mitochondrial antioxidant capacity and glutathione synthesis and recycling in glucose media, whereas its effects in galactose media were primarily focused on glutathione homeostasis. In conclusion, our findings underscore the critical influence of cellular metabolic status on the antioxidant capacity of redox-active molecules such as (PhSe)2., This work was supported by MCIN/AEI/10.13039/501100011033/FEDER, UE, [grant numbers RTI2018-093864-B-I00 and PID2021-122765OB-I00]; the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie [grant agreement 721236-TREATMENT]; EU Horizon Europe Program under the EIC Pathfinder DiBaN [GA-101162517]; the Brazilian institutions: Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Fundação de Apoio à Pesquisa do Distrito Federal (FAPDF) [grants 00193-00000884/2021-89 and 00193-00002348/2022-07]; and Instituto Nacional de Ciência e Tecnologia e Neuro-ImunoModulação (INCT-NIM) [grant 485489/2014-1]. The authors are grateful to Smart Servier Biology Medicine for the available figures., No

Objectives: Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. Protein tyrosine phosphatase 1B (PTP1B), a negative modulator of insulin and cytokine signaling, is a therapeutic target for type 2 diabetes and obesity. We investigated the impact of PTP1B deficiency during NAFLD, particularly in non-alcoholic steatohepatitis (NASH).
Methods: NASH features were evaluated in livers from wild-type (PTP1BWT) and PTP1B-deficient (PTP1BKO) mice fed methionine/cholinedeficient diet (MCD) for 8 weeks. A recovery model was established by replacing MCD to chow diet (CHD) for 2e7 days. Non-parenchymal liver cells (NPCs) were analyzed by flow cytometry. Oval cells markers were measured in human and mouse livers with NASH, and in oval cells from PTP1BWT and PTP1BKO mice.
Results: PTP1BWT mice fed MCD for 8 weeks exhibited NASH, NPCs infiltration, and elevated Fgf21, Il6 and Il1b mRNAs. These parameters
decreased after switching to CHD. PTP1B deficiency accelerated MCD-induced NASH. Conversely, after switching to CHD, PTP1BKO mice rapidly
reverted NASH compared to PTP1BWT mice in parallel to the normalization of serum triglycerides (TG) levels. Among NPCs, a drop in cytotoxic
natural killer T (NKT) subpopulation was detected in PTP1BKO livers during recovery, and in these conditions M2 macrophage markers were upregulated. Oval cells markers (EpCAM and cytokeratin 19) significantly increased during NASH only in PTP1B-deficient livers. HGF-mediated
signaling and proliferative capacity were enhanced in PTP1BKO oval cells. In NASH patients, oval cells markers were also elevated.
Conclusions: PTP1B elicits a dual role in NASH progression and reversion. Additionally, our results support a new role for PTP1B in oval cell
proliferation during NAFLD.

Aims: Oxidative stress is known to induce early replicative senescence. Senescence has been proposed to work as a barrier to immortalization and tumor development. Here, we aimed to evaluate the impact of the loss of peroxisome proliferator activated receptor γ co-activator 1α (PGC-1α), a master regulator of oxidative metabolism and mitochondrial reactive oxygen species (ROS) generation, on replicative senescence and immortalization in mouse embryonic fibroblasts (MEFs). Results: We found that primary MEFs lacking PGC-1α showed higher levels of ROS than wild-type MEFs at all cell passages tested. The elevated production of ROS was associated with higher levels of oxidative DNA damage and the increased formation of DNA double-strand breaks. Evaluation of the induction of DNA repair systems in response to γ-radiation indicated that the loss of PGC-1α also resulted in a small but significant reduction in their activity. DNA damage induced the early activation of senescence markers, including an increase in the number of β-galactosidase-positive cells, the induction of p53 phosphorylation, and the increase in p16 and p19 protein. These changes were, however, not sufficient to reduce proliferation rates of PGC-1α-deficient MEFs at any cell passage tested. Moreover, PGC-1α-deficient cells escaped replicative senescence. Innovation & conclusion: PGC-1α plays an important role in the control of cellular senescence and immortalization.

Second-generation antipsychotics (SGAs) are a mainstay therapy for schizophrenia. SGA-treated patients present higher risk for weight gain, dyslipidemia and hyperglycemia. Herein, we evaluated the effects of olanzapine (OLA), widely prescribed SGA, in mice focusing on changes in body weight and energy balance. We further explored OLA effects in protein tyrosine phosphatase-1B deficient (PTP1B-KO) mice, a preclinical model of leptin hypersensitivity protected against obesity.
Wild-type (WT) and PTP1B-KO mice were fed an OLA-supplemented diet (5 mg/kg/day, 7 months) or treated with OLA via intraperitoneal (i.p.) injection or by oral gavage (10 mg/kg/day, 8 weeks). Readouts of the crosstalk between hypothalamus and brown or subcutaneous white adipose tissue (BAT and iWAT, respectively) were assessed. The effects of intrahypothalamic administration of OLA with adenoviruses expressing constitutive active AMPKα1 in mice were also analyzed.
Both WT and PTP1B-KO mice receiving OLA-supplemented diet presented hyperphagia, but weight gain was enhanced only in WT mice. Unexpectedly, all mice receiving OLA via i.p. lost weight without changes in food intake, but with increased energy expenditure (EE). In these mice, reduced hypothalamic AMPK phosphorylation concurred with elevations in UCP-1 and temperature in BAT. These effects were also found by intrahypothalamic OLA injection and were abolished by constitutive activation of AMPK in the hypothalamus. Additionally, OLA i.p. treatment was associated with enhanced Tyrosine Hydroxylase (TH)-positive innervation and less sympathetic neuron-associated macrophages in iWAT. Both central and i.p. OLA injections increased UCP-1 and TH in iWAT, an effect also prevented by hypothalamic AMPK activation. By contrast, in mice fed an OLA-supplemented diet, BAT thermogenesis was only enhanced in those lacking PTP1B. Our results shed light for the first time that a threshold of OLA levels reaching the hypothalamus is required to activate the hypothalamus BAT/iWAT axis and, therefore, avoid weight gain.
Our results have unraveled an unexpected metabolic rewiring controlled by hypothalamic AMPK that avoids weight gain in male mice treated i.p. with OLA by activating BAT thermogenesis and iWAT browning and a potential benefit of PTP1B inhibition against OLA-induced weight gain upon oral treatment.