Hu Antigen R (HuR) Protein Structure, Function and Regulation in Hepatobiliary Tumors

Hepatobiliary tumors are a group of primary malignancies encompassing the liver, the intra- and extra-hepatic biliary tracts, and the gall bladder. Within the liver, hepatocellular carcinoma (HCC) is the most common type of primary cancer, which is, also, representing the third-most recurrent cause of cancer-associated death and the sixth-most prevalent type of tumor worldwide, nowadays. Although less frequent, cholangiocarcinoma (CCA) is, currently, a fatal cancer with limited therapeutic options. Here, we review the regulatory role of Hu antigen R (HuR), a ubiquitous member of the ELAV/Hu family of RNA-binding proteins (RBPs), in the pathogenesis, progression, and treatment of HCC and CCA. Overall, HuR is proposed as a valuable diagnostic and prognostic marker, as well as a therapeutic target in hepatobiliary cancers. Therefore, novel therapeutic approaches that can selectively modulate HuR function appear to be highly attractive for the clinical management of these types of tumors.España Ministerio de Ciencia, Innovación y Universidades (MICINN), integrated in the Plan Estatal de Investigación Científica y Técnica e Innovación, grant numbers PID2020-117116RB-I00 (to M.L.M.-C.) and PGC2018-096049-B-I00Gobierno de Andalucía, grant numbers BIO-198, US-1254317, US-1257019, P18-FR-3487, and P18-HO-4091Fondo Europeo de Desarrollo Regional (FEDER) (to MLM-C and I.D.-M.); and the La Caixa Foundation Program, grant number HR17-00601 (to M.L.M.-C.).España Ministerio de Educación, Cultura y Deporte, grant number FPU016/0151

Micelle carriers based on dendritic macromolecules containing bis-MPA and glycine for antimalarial drug delivery

Biomaterials for antimalarial drug transport still need to be investigated in order to attain nanocarriers that can tackle essential issues related to malaria treatment, e.g. complying with size requirements and targeting specificity for their entry into Plasmodium-infected red blood cells (pRBCs), and limiting premature drug elimination or drug resistance evolution. Two types of dendritic macromolecule that can form vehicles suitable for antimalarial drug transport are herein explored. A new hybrid dendritic-linear-dendritic block copolymer based on Pluronic\xC2\xAE F127 and amino terminated 2,2'-bis(glycyloxymethyl)propionic acid dendrons with a poly(ester amide) skeleton (HDLDBC-bGMPA) and an amino terminated dendronized hyperbranched polymer with a polyester skeleton derived from 2,2'-bis(hydroxymethyl)propionic acid (DHP-bMPA) have provided self-assembled and unimolecular micelles. Both types of micelle carrier are biocompatible and exhibit appropriate sizes to enter into pRBCs. Targeting studies have revealed different behaviors for each nanocarrier that may open new perspectives for antimalarial therapeutic approaches. Whereas DHP-bMPA exhibits a clear targeting specificity for pRBCs, HDLDBC-bGMPA is incorporated by all erythrocytes. It has also been observed that DHP-bMPA and HDLDBC-bGMPA incorporate into human umbilical vein endothelial cells with different subcellular localization, i.e. cytosolic and nuclear, respectively. Drug loading capacity and encapsulation efficiencies for the antimalarial compounds chloroquine, primaquine and quinacrine ranging from 30% to 60% have been determined for both carriers. The resulting drug-loaded nanocarriers have been tested for their capacity to inhibit Plasmodium growth in in vitro and in vivo assays

Community structure of woody plants on islands along a bioclimatic gradient

Peer reviewe

Methionine Cycle Rewiring by Targeting miR-873-5p Modulates Ammonia Metabolism to Protect the Liver from Acetaminophen

Drug-induced liver injury (DILI) development is commonly associated with acetaminophen (APAP) overdose, where glutathione scavenging leads to mitochondrial dysfunction and hepatocyte death. DILI is a severe disorder without effective late-stage treatment, since N-acetyl cysteine must be administered 8 h after overdose to be efficient. Ammonia homeostasis is altered during liver diseases and, during DILI, it is accompanied by decreased glycine N-methyltransferase (GNMT) expression and S-adenosylmethionine (AdoMet) levels that suggest a reduced methionine cycle. Anti-miR-873-5p treatment prevents cell death in primary hepatocytes and the appearance of necrotic areas in liver from APAP-administered mice. In our study, we demonstrate a GNMT and methionine cycle activity restoration by the anti-miR-873-5p that reduces mitochondrial dysfunction and oxidative stress. The lack of hyperammoniemia caused by the therapy results in a decreased urea cycle, enhancing the synthesis of polyamines from ornithine and AdoMet and thus impacting the observed recovery of mitochondria and hepatocyte proliferation for regeneration. In summary, anti-miR-873-5p appears to be an effective therapy against APAP-induced liver injury, where the restoration of GNMT and the methionine cycle may prevent mitochondrial dysfunction while activating hepatocyte proliferative response.We thank Ministerio de Ciencia e Innovación, Programa Retos-Colaboración RTC2019- 007125-1 (for J.S. and M.L.M.-C.); Instituto de Salud Carlos III: Proyectos de Investigación en Salud DTS20/00138 (for J.S. and M.L.M.-C.), PI20/00690 (for R.J.) and PT20/000127 (for M.I.L.); CIBERehd: EHD21TRF01/2022 (to M.L.M.-C.); Departamento de Industria del Gobierno Vasco (for M.L.M.-C.); Ministerio de Ciencia, Innovación y Universidades MICINN: PID2020-117116RB-I00 and RTI2018- 096759-1-100 integrado en el Plan Estatal de Investigación Cientifica y Técnica y Innovación, cofinanciado con Fondos FEDER (for M.L.M.-C. and T.C.D., respectively); BIOEF (Basque Foundation for Innovation and Health Research); Asociación Española contra el Cáncer (AECC) (to M.L.M.-C., T.C.D.); AECC: GCTRA18006CARR (to A.C.); Fundación Científica de la Asociación Española Contra el Cancer (AECC Scientific Foundation) Rare Tumor Calls 2017 (for M.L.M.); La Caixa Foundation Program (for M.L.M.); BFU2015-70067-REDC, BFU2016-77408-R and BES-2017-080435 (MINECO/FEDER, UE); Ministerio de Ciencia, Innovación y universidades PID2019-108787RB-100 (to A.C.), PID2019- 109055RB-I00 (L.A.M.-C.), PID2020-117941RB-100 (to F.J.C.); Spanish Ministry of Economy and Competitiveness Grants BFU2013-47531-R and BFU2016-77408-R (L.A.M.-C.) and the FIGHT-CNNM2 project from the EJP RD Joint Transnational Call (JTC2019) (Ref. AC19/00073) (for L.A.M.-C.); Comunidad de Madrid: EXOHEP-CM S2017/BMD-3727 and NanoLiver-CM Y2018/NMT-4949 co-funded by European Structural and Investment Fund and COST Action CA17112 (to F.J.C.); Vencer el Cáncer Foundation (to A.C.); European Research Council: Consolidator Grant 819242 (to A.C.); CIBERONC and CIBERehd were funded by the Instituto de Salud Carlos III and Cofunded by FEDER funds. Partial funding for open access charge: Universidad de Málag

Glutamate and GABA in appetite regulation

Appetite is regulated by a coordinated interplay between gut, adipose tissue and brain. A primary site for the regulation of appetite is the hypothalamus where interaction between orexigenic neurons, expressing Neuropeptide Y/Agouti-related protein, and anorexigenic neurons, expressing Pro-opiomelanocortin cocaine/Amphetamine-related transcript, controls energy homeostasis. Within the hypothalamus, several peripheral signals have been shown to modulate the activity of these neurons, including the orexigenic peptide ghrelin and the anorexigenic hormones insulin and leptin. In addition to the accumulated knowledge on neuropeptide signaling, presence and function of amino acid neurotransmitters in key hypothalamic neurons brought a new light into appetite regulation. Therefore, the principal aim of this review will be to describe the current knowledge of the role of amino acid neurotransmitters in the mechanism of neuronal activation during appetite regulation and the associated neuronal-astrocytic metabolic coupling mechanisms.Glutamate and GABA dominate synaptic transmission in the hypothalamus and administration of their receptors agonists into hypothalamic nuclei stimulates feeding. By using 13C High-Resolution Magic Angle Spinning Nuclear Magnetic Resonance spectroscopy based analysis, the Cerdán group has shown that increased neuronal firing in mice hypothalamus, as triggered by appetite during the feeding-fasting paradigm, may stimulate the use of lactate as neuronal fuel leading to increased astrocytic glucose consumption and glycolysis. Moreover, fasted mice showed increased hypothalamic [2-13C]GABA content, which may be explained by the existence of GABAergic neurons in key appetite regulation hypothalamic nuclei. Interestingly, increased [2-13C]GABA concentration in the hypothalamus of fasted animals appears to result mainly from reduction in GABA metabolizing pathways, rather than increased GABA synthesis by augmented activity of the glutamate-glutamine-

Resistência à insulina e diabetes : estudos de ressonância magnética do metabolismo hepático da glicose e lipídico

Tese de doutoramento em Bioquímica (Biofísida Celular) apresentada à Fac. Ciências da Universidade de CoimbraNas últimas décadas, a incidência de resistência à insulina e de diabetes tipo 2 têm aumentado principalmente devido a alterações de estilos de vida e na dieta. A diabetes pós-transplante é igualmente um tópico de crescente interesse considerando os números elevados e aumento das taxas de sobrevivência actuais de transplantes de orgãos. O fígado é um orgão profundamente envolvido na regulação da homeostase corporal da glicose e lipídica. Como tal, alterações hepáticas do metabolismo da glicose e lipídico poderão ocupar um papel central no desenvolvimento de patologias como a resistência à insulina, a diabetes tipo 2 ou a diabetes pós-transplante. Nesta Tese procedeu-se à avaliação do metabolismo hepático da glicose e lipídico em modelos animais e em pacientes com resistência à insulina, diabetes tipo 2 ou diabetes pós-transplante, utilizandose marcadores de isótopos estáveis e análise por espectroscopia de Ressonância Magnética Nuclear (RMN). Adicionalmente, foram desenvolvidas novas metodologias para o estudo do metabolismo hepático da glicose em situações de jejum e em condições mais dinâmicas, e procedeu-se a uma análise integrativa do metabolismo hepático da glicose e lipídico. No Capítulo 1, o metabolismo corporal da glicose e lípidos foi revisto e integrado focando as alterações do metabolismo hepático associadas à resistência à insulina e à diabetes. Os marcadores de isótopos estáveis e as técnicas de Ressonância Magnética assim como a sua aplicação à avaliação do metabolismo hepático foram introduzidos. No Capítulo 2, as fontes endógenas de glicose durante o jejum foram avaliadas em ratos controlo e modelos animais de resistência à insulina induzida através de uma dieta rica em gordura. Para tal, utilizou-se [3,4-13C2]glicose e água deuterada (2H2O) em combinação com espectroscopia de RMN de carbono 13 (13C) e deutério (2H). A produção endógena de glicose resultante da gluconeogénese e da glicogenólise determinada nos animais alimentados com uma dieta rica em gordura foi essencialmente idêntica à dos controlos com uma dieta normal. Os fluxos normais de produção endógena de glicose encontrados nos animais sujeitos a uma dieta rica em gordura sugerem que as alterações dos fluxos hepáticos da glicose não estão envolvidas, pelo menos nos estádios iniciais, no desenvolvimento da resistência à insulina como consequência de uma dieta rica em gordura. No Capítulo 3, as fontes hepáticas de glicose durante o jejum foram quantificadas em pacientes com transplante renal e terapia imunossupressora baseada em Ciclosporina A (CsA). Foi proposta uma nova análise Bayesiana dos sinais de RMN de 2H das posições 2 e 5 da monoacetona glicose derivada do glucuronado de paracetamol urinário, após ingestão de 2H2O e paracetamol, para a quantificação relativa das fontes gliconeogénicas e glicogenolíticas de produção hepática de glicose. Nos pacientes com transplante renal, a contribuição da gliconeogénese para a produção hepática de glicose é significativamente superior nos casos estabelecidos de diabetes pós-transplante. Esta alteração metabólica está principalmente relacionada com a crescente adiposidade e aumento do índice de massa corporal. Por outro lado, o tratamento com CsA per se provocou alterações modestas nas fontes hepáticas de glicose, não estando associado com a hiperglicémia ou hiperinsulinémia em jejum. O destino de uma carga de glicose intra-peritoneal em ratos saudáveis foi estudado no Capítulo 4, utilizando a análise por espectroscopia de RMN de 13C e 2H após administração de [U-13C]glicose e 2H2O. A contribuição da produção hepática de glicose para a glicose total foi determinada a partir do enriquecimento em 2H da posição 2 da glicose plasmática em relação ao enriquecimento da água plasmática. Por outro lado, a contribuição da gliconeogénese foi determinada pela quantificação da razão entre os enriquecimentos em 2H da posição 5 do glicose plasmática e da água plasmática. As contribuições da carga intra-peritoneal de glicose e da glicose “reciclada” através do ciclo de Cori para a glicose total foram estimadas a partir dos níveis de enriquecimento dos respectivos isotopómeros de [U-13C]glicose presentes no plasma em relação ao enriquecimento inicial da carga de glicose. As fontes de glicose após a prova de tolerância à glicose intra-peritoneal foram avaliadas em animais sob uma dieta rica em gordura e com tratamento baseado em CsA. Ambos os grupos de animais mostraram intolerância à glicose relativamente aos controlos. Com a dieta rica em gordura, a produção hepática de glicose não foi alterada, tendo-se observado um padrão de secreção normal de insulina pelas células-b pancreáticas. Como tal, a hiperglicémia pós-prandial observada provavelmente reflecte uma ligeira resistência à insulina a nível dos orgãos periféricos em vez de produção hepática de glicose alterada. Por outro lado, nos animais tratados com CsA, a intolerância à glicose observada está associada com um aumento da contribuição da produção hepática de glicose. O Capítulo 5 focou o metabolismo hepático pós-prandial de glicose após uma prova de tolerância à glicose oral (PTGO) em pessoas saudáveis. Foram avaliadas as contribuições relativas da via directa e indirecta de síntese de glicogénio durante a PTGO utilizando dois marcadores de isótopos estáveis ([U-13C]glicose e [U-2H7]glicose), após ingestão de mentol e análise por espectroscopia de RMN dos enriquecimentos em 13C e 2H da glicose plasmática e do glucuronado de mentol urinário. As trocas existentes dos meios de carbono e de hidrogénio envolvidos foram reveladas e quantificadas. Durante a PTGO em pessoas saudáveis, cerca de metade da síntese de glicogénio hepático derivou de precursores de 3 carbonos (via indirecta) e não directamente da glicose oral fornecida. Durante a PTGO, ~20% do fluxo da via directa esteve envolvido em trocas promovidas pela transaldolase, pelo que os valores derivados a partir do marcador de [U-13C]glicose resultam numa subestima das contribuições da via directa. As fontes de triglicéridos hepáticos em ratos saudáveis e alimentados com uma dieta rica em gordura foram determinadas no Capítulo 6 utilizando uma nova metodologia baseada em 2H2O e análise por espectroscopia de RMN de 2H juntamente com espectroscopia de Ressonância Magnética de protão (1H) in vivo para avaliação do conteúdo de triglicéridos no fígado. Em ratos saudáveis, os níveis de triglicéridos hepáticos podem ser rapidamente regulados alterando o conteúdo em gordura da dieta. Estas mudanças podem ser efectivamente monitorizadas usando espectroscopia de Ressonância Magnética de 1H in vivo. Nos ratos com uma dieta rica em gordura, quase a totalidade dos triglicéridos hepáticos derivaram dos lípidos da dieta com uma contribuição residual da lipogénese de novo hepática. Finalmente, o Capítulo 7 apresenta as conclusões gerais onde todos os resultados descritos nesta Tese foram discutidos e integrados.In the last decades, insulin resistance (IR) and type 2 diabetes (T2D) are becoming more prevalent due to alterations in dietary and life-styles. Posttransplant diabetes mellitus (PTDM) has also become a subject of interest and importance in the wake of increased numbers and survival rates of solid organ transplantations. The liver is deeply involved in the regulation of whole body glucose and lipid homeostasis and hepatic glucose and lipid metabolic disruptions may play a central role in the onset of IR, T2D and PTDM. Changes in hepatic glucose and lipid fluxes using stable isotope tracers and Nuclear Magnetic Resonance (NMR) analysis both in animal models and patients with IR, T2D or PTDM were addressed in this Thesis. Moreover, further developments of techniques for the study of hepatic glucose metabolism from fasting to dynamic situations were challenged as well as for the integrated analysis of hepatic glucose and lipid metabolism. In Chapter 1, whole body glucose and lipid metabolism were reviewed and integrated focusing on hepatic metabolic disruptions associated with IR and diabetes. Moreover, the stable isotope tracers and Magnetic Resonance techniques and their applications to hepatic metabolism evaluation were introduced. In Chapter 2, fasting sources of endogenous glucose production (EGP) were evaluated in control and high fat (HF) diet induced-IR animal models by using [3,4-13C2]glucose and deuterated water (2H2O) combined with carbon 13 (13C) and deuterium (2H) NMR spectroscopy. Postabsorptive EGP from gluconeogenesis and glycogenolysis in HF diet-fed animals was essentially identical to the normally fed controls. The normal EGP rates found in HF dietfed animals suggest that altered hepatic glucose fluxes are not involved in the development of IR secondary to HF diet feeding, at least in the early stages. In Chapter 3, sources of fasting hepatic glucose production (HGP) were quantified in kidney transplant patients, undergoing cyclosporine A (CsA) immunosuppressant therapy. A novel Bayesian analysis of the position 2 and 5 2H NMR signals of monoacetone glucose derived from urinary acetaminophen glucuronide following 2H2O and acetominophen ingestion was proposed for quantification of gluconeogenesis and glycogenolysis relative contributions to HGP. For kidney transplant patients, the gluconeogenic contribution to HGP was significantly increased in the setting of PTDM. This metabolic alteration was found to be most strongly associated with adiposity and increased body mass index whereas CsA treatment per se provoked only modest alterations of HGP sources and was not shown to be associated with fasting hyperglycemia or hyperinsulinemia. The fate of an intraperitoneal (i.p.) glucose load by 13C and 2H NMR analysis following administration of [U-13C]glucose and 2H2O in healthy, HF diet-fed and CsA-treated rodents was addressed in Chapter 4. The contribution of HGP to total glucose was determined from the 2H-enrichment of plasma glucose position 2 relative to that of plasma water and gluconeogenic sources were quantified from the 2H-enrichment level at position 5 of plasma glucose relative to that of plasma water. The i.p. glucose load and “recycled” glucose from Cori cycle contributions to total glucose were estimated from the enrichment level of plasma [U- 13C]glucose respective isotopomers relative to that of the load. Sources of total glucose after an i.p. glucose load were further evaluated in HF diet-fed and CsA-treated animals. Both groups of animals showed impaired glucose tolerance (IGT) relative to controls. HF diet did not promote impaired contribution from HGP and insulin secretion by pancreatic b-cells was not affected. Thus, the IGT of HF diet-fed animals can be attributed to decreased whole body glucose disposal secondary to peripheral insulin resistance rather than impaired HGP suppression. With CsA-treated animals the observed IGT was associated with a higher HGP contribution to plasma glucose levels, suggesting that in this model impaired HGP suppression was a significant component of IGT. Chapter 5 focused on postprandial hepatic glucose metabolism after a glucose load in healthy humans. Direct and indirect pathway contributions to hepatic glycogen synthesis during an oral glucose tolerance test (OGTT) were assessed by means of two isotopic tracers ([U-13C]glucose and [U-2H7]glucose) following ingestion of peppermint oil and NMR analysis of plasma glucose and menthol glucuronide enrichments. Exchanges of both carbon and hydrogen moieties during the direct pathway metabolism of glucose were further revealed and quantified. During an OGTT in healthy humans, half of the hepatic glycogen synthesis was derived from 3-carbon precursors (indirect pathway) rather than directly from the glucose load. Furthermore, during an OGTT, ~20% of the direct pathway flux was involved in transaldolase exchange, hence the values derived from the [U-13C]glucose tracer resulted in underestimates of the direct pathway contribution. Sources of hepatic triglycerides accumulation in healthy and HF diet-fed rats, for evaluation of hepatic triglycerides content by using a novel 2H2O and 2H NMR methodology combined with in vivo proton (1H) Magnetic Resonance Spectroscopy (MRS), were determined in Chapter 6. In healthy rats, hepatic triglyceride levels can be acutely raised or lowered by altering the dietary fat content and these changes can be effectively monitored by 1H MRS. During HF diet feeding, essentially all of the hepatic triglycerides were derived from dietary lipid with very little contribution from hepatic de novo lipogenesis. Finally, Chapter 7 presents the concluding remarks where all the results described in this Thesis were integrated and further discussed

flock.uc.pt – A Web Platform for Online Educational Modules with Online Experiments

Emerging technologies provide the necessary means to develop online learning programs with online experimental setups through web platforms for educational and training purposes. Combining design techniques with virtual and augmented reality, templates, contents and interfaces can improve usersâ?? analytical capabilities of perception and cognition. In addition, it will allow the development of more attractive online courses, while promoting the learning process. This paper briefly describes some relevant features of the platform flock.uc.pt under development at the University of Coimbra, including some application examples. The authorsâ?? intension is to demonstrate the main characteristics of this platform presenting some examples of online educational modules