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Endothelial adenosine monophosphate-activated protein kinase (AMPK) plays a critical role in the regulation of vascular tone through stimulating nitric oxide (NO) release in endothelial cells. Since obesity leads to endothelial dysfunction and AMPK dysregulation, AMPK activation might be an important strategy to restore vascular function in cardiometabolic alterations. Here, we report the identification of a novel AMPK modulator, the indolic derivative IND6, which shows affinity for AMPKα1β1γ1, the primary AMPK isoform in human EA.Hy926 endothelial cells. IND6 shows inhibitory action of the enzymatic activity in vitro, but increases the levels of p-Thr174AMPK, p-Ser1177eNOS and p-Ser79ACC in EA.Hy926. This paradoxical finding might be explained by the ability of IND6 to act as a mixed-type inhibitor, but also to promote the enzyme activation by adopting two distinct binding modes at the ADaM site. Moreover, functional assays reveal that IND6 increased the eNOS-dependent production of NO and elicited a concentration-dependent vasodilation of endothelium-intact rat aorta due to AMPK and eNOS activation, demonstrating a functional activation of the AMPK–eNOS–NO endothelial pathway. This kinase inhibition profile, combined with the paradoxical AMPK activation in cells and arteries, suggests that these new chemical entities may constitute a valuable starting point for the development of new AMPK modulators with therapeutic potential for the treatment of vascular complications associated with obesity., his research was funded by Fundación Eugenio Rodríguez Pascual, the Spanish Ministerio de Economía y Competitividad (MDM-2017-0767), the Spanish Ministerio de Ciencia Innovación y Universidades (RTI2018- 095544-B-I00) and the Generalitat de Catalunya (2017SGR1746) for financial support. Computational resources from the Barcelona Supercomputing Center (BSC; BCV-2019-1-0009 and BCV-2019-2-0017) and the Consorci de Serveis Universitaris de Catalunya (CSUC; Molecular Recognition project) are acknowledged. E.A. thanks AGAUR (Generalitat of Catalunya; 2019FI_B2_00001) for her fellowship. M.S.G. thanks Universidad Com- plutense de Madrid for her fellowship.

The liver displays a remarkable regenerative capacity triggered upon tissue injury or resection. However, liver regeneration can be overwhelmed by excessive parenchymal destruction or diminished by pre-existing conditions hampering repair. Fibroblast growth factor 19 (FGF19, rodent FGF15) is an enterokine that regulates liver bile acid and lipid metabolism, and stimulates hepatocellular protein synthesis and proliferation. FGF19/15 is also important for liver regeneration after partial hepatectomy (PH). Therefore recombinant FGF19 would be an ideal molecule to stimulate liver regeneration, but its applicability may be curtailed by its short half-life. We developed a chimaeric molecule termed Fibapo in which FGF19 is covalently coupled to apolipoprotein A-I. Fibapo retains FGF19 biological activities but has significantly increased half-life and hepatotropism. Here we evaluated the proregenerative activity of Fibapo in two clinically relevant models where liver regeneration may be impaired: acetaminophen (APAP) poisoning, and PH in aged mice. The only approved therapy for APAP intoxication is N-acetylcysteine (NAC) and no drugs are available to stimulate liver regeneration. We demonstrate that Fibapo reduced liver injury and boosted regeneration in APAPintoxicated mice. Fibapo improved survival of APAP-poisoned mice when given at later time points, when NAC is ineffective. Mechanistically, Fibapo accelerated recovery of hepatic glutathione levels, potentiated cell growth-related pathways and increased functional liver mass. When Fibapo was administered to old mice prior to PH, liver regeneration was markedly increased. The exacerbated injury developing in these mice upon PH was attenuated, and the hepatic biosynthetic capacity was enhanced. Fibapo reversed metabolic and molecular alterations that impede regeneration in aged livers. It reduced liver steatosis and downregulated p21 and hepatocyte nuclear factor 4 α (Hnf4α) levels, whereas it stimulated Foxm1b gene expression. Together our findings indicate that FGF19 variants retaining the metabolic and growth-promoting effects of this enterokine may be valuable for the stimulation of liver regeneration., Work in the authors' laboratory is supported by CIBERehd and Grants from Instituto de Salud Carlos III (ISCIII) co-financed by 'Fondo Europeo de Desarrollo Regional' (FEDER) 'Una manera de hacer Europa', numbers: FIS PI13/00359, PI13/00385 and PI16/01126. Grants SAF2015-66515-R, SAF201569944-R, SAF 2016-75972 R from Ministerio de Economía y Competitividad, and the center grant P50AA011999 funded by NIAAA. 'Ramón y Cajal-I3' contract to MUL, Mineco-FPI Fellowship to MB-V. Marie Curie EU contract to MGF-B. Fundación M Torres; Fundación Eugenio Rodríguez Pascual; Fundación Mario Losantos; Fundación Familia Puig-Infante and Fundación Bancaria La CaixaHepacare Project.

New subventricular zone (SVZ)-derived neuroblasts that migrate via the rostral migratory stream are continuously added to the olfactory bulb (OB) of the adult rodent brain. Anosmin-1 (A1) is an extracellular matrix protein that binds to FGF receptor 1 (FGFR1) to exert its biological effects. When mutated as in Kallmann syndrome patients, A1 is associated with severe OB morphogenesis defects leading to anosmia and hypogonadotropic hypogonadism. Here, we show that A1 over-expression in adult mice strongly increases proliferation in the SVZ, mainly with symmetrical divisions, and produces substantial morphological changes in the normal SVZ architecture, where we also report the presence of FGFR1 in almost all SVZ cells. Interestingly, for the first time we show FGFR1 expression in the basal body of primary cilia in neural progenitor cells. Additionally, we have found that A1 over-expression also enhances neuroblast motility, mainly through FGFR1 activity. Together, these changes lead to a selective increase in several GABAergic interneuron populations in different OB layers. These specific alterations in the OB would be sufficient to disrupt the normal processing of sensory information and consequently alter olfactory memory. In summary, this work shows that FGFR1-mediated A1 activity plays a crucial role in the continuous remodelling of the adult OB., This research was supported by grants from the Spanish Ministerio de Economía, Innovación y Competitividad MINECO (SAF2009-07842, ADE10-0010, RD07-0060-2007, RD12-0032-12 and SAF2012-40023 to FdC; and BFU2010-18284 to JMG-V), FISCAM (Gobierno de Castilla-La Mancha, Spain—Grant Number PI2007-66), the Junta de Castilla y León (Spain, to EW), and from the Fundación Eugenio Rodríguez Pascual (Spain) to FdC. DGG and VMB were PhD students hired by Gobierno de Castilla-La Mancha (MOV2010-JI/11 and MOV2007-JI/19, respectively). FdCS is a CSIC staff scientist in special permission hired by SESCAM (Gobierno de Castilla-La Mancha, Spain). PFE was a researcher hired by SESCAM (Gobierno de Castilla-La Mancha) and ADE10-0010.

During development of the central nervous system, anosmin-1 (A1) works as a chemotropic cue contributing to axonal outgrowth and collateralization, as well as modulating the migration of different cell types, fibroblast growth factor receptor 1 (FGFR1) being the main receptor involved in all these events. To further understand the role of A1 during development, we have analysed the over-expression of human A1 in a transgenic mouse line. Compared with control mice during development and in early adulthood, A1 over-expressing transgenic mice showed an enhanced oligodendrocyte precursor cell (OPC) proliferation and a higher number of OPCs in the subventricular zone and in the corpus callosum (CC). The migratory capacity of OPCs from the transgenic mice is increased in vitro due to a higher basal activation of ERK1/2 mediated through FGFR1 and they also produced more myelin basic protein (MBP). In vivo, the over-expression of A1 resulted in an elevated number of mature oligodendrocytes with higher levels of MBP mRNA and protein, as well as increased levels of activation of the ERK1/2 proteins, while electron microscopy revealed thicker myelin sheaths around the axons of the CC in adulthood. Also in the mature CC, the nodes of Ranvier were significantly longer and the conduction velocity of the nerve impulse in vivo was significantly increased in the CC of A1 over-expressing transgenic mice. Altogether, these data confirmed the involvement of A1 in oligodendrogliogenesis and its relevance for myelination., This work was supported by grants from the Spanish Ministerio de Economía y Competitividad-MINECO (Grant Numbers SAF2009-07842, SAF2012-40023, RD07-0060-2007 and RD12-0032-12 Red Española de Esclerosis Múltple) and the “Fundación Eugenio Rodríguez Pascual” to FdC; the Fundación para la Investigación Socio-sanitaria de Castilla La-Mancha FISCAM (Grant Number PI2007-66 to FdC and PI2009-29 to PFE); from the Spanish MINECO (Grant Numbers BFU2008-00899 and BFU2008-03390) and the Junta de Andalucía (Grant Numbers BIO-122, CVI-02487 and P07-CVI-02686) to JMDG and AGM; and from the Spanish MINECO (Grant Number BFU-2009-08404) to RL. VMB was hired under FISCAM (MOV2007-JI/19), SAF2009-07842 and RD07-0060-2007; PFE was hired by the Servicio de Salud de Castilla La-Mancha (SESCAM, Gobierno de Castilla-La Mancha) and under grant ADE10-0010 from MINECO to FdC. FdC is a CSIC Staff Scientist on leave of absence and currently hired by SESCAM.

[Background]: Hyperglycemia is acknowledged as a pro-inflammatory condition and a major cause of vascular damage. Nevertheless, we have previously described that high glucose only promotes inflammation in human vascular cells previously primed with pro-inflammatory stimuli, such as the cytokine interleukin (IL)1β. Here, we aimed to identify the cellular mechanisms by which high glucose exacerbates the vascular inflammation induced by IL1β. [Methods]: Cultured human aortic smooth muscle cells (HASMC) and isolated rat mesenteric microvessels were treated with IL1β in medium containing 5.5-22 mmol/L glucose. Glucose uptake and consumption, lactate production, GLUT1 levels, NADPH oxidase activity and inflammatory signalling (nuclear factor-ΚB activation and inducible nitric oxide synthase expression) were measured in HASMC, while endothelium-dependent relaxations to acetylcholine were determined in rat microvessels. Pharmacological inhibition of IL1 receptors, NADPH oxidase and glucose-6-phosphate dehydrogenase (G6PD), as well as silencing of G6PD, were also performed. Moreover, the pentose phosphate pathway (PPP) activity and the levels of reduced glutathione were determined. [Results]: We found that excess glucose uptake in HASMC cultured in 22 mM glucose only occurred following activation with IL1β. However, the simple entry of glucose was not enough to be deleterious since over-expression of the glucose transporter GLUT1 or increased glucose uptake following inhibition of mitochondrial respiration by sodium azide was not sufficient to trigger inflammatory mechanisms. In fact, besides allowing glucose entry, IL1β activated the PPP, thus permitting some of the excess glucose to be metabolized via this route. This in turn led to an over-activation NADPH oxidase, resulting in increased generation of free radicals and the subsequent downstream pro-inflammatory signalling. Moreover, in rat mesenteric microvessels high glucose incubation enhanced the endothelial dysfunction induced by IL1β by a mechanism which was abrogated by the inhibition of the PPP. [Conclusions]: A pro-inflammatory stimulus like IL1β transforms excess glucose into a vascular deleterious agent by causing an increase in glucose uptake and its subsequent diversion into the PPP, promoting the pro-oxidant conditions required for the exacerbation of pro-oxidant and pro-inflammatory pathways. We propose that over-activation of the PPP is a crucial mechanism for the vascular damage associated to hyperglycemia., This study was supported with grants from Plan Nacional de I+ D (SAF2014-52762-R), Sociedad Española de Farmacología-Almirall, Fundación Eugenio Rodríguez Pascual, and ISCIII (RETICEF-R12/0043/0021) and Junta de Castilla y León (SA003U13). LV was supported by a fellowship from CONACYT (Mexico). TR is the recipient of a Marie Curie Intra-European Fellowship (2012-IEF-328793 ADDIO).

[Background] P19 H-Ras, a second product derived from the H-Ras gene by alternative splicing, induces a G1/S phase delay, thereby maintaining cells in a reversible quiescence state. When P21 H-Ras is mutated in tumour cells, the alternative protein P19 H-Ras is also mutated. The H-Ras mutation Q61L is frequently detected in different tumours, which acts as constitutive activator of Ras functions and is considered to be a strong activating mutant. Additionally, a rare congenital disorder named Costello Syndrome, is described as a H-Ras disorder in children, mainly due to mutation G12S in p19 and p21 H-Ras proteins, which is present in 90 % of the Costello Syndrome patients. Our aim is to better understand the role of p19 and p21 H-Ras proteins in the cancer and Costello Syndrome development, concerning the miRNAs expression., [Methods] Total miRNAs expression regulated by H-Ras proteins were first analyzed in human miRNA microarrays assays. Previously selected miRNAs, were further analyzed in developed cell lines containing H-Ras protein mutants, that included the G12S Costello Syndrome mutant, with PCR Real-Time Taq Man miRNA Assays primers., [Results] This study describes how p19 affects the RNA world and shows that: i) miR-342, miR-206, miR-330, miR-138 and miR-99b are upregulated by p19 but not by p19W164A mutant; ii) anti-miR-206 can restore the G2 phase in the presence of p19; iii) p19 and p21Q61L regulate their own alternative splicing; iv) miR-206 and miR-138 are differentially regulated by p19 and p21 H-Ras and v) P19G12S Costello mutants show a clear upregulation of miR-374, miR-126, miR-342, miR-330, miR-335 and let-7., [Conclusions] These results allow us to conclude that the H-Ras G12S mutation plays an important role in miRNA expression and open up a new line of study to understand the consequences of this mutation on Costello syndrome. Furthermore, they suggest that oncogenes may have a sufficiently important impact on miRNA expression to promote the development of numerous cancers., This work was supported by the Plan Nacional (MEC) BFU2005-00701, FIS PI080007 and the Fundación Eugenio Rodríguez Pascual. M.C. was funded by an Fmed MMA fellowship. We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI)