Prince Felipe Research Center

Valencia, Spain

Prince Felipe Research Center

Valencia, Spain
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Alfonso-Loeches S.,Prince Felipe Research Center | Urena-Peralta J.R.,Prince Felipe Research Center | Morillo-Bargues M.J.,Prince Felipe Research Center | La Cruz J.O.-D.,Prince Felipe Research Center | Guerri C.,Prince Felipe Research Center
Frontiers in Cellular Neuroscience | Year: 2014

Toll-like receptors (TLRs) and NOD-like receptors (NLRs) are innate immunity sensors that provide an early/effective response to pathogenic or injury conditions. We have reported that ethanol-induced TLR4 activation triggers signaling inflammatory responses in glial cells, causing neuroinflammation and brain damage. However, it is uncertain if ethanol is able to activate NLRs/inflammasome in astroglial cells, which is the mechanism of activation, and whether there is crosstalk between both immune sensors in glial cells. Here we show that chronic ethanol treatment increases the co-localization of caspase-1 with GFAP+ cells, and up-regulates IL-1β and IL-18 in the frontal medial cortex in WT, but not in TLR4 knockout mice. We further show that cultured cortical astrocytes expressed several inflammasomes (NLRP3, AIM2, NLRP1, and IPAF), although NLRP3 mRNA is the predominant form. Ethanol, as ATP and LPS treatments, up-regulates NLRP3 expression, and causes caspase-1 cleavage and the release of IL-1β and IL-18 in astrocytes supernatant. Ethanol-induced NLRP3/caspase-1 activation is mediated by mitochondrial (m) reactive oxygen species (ROS) generation because when using a specific mitochondria ROS scavenger, the mito-TEMPO (500μM) or NLRP3 blocking peptide (4μg/ml) or a specific caspase-1 inhibitor, Z-YVAD-FMK (10μM), abrogates mROS release and reduces the up-regulation of IL-1β and IL-18 induced by ethanol or LPS or ATP. Confocal microscopy studies further confirm that ethanol, ATP or LPS promotes NLRP3/caspase-1 complex recruitment within the mitochondria to promote cell death by caspase-1-mediated pyroptosis, which accounts for≈73% of total cell death (≈22%) and the remaining (≈25%) die by caspase-3-dependent apoptosis. Suppression of the TLR4 function abrogates most ethanol effects on NLRP3 activation and reduces cell death. These findings suggest that NLRP3 participates, in ethanol-induced neuroinflammation and highlight the NLRP3/TLR4 crosstalk in ethanol-induced brain injury. © 2014 Alfonso-Loeches, Ureña-Peralta, Morillo-Bargues, Oliver-De La Cruz and Guerri.

Mathur D.,University of Valencia | Cano A.H.,Prince Felipe Research Center
Annals of Neurosciences | Year: 2012

Ever since the pathology of multiple sclerosis (MS) was described, it has been debated whether immune-mediated myelin damage is a primary or secondary event to axon degeneration. Traditionally the disease is considered to ensue as a consequence of demyelination followed by neuronal loss. Recent in vivo findings, however, demonstrate axons to be the primary target of the disease process. Animal models of MS are greatly instrumental in studying the pathogenesis of the disease and understanding the sequence of events that take place in neurodegenerative disorders.

Sencadas V.,University of Minho | Martins P.,University of Minho | Pitaes A.,University of Minho | Pitaes A.,Polytechnic University of Valencia | And 5 more authors.
Langmuir | Year: 2011

This work reports on the nucleation of the β-phase of poly(vinylidene fluoride) (PVDF) by incorporating CoFe2O4 and NiFe 2O4 nanoparticles, leading in this way to the preparation of magnetoelectric composites. The fraction of filler nanoparticles needed to produce the same β- to α-phase ratio in crystallized PVDF is 1 order of magnitude lower in the cobalt ferrite nanoparticles. The interaction between nanoparticles and PVDF chains induce the all-trans conformation in PVDF segments, and this structure then propagates in crystal growth. The nucleation kinetics is enhanced by the presence of nanoparticles, as corroborated by the increasing number of spherulites with increasing nanoparticle content and by the variations of the Avrami's exponent. Further, the decrease of the crystalline fraction of PVDF with increasing nanoparticle content indicates that an important fraction of polymer chains are confined in interphases with the filler particle. © 2011 American Chemical Society.

Aguilar-Gallardo C.,Prince Felipe Research Center | Rutledge E.C.,Prince Felipe Research Center | Martinez-Arroyo A.M.,Prince Felipe Research Center | Hidalgo J.J.,University of Valencia | And 3 more authors.
Stem Cell Reviews and Reports | Year: 2012

Understanding the genetic and molecular mechanisms of ovarian cancer has been the focus of research efforts working toward the greater goal of improving cancer therapy for patients with residual disease after initial treatment with conventional surgery and neoadjuvant chemotherapy. The focus of this review will be centered on new therapeutic strategies based on Cancer Stem Cells studies of chemoresistant subpopulations, the prevention of metastasis, and individualized therapy in order to find the most successful combination of treatments to effectively treat human ovarian cancer. We reviewed recent literature (1993-2011) of novel treatment approaches to ovarian cancer stem cells. As the focus of ovarian cancer investigation has centered on the cancer stem cell model and the complexities that it presents in the development of effective treatments, the future of treating ovarian cancer lies in utilizing individualized treatment systems that include enhancing existing treatments, aiming for novel therapy targets, managing the plasticity of stem cells to induce cellular differentiation, and regulating oncogenic signaling pathways. © 2012 Springer Science+Business Media, LLC.

Peris B.,Prince Felipe Research Center | Gonzalez-Granero S.,University of Valencia | Ballester-Lurbe B.,CEU Cardenal Herrera University | Garcia-Verdugo J.-M.,University of Valencia | And 4 more authors.
Journal of Neurochemistry | Year: 2012

Proper development of neuronal networks relies on the polarization of the neurons, thus the establishment of two compartments, axons and dendrites, whose formation depends on cytoskeletal rearrangements. Rnd proteins are regulators of actin organization and they are important players in several aspects of brain development as neurite formation, axon guidance and neuron migration. We have recently demonstrated that mice lacking RhoE/Rnd3 expression die shortly after birth and have neuromotor impairment and neuromuscular alterations, indicating an abnormal development of the nervous system. In this study, we have further investigated the specific role played by RhoE in several aspects of neuronal development by using hippocampal neuron cultures. Our findings show that neurons from a mice lacking RhoE expression exhibit a decrease in the number and the total length of the neurites. We also show that RhoE-deficient neurons display a reduction in axon outgrowth and a delay in the process of neuronal polarization. In addition, our results suggest an involvement of the RHOA/ROCK/LIMK/COFILIN signaling pathway in the neuronal alterations induced by the lack of RhoE. These findings support our previous report revealing the important role of RhoE in the normal development of the nervous system and may provide novel therapeutic targets in neurodegenerative disorders. © 2012 The Authors Journal of Neurochemistry © 2012 International Society for Neurochemistry.

Aguilar-Gallardo C.,Prince Felipe Research Center | Simon C.,Prince Felipe Research Center | Simon C.,University of Valencia
Seminars in Reproductive Medicine | Year: 2013

The stem cell field owes a great deal to the previous work conducted by embryologists and researchers devoted to reproductive medicine. The time is coming when this emerging field will pay off in the reproductive sciences by offering new avenues of understanding gametogenesis and early embryonic development. Human embryonic stem cells are pluripotent cells that proliferate in vitro while maintaining an undifferentiated state, and they are capable of differentiating into most cell types under appropriate conditions. Embryo-friendly approaches have been developed as new methods of obtaining human embryonic stem cells without destroying the embryo. Somatic stem cells have been identified and isolated from numerous adult organs and tissues to create a multipotent and autologous source of cells with established medical indications. Cell reprogramming is now a scientific fact, and induced pluripotent cells, a new pluripotent cell type, have been generated by the overexpression of specific genes from a myriad of differentiated adult cell types. Cancer is now considered a stem cell disease. Cancer stem cells share numerous features with normal stem cells including hallmarks properties such as self-renewal and undifferentiation. Therefore, the actual focus of ovarian cancer research on the cancer stem cell model should generate efficient and personalized treatment designs to improve treatment efficiency. © 2013 by Thieme Medical Publishers, Inc.

Cervello I.,University of Valencia | Mas A.,University of Valencia | Gil-Sanchis C.,University of Valencia | Peris L.,University of Valencia | And 5 more authors.
PLoS ONE | Year: 2011

Endometrial regeneration is mediated, at least in part, by the existence of a specialized somatic stem cell (SSC) population recently identified by several groups using the side population (SP) technique. We previously demonstrated that endometrial SP displays genotypic, phenotypic and the functional capability to develop human endometrium after subcutaneous injection in NOD-SCID mice. We have now established seven human endometrial SP (hESP) cell lines (ICE 1-7): four from the epithelial and three from the stromal fraction, respectively. SP cell lines were generated under hypoxic conditions based on their cloning efficiency ability, cultured for 12-15 passages (20 weeks) and cryopreserved. Cell lines displayed normal 46XX karyotype, intermediate telomerase activity pattern and expressed mRNAs encoding proteins that are considered characteristic of undifferentiated cells (Oct-4, GDF3, DNMT3B, Nanog, GABR3) and those of mesodermal origin (WT1, Cardiac Actin, Enolase, Globin, REN). Phenotype analysis corroborated their epithelial (CD9+) or stromal (vimentin+) cell origin and mesenchymal (CD90+, CD73+ and CD45-) attributes. Markers considered characteristic of ectoderm or endoderm were not detected. Cells did not express either estrogen receptor alpha (ERα) or progesterone receptor (PR). The hESP cell lines were able to differentiate in vitro into adipocytes and osteocytes, which confirmed their mesenchymal origin. Finally, we demonstrated their ability to generate human endometrium when transplanted beneath the renal capsule of NOD-SCID mice. These findings confirm that SP cells exhibit key features of human endometrial SSC and open up new possibilities for the understanding of gynecological disorders such as endometriosis or Asherman syndrome. Our cell lines can be a valuable model to investigate new targets for endometrium proliferation in endometriosis. © 2011 Cervelló et al.

Nueda M.J.,University of Alicante | Tarazona S.,Prince Felipe Research Center | Tarazona S.,Polytechnic University of Valencia | Conesa A.,Prince Felipe Research Center
Bioinformatics | Year: 2014

Motivation: The widespread adoption of RNA-seq to quantitatively measure gene expression has increased the scope of sequencing experimental designs to include time-course experiments. maSigPro is an R package specifically suited for the analysis of time-course gene expression data, which was developed originally for microarrays and hence was limited in its application to count data.Results: We have updated maSigPro to support RNA-seq time series analysis by introducing generalized linear models in the algorithm to support the modeling of count data while maintaining the traditional functionalities of the package. We show a good performance of the maSigPro-GLM method in several simulated time-course scenarios and in a real experimental dataset.Availability and implementation: The package is freely available under the LGPL license from the Bioconductor Web site ( © 2014 The Author.

Galan A.,Prince Felipe Research Center | Simon C.,Prince Felipe Research Center
Methods in Molecular Biology | Year: 2012

In this chapter, we describe the derivation and characterization of nine hIn this chapter, we describe the derivation and characterization of nine human embryonic stem cells (hESC) (VAL-3 to-11B) from different developmental embryo stages (inner cell mass from a blastocyst, morula, and blastomere from a 3-day embryo) under xeno-free conditions providing the necessary protocols and techniques to carry out their derivation, characterization, and propagation. © 2012 Springer Science+Business Media, LLC.

Galan A.,Prince Felipe Research Center | Simon C.,Prince Felipe Research Center
Methods in molecular biology (Clifton, N.J.) | Year: 2010

Human embryonic stem cells (hESC) involve long-term cultures that must remain undifferentiated. The real-time PCR (RT-PCR) technique allows the relative quantification of target genes, including undifferentiation and differentiation markers when referred to a housekeeping control with the addition of a calibrator that serves as an internal control to compare different lots of reactions during the time. The main aspects will include a minimal number of cells to be analyzed, genes to be tested, and how to choose the appropriate calibrator sample and the reference gene. In this chapter, we present how to apply the RT-PCR technique, protocols for its performance, experimental set-up and software analysis, as of the gene expression of hESC lines in consecutive passages for long-term culture surveillance.

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