Imatge de diagramació Imatge de diagramació Imatge de diagramació Imatge de diagramació
Logo Universitat de Barcelona Facultat de Farmàcia Imatge de diagramació
Inici Facultat Inici UB   
Imatge de diagramació
Recerca Imatge de diagramació
Imatge de diagramació Imatge de diagramació
Imatge de diagramació Imatge de diagramació

 

 

 

 

Imatge de diagramació
imatge de diagramació
Índex
Comissió de recerca
Convocatòries i ajuts a la recerca
Conferències de recerca
Oficina de recerca
Serveis de suport a la recerca
Liníes de recerca
Enllaços d'interés
 
 
Icona d'informació Per a més informació:

Facultat de Farmàcia
Oficina de Recerca
Edifici A - Primer pis

Avinguda Joan XXIII s/n
08028 - Barcelona
Telèfon: 93 402 1093
Fax: 93 403 1886

Imatge de diagramació

 

 

 

 

 

 

 

 

Molecular phylogenetics and temporal diversification in the genus Aeromonas based on the sequences of five housekeeping genes

J. Gaspar Lorén, Maribel Farfán,* M. Carmen Fusté

* Departament de Microbiologia i Parasitologia Sanitàries, Unitat de Microbiologia

Abstract

Several approaches have been developed to estimate both the relative and absolute rates of speciation and extinction within clades based on molecular phylogenetic reconstructions of evolutionary relationships, according to an underlying model of diversification. However, the macroevolutionary models established for eukaryotes have scarcely been used with prokaryotes. We have investigated the rate and pattern of cladogenesis in the genus Aeromonas ( ?-Proteobacteria , Proteobacteria , Bacteria) using the sequences of five housekeeping genes and an uncorrelated relaxed-clock approach. To our knowledge, until now this analysis has never been applied to all the species described in a bacterial genus and thus opens up the possibility of establishing models of speciation from sequence data commonly used in phylogenetic studies of prokaryotes. Our results suggest that the genus Aeromonas began to diverge between 248 and 266 million years ago, exhibiting a constant divergence rate through the Phanerozoic, which could be described as a pure birth process.

Article complet

Grup de recerca

 

Stability and immunogenicity properties of the gene-silencing polypurine reverse Hoogsteen hairpins  

Xenia Villalobos, Laura Rodríguez, Jeanne Prévot, Carlota Oleaga, Carlos J. Ciudad, Véronique Noé*

* Departament de Bioquímica i Biologia Molecular

Abstract

Gene silencing by either small-interference RNAs (siRNA) or antisense oligodeoxynucleotides (aODN) is widely used in biomedical research. However, their use as therapeutic agents is hindered by two important limitations: their low stability and the activation of the innate immune response. Recently, we developed a new type of molecule to decrease gene expression named polypurine reverse Hoogsteen hairpins (PPRHs) that bind to polypyrimidine targets in the DNA. Herein, stability experiments performed in mouse, human, and fetal calf serum and in PC3 cells revealed that the half-life of PPRHs is much longer than that of siRNAs in all cases. Usage of PPRHs with a nicked-circular structure increased the binding affinity to their target sequence and their half-life in FCS when bound to the target. Regarding the innate immune response, we determined that the levels of the transcription factors IRF3 and its phosphorylated form, as well as NF-?B were increased by siRNAs and not by PPRHs; that the expression levels of several proin?ammatory cytokines including IL-6, TNF-a, IFN-a, IFN-ß, IL-1ß, and IL-18 were not signi?cantly increased by PPRHs; and that the cleavage and activation of the proteolytic enzyme caspase-1 was not triggered by PPRHs. These determinations indicated that PPRHs, unlike siRNAs, do not activate the innate in?ammatory response.

Article complet

Grup de recerca

 

Polypurine reverse Hoogsteen hairpins as a gene therapy tool against survivin in human prostate cancer PC3 cells in vitro and in vivo  

Laura Rodríguez, Xenia Villalobos, Sheila Dakhel, Laura Padilla, Rosa Hervas, Jose Luis Hernández, Carlos J. Ciudad,* Véronique Noé

* Departament de Bioquímica i Biologia Molecular

Abstract

As a new approach for gene therapy, we recently developed a new type of molecule called polypurine reverse Hoogsteen hairpins (PPRHs). We decided to explore the in vitro and in vivo effect of PPRHs in cancer choosing survivin as a target since it is involved in apoptosis, mitosis and angiogenesis, and overexpressed in different tumors. We designed four PPRHs against the survivin gene, one of them directed against the template strand and three against different regions of the coding strand. These PPRHs were tested in PC3 prostate cancer cells in an in vitro screening of cell viability and apoptosis. PPRHs against the promoter sequence were the most effective and caused a decrease in survivin mRNA and protein levels. We con?rmed the binding between the selected PPRHs and their target sequences in the survivin gene. In addition we determined that both the template- and the coding-PPRH targeting the survivin promoter were interfering with the binding of transcription factors Sp1 and GATA-3, respectively. Finally, we conducted two in vivo ef?cacy assays using the Coding-PPRH against the survivin promoter and performing two routes of administration, namely intratumoral and intravenous, in a subcutaneous xenograft tumor model of PC3 prostate cancer cells. The results showed that the chosen Coding-PPRH proved to be effective in decreasing tumor volume, and reduced the levels of survivin protein and the formation of blood vessels. These ?ndings represent the preclinical proof of principle of PPRHs as a new silencing tool for cancer gene therapy .

 

Article complet

Grup de recerca

 

3 - Azatetracyclo[5.2.1.1 5,8 .0 1,5 ]undecane derivatives: from wild-type inhibitors of the M2 ion channel of influenza A virus to derivatives with potent activity against the V27A mutant  

Matias Rey-Carrizo, Eva Torres, Chunlong Ma, Marta Barniol-Xicota, Jun Wang, Yibing Wu, Lieve Naesens, William F. DeGrado, Robert A. Lamb, Lawrence H. Pinto, and Santiago Va´zquez

* Departament de Farmacologia i Química Terapèutica, Unitat de Química Farmacèutica

Abstract

We have synthesized and characterized a series of compounds containing the 3-azatetracyclo[5.2.1.1 5,8 .0 1,5 ]undecane scaffold designed as analogues of amantadine, an inhibitor of the M2 proton channel of influenza A virus. Inhibition of the wild-type (WT) M2 channel and the amantadine-resistant A/M2-S31N and A/M2-V27A mutant ion channels were measured in Xenopus oocytes using two-electrode voltage clamp (TEV) assays. Most of the novel compounds inhibited the WT ion channel in the low micromolar range. Of note, several compounds inhibited the A/M2 V27A mutant ion channel, one of them with submicromolar IC 50 . None of the compounds was found to inhibit the S31N mutant ion channel. The antiviral activity of three novel dual WT and A/M2-V27A channels inhibitors was confirmed by influenza virus yield assays .

 

Article complet

Grup de recerca

 

Arabidopsis semidwarfs evolved from independent mutations in GA20ox1 , ortholog to green revolution dwarf alleles in rice and barley  

Luis Barboza, Sigi Effgen, Carlos Alonso-Blanco, Rik Kooke, Joost J. B. Keurentjes, Maarten Koornneef, and Rubén Alcázar*

* Departament de Productes Naturals, Biologia Vegetal i Edafologia, Unitat de Fisiologia Vegetal

Abstract

Understanding the genetic bases of natural variation for developmental and stress-related traits is a major goal of current plant biology. Variation in plant hormone levels and signaling might underlie such phenotypic variation occurring even within the same species. Here we report the genetic and molecular basis of semidwarf individuals found in natural Arabidopsis thaliana populations. Allelism tests demonstrate that independent loss-of-function mutations at GA locus 5 ( GA5 ), which encodes gibberellin 20-oxidase 1 ( GA20ox1 ) involved in the last steps of gibberellin biosynthesis, are found in different populations from southern, western, and northern Europe; central Asia; and Japan . Sequencing of GA5 identified 21 different loss-of-function alleles causing semidwarfness without any obvious general tradeoff affecting plant performance traits. GA5 shows signatures of purifying selection, whereas GA5 loss-of-function alleles can also exhibit patterns of positive selection in specific populations as shown by Fay and Wu's H statistics. These results suggest that antagonistic pleiotropy might underlie the occurrence of GA5 loss-of-function mutations in nature. Furthermore, because GA5 is the ortholog of rice SD1 and barley Sdw1 / Denso green revolution genes, this study illustrates the occurrence of conserved adaptive evolution between wild A. thaliana and domesticated plants.

Article complet

Grup de recerca

 

Therapeutic targeting of tumor growth and angiogenesis with a novel anti-S100A4 monoclonal antibody

Jose Luis Hernández, Laura Padilla, Sheila Dakhel, Toni Coll, Rosa Hervas, Jaume Adan, Marc Masa, Francesc Mitjans, Josep Maria Martinez, Silvia Coma, Laura Rodríguez,* Véronique Noé,* Carlos J. Ciudad,* Francesc Blasco, Ramon Messeguer

* Departament de Bioquímica i Biologia Molecular.

 

Abstract

S100A4, a member of the S100 calcium-binding protein family secreted by tumor and stromal cells, supports tumorigenesis by stimulating angiogenesis. We demonstrated that S100A4 synergizes with vascular endothelial growth factor (VEGF), via the RAGE receptor, in promoting endothelial cell migration by increasing KDR expression and MMP-9 activity. In vivo overexpression of S100A4 led to a significant increase in tumor growth and vascularization in a human melanoma xenograft M21 model. Conversely, when silencing S100A4 by shRNA technology, a dramatic decrease in tumor development of the pancreatic MiaPACA-2 cell line was observed. Based on these results we developed 5C3, a neutralizing monoclonal antibody against S100A4. This antibody abolished endothelial cell migration, tumor growth and angiogenesis in immunodeficient mouse xenograft models of MiaPACA-2 and M21-S100A4 cells. It is concluded that extracellular S100A4 inhibition is an attractive approach for the treatment of human cancer.

 

Article complet

Grup de recerca

 

 

Mixed protein–DNA gel particles for DNA delivery: Role of protein composition and preparation method on biocompatibility  

M.C. Morán ,* D.R. Nogueira, M.P. Vinardell, M.G. Miguel, B. Lindman

* Departament de Fisiologia.

Abstract

Mixtures of two cationic proteins were used to prepare protein–DNA gel particles, employing associative phase separation and interfacial diffusion ( Morán et al., 2009a ). By mixing the two proteins, we have obtained particles that displayed higher loading efficiency and loading capacity values than those obtained in single-protein systems. However, nothing is known about the adverse effects on haemo-compatibility and cytotoxicity of these protein–DNA gel particles. Here, we examined the interaction of protein–DNA gel particles obtained by two different preparation methods, and their components, with red blood cells and established cells. From a haemolytic point of view, these protein–DNA gel particles were demonstrated to be promising long-term blood-contacting medical devices. Safety evaluation with the established cell lines revealed that, in comparison with proteins in solution, the cytotoxicity was reduced when administered in the protein–DNA systems. In comparison with large-sized particles, the cytotoxic responses of small-sized protein–DNA gel particles showed to be strongly dependent of both the protein composition and the cell line being the tumour cell line HeLa more sensitive to the deleterious effects of the mixed protein-based particles. The observed trends in haemolysis and cell viabilities were in agreement with the degree of complexation values obtained for the protein–DNA gel particles prepared by both preparation methods.

Article complet

Grup de recerca

 

Establishment of an in vitro photoassay using THP-1 cells and IL-8 to discriminate photoirritants from photoallergens

V. Martínez, V. Galbiati, E. Corsini, R. Martín-Venegas, M.P. Vinardell, M. Mitjans*

* Departament de Fisiologia.

Abstract

At present, there are no in vivo or in vitro methods developed which has been adopted by regulatory authorities to assess photosensitization induced by chemicals. Recently, we have proposed the use of THP-1 cells and IL-8 release to identify the potential of chemicals to induce skin sensitization. Based on the assumption that sensitization and photosensitization share common mechanisms, the aim of this work was to explore the THP-1 model as an in vitro model to identify photoallergenic chemicals.

THP-1 cells were exposed to 7 photoallergens and 3 photoirritants and irradiated with UVA light or kept in dark. Non phototoxic allergens or irritants were also included as negative compounds. Following 24 h of incubation, cytotoxicity and IL-8 release were measured. At subtoxic concentrations, photoallergens produced a dose-related increase in IL-8 release after irradiation. Some photoirritants also produced a slight increase in IL-8 release. However, when the overall stimulation indexes of IL-8 were calculated for each chemical, 6 out of 7 photoallergens tested reached a stimulation index above 2, while the entire set of negative compounds had stimulation indexes below 2. Our data suggest that this assay may become a useful cell-based in vitro test for evaluating the photosensitizing potential of chemicals.

Article complet

Grup de recerca

 

Draft genome sequence of the Aeromonas diversa type strain

Maribel Farfán, Nino Spataro, Ariadna Sanglas, Vicenta Albarral, J. Gaspar Lorén, Elena Bosch, M. Carmen Fusté*

*Departament de Microbiologia i Parasitologia Sanitàries, Unitat de Microbiologia.

Abstract

We present here the first genome sequence of the Aeromonas diversa type strain (CECT 4254 T ). This strain was isolated from the leg wound of a patient in New Orleans (Louisiana) and was originally described as enteric group 501 and distinguished from A. schubertii by DNA-DNA hybridization and phenotypical characterization.

 

Article complet

Grup de recerca

 

Draft genome sequence of Aeromonas molluscorum strain 848T T , isolated from bivalve molluscs

Nino Spataro, Maribel Farfán, Vicenta Albarral, Ariadna Sanglas, J. Gaspar Lorén, M. Carmen Fusté,* Elena Bosch

*Departament de Microbiologia i Parasitologia Sanitàries, Unitat de Microbiologia.

Abstract

We report here the draft genome sequence of Aeromonas molluscorum 848T, the type strain of this Aeromonas species, which was isolated from wedge shells ( Donax trunculus ) obtained from a retail market in Barcelona, Spain, in 1997.

 

Article complet

Grup de recerca

 

 

Impact of physical parameters on particle size and reaction yield when using the ionic gelation method to obtain cationic polymeric chitosan–tripolyphosphate nanoparticles  

A. Fàbregas,* M. Miñarro, E. García-Montoya, P. Pérez-Lozano, C. Carrillo, R. Sarrate, N. Sánchez, J.R. Tic ó , J.M. Suñé-Negre

* Departament de Farmàcia i Tecnologia Farmacèutica.

Abstract

Ionic gelation is the most frequently used method to obtain chitosan-tripolyphosphate nanoparticles due to its simplicity and because it does not generate waste solvents in the samples prepared.

This paper presents a study of the physical factors involved in this method for obtaining nanoparticles in order to determine which of them significantly influences the particle size of polymeric nanoparticles made from low-molecular-weight chitosan, without any additional chemical treatment, with the aim of standardising and optimising the method conditions, in addition to establishing the reaction yield.

The results indicate that stirring speed during ionic gelation reaction is decisive for the size of the nanoparticles obtained. Furthermore, it thus follows that the stirring speed during ionic gelation significantly affects reaction yield, and therefore, by manipulating this parameter a greater proportion of nanoparticles of a given size range can be obtained.

 

Article complet

Grup de recerca

 

 

Thioflavin-T excimer formation upon interaction with amyloid fibers  

Raimon Sabate,* Luis Rodriguez-Santiago, Mariona Sodupe, Sven J. Saupe and Salvador Ventura

* Departament de Fisicoquímica.

Abstract

The molecular mechanism of the Thioflavin-T (Th-T) binding to amyloids remains unknown. By combining experimental analysis of Th-T excitation and emission spectra with theoretical calculations we suggest that Th-T fluorescence changes upon interaction with amyloids may arise from the formation of an excimer with an oblique angle of approx. 120 degrees.

Article complet

Grup de recerca

 

The redox state of cytochrome c modulates resistance to methotrexate in human MCF7 breast cancer cells

Susana Barros, Núria Mencia, Laura Rodríguez, Carlota Oleaga, Conceiçao Santos, Verónique Noé, Carlos J. Ciudad*

Departament de Bioquímica i Biologia Molecular

Abstract

Background: Methotrexate is a chemotherapeutic agent used to treat a variety of cancers. However, the occurrence of resistance limits its effectiveness. Cytochrome c in its reduced state is less capable of triggering the apoptotic cascade. Thus, we set up to study the relationship among redox state of cytochrome c, apoptosis and the development of resistance to methotrexate in MCF7 human breast cancer cells.

Results: Cell incubation with cytochrome c-reducing agents, such as tetramethylphenylenediamine, ascorbate or reduced glutathione, decreased the mortality and apoptosis triggered by methotrexate. Conversely, depletion of glutathione increased the apoptotic action of methotrexate, showing an involvement of cytochrome c redox state in methotrexate-induced apoptosis. Methotrexate-resistant MCF7 cells showed increased levels of endogenous reduced glutathione and a higher capability to reduce exogenous cytochrome c. Using functional genomics we detected the overexpression of GSTM1 and GSTM4 in methotrexate-resistant MCF7 breast cancer cells, and determined that methotrexate was susceptible of glutathionylation by GSTs. The inhibition of these GSTM isoforms caused an increase in methotrexate cytotoxicity in sensitive and resistant cells.

Conclusions: We conclude that overexpression of specific GSTMs, GSTM1 and GSTM4, together with increased endogenous reduced glutathione levels help to maintain a more reduced state of cytochrome c which, in turn, would decrease apoptosis, thus contributing to methotrexate resistance in human MCF7 breast cancer cells.

 

Article complet

Grup de recerca

 

Cocoa flavanol metabolites activate HNF-3 beta , Sp1, and NFY-mediated transcription of apolipoprotein AI in human cells

Carlota Oleaga, Carlos J. Ciudad, Maria Izquierdo-Pulido and Véronique Noé *

* Departament de Bioquímica i Biologia Molecular.

Abstract

Scope: To identify the mechanisms by which cocoa induces HDL levels and since apolipoprotein AI (ApoAI) is the major protein in HDLs, we analyzed, upon incubation with cocoa metabolites, ApoAI mRNA levels, its transcriptional regulation, and the levels of the transcription factors involved in this process.

Methods and results: Epicatechin and cocoa metabolites caused an increase in ApoAI expression in HepG2 cells. Electrophoretic mobility shift assays revealed the involvement of Sites A and B of the ApoAI promoter in the induction of ApoAI mRNA upon incubation with cocoa metabolites. Using supershift assays, we demonstrated the binding of HNF-3beta, HNF-4, ER-alpha, and RXR-alpha to Site A and the binding of HNF-3beta, NFY, and Sp1 to Site B. Luciferase assays performed with a construct containing Site B confirmed its role in the upregulation of ApoAI by cocoa metabolites. Incubation with 3-methyl-epicatechin led to an increase in HNF-3beta mRNA, HNF-3beta, ER-alpha, Sp1, and NFY protein levels and the activation of ApoAI transcriptionmediated by NFY, Sp1, and ER-alpha.

Conclusion: The activation of ApoAI transcription through Site B by cocoa flavanol metabolites is mainly mediated by an increase in HNF-3beta, with a significant contribution of Sp1 and NFY, as a mechanism for the protective role of these compounds in cardiovascular diseases.

 

Article complet

Grup de recerca

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 
 

 

 

 

 

 

 

 

 
Imatge de diagramació  
Imatge de diagramació Imatge de diagramació Imatge de diagramació Imatge de diagramació
  © Universitat de Barcelona Edició: Secretaria d'Estudiants i Docència Facultat de Farmàcia
Última actualització o validació:10.03.2014