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3. EC50 and CC50 of 39 antiviral drug candidates

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

4. Gating strategy for the identification of cell subpopulations in the human lung tissue model

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

5. S4 Fig - Evaluation of SARS-CoV-2 entry, inflammation and new therapeutics in human lung tissue cells

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

6. Antiviral drug candidates for entry inhibition of SARS-CoV-2

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

7. Gating strategy for the identification of anti-inflammatory effects of selected compounds

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

8. Optimization of lung tissue enzymatic digestion visualized by t-distributed Stochastic Neighbor Embedding (tSNE), and representative SPC and ACE2 expression

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Published
2022

9. Visual summary of the HLT model

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Abstract

Ex vivo physiological systems for the study of SARS-CoV-2-host interactions are scarce. We establish a method using primary human lung tissue (HLT) cells for the rapid analysis of cell tropism and identification of therapeutics. Main findings: i) HLT cells preserve main cell subpopulations, including alveolar type-II cells, and expression of SARS-CoV-2 entry factors ACE2, CD147, TMPRSS2 and AXL. ii) HLT cells are readily susceptible to SARS-CoV-2 infection without the need of cell isolation or further cell differentiation. iii) Antiviral testing in HLT cells allows the rapid identification of new drug candidates against SARS-CoV-2 variants, missed by conventional systems. iv) Local inflammation is supported in HLT cells and offers the identification of relevant anti-inflammatory compounds for SARS-CoV-2 infection.

Published
2022

10. Evaluation of SARS-CoV-2 entry, inflammation and new therapeutics in human lung tissue cells

Author

Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Fundació La Marató de TV3, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Gilead Sciences, Taller Argal, Vall d'Hebron Research Institute, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Fundació La Marató de TV3, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Gilead Sciences, Taller Argal, Vall d'Hebron Research Institute, Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Abstract

The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions directly from primary human lung tissues (HLT), we have developed a rapid platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. The direct use of HLT cells, without long-term cell culture and in vitro differentiation approaches, preserves main immune and structural cell populations, including the most susceptible cell targets for SARS-CoV-2; alveolar type II (AT-II) cells, while maintaining the expression of proteins involved in viral infection, such as ACE2, TMPRSS2, CD147 and AXL. Further, antiviral testing of 39 drug candidates reveals a highly reproducible method, suitable for different SARS-CoV-2 variants, and provides the identification of new compounds missed by conventional systems, such as VeroE6. Using this method, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a relevant and rapid method for the study of SARS-CoV-2.

Published
2022

11. Human lung for the study of SARS-CoV-2

Author

Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Red Española de Investigación en SIDA, Fundació La Marató de TV3, Gilead Sciences, Taller Argal, Buzón, María José, and Buzón, María José [0000-0003-4427-9413]

Abstract

The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions directly from primary human lung tissues (HLT), we have developed a rapid platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. The direct use of HLT cells, without long-term cell culture and in vitro differentiation approaches, preserves main immune and structural cell populations, including the most susceptible cell targets for SARS-CoV-2; alveolar type II (AT-II) cells, while maintaining the expression of proteins involved in viral infection, such as ACE2, TMPRSS2, CD147 and AXL. Further, antiviral testing of 39 drug candidates reveals a highly reproducible method, suitable for different SARS-CoV-2 variants, and provides the identification of new compounds missed by conventional systems, such as VeroE6. Using this method, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a relevant and rapid method for the study of SARS-CoV-2., This work was primarily supported by a grant from the Health Department of the Government of Catalonia (DGRIS 1_5). This work was additionally supported in part by the Spanish Health Institute Carlos III (ISCIII, PI17/01470; PI19CIII/00004; PI21CIII/00025 and COV20-00679 (MPY 222-20)), the Spanish Secretariat of Science and Innovation and FEDER funds (grant RTI2018-101082-B-I00 [MINECO/FEDER]), the Spanish AIDS network Red Temática Cooperativa de Investigación en SIDA (RD16/0025/0007 and RD16CIII/0002/0001), the European Regional Development Fund (ERDF), the Fundació La Marató TV3 (grants 201805-10FMTV3 and 201814- 10FMTV3), the Gilead fellowships GLD19/00084 and GLD18/00008 and the Becas Taller Argal 2020. M.J.B is supported by the Miguel Servet program funded by the Spanish Health Institute Carlos III (CP17/00179). N.M. is supported by a Ph.D. fellowship from the Vall d’Hebron Institut de Recerca (VHIR). The funders had no role in study design, data collection and analysis, the decision to publish, or preparation of the manuscript.

Published
2021

12. Novel Human Lung Tissue Model for the Study of SARS-CoV-2 Entry, Inflammation and New Therapeutics

Author

Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Fundació La Marató de TV3, Gilead Sciences, Buzón, María José [0000-0003-4427-9413], Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Fundació La Marató de TV3, Gilead Sciences, Buzón, María José [0000-0003-4427-9413], Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Abstract

The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions from primary human lung tissues (HLT), we have developed a platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. We show that the HLT model preserves its main cell populations, maintains the expression of proteins required for SARS-CoV-2 infection, and identifies alveolar type II (AT-II) cells as the most susceptible cell targets for SARS-CoV-2 in the human lung. Antiviral testing of 39 drug candidates revealed a highly reproducible system, and provided the identification of new compounds missed by conventional systems such as VeroE6. Using this model, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a novel and relevant physiological model for the study of SARS-CoV-2.

Published
2021

13. Evaluation of SARS-CoV-2 Entry, Inflammation and New Therapeutics in Human Lung Tissue Cells

Author

Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Red Española de Investigación en SIDA, Fundació La Marató de TV3, Gilead Sciences, Taller Argal, Buzón, María José [0000-0003-4427-9413], Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, Buzón, María José, Generalitat de Catalunya, Instituto de Salud Carlos III, European Commission, Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Red Española de Investigación en SIDA, Fundació La Marató de TV3, Gilead Sciences, Taller Argal, Buzón, María José [0000-0003-4427-9413], Grau-Expósito, Judith, Perea, David, Suppi, Marina, Massana, Nuria, Vergara, Ander, Soler, María José, Trinité, Benjamin, Blanco, Julià, García-Pérez, Javier, Alcamí, José, Serrano-Mollar, Anna, Rosado, Joel, Falcó, Vicenç, Genesca, Meritxell, and Buzón, María José

Abstract

The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions directly from primary human lung tissues (HLT), we have developed a rapid platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. The direct use of HLT cells, without long-term cell culture and in vitro differentiation approaches, preserves main immune and structural cell populations, including the most susceptible cell targets for SARS-CoV-2; alveolar type II (AT-II) cells, while maintaining the expression of proteins involved in viral infection, such as ACE2, TMPRSS2, CD147 and AXL. Further, antiviral testing of 39 drug candidates reveals a highly reproducible method, suitable for different SARS-CoV-2 variants, and provides the identification of new compounds missed by conventional systems, such as VeroE6. Using this method, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a relevant and rapid method for the study of SARS-CoV-2.

Published
2021

14. Beyond the scavenging of reactive oxygen species (ROS): Direct effect of cerium oxide nanoparticles in reducing fatty acids content in an In vitro model of hepatocellular steatosis

Author

Ministerio de Economía y Competitividad (España), European Commission, Generalitat de Catalunya, Instituto de Salud Carlos III, Natural Science Foundation of Guangdong Province, Wuyi University, Parra-Robert, Marina, Casals, Eudald, Massana, Nuria, Zeng, Muling, Perramón, Meritxell, Fernández-Varo, Guillermo, Morales-Ruiz, Manuel, Puntes, Víctor F., Jiménez, Wladimiro, Casals, Gregori, Ministerio de Economía y Competitividad (España), European Commission, Generalitat de Catalunya, Instituto de Salud Carlos III, Natural Science Foundation of Guangdong Province, Wuyi University, Parra-Robert, Marina, Casals, Eudald, Massana, Nuria, Zeng, Muling, Perramón, Meritxell, Fernández-Varo, Guillermo, Morales-Ruiz, Manuel, Puntes, Víctor F., Jiménez, Wladimiro, and Casals, Gregori

Abstract

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic accumulation of lipids. Antisteatotic effects of cerium oxide nanoparticles (CeO2NPs) have recently been shown in animal models of liver disease. However, it is unclear whether the activity of CeO2NPs is related solely to the decrease in oxidative stress or, in addition, they directly decrease liver fatty acid accumulation. To address this question, in this work, we used an in vitro model of hepatocellular steatosis, exposing HepG2 cells to oleic and palmitic acid. Cell uptake of CeO2NPs and their effect on oxidative stress and viability of hepatic cells cultured with H2O2 were also evaluated. Results show that CeO2NPs were uptaken by HepG2 cells and reduced oxidative stress and improved cell viability. Treatment with oleic and palmitic acid increased lipogenesis and the content of different fatty acids. CeO2NPs reduced palmitic and stearic acid and most fatty acids consisting of more than 18 carbon atoms. These effects were associated with significant changes in elongase and desaturase activity. In conclusion, CeO2NPs directly protected HepG2 cells from cell injury in oxidative stress conditions and reduced fatty acid content in steatotic conditions by inducing specific changes in fatty acid metabolism, thus showing potential in the treatment of NAFLD.

Published
2019

15. Beyond the Scavenging of Reactive Oxygen Species (ROS): Direct Effect of Cerium Oxide Nanoparticles in Reducing Fatty Acids Content in an In Vitro Model of Hepatocellular Steatosis

Author

Parra-Robert, Marina, primary, Casals, Eudald, additional, Massana, Nuria, additional, Zeng, Muling, additional, Perramón, Meritxell, additional, Fernández-Varo, Guillermo, additional, Morales-Ruiz, Manuel, additional, Puntes, Víctor, additional, Jiménez, Wladimiro, additional, and Casals, Gregori, additional

Published
2019
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16. Caracterización estructural y funcional de la ARN polimerasa dependiente de ARN del virus de Thosea asigna

Author

Ferrero, Diego Sebastián, Rodríguez Aguirre, José Francisco, Verdaguer Massana, Nuria, and UAM. Departamento de Biología Molecular

Subjects
ARN polimerasas - Tesis Doctorales, Insectos - Parásitos - Tesis Doctorales, Biología y Biomedicina / Biología
Abstract

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 22-07-2013, Thosea asigna virus (TaV) is an insect-restricted virus with a monosegmented positivesense, single stranded RNA (ssRNA) genome enclosed within an icosahedral T = 4 capsid. TaV belongs to the recently created Permutotetraviridae family, which includes non-enveloped viruses with a particular organization of sequence motifs within their RNA dependent RNA polymerases (RdRp). All polymerases are broadly conserved in viruses following a right hand architecture with fingers, palm and thumb subdomains, and also conserved six ordered sequence motifs (A-B-C-D-E-F), four of them located into the palm subdomain (A to D) and two (E-F) that are only present in RdRps. Previous sequence analyses showed that, similarly to what had been found for members of the Birnaviridae family, the TaV RpRd does not follow the canonical organization (C-A-B-D) of the palm subdomain. Given the rarity of these atypical polymerases, the structural and biochemical information about them is rather scarce. For this reason, we performed a comprehensive study of the RdRp of TaV. We have solved the structure of a recombinant version of the TaV RdRp domain purified from insect cells by X-ray crystallography. The exhaustive analysis of the atomic structure allowed the identification of the conservation of the overall fold of the polymerase which shows a surprisingly high degree of similarity to its Flavivirus counterpart. We have also identified several structural elements that might play a role in the regulation of the polymerase activity, i.e. the 30 N-terminal residues and the extense loop that block the active site cavity that may act as negative regulator elements. This finding suggested that these elements may undergo a structural rearrangement, thus allowing the polymerase to be active by a de novo initiation mechanism as confirmed by biochemical analyses presented here. The use of mutant versions of the TaV RdRp provided insight into general mechanisms controlling RdRp activity and about the regulatory role played by their own structural elements. In addition, electron microscopy images led to the generation of a three-dimensional reconstruction of the complete protein encoded by TaV ORF1. This provided relevant structural information concerning the C-terminal domain of the polypeptide. Results gathered by confocal microscopy analysis of a series of mutant versions of the protein expressed in insect cell indicate that the C-terminal domain of the TaV ORF protein is responsible for the formation of veshicle-like structures, and probably for its interaction with an as yet unindentified membranous compartment, within the cell cytoplasm where the viral replication machinery is located. According to the results presented here, TaV can be related to Flaviviridae family considering the structural and functional similarity

Published
2013

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