Staff member

Núria Montserrat Pulido

Group Leader / ICREA Research Professor
Pluripotency for organ regeneration
+34 934 031 391
CV Summary

Dra. Nuria Montserrat became interested in organ regeneration and stem cells during her master and PhD training that finished in 2006. The same year she got a Postdoctoral fellowship from the Fundaçao para a Ciência e Tecnología (Portugal). In 2007 she moved as a post-doctoral researcher at the Hospital of Santa Creu i Sant Pau in Barcelona.

In 2008 she joined the Center of Regenerative Medicine of Barcelona (CMRB) thanks to the support of a Juan de la Cierva fellowship under the direction of Dr. Izpisúa Belmonte. In 2010 she first co-authored how to reprogram cord blood stem cells for the first time (Nature Protocols, 2010). Then she first-coauthor the first work deriving iPSCs with new factors (Cell Stem Cell, 2013). She also collaborated in projects aimed to characterize the genomic integrity of human iPSCs as well as their differentiation towards different lineages for applications in human disease modeling (Stem Cells, 2011; Nature, 2012; Nature, 2012, Nature Communications, 2014). She has first co-authored how the reactivation of endogenous pathways can be artificially reactivated and promote heart regeneration in mammals (Cell Stem Cell, 2014).

Her expertise in the fields of somatic reprogramming and organ regeneration was recognized with an awarded an ERC Starting Grant by 2014 which allowed her to become Junior group leader at the Institute of Bioengineering of Catalonia (IBEC).

In January 2015 she got a Ramon y Cajal fellowship (first ranked candidate, 100/100 points) and since 2019 she is an ICREA Research Professor and Senior Group Leader at IBEC. During these years her findings in the field of Regenerative Medicine led to the derivation, for the first time, of cardiac grafts from human pluripotent stem cells and decellularized cardiac myocardium (Biomaterials, 2016), and the derivation of renal analogues with 3D bioprinting (Materials Today, 2017) and vascularized kidney organoids from hPSCs (Nature Materials, 2019).

She has recently co-led on the application of kidney organoid technology to model SARS-CoV-2 infections (Cell, 2020) identifying a therapeutic compound that nowadays is under clinical trial in COVID19 patients (The Lancet Respiratory Medicine, 2020; EMBO Molecular Medicine, 2020). Their Cell paper has been highlighted as a Research Highlight in both Nature Reviews Nephrology and in the Nature journal, also attracting remarkable attention (more than 200 national press releases, and 30 radio/television) being awarded as the “Best biomedical research publication 2020” (Constantes y Vitales Prize).
With the aim to further understand on the central role of metabolism in renal fate and damage protection she has recently co-led the first work on the identification of metabolic regulators protecting the renal tubule from acute injury exploiting kidney organoid technology (Cell Metabolism, 2020). In sum, from 2015, her work has attracted almost 8 M€ of direct competitive funding and around 1.2 M€ in personnel related grants, from both Spanish and European institutions.

All these efforts have been recently awarded with the prestigious EMBO Young Investigator Prize. In December 2020 the ERC has recognized all these efforts and she have been awarded with the prestigious ERC Consolidator Grant to study the interplay between mechanobiology and metabolism during human kidney development and disease.

Her commitment to scientific dissemination and communication has allowed her to participate in more than 300 outreach activities to promote our research activities to the general public, with a particular focus to young girls and woman in science. For this reason, she was selected as Commissioner of the first City and Science Biennial of Barcelona in 2019 being re-elected for the second edition (June, 2021).

Staff member publications

Selfa, Idoia Lucía, Gallo, Maria, Montserrat, Nuria, Garreta, Elena, (2021). Directed differentiation of human pluripotent stem sells for the generation of high-order kidney organoids Programmed Morphogenesis: Methods and Protocols (ed. Ebrahimkhani M.R., Hislop J.), Springer US (New York, NY) 2258, 171-192

Our understanding in the inherent properties of human pluripotent stem cells (hPSCs) have made possible the development of differentiation procedures to generate three-dimensional tissue-like cultures, so-called organoids. Here we detail a stepwise methodology to generate kidney organoids from hPSCs. This is achieved through direct differentiation of hPSCs in two-dimensional monolayer culture toward the posterior primitive streak fate, followed by induction of intermediate mesoderm-committed cells, which are further aggregated and cultured in three-dimensions to generate kidney organoids containing segmented nephron-like structures in a process that lasts 20 days. We also provide a concise description on how to assess renal commitment during the time course of kidney organoid generation. This includes the use of flow cytometry and immunocytochemistry analyses for the detection of specific renal differentiation markers.

Kyndiah, A., Leonardi, F., Tarantino, C., Cramer, T., Millan-Solsona, R., Garreta, E., Montserrat, N., Mas-Torrent, M., Gomila, G., (2020). Bioelectronic recordings of cardiomyocytes with accumulation mode electrolyte gated organic field effect transistors Biosensors and Bioelectronics 150, 111844

Organic electronic materials offer an untapped potential for novel tools for low-invasive electrophysiological recording and stimulation devices. Such materials combine semiconducting properties with tailored surface chemistry, elastic mechanical properties and chemical stability in water. In this work, we investigate solution processed Electrolyte Gated Organic Field Effect Transistors (EGOFETs) based on a small molecule semiconductor. We demonstrate that EGOFETs based on a blend of soluble organic semiconductor 2,8-Difluoro-5,11-bis(triethylsilylethynyl)anthradithiophene (diF-TES-ADT) combined with an insulating polymer show excellent sensitivity and long-term recording under electrophysiological applications. Our devices can stably record the extracellular potential of human pluripotent stem cell derived cardiomyocyte cells (hPSCs-CMs) for several weeks. In addition, cytotoxicity tests of pharmaceutical drugs, such as Norepinephrine and Verapamil was achieved with excellent sensitivity. This work demonstrates that organic transistors based on organic blends are excellent bioelectronics transducer for extracellular electrical recording of excitable cells and tissues thus providing a valid alternative to electrochemical transistors.

Keywords: Bioelectronics, Cardiac cells, Organic electronics, Organic field effect transistors, Organic semiconducting blend

Lynch, Cian J., Bernad, Raquel, Martínez-Val, Ana, Shahbazi, Marta N., Nóbrega-Pereira, Sandrina, Calvo, Isabel, Blanco-Aparicio, Carmen, Tarantino, Carolina, Garreta, Elena, Richart-Ginés, Laia, Alcazar, Noelia, Graña-Castro, Osvaldo, Gómez-Lopez, Gonzalo, Aksoy, Irene, Muñoz-Martín, Maribel, Martinez, Sonia, Ortega, Sagrario, Prieto, Susana, Simboeck, Elisabeth, Camasses, Alain, Stephan-Otto Attolini, Camille, Fernandez, Agustin F., Sierra, Marta I., Fraga, Mario F., Pastor, Joaquin, Fisher, Daniel, Montserrat, Nuria, Savatier, Pierre, Muñoz, Javier, Zernicka-Goetz, Magdalena, Serrano, Manuel, (2020). Global hyperactivation of enhancers stabilizes human and mouse naive pluripotency through inhibition of CDK8/19 Mediator kinases Nature Cell Biology 22, (10), 1223-1238

Pluripotent stem cells (PSCs) transition between cell states in vitro, reflecting developmental changes in the early embryo. PSCs can be stabilized in the naive state by blocking extracellular differentiation stimuli, particularly FGF–MEK signalling. Here, we report that multiple features of the naive state in human and mouse PSCs can be recapitulated without affecting FGF–MEK signalling or global DNA methylation. Mechanistically, chemical inhibition of CDK8 and CDK19 (hereafter CDK8/19) kinases removes their ability to repress the Mediator complex at enhancers. CDK8/19 inhibition therefore increases Mediator-driven recruitment of RNA polymerase II (RNA Pol II) to promoters and enhancers. This efficiently stabilizes the naive transcriptional program and confers resistance to enhancer perturbation by BRD4 inhibition. Moreover, naive pluripotency during embryonic development coincides with a reduction in CDK8/19. We conclude that global hyperactivation of enhancers drives naive pluripotency, and this can be achieved in vitro by inhibiting CDK8/19 kinase activity. These principles may apply to other contexts of cellular plasticity.

Zoufaly, Alexander, Poglitsch, Marko, Aberle, Judith H., Hoepler, Wolfgang, Seitz, Tamara, Traugott, Marianna, Grieb, Alexander, Pawelka, Erich, Laferl, Hermann, Wenisch, Christoph, Neuhold, Stephanie, Haider, Doris, Stiasny, Karin, Bergthaler, Andreas, Puchhammer-Stoeckl, Elisabeth, Mirazimi, Ali, Montserrat, Nuria, Zhang, Haibo, Slutsky, Arthur S., Penninger, Josef M., (2020). Human recombinant soluble ACE2 in severe COVID-19 The Lancet Respiratory Medicine Online first,

Angiotensin converting enzyme 2 (ACE2) is the crucial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor and protects multiple tissues, including the lung, from injury as a regulator of the renin–angiotensin system.1 Therefore, ACE2 has become the focus of COVID-19 research and a plethora of drug development efforts. Among the novel compounds under development is human recombinant soluble ACE2 (hrsACE2 [APN01; Apeiron Biologics, Vienna, Austria]), which has two mechanisms of action that theoretically should be of benefit in COVID-19.2 The first involves binding the viral spike protein and thereby neutralising SARS-CoV-2,3 and the second is minimising injury to multiple organs, including the lungs, kidneys, and heart, because of unabated renin–angiotensin system hyperactivation and increased angiotensin II concentrations.4, 5, 6 hrsACE2 has been tested in 89 patients, namely in healthy volunteers in phase 1 studies and in patients with acute respiratory distress syndrome (ARDS) in phase 2 clinical studies, with a good safety profile.7, 8 Moreover, hrsACE2 can reduce SARS-CoV-2 load by a factor of 1000–5000 in in-vitro cell-culture experiments and engineered organoids, directly demonstrating that ACE2 can effectively neutralise SARS-CoV-2.3 We describe in this Case Report the first course of treatment with hrsACE2 of a patient with severe COVID-19.

Monteil, Vanessa, Dyczynski, Matheus, Lauschke, Volker M., Kwon, Hyesoo, Wirnsberger, Gerald, Youhanna, Sonia, Zhang, Haibo, Slutsky, Arthur S., Hurtado del Pozo, Carmen, Horn, Moritz, Montserrat, Nuria, Penninger, Josef M., Mirazimi, Ali, (2020). Human soluble ACE2 improves the effect of remdesivir in SARS-CoV-2 infection EMBO Molecular Medicine , e13426

There is a critical need for safe and effective drugs for COVID-19. Only remdesivir has received authorization for COVID-19 and has been shown to improve outcomes but not decrease mortality. However, the dose of remdesivir is limited by hepatic and kidney toxicity. ACE2 is the critical cell surface receptor for SARS-CoV-2. Here, we investigated additive effect of combination therapy using remdesivir with recombinant soluble ACE2 (high/low dose) on Vero E6 and kidney organoids, targeting two different modalities of SARS-CoV-2 life cycle: cell entry via its receptor ACE2 and intracellular viral RNA replication. This combination treatment markedly improved their therapeutic windows against SARS-CoV-2 in both models. By using single amino-acid resolution screening in haploid ES cells, we report a singular critical pathway required for remdesivir toxicity, namely, Adenylate Kinase 2. The data provided here demonstrate that combining two therapeutic modalities with different targets, common strategy in HIV treatment, exhibit strong additive effects at sub-toxic concentrations. Our data lay the groundwork for the study of combinatorial regimens in future COVID-19 clinical trials.

Monteil, Vanessa, Kwon, Hyesoo, Prado, Patricia, Hagelkrüys, Astrid, Wimmer, Reiner A., Stahl, Martin, Leopoldi, Alexandra, Garreta, Elena, Hurtado Del Pozo, Carmen, Prosper, Felipe, Romero, Juan Pablo, Wirnsberger, Gerald, Zhang, Haibo, Slutsky, Arthur S., Conder, Ryan, Montserrat, Nuria, Mirazimi, Ali, Penninger, Josef M., (2020). Inhibition of SARS-CoV-2 infections in engineered human tissues using clinical-grade soluble human ACE2 Cell 181, (4), 905-913.e7

We have previously provided the first genetic evidence that angiotensin converting enzyme 2 (ACE2) is the critical receptor for severe acute respiratory syndrome coronavirus (SARS-CoV), and ACE2 protects the lung from injury, providing a molecular explanation for the severe lung failure and death due to SARS-CoV infections. ACE2 has now also been identified as a key receptor for SARS-CoV-2 infections, and it has been proposed that inhibiting this interaction might be used in treating patients with COVID-19. However, it is not known whether human recombinant soluble ACE2 (hrsACE2) blocks growth of SARS-CoV-2. Here, we show that clinical grade hrsACE2 reduced SARS-CoV-2 recovery from Vero cells by a factor of 1,000-5,000. An equivalent mouse rsACE2 had no effect. We also show that SARS-CoV-2 can directly infect engineered human blood vessel organoids and human kidney organoids, which can be inhibited by hrsACE2. These data demonstrate that hrsACE2 can significantly block early stages of SARS-CoV-2 infections.

Keywords: COVID-19, Angiotensin converting enzyme 2, Blood vessels, Human organoids, Kidney, Severe acute respiratory syndrome coronavirus, Spike glycoproteins, Treatment

Dhillon, Poonam, Park, Jihwan, Hurtado del Pozo, Carmen, Li, Lingzhi, Doke, Tomohito, Huang, Shizheng, Zhao, Juanjuan, Kang, Hyun Mi, Shrestra, Rojesh, Balzer, Michael S., Chatterjee, Shatakshee, Prado, Patricia, Han, Seung Yub, Liu, Hongbo, Sheng, Xin, Dierickx, Pieterjan, Batmanov, Kirill, Romero, Juan P., Prósper, Felipe, Li, Mingyao, Pei, Liming, Kim, Junhyong, Montserrat, Nuria, Susztak, Katalin, (2020). The nuclear receptor ESRRA protects from kidney disease by coupling metabolism and differentiation Cell Metabolism In Press, Corrected Proof

Kidney disease is poorly understood because of the organ’s cellular diversity. We used single-cell RNA sequencing not only in resolving differences in injured kidney tissue cellular composition but also in cell-type-specific gene expression in mouse models of kidney disease. This analysis highlighted major changes in cellular diversity in kidney disease, which markedly impacted whole-kidney transcriptomics outputs. Cell-type-specific differential expression analysis identified proximal tubule (PT) cells as the key vulnerable cell type. Through unbiased cell trajectory analyses, we show that PT cell differentiation is altered in kidney disease. Metabolism (fatty acid oxidation and oxidative phosphorylation) in PT cells showed the strongest and most reproducible association with PT cell differentiation and disease. Coupling of cell differentiation and the metabolism was established by nuclear receptors (estrogen-related receptor alpha [ESRRA] and peroxisomal proliferation-activated receptor alpha [PPARA]) that directly control metabolic and PT-cell-specific gene expression in mice and patient samples while protecting from kidney disease in the mouse model.

Keywords: Single-cell RNA sequencing, Single-cell ATAC sequencing, Kidney, Fibrosis, Organoids, Fatty-acid oxidation, PPARA, ESRRA, Proximal tubule cells, Chronic kidney disease

Garreta, Elena, Kamm, Roger D., Chuva de Sousa Lopes, Susana M., Lancaster, Madeline A., Weiss, Ron, Trepat, Xavier, Hyun, Insoo, Montserrat, Nuria, (2020). Rethinking organoid technology through bioengineering Nature Materials ahead

In recent years considerable progress has been made in the development of faithful procedures for the differentiation of human pluripotent stem cells (hPSCs). An important step in this direction has also been the derivation of organoids. This technology generally relies on traditional three-dimensional culture techniques that exploit cell-autonomous self-organization responses of hPSCs with minimal control over the external inputs supplied to the system. The convergence of stem cell biology and bioengineering offers the possibility to provide these stimuli in a controlled fashion, resulting in the development of naturally inspired approaches to overcome major limitations of this nascent technology. Based on the current developments, we emphasize the achievements and ongoing challenges of bringing together hPSC organoid differentiation, bioengineering and ethics. This Review underlines the need for providing engineering solutions to gain control of self-organization and functionality of hPSC-derived organoids. We expect that this knowledge will guide the community to generate higher-grade hPSC-derived organoids for further applications in developmental biology, drug screening, disease modelling and personalized medicine.

Cilloni, Daniela, Petiti, Jessica, Campia, Valentina, Podestà , Marina, Squillario, Margherita, Montserrat, Nuria, Bertaina, Alice, Sabatini, Federica, Carturan, Sonia, Berger, Massimo, Saglio, Francesco, Bandini, Giuseppe, Bonifazi, Francesca, Fagioli, Franca, Moretta, Lorenzo, Saglio, Giuseppe, Verri, Alessandro, Barla, Annalisa, Locatelli, Franco, Frassoni, Francesco, (2020). Transplantation induces profound changes in the transcriptional asset of hematopoietic stem cells: Identification of specific signatures using machine learning techniques Journal of Clinical Medicine 9, (6), 1670

During the phase of proliferation needed for hematopoietic reconstitution following transplantation, hematopoietic stem/progenitor cells (HSPC) must express genes involved in stem cell self-renewal. We investigated the expression of genes relevant for self-renewal and expansion of HSPC (operationally defined as CD34+ cells) in steady state and after transplantation. Specifically, we evaluated the expression of ninety-one genes that were analyzed by real-time PCR in CD34+ cells isolated from (i) 12 samples from umbilical cord blood (UCB); (ii) 15 samples from bone marrow healthy donors; (iii) 13 samples from bone marrow after umbilical cord blood transplant (UCBT); and (iv) 29 samples from patients after transplantation with adult hematopoietic cells. The results show that transplanted CD34+ cells from adult cells acquire an asset very different from transplanted CD34+ cells from cord blood. Multivariate machine learning analysis (MMLA) showed that four specific gene signatures can be obtained by comparing the four types of CD34+ cells. In several, but not all cases, transplanted HSPC from UCB overexpress reprogramming genes. However, these remarkable changes do not alter the commitment to hematopoietic lineage. Overall, these results reveal undisclosed aspects of transplantation biology.

Keywords: Hematopoietic stem/progenitor cell, Cord blood, Stem cell transplantation

Garreta, Elena, Prado, Patricia, Tarantino, Carolina, Oria, Roger, Fanlo, Lucia, Martí, Elisa, Zalvidea, Dobryna, Trepat, Xavier, Roca-Cusachs, Pere, Gavaldà -Navarro, Aleix, Cozzuto, Luca, Campistol, Josep M., Izpisúa Belmonte, Juan Carlos, Hurtado del Pozo, Carmen, Montserrat, Nuria, (2019). Fine tuning the extracellular environment accelerates the derivation of kidney organoids from human pluripotent stem cells Nature Materials 18, 397-405

The generation of organoids is one of the biggest scientific advances in regenerative medicine. Here, by lengthening the time that human pluripotent stem cells (hPSCs) were exposed to a three-dimensional microenvironment, and by applying defined renal inductive signals, we generated kidney organoids that transcriptomically matched second-trimester human fetal kidneys. We validated these results using ex vivo and in vitro assays that model renal development. Furthermore, we developed a transplantation method that utilizes the chick chorioallantoic membrane. This approach created a soft in vivo microenvironment that promoted the growth and differentiation of implanted kidney organoids, as well as providing a vascular component. The stiffness of the in ovo chorioallantoic membrane microenvironment was recapitulated in vitro by fabricating compliant hydrogels. These biomaterials promoted the efficient generation of renal vesicles and nephron structures, demonstrating that a soft environment accelerates the differentiation of hPSC-derived kidney organoids.

Sample, Matthew, Boulicault, Marion, Allen, Caley, Bashir, Rashid, Hyun, Insoo, Levis, Megan, Lowenthal, Caroline, Mertz, David, Montserrat, Nuria, Palmer, Megan J., Saha, Krishanu, Zartman, Jeremiah, (2019). Multi-cellular engineered living systems: building a community around responsible research on emergence Biofabrication 11, (4), 043001

Ranging from miniaturized biological robots to organoids, multi-cellular engineered living systems (M-CELS) pose complex ethical and societal challenges. Some of these challenges, such as how to best distribute risks and benefits, are likely to arise in the development of any new technology. Other challenges arise specifically because of the particular characteristics of M-CELS. For example, as an engineered living system becomes increasingly complex, it may provoke societal debate about its moral considerability, perhaps necessitating protection from harm or recognition of positive moral and legal rights, particularly if derived from cells of human origin. The use of emergence-based principles in M-CELS development may also create unique challenges, making the technology difficult to fully control or predict in the laboratory as well as in applied medical or environmental settings. In response to these challenges, we argue that the M-CELS community has an obligation to systematically address the ethical and societal aspects of research and to seek input from and accountability to a broad range of stakeholders and publics. As a newly developing field, M-CELS has a significant opportunity to integrate ethically responsible norms and standards into its research and development practices from the start. With the aim of seizing this opportunity, we identify two general kinds of salient ethical issues arising from M-CELS research, and then present a set of commitments to and strategies for addressing these issues. If adopted, these commitments and strategies would help define M-CELS as not only an innovative field, but also as a model for responsible research and engineering.

Keywords: Ethics, Society, Governance, Emergence, Moral considerability, Responsible innovation

Garreta, Elena, Montserrat, Nuria, Belmonte, Juan Carlos Izpisua, (2018). Kidney organoids for disease modeling Oncotarget 9, (16), 12552-12553

Garreta, Elena, Sanchez, Sonia, Lajara, Jeronimo, Montserrat, Nuria, Belmonte, Juan Carlos Izpisua, (2018). Roadblocks in the path of iPSC to the vlinic Current Transplantation Reports 5, (1), 14-18

PURPOSE OF REVIEW: The goal of this paper is to highlight the major challenges in the translation of human pluripotent stem cells into a clinical setting. RECENT FINDINGS: Innate features from human induced pluripotent stem cells (hiPSCs) positioned these patient-specific cells as an unprecedented cell source for regenerative medicine applications. Immunogenicity of differentiated iPSCs requires more research towards the definition of common criteria for the evaluation of innate and host immune responses as well as in the generation of standardized protocols for iPSC generation and differentiation. The coming years will resolve ongoing clinical trials using both human embryonic stem cells (hESCs) and hiPSCs providing exciting information for the optimization of potential clinical applications of stem cell therapies. SUMMARY: Rapid advances in the field of iPSCs generated high expectations in the field of regenerative medicine. Understanding therapeutic applications of iPSCs certainly needs further investigation on autologous/allogenic iPSC transplantation.

Latorre, Ernest, Kale, Sohan, Casares, Laura, Gómez-González, Manuel, Uroz, Marina, Valon, Léo, Nair, Roshna V., Garreta, Elena, Montserrat, Nuria, del Campo, Aránzazu, Ladoux, Benoit, Arroyo, Marino, Trepat, Xavier, (2018). Active superelasticity in three-dimensional epithelia of controlled shape Nature 563, (7730), 203-208

Fundamental biological processes are carried out by curved epithelial sheets that enclose a pressurized lumen. How these sheets develop and withstand three-dimensional deformations has remained unclear. Here we combine measurements of epithelial tension and shape with theoretical modelling to show that epithelial sheets are active superelastic materials. We produce arrays of epithelial domes with controlled geometry. Quantification of luminal pressure and epithelial tension reveals a tensional plateau over several-fold areal strains. These extreme strains in the tissue are accommodated by highly heterogeneous strains at a cellular level, in seeming contradiction to the measured tensional uniformity. This phenomenon is reminiscent of superelasticity, a behaviour that is generally attributed to microscopic material instabilities in metal alloys. We show that in epithelial cells this instability is triggered by a stretch-induced dilution of the actin cortex, and is rescued by the intermediate filament network. Our study reveals a type of mechanical behaviour—which we term active superelasticity—that enables epithelial sheets to sustain extreme stretching under constant tension.

Hernandez-Benitez, R., Llanos Martinez-Martinez, M., Lajara, J., Magistretti, P., Montserrat, N., Izpisua Belmonte, Juan Carlos, (2018). At the heart of genome editing and cardiovascular diseases Circulation Research 123, (2), 221-223

Cardiovascular disease (CVD) is still the leading cause of death worldwide, but the knowledge and technologies for counteracting this disease may already be in our hands. Scientific advances over the past few years, such as the isolation and differentiation of induced pluripotent stem cells, and the development of gene-editing tools, have enabled us to model CVD, but more importantly, may represent tools for CVD early diagnosis, patient stratification, and treatment.

Niederberger, Craig, Pellicer, Antonio, Cohen, Jacques, Gardner, David K., Palermo, Gianpiero D., O'Neill, Claire L., Chow, Stephen, Rosenwaks, Zev, Cobo, Ana, Swain, Jason E., Schoolcraft, William B., Frydman, René, Bishop, Lauren A., Aharon, Davora, Gordon, Catherine, New, Erika, Decherney, Alan, Tan, Seang Lin, Paulson, Richard J., Goldfarb, James M., Brännström, Mats, Donnez, Jacques, Silber, Sherman, Dolmans, Marie-Madeleine, Simpson, Joe Leigh, Handyside, Alan H., Munné, Santiago, Eguizabal, Cristina, Montserrat, Nuria, Izpisua Belmonte, Juan Carlos, Trounson, Alan, Simon, Carlos, Tulandi, Togas, Giudice, Linda C., Norman, Robert J., Hsueh, Aaron J., Sun, Yingpu, Laufer, Neri, Kochman, Ronit, Eldar-Geva, Talia, Lunenfeld, Bruno, Ezcurra, Diego, D'Hooghe, Thomas, Fauser, Bart C. J. M., Tarlatzis, Basil C., Meldrum, David R., Casper, Robert F., Fatemi, Human M., Devroey, Paul, Galliano, Daniela, Wikland, Matts, Sigman, Mark, Schoor, Richard A., Goldstein, Marc, Lipshultz, Larry I., Schlegel, Peter N., Hussein, Alayman, Oates, Robert D., Brannigan, Robert E., Ross, Heather E., Pennings, Guido, Klock, Susan C., Brown, Simon, Van Steirteghem, André, Rebar, Robert W., LaBarbera, Andrew R., (2018). Forty years of IVF Fertility and Sterility 110, (2), 185-324

This monograph, written by the pioneers of IVF and reproductive medicine, celebrates the history, achievements, and medical advancements made over the last 40 years in this rapidly growing field.

Hurtado del Pozo, Carmen, Garreta, Elena, Izpisúa Belmonte, Juan Carlos, Montserrat, Nuria, (2018). Modeling epigenetic modifications in renal development and disease with organoids and genome editing Disease Models & Mechanisms 11, (11), 035048

Understanding epigenetic mechanisms is crucial to our comprehension of gene regulation in development and disease. In the past decades, different studies have shown the role of epigenetic modifications and modifiers in renal disease, especially during its progression towards chronic and end-stage renal disease. Thus, the identification of genetic variation associated with chronic kidney disease has resulted in better clinical management of patients. Despite the importance of these findings, the translation of genotype–phenotype data into gene-based medicine in chronic kidney disease populations still lacks faithful cellular or animal models that recapitulate the key aspects of the human kidney. The latest advances in the field of stem cells have shown that it is possible to emulate kidney development and function with organoids derived from human pluripotent stem cells. These have successfully recapitulated not only kidney differentiation, but also the specific phenotypical traits related to kidney function. The combination of this methodology with CRISPR/Cas9 genome editing has already helped researchers to model different genetic kidney disorders. Nowadays, CRISPR/Cas9-based approaches also allow epigenetic modifications, and thus represent an unprecedented tool for the screening of genetic variants, epigenetic modifications or even changes in chromatin structure that are altered in renal disease. In this Review, we discuss these technical advances in kidney modeling, and offer an overview of the role of epigenetic regulation in kidney development and disease.

Garreta, E., González, F., Montserrat, N., (2018). Studying kidney disease using tissue and genome engineering in human pluripotent stem cells Nephron 138, 48-59

Kidney morphogenesis and patterning have been extensively studied in animal models such as the mouse and zebrafish. These seminal studies have been key to define the molecular mechanisms underlying this complex multistep process. Based on this knowledge, the last 3 years have witnessed the development of a cohort of protocols allowing efficient differentiation of human pluripotent stem cells (hPSCs) towards defined kidney progenitor populations using two-dimensional (2D) culture systems or through generating organoids. Kidney organoids are three-dimensional (3D) kidney-like tissues, which are able to partially recapitulate kidney structure and function in vitro. The current possibility to combine state-of-the art tissue engineering with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated systems 9 (Cas9)-mediated genome engineering provides an unprecedented opportunity for studying kidney disease with hPSCs. Recently, hPSCs with genetic mutations introduced through CRISPR/Cas9-mediated genome engineering have shown to produce kidney organoids able to recapitulate phenotypes of polycystic kidney disease and glomerulopathies. This mini review provides an overview of the most recent advances in differentiation of hPSCs into kidney lineages, and the latest implementation of the CRISPR/Cas9 technology in the organoid setting, as promising platforms to study human kidney development and disease.

Keywords: Clustered regularly interspaced short palindromic repeats/CRISPR-associated systems 9, Disease modeling, Gene editing, Human pluripotent stem cells, Kidney genetics, Tissue engineering

Garreta, Elena, Oria, Roger, Tarantino, Carolina, Pla-Roca, Mateu, Prado, Patricia, Fernández-Avilés, Francisco, Campistol, Josep Maria, Samitier, Josep, Montserrat, Nuria, (2017). Tissue engineering by decellularization and 3D bioprinting Materials Today , 20, (4), 166-178

Discarded human donor organs have been shown to provide decellularized extracellular matrix (dECM) scaffolds suitable for organ engineering. The quest for appropriate cell sources to satisfy the need of multiple cells types in order to fully repopulate human organ-derived dECM scaffolds has opened new venues for the use of human pluripotent stem cells (hPSCs) for recellularization. In addition, three-dimensional (3D) bioprinting techniques are advancing towards the fabrication of biomimetic cell-laden biomaterial constructs. Here, we review recent progress in decellularization/recellularization and 3D bioprinting technologies, aiming to fabricate autologous tissue grafts and organs with an impact in regenerative medicine.

Climent, A. M., Hernandez-Romero, I., Guillem, M. S., Montserrat, N., Fernandez, M. E., Atienza, F., Fernandez-Aviles, F., (2017). High resolution microscopic optical mapping of anatomical and functional reentries in human cardiac cell cultures IEEE Conference Publications Computing in Cardiology Conference (CinC), 2016 , IEEE (Vancouver, Canada) 43, 233-236

Anatomical and/or functional reentries have been proposed as one of the main mechanism of perpetuation of cardiac fibrillation processes. However, technical limitations have difficult the characterization of those reentries and are hampering the development of effective anti-arrhythmic treatments. The goal of this study is to present a novel technology to map with high resolution the center of fibrillation drivers in order to characterize the mechanisms of reentry. Cell cultures of human cardiac-like cells differentiated from pluripotent stem cells were analyzed with a novel microscopic optical mapping system. The pharmacological response to verapamil administration of each type of reentry was analyzed. In all analyzed cell cultures, a reentry was identified as the mechanism of maintenance of the arrhythmia. Interestingly, the administration of verapamil produced opposite effects on activation rate depending on the mechanisms of reentry (i.e. anatomical or functional). Microscopic optical mapping of reentries allows the identification of perpetuation mechanisms which has been demonstrated to be linked with different pharmacological response.

Keywords: Stem cells, Rotors, Microscopy, Optical filters, Calcium, Optical microscopy, Biomedical optical imaging

Garreta, Elena, Marco, Andrés, Eguizábal, Cristina, Tarantino, Carolina, Samitier, Mireia, Badiola, Maider, Gutiérrez, Joaquín, Samitier, Josep, Montserrat, Nuria, (2017). Pluripotent stem cells and skeletal muscle differentiation: Challenges and immediate applications The Plasticity of Skeletal Muscle: From Molecular Mechanism to Clinical Applications (ed. Sakuma, Kunihiro), Springer Singapore (Singapore, Singapore) 2018, 1-35

Recent advances in the generation of skeletal muscle derivatives from pluripotent stem cells (PSCs) provide innovative tools for muscle development, disease modeling, and cell replacement therapies. Here, we revise major relevant findings that have contributed to these advances in the field, by the revision of how early findings using mouse embryonic stem cells (ESCs) set the bases for the derivation of skeletal muscle cells from human pluripotent stem cells (hPSCs) and patient-derived human-induced pluripotent stem cells (hiPSCs) to the use of genome editing platforms allowing for disease modeling in the petri dish.

Keywords: Pluripotent stem cells, Differentiation, Genome editing, Disease modeling

Xia, Yun, Montserrat, Nuria, Campistol, Josep M., Izpisua Belmonte, Juan Carlos, Remuzzi, Giuseppe, Williams, David F., (2017). Lineage reprogramming toward kidney regeneration Kidney Transplantation, Bioengineering and Regeneration (ed. Orlando, G., Remuzzi, Giuseppe, Williams, David F.), Academic Press (London, UK) , 1167-1175

We have known for decades that it is possible to switch the phenotype of one somatic cell type into another. Such epigenetic rewiring processes can be artificially managed and even reversed by using a defined set of transcription factors. Lineage reprogramming is very often defined as a process of converting one cell type into another without going through a pluripotent state, providing great promise for regenerative medicine. However, the identification of key transcription factors for lineage reprogramming is limited, due to the exhaustive and expensive experimental processes. Accumulating knowledge of genetic and epigenetic regulatory networks that are critical for defining a specific lineage provides unprecedented opportunities to model and predict pioneering factors that may drive directional lineage reprogramming to obtain the desired cell type.

Keywords: Reprogramming, Pluripotency, Differentiation, Lineage specification, Epigenetic regulatory network, Regeneration

Garreta, Elena, Marco, Andres, Izpisua Belmonte, Juan Carlos, Montserrat, Nuria, (2016). Genome editing in human pluripotent stem cells: a systematic approach unrevealing pancreas development and disease Stem Cell Investigation , 4, (11), 1-4

Although mouse models have represented a major tool for understanding and predicting molecular mechanisms responsible for several human genetic diseases, still species-specific differences between mouse and humans in their biochemical and physiological characteristics represent a major hurdle when translating promising findings into the human setting (1). For instance, in several types of maturity onset diabetes of the young (MODY; autosomal dominant), mice with heterozygous mutations do not develop diabetes (2). In this regard, the derivation of human embryonic stem cells (hESCs) in 1998 represented an unprecedented opportunity for human disease modelling, and a promising source for cell replacement therapies (3). Later on, the possibility to generate patient-derived induced pluripotent stem cells (iPSCs) has opened new venues for the potential translation of stem-cell related studies into the clinic (4).

Garreta, E., de Oñate, L., Fernández-Santos, M. E., Oria, R., Tarantino, C., Climent, A. M., Marco, A., Samitier, M., Martínez, Elena, Valls-Margarit, M., Matesanz, R., Taylor, D. A., Fernández-Avilés, F., Izpisua Belmonte, J. C., Montserrat, N., (2016). Myocardial commitment from human pluripotent stem cells: Rapid production of human heart grafts Biomaterials 98, 64-78

Genome editing on human pluripotent stem cells (hPSCs) together with the development of protocols for organ decellularization opens the door to the generation of autologous bioartificial hearts. Here we sought to generate for the first time a fluorescent reporter human embryonic stem cell (hESC) line by means of Transcription activator-like effector nucleases (TALENs) to efficiently produce cardiomyocyte-like cells (CLCs) from hPSCs and repopulate decellularized human heart ventricles for heart engineering. In our hands, targeting myosin heavy chain locus (MYH6) with mCherry fluorescent reporter by TALEN technology in hESCs did not alter major pluripotent-related features, and allowed for the definition of a robust protocol for CLCs production also from human induced pluripotent stem cells (hiPSCs) in 14 days. hPSCs-derived CLCs (hPSCs-CLCs) were next used to recellularize acellular cardiac scaffolds. Electrophysiological responses encountered when hPSCs-CLCs were cultured on ventricular decellularized extracellular matrix (vdECM) correlated with significant increases in the levels of expression of different ion channels determinant for calcium homeostasis and heart contractile function. Overall, the approach described here allows for the rapid generation of human cardiac grafts from hPSCs, in a total of 24 days, providing a suitable platform for cardiac engineering and disease modeling in the human setting.

Keywords: Cardiac function, Extracellular matrix, Gene targeting, Pluripotent stem cells

Eguizabal, C., Herrera, L., De Oñate, L., Montserrat, N., Hajkova, P., Izpisua Belmonte, J. C., (2016). Characterization of the epigenetic changes during human gonadal primordial germ cells reprogramming Stem Cells , 34, (9), 2418-2428

Abstract: Epigenetic reprogramming is a central process during mammalian germline development. Genome-wide DNA demethylation in primordial germ cells (PGCs) is a prerequisite for the erasure of epigenetic memory, preventing the transmission of epimutations to the next generation. Apart from DNA demethylation, germline reprogramming has been shown to entail reprogramming of histone marks and chromatin remodelling. Contrary to other animal models, there is limited information about the epigenetic dynamics during early germ cell development in humans. Here, we provide further characterization of the epigenetic configuration of the early human gonadal PGCs. We show that early gonadal human PGCs are DNA hypomethylated and their chromatin is characterized by low H3K9me2 and high H3K27me3 marks. Similarly to previous observations in mice, human gonadal PGCs undergo dynamic chromatin changes concomitant with the erasure of genomic imprints. Interestingly, and contrary to mouse early germ cells, expression of BLIMP1/PRDM1 persists in through all gestational stages in human gonadal PGCs and is associated with nuclear lysine-specific demethylase-1. Our work provides important additional information regarding the chromatin changes associated with human PGCs development between 6 and 13 weeks of gestation in male and female gonads.

Keywords: Epigenetic, Human primordial germ cells, Reprograming

Montserrat, N., Garreta, E., Izpisua Belmonte, J. C., (2016). Regenerative strategies for kidney engineering FEBS Journal , 283, (18), 3303-3324

The kidney is the most important organ for water homeostasis and waste excretion. It performs several important physiological functions for homeostasis: it filters the metabolic waste out of circulation, regulates body fluid balances, and acts as an immune regulator and modulator of cardiovascular physiology. The development of in vitro renal disease models with pluripotent stem cells (both human embryonic stem cells and induced pluripotent stem cells) and the generation of robust protocols for in vitro derivation of renal-specific-like cells from patient induced pluripotent stem cells have just emerged. Here we review major findings in the field of kidney regeneration with a major focus on the development of stepwise protocols for kidney cell production from human pluripotent stem cells and the latest advances in kidney bioengineering (i.e. decellularized kidney scaffolds and bioprinting). The possibility of generating renal-like three-dimensional structures to be recellularized with renal-derived induced pluripotent stem cells may offer new avenues to develop functional kidney grafts on-demand.

Keywords: Induced pluripotent stem cells, Kidney disease, Kidney engineering, Pluripotent stem cells, Renal differentiation

Reddy, Pradeep, Ocampo, Alejandro, Suzuki, Keiichiro, Luo, Jinping, Bacman, Sandra , Williams, Sion, Sugawara, Atsushi, Okamura, Daiji, Tsunekawa, Yuji, Wu, Jun, Lam, David, Xiong, Xiong, Montserrat, Nuria, Esteban, Concepcion, Liu, Guang-Hui, Sancho-Martinez, Ignacio, Manau, Dolors, Civico, Salva, Cardellach, Francesc, del Mar O'Callaghan, Maria, Campistol, Jaime, Zhao, Huimin, Campistol, Josep, Moraes, Carlos, Izpisua Belmonte, Juan Carlos, (2015). Selective elimination of mitochondrial mutations in the germline by genome editing Cell 161, (3), 459-469

Mitochondrial diseases include a group of maternally inherited genetic disorders caused by mutations in mtDNA. In most of these patients, mutated mtDNA coexists with wild-type mtDNA, a situation known as mtDNA heteroplasmy. Here, we report on a strategy toward preventing germline transmission of mitochondrial diseases by inducing mtDNA heteroplasmy shift through the selective elimination of mutated mtDNA. As a proof of concept, we took advantage of NZB/BALB heteroplasmic mice, which contain two mtDNA haplotypes, BALB and NZB, and selectively prevented their germline transmission using either mitochondria-targeted restriction endonucleases or TALENs. In addition, we successfully reduced human mutated mtDNA levels responsible for Leber?s hereditary optic neuropathy (LHOND), and neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP), in mammalian oocytes using mitochondria-targeted TALEN (mito-TALENs). Our approaches represent a potential therapeutic avenue for preventing the transgenerational transmission of human mitochondrial diseases caused by mutations in mtDNA. Mitochondrial diseases include a group of maternally inherited genetic disorders caused by mutations in mtDNA. In most of these patients, mutated mtDNA coexists with wild-type mtDNA, a situation known as mtDNA heteroplasmy. Here, we report on a strategy toward preventing germline transmission of mitochondrial diseases by inducing mtDNA heteroplasmy shift through the selective elimination of mutated mtDNA. As a proof of concept, we took advantage of NZB/BALB heteroplasmic mice, which contain two mtDNA haplotypes, BALB and NZB, and selectively prevented their germline transmission using either mitochondria-targeted restriction endonucleases or TALENs. In addition, we successfully reduced human mutated mtDNA levels responsible for Leber?s hereditary optic neuropathy (LHOND), and neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP), in mammalian oocytes using mitochondria-targeted TALEN (mito-TALENs). Our approaches represent a potential therapeutic avenue for preventing the transgenerational transmission of human mitochondrial diseases caused by mutations in mtDNA.

de Oñate, L., Garreta, E., Tarantino, C., Martínez, Elena, Capilla, E., Navarro, I., Gutiérrez, J., Samitier, J., Campistol, J.M., Muñoz-Cánovas, P., Montserrat, N., (2015). Research on skeletal muscle diseases using pluripotent stem cells Muscle Cell and Tissue (ed. Sakuma, K.), InTech (Rijeka, Croatia) , 333-357

The generation of induced pluripotent stem cells (iPSCs), especially the generation of patient-derived pluripotent stem cells (PSCs) suitable for disease modelling in vitro, opens the door for the potential translation of stem-cell related studies into the clinic. Successful replacement, or augmentation, of the function of damaged cells by patient-derived differentiated stem cells would provide a novel cell-based therapy for skeletal muscle-related diseases. Since iPSCs resemble human embryonic stem cells (hESCs) in their ability to generate cells of the three germ layers, patient-specific iPSCs offer definitive solutions for the ethical and histo-incompatibility issues related to hESCs. Indeed human iPSC (hiPSC)-based autologous transplantation is heralded as the future of regenerative medicine. Interestingly, during the last years intense research has been published on disease-specific hiPSCs derivation and differentiation into relevant tissues/organs providing a unique scenario for modelling disease progression, to screen patient-specific drugs and enabling immunosupression-free cell replacement therapies. Here, we revise the most relevant findings in skeletal muscle differentiation using mouse and human PSCs. Finally and in an effort to bring iPSC technology to the daily routine of the laboratory, we provide two different protocols for the generation of patient-derived iPSCs.

Keywords: Pluripotent stem cells, Myogenic differentiation, Disease modelling, Patient-specific induced pluripotent stem cells, Muscular dystrophy

Garcia, A., Hortigüela, V., Lagunas, A., Cortina, C., Montserrat, N., Samitier, J., Martinez, Elena, (2014). Protein patterning on hydrogels by direct microcontact printing: application to cardiac differentiation RSC Advances 4, (55), 29120-29123

An extended microcontact printing technique to chemically pattern hydrogels is reported. The procedure employs standard polydimethylsiloxane stamps and requires minor pre-processing of the hydrogels by freeze-drying. Micropatterned Matrigel[trade mark sign] and gelatin hydrogels induce NIH-3T3 cell alignment and elongation. Furthermore, human embryonic stem cells cultured on fibronectin-patterned hydrogels display beating foci earlier than those cultured on non-patterned substrates.