Papers by Margarida Serra
The increasing usage of composite materials in the aerospace industry and the associated need for... more The increasing usage of composite materials in the aerospace industry and the associated need for more accurate estimations of structural response motivated the development of a user-defined element (UEL) for static and free vibration analysis of composite laminated plates. The user element developed is based in an equivalent single-layer (ESL) theory, specifically in the third-order shear deformation theory (TSDT) of Reddy. Both C and C continuity interpolation functions are used. The implementation of the UEL is done in Abaqus, a commercial finite element analysis (FEA) software, which provides standard procedures for users to define their own elements and make use of its solver and interface to easily visualize and interpret solutions. As a preliminary phase, results for an isotropic plate are presented and validated through comparison with solutions obtained using a finite element available in Abaqus library. Then, numerical applications of composite laminates are shown and comp...
Margarida Serra, ITQB-NOVA, Portugal [email protected] Bárbara Cunha , iBET/ITQB-NOVA, Portugal Tiag... more Margarida Serra, ITQB-NOVA, Portugal [email protected] Bárbara Cunha , iBET/ITQB-NOVA, Portugal Tiago Aguiar, iBET/ITQB-NOVA, Portugal Sofia B. Carvalho, iBET/ITQB-NOVA, Portugal Marta M. Silva, iBET/ITQB-NOVA, Portugal Ricardo A. Gomes, iBET/ITQB-NOVA, Portugal Manuel J. T. Carrondo, iBET/ITQB-NOVA, Portugal Patrícia Gomes-Alves, iBET/ITQB-NOVA, Portugal Cristina Peixoto, iBET/ITQB-NOVA, Portugal Paula M. Alves, iBET/ITQB-NOVA, Portugal
Maria João Sebastião, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Q... more Maria João Sebastião, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Química e Biológica António Xavier, UNL, Oeiras, Portugal [email protected] Ivo Reis, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Química e Biológica António Xavier, UNL, Oeiras, Portugal Itziar Palacios, Coretherapix, S.L.U. (Tigenix Group), Tres Cantos, Spain. Margarida Serra, iBET, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Química e Biológica António Xavier, UNL, Oeiras, Portugal Patrícia Gomes-Alves, iBET, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Química e Biológica António Xavier, UNL, Oeiras, Portugal Paula Marques Alves, iBET, Instituto de Biologia Experimental e Tecnológica, Instituto de Tecnologia Química e Biológica António Xavier, UNL, Oeiras, Portugal
Difficulties in maturing human induced pluripotent stem cells derived cardiomyocytes (hiPSC-CMs) ... more Difficulties in maturing human induced pluripotent stem cells derived cardiomyocytes (hiPSC-CMs) to an adult-like state is still an obstacle hindering their clinical use. Cardiac tissues grown from hiPSC-CMs are still subjected to a microenvironment distinct from physiological conditions. In Nature, the extracellular matrix (ECM) serves as a template to build tissues, being a key regulator in tissue development, homeostasis and injury. Exposing hiPSC-CMs to an ECM-mimicking microenvironment may guide their maturation. In this work, human ECM was extracted from cardiac tissue through physical and chemical-based decellularization approaches. Tissue milling and freeze-drying resulted in tissue cellularity reduction and maintenance of ECM composition, even though tissue architecture is not preserved. By using detergents, consistent reductions in cellularity amongst different donors was only achieved with exposure times comparable with organ perfusion decellularization. ECM composition a...
Cardiovascular Research
CardioVascular Research Initiative, ERC 3D-JOINT, ERC Design-2Heal Background: Classical cell the... more CardioVascular Research Initiative, ERC 3D-JOINT, ERC Design-2Heal Background: Classical cell therapy approaches for the injured heart still have therapeutic promise, but are limited by poor cell retention. Engineering native myocardial tissue, thereby recapitulating the structural organization, has therapeutic potential but is still a challenge. Recently, we developed a microfiber scaffold of a biodegradable polymer in combination with cardiac progenitor cells by using a novel technique, called Melt Electrowriting (MEW). Although successful, the limited dynamic mechanical properties of these scaffolds do not allow for deformation under heart contraction and consequently hinders functional integration. Purpose: We hypothesize that the fabrication of fiber scaffolds with non-linear hexagonal geometries will allow for variable mechanical properties. Moreover, combining these novel fiber scaffolds with human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) will enhance iPSC-CM function, thereby creating a compatible and clinically-relevant heart patch for cardiac regeneration. Methods: Using a custom-made MEW device, poly (e-caprolactone) (PCL) based scaffolds were fabricated with an internal hexagonal architecture. Scaffolds were manufactured with equilateral hexagons with different side lengths and then mechanically characterized under quasi static and dynamic loading conditions. Subsequently, scaffolds were seeded with iPSC-CMs encapsulated in a cardiac ECM-like hydrogel to create a functional cardiac patch. iPSC-CM viability, alignment, maturation and function were determined after 7 and 14 days in culture. To show minimally invasive application of the hexagonal scaffolds, injectability and shape-recovery was assessed in a large-animal (porcine) model. Results: Scaffolds with hexagonal geometries were successfully manufactured for the first time using MEW. The smallest hexagon pore resolution achievable was 400 mm (for microfibers ؼ20-25 mm and scaffold thickness ¼ 300 mm). The hexagonal architecture allowed for better deformation recoverability (50% strain), compared to the linear architecture (5%). Additionally, hexagonal scaffolds showed improved recovery under dynamic loading conditions. The hexagonal cardiac patches showed increased beating rate (1.5 fold), enhanced cell alignment, sarcomere content, and Cx43 organization, and an increase in cardiac marker expression (cardiac Actin 1.5 fold, SERCA2a 2.1 fold, and PPARGC1a 3.1 fold), indicative of enhanced iPSC-CM maturation. Importantly, we showed successful minimally invasive epicardial delivery of the hexagonal patches on a porcine heart. Conclusion: The creation of MEW scaffolds with stretchable geometries composed of microfibers allows for the generation of in vitro cardiac constructs that better mimic the mechanical environment of endogenous cardiac tissue. By combining these flexible fiber scaffolds with clinically relevant human iPSC-CMs, we have produced a heart patch with superior features for cardiac tissue engineering.
Margarida Serra, iBET/ITQB-NOVA, Portugal [email protected] Pedro Vicente, iBET/ITQB-NOVA, Oeiras, P... more Margarida Serra, iBET/ITQB-NOVA, Portugal [email protected] Pedro Vicente, iBET/ITQB-NOVA, Oeiras, Portugal Catarina Gomes, iBET/ITQB-NOVA, Oeiras, Portugal Joana Almeida, iBET/ITQB-NOVA, Oeiras, Portugal Daniel AM Pais, iBET/ITQB-NOVA, Oeiras, Portugal Bernardo Abecasis, iBET/ITQB-NOVA, Oeiras, Portugal Marta Paiva, iBET/ITQB-NOVA, Oeiras, Portugal Inês A Isidro, iBET/ITQB-NOVA, Oeiras, Portugal Paula M Alves, iBET/ITQB-NOVA, Oeiras, Portugal Rebeca Mertinez-Turrillas, CIMA/UNAV, Pamplona, Spain Juan R. Rodriguez-Madoz, CIMA/UNAV, Pamplona, Spain Anders Aspegren, Takara Bio Europe AB, Gothenburg, Sweden Ibrahim Domian, Harvard Stem Cell Institute/Harvard Medical School, Cambridge, USA
Martina Micheletti, Department of Biochemical Engineering, UCL [email protected] Jasmin Sama... more Martina Micheletti, Department of Biochemical Engineering, UCL [email protected] Jasmin Samaras, Department of Biochemical Engineering, UCL Andrea Ducci, Department of Mechanical Engineering, UCL Claudia Correia, Instituto de Biologia Experimental e Tecnologica (iBET), Oeiras, Portugal Paula Alves, Instituto de Biologia Experimental e Tecnologica (iBET), Oeiras, Portugal Margarida Serra, Instituto de Biologia Experimental e Tecnologica (iBET), Oeiras, Portugal
Margarida Serra, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universida... more Margarida Serra, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa ; iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal [email protected] Cláudia Correia, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa ; iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal Alexey Koshkin, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa ; iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal Patrícia Duarte, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa ; iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal Ana Teixeira, ITQB-UNL, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa ; iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal Paula M Alves, ITQB-UNL, Instituto d...
Advanced materials, 2021
Worldwide, over 26 million patients suffer from heart failure (HF). One strategy aspiring to prev... more Worldwide, over 26 million patients suffer from heart failure (HF). One strategy aspiring to prevent or even to reverse HF is based on the transplantation of cardiac tissue-engineered (cTE) constructs. These patient-specific constructs aim to closely resemble the native myocardium and, upon implantation on the diseased tissue, support and restore cardiac function, thereby preventing the development of HF. However, cTE constructs off-the-shelf availability in the clinical arena critically depends on the development of efficient preservation methodologies. Short- and long-term preservation of cTE constructs would enable transportation and direct availability. Herein, currently available methods, from normothermic- to hypothermic- to cryopreservation, for the preservation of cardiomyocytes, whole-heart, and regenerative materials are reviewed. A theoretical foundation and recommendations for future research on developing cTE construct specific preservation methods are provided. Current...
Human embryonic stem cells (hESCs) have an enormous potential as a source for cell replacement th... more Human embryonic stem cells (hESCs) have an enormous potential as a source for cell replacement therapies, tissue engineering, and in vitro toxicology applications. The lack of standardized and robust bioprocesses for hESC expansion has hindered the application of hESCs and their derivatives in clinical settings. We developed a robust and well-characterized bioprocess for hESC expansion under fully defined conditions and explored the potential of transcriptomic and metabolomic tools for a more comprehensive assessment of culture system impact on cell proliferation, metabolism, and phenotype. Two different hESC lines (feeder-dependent and feeder-free lines) were efficiently expanded on xeno-free microcarriers in stirred culture systems. Both hESC lines maintained the expression of stemness markers such as Oct-4, Nanog, SSEA-4, and TRA1-60 and the ability to spontaneously differentiate into the three germ layers. Whole-genome transcriptome profiling revealed a phenotypic convergencebet...
We relied on the aggregation of hPSC-derived cardiac progenitors to establish a scalable differen... more We relied on the aggregation of hPSC-derived cardiac progenitors to establish a scalable differentiation protocol capable of generating highly pure CM aggregate cultures. Whole-transcriptome analysis and 13C-metabolic flux analysis demonstrate at both molecular and fluxome levels that a 3D culture environment enhances metabolic maturation of hPSC-CMs. When compared to 2D, 3D cultures of hPSC-CMs displayed down-regulation of genes involved in glycolysis and lipid biosynthesis and increased expression of genes involved in OXPHOS. Accordingly, 3D hPSC-CMs had lower fluxes through glycolysis and fatty acid synthesis and increased TCAcycle activity.
Current Opinion in Biotechnology
Stem cells hold outstanding potential to model and treat disease and are valuable tools in pharma... more Stem cells hold outstanding potential to model and treat disease and are valuable tools in pharmacology and toxicology. Characterization of stem cells and derivatives still poses many challenges to ensure safe, efficacious, and reliable therapies. Regulatory agencies have defined key mandatory attributes related to identity, purity, sterility, and genomic integrity, however robust analytics to determine cell's potency are still a major challenge, in most cases assessed case-by-case. Importantly, the application of high-throughput 'omic tools is opening new perspectives on stem cell's research and development. Here, analytical methodologies currently employed to characterize stem cells' quality attributes are discussed, with special focus on 'omics as relevant tools for definition of cell's mechanism of action, and for potency assay development and assessment.
Scientific Reports
The extracellular matrix (ECM) of engineered human cardiac tissues corresponds to simplistic biom... more The extracellular matrix (ECM) of engineered human cardiac tissues corresponds to simplistic biomaterials that allow tissue assembly, or animal derived off-the-shelf non-cardiac specific matrices. Decellularized ECM from human cardiac tissue could provide a means to improve the mimicry of engineered human cardiac tissues. Decellularization of cardiac tissue samples using immersion-based methods can produce acceptable cardiac ECM scaffolds; however, these protocols are mostly described for animal tissue preparations. We have tested four methods to decellularize human cardiac tissue and evaluated their efficiency in terms of cell removal and preservation of key ECM components, such as collagens and sulfated glycosaminoglycans. Extended exposure to decellularization agents, namely sodium dodecyl sulfate and Triton-X-100, was needed to significantly remove DNA content by approximately 93% in all human donors. However, the biochemical composition of decellularized tissue is affected, and...
Antioxidants
Bravo de Esmolfe (BE) is a traditional Portuguese apple highly appreciated by consumers due to it... more Bravo de Esmolfe (BE) is a traditional Portuguese apple highly appreciated by consumers due to its peculiar flavor and aroma. This apple contains higher concentration of phenolic compounds than other cultivars and is thus considered a rich source of antioxidants. Its sensorial and functional properties have attracted farmers’ associations to increase BE production. However, a large quantity of apples is wasted due to storage/transportation procedures that impact BE’s quality attributes. In this work, we applied high-pressure extraction methodologies to generate antioxidant-rich fractions from BE residues aiming at adding high value to these agro-food by-products. We performed a first extraction step using supercritical CO2, followed by a second extraction step where different CO2 + ethanol mixtures (10–100% v/v) were tested. All experiments were carried out at 25 MPa and 50 °C. Extracts were characterized in terms of global yield, phenolic content and antioxidant activity using chem...
npj Regenerative Medicine
The adult heart is a vital and highly specialized organ of the human body, with limited capabilit... more The adult heart is a vital and highly specialized organ of the human body, with limited capability of self-repair and regeneration in case of injury or disease. Engineering biomimetic cardiac tissue to regenerate the heart has been an ambition in the field of tissue engineering, tracing back to the 1990s. Increased understanding of human stem cell biology and advances in process engineering have provided an unlimited source of cells, particularly cardiomyocytes, for the development of functional cardiac muscle, even though pluripotent stem cell-derived cardiomyocytes poorly resemble those of the adult heart. This review outlines key biology-inspired strategies reported to improve cardiomyocyte maturation features and current biofabrication approaches developed to engineer clinically relevant cardiac tissues. It also highlights the potential use of this technology in drug discovery science and disease modeling as well as the current efforts to translate it into effective therapies th...
Biotechnology and Bioengineering
Hepatocyte‐like cells derived from human‐induced pluripotent stem cells (hiPSC‐HLC) are expected ... more Hepatocyte‐like cells derived from human‐induced pluripotent stem cells (hiPSC‐HLC) are expected to have important applications in drug screening and regenerative medicine. However, hiPSC‐HLC are difficult to produce on a large‐scale to obtain relevant numbers for such applications. The aim of this study was to implement a novel integrated strategy for scalable production of hiPSC‐HLC and demonstrate the applicability of dielectric spectroscopy to monitor hiPSC expansion/differentiation processes. We cultured hiPSC as three‐dimensional (3D) aggregates in stirred‐tank bioreactors (STB) operated in perfusion with an in situ capacitance probe. Dissolved oxygen concentration and dilution rate were controlled along the process and after 5 days of cell expansion, the hepatic differentiation was integrated in sequential steps for 28 days. The hiPSC were able to grow as 3D aggregates and the expression of hepatic markers and albumin production after differentiation confirmed that hepatocyte differentiation improved when compared to 2D culture. These hiPSC‐HLC exhibited functional characteristics of hepatocytes including glycogen storage and drug metabolization capacity. Our results also show a good correlation between the cell permittivity measured online and the aggregate biovolume measured by standard offline methods, demonstrating for the first time the potential of dielectric spectroscopy to monitor hiPSC expansion and differentiation in STB.
ACS Applied Bio Materials
Human induced pluripotent stem cells (hiPSC) possess significant therapeutic potential due to the... more Human induced pluripotent stem cells (hiPSC) possess significant therapeutic potential due to their high self-renewal capability and potential to differentiate into specialized cells such as cardiomyocytes. However, generated hiPSCderived cardiomyocytes (hiPSC-CM) are still immature, with phenotypic and functional features resembling the fetal rather than their adult counterparts, which limits their application in cell-based therapies, in vitro cardiac disease modeling, and drug cardiotoxicity screening. Recent discoveries have demonstrated the potential of the extracellular matrix (ECM) as a critical regulator in development, homeostasis, and injury of the cardiac microenvironment. Within this context, this work aimed to assess the impact of human cardiac ECM in the phenotype and maturation features of hiPSC-CM. Human ECM was isolated from myocardium tissue through a physical decellularization approach. The cardiac tissue decellularization process reduced DNA content significantly while maintaining ECM composition in terms of sulfated glycosaminoglycans (s-GAG) and collagen content. These ECM particles were successfully incorporated in three-dimensional (3D) hiPSC-CM aggregates (CM+ECM) with no impact on viability and metabolic activity throughout 20 days in 3D culture conditions. Also, CM+ECM aggregates displayed organized and longer sarcomeres, with improved calcium handling when compared to hiPSC-CM aggregates. This study shows that human cardiac ECM functionalization of hiPSC-based cardiac tissues improves cardiomyocyte maturation. The knowledge generated herein provides essential insights to streamline the application of ECM in the development of hiPSC-based therapies targeting cardiac diseases.
Cardiovascular Research, Apr 1, 2018
CardioVascular Research Initiative, ERC 3D-JOINT, ERC Design-2Heal Background: Classical cell the... more CardioVascular Research Initiative, ERC 3D-JOINT, ERC Design-2Heal Background: Classical cell therapy approaches for the injured heart still have therapeutic promise, but are limited by poor cell retention. Engineering native myocardial tissue, thereby recapitulating the structural organization, has therapeutic potential but is still a challenge. Recently, we developed a microfiber scaffold of a biodegradable polymer in combination with cardiac progenitor cells by using a novel technique, called Melt Electrowriting (MEW). Although successful, the limited dynamic mechanical properties of these scaffolds do not allow for deformation under heart contraction and consequently hinders functional integration. Purpose: We hypothesize that the fabrication of fiber scaffolds with non-linear hexagonal geometries will allow for variable mechanical properties. Moreover, combining these novel fiber scaffolds with human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) will enhance iPSC-CM function, thereby creating a compatible and clinically-relevant heart patch for cardiac regeneration. Methods: Using a custom-made MEW device, poly (e-caprolactone) (PCL) based scaffolds were fabricated with an internal hexagonal architecture. Scaffolds were manufactured with equilateral hexagons with different side lengths and then mechanically characterized under quasi static and dynamic loading conditions. Subsequently, scaffolds were seeded with iPSC-CMs encapsulated in a cardiac ECM-like hydrogel to create a functional cardiac patch. iPSC-CM viability, alignment, maturation and function were determined after 7 and 14 days in culture. To show minimally invasive application of the hexagonal scaffolds, injectability and shape-recovery was assessed in a large-animal (porcine) model. Results: Scaffolds with hexagonal geometries were successfully manufactured for the first time using MEW. The smallest hexagon pore resolution achievable was 400 mm (for microfibers ؼ20-25 mm and scaffold thickness ¼ 300 mm). The hexagonal architecture allowed for better deformation recoverability (50% strain), compared to the linear architecture (5%). Additionally, hexagonal scaffolds showed improved recovery under dynamic loading conditions. The hexagonal cardiac patches showed increased beating rate (1.5 fold), enhanced cell alignment, sarcomere content, and Cx43 organization, and an increase in cardiac marker expression (cardiac Actin 1.5 fold, SERCA2a 2.1 fold, and PPARGC1a 3.1 fold), indicative of enhanced iPSC-CM maturation. Importantly, we showed successful minimally invasive epicardial delivery of the hexagonal patches on a porcine heart. Conclusion: The creation of MEW scaffolds with stretchable geometries composed of microfibers allows for the generation of in vitro cardiac constructs that better mimic the mechanical environment of endogenous cardiac tissue. By combining these flexible fiber scaffolds with clinically relevant human iPSC-CMs, we have produced a heart patch with superior features for cardiac tissue engineering.
Uploads
Papers by Margarida Serra