iBB

Understanding the mechano–biological coupling mechanisms of biomaterials for tissue engineering is of major importance to assure proper scaffold performance in situ. Therefore, it is of paramount importance to establish correlations between biomaterials, their processing conditions, and their mechanical behaviour, as well as their biological performance. In a collaborative work between CDRSP-Politécnico de Leiria and SCERG-iBB  (João C. Silva and Frederico Ferreira), it was possible to infer a correlation between the addition of different concentrations of graphene nanoparticles (GPN) in three-dimensional poly(ε-caprolactone) (PCL)-based scaffolds, their extrusion-based processing parameters, and the lamellar crystal orientation observed in the different scaffolds through small-angle X-ray scattering experiments. Moreover, in vitro cell culture studies performed at SCERG-iBB demonstrated the suitability and potential of these novel 3D PCL/GPN scaffolds for tissue engineering applications. The results of this study were just published in Polymers.

The research project “InSilico4OCReg - In silico models guiding in vitro biophysical stimulation of biomimetic hierarchical scaffolds: a computational modelling approach towards functional osteochondral regeneration” was recommended for funding by FCT (250,000 euros in the 2021 Call for SR&TD Project Grants). The project started in January 2022 and aims to develop an innovative combined in silico modelling-in vitro experimental system which, inspired by the properties of the native osteochondral tissue, will be able to optimize tissue engineering strategies towards the production of implants with improved functionality and mechanical properties. The multidisciplinary team of InSilico4OCReg includes researchers from iBB, CDRSP-Politécnico de Leiria, Rensselaer Polytechnic Institute (Troy, NY-USA), and an orthopedic surgeon (Hospital dos Lusíadas). The project, which falls within the scientific area of Mechanical Engineering-Engineering Systems, is headed by João Carlos Silva (PI, SCERG-iBB) and Prof. Paula Pascoal-Faria (co-PI, CDRSP-Politécnico de Leiria).

"This week in the Portuguese newspaper "Público" you will find a summary of the Algae2Fish project, where you can read about the reasons behind the project and its perspectives for the next two years. The team, composed by professor Frederico Ferreira, Dr. Paola Sanjuan Alberte, Dr. Carlos Rodrigues, and Msc Diana Marques, with the support of The Good Food Institute, is currently working to produce the first cell-cultured sea bass fillet!"

Engineering brain organoids from human induced pluripotent stem cells (hiPSCs) is a powerful tool for modeling brain development and neurological disorders. Rett syndrome (RTT), a rare neurodevelopmental disorder, can greatly benefit from this technology, since it affects multiple neuronal subtypes in forebrain sub-regions. SCERG-iBB researchers have recently established dorsal and ventral forebrain organoids from control and RTT patient-specific hiPSCs recapitulating the 3D organization and functional network complexity of this brain region. The data obtained revealed a premature development of the deep-cortical layer, associated to the formation of TBR1 and CTIP2 neurons, and a lower expression of neural progenitor/proliferative cells in RTT dorsal organoids. Moreover, calcium imaging and electrophysiology analysis demonstrated functional defects of RTT neurons. Additionally, assembly of RTT dorsal and ventral organoids revealed impairments of interneuron’s migration. Overall, these models provide a better understanding of RTT during early stages of neural development, demonstrating a great potential for personalized diagnosis and drug screening. The paper was published in Frontiers in Cell Development Biology.

The project "DentalBioMatrix: Exploiting the power of decellularized extracellular matrix to fabricate hierarchical biomimetic scaffolds to regenerate functional periodontal tissues" has been recommended for funding by FCT (2020 Call for SR&TD Project Grants). The goal of DentalBioMatrix is to develop novel bioengineering strategies to regenerate periodontium, by engineering hierarchically designed compartmentalized systems to meet the different characteristics of the tissues involved in periodontal defects. Decellularized extracellular matrix (dECM) will be used to create complex dECM-derived scaffolds to recreate the different niches of periodontium. Collectively, this approach will potentially provide the opportunity to harness the properties of periodontal tissue ECM, recreating a local niche at the tooth interface that can promote periodontal regeneration, especially important for osteoporotic and older patients with compromised ECM. The project, which falls within the scientific area of Medical Biotechnology, is headed by Marta Carvalho from SCERG-iBB.

Cell-based therapies have been showing unprecedented therapeutic potential, already changing the landscape of medical care. Extracellular vesicles are nanoparticles naturally secreted by cells, which are important mediators of intercellular communication in our organism and are able to mediate therapeutic effects from their cells of origin. These vesicles can also be used as drug delivery vehicles to target multiple diseases. In a paper recently published in Frontiers in Cell and Developmental Biology, iBB researchers in collaboration with iMM Lisboa, CQE-IST and the companies PBS Biotech and AventaCell Biomedical, developed a new strategy for the robust and scalable production of extracellular vesicles from mesenchymal stromal cells, using bioreactors and an animal serum-free cell culture supplement. This strategy, developed within the frame of the PhD Program in Bioengineering of Miguel de Almeida Fuzeta, is a relevant step towards the large scale production of extracellular vesicles form different human tissue sources, which are promising tools for the development of new therapies against a variety of diseases, from cardiovascular diseases to cancer.

Affinity-triggered hydrogels were created using one of the most selective and high-affinity pairs found in Nature, avidin-biotin. The multimerization of biotin was studied by conjugation into different multi-arm polyethylene glycol molecules. Depending on the multimerization, assemblies with tunable affinity constant were obtained leading to hydrogels with different mechanical properties and controllable erosion time and profiles. The results showed that mimicking natural multivalency gave rise to robust biocompatible hydrogel with applications in tissue engineering and stem cell research. The study results from ongoing collaboration between Cecília Roque (FCT-NOVA) and Tiago Fernandes (DBE-Técnico) and was recently published in the ACS journal Biomacromolecules.

In a new paper published in Frontiers in Bioengineering and Biotechnology, SCERG-iBB researchers in collaboration with colleagues from the Institute of Molecular Medicine (iMM) describe a novel differentiation strategy that uses defined medium to generate Purkinje cells, granule cells, interneurons, and deep cerebellar nuclei projection neurons, that self-formed and matured into electrically active cells. This research is expected to result in better models for the study of cerebellar dysfunctions and represent an important advancement towards the development of autologous replacement strategies for treating cerebellar degenerative diseases.

Mariana Branco, a PhD candidate at the Stem Cell Engineering Research Group of iBB (SCERG-iBB), was awarded with the prize of best oral presentation at the 11th International Meeting of the Portuguese Society for Stem Cells and Cell Therapies (SPCE-TC) that was held during the last 11 – 13th of October at Faculdade de Farmácia of Universidade de Lisboa. The oral communication was entitled “Transcriptomic analysis of 3D cardiac differentiation of human induced pluripotent stem cells reveals faster maturation of cardiomyocytes compared to 2D culture”. Congratulations Mariana!

Bioreactors that provide different biophysical stimuli have been used in tissue engineering approaches aimed at enhancing the quality of the cartilage tissue generated. However, such systems are often highly complex, costly and not very versatile. In a recent study published in Biotechnology Journal, researchers from SCERG-iBB working with colleagues from the Polytechnic Institute of Leiria and Rensselaer Polytechnic Institute (USA) developed a novel, cost-effective and customizable perfusion bioreactor fabricated by additive manufacturing (AM) to study of the effect of fluid flow on the chondrogenic differentiation of human bone-marrow mesenchymal stem/stromal cells (hBMSCs) in 3D porous poly (ε-caprolactone) (PCL) scaffolds. Results suggest that the chondrogenic differentiation of hBMSCs was enhanced in cell-scaffold constructs cultured under perfusion and highlights the potential of customizable AM platforms for developing more reliable in vitro models and improved personalized cartilage repair strategies.

Phenylketonuria (PKU) is a recessive genetic disorder of amino-acid metabolism, where impaired phenylalanine hydroxylase function in the liver of patients leads to the accumulation of neurotoxic phenylalanine levels in the brain. Despite the current knowledge, the chronic effect of PKU in the brain is still poorly understood. In a recent publication in Frontiers in Molecular Neurosciences, DBE faculty and SCERG-iBB researcher Tiago Fernandes, discusses the need for better predictive models, able to recapitulate specific mechanisms of this disease. New exciting in vitro platforms to model specific PKU-derived neuronal impairment are presented in a attempt to understand the impact of phenylalanine in the brain of patients, and ultimately contribute to the understanding of this disease.

The project Algae2Fish led by Frederico Ferreira from SCERG-iBB has received funding from the Global Food Institute with the aim of developing a boneless cultivated seabass fillet. 3D printing technology will be used to produce scaffolds that will give the fillet structure, replicating the fibrous texture of fish. The scaffolds will be formed using material from algae and plants, with the algae contributing valuable omega-3 fatty acids such as those found in conventional fish. Read more about the project here.

Endogenous human brain tissue is not easily available for studying neurodevelopment and neurodegenerative diseases. However, human pluripotent stem cells (PSCs) have been used to generate a variety of glial and neuronal cells of the central nervous system. Still, reproducible protocols for generating in vitro models of the human cerebellum are scarce. In this context, Silva et al. describe the scalable production of human PSC-derived cerebellar organoids using single-use vertical-wheel bioreactors. The transcriptomic profile of cerebellar organoids derived under dynamic conditions demonstrates a faster cerebellar differentiation combined with significant enrichment of extracellular matrix and upregulation of transcripts involved in angiogenesis when compared with the static protocol. The authors anticipate that large-scale production of cerebellar organoids may help developing models for drug screening, toxicological tests and studying pathological pathways involved in cerebellar degeneration.

João Carlos Silva and Frederico Castelo Ferreira from SCERG-iBB are guest-editing a special issue for the open-access journal “Polymers” entitled “Advanced Polymeric Scaffolds for Stem Cell Engineering and Regenerative Medicine”. Polymer scaffolds play a crucial role in tissue engineering and regenerative medicine applications since they can closely mimic the architecture of a native extracellular matrix (ECM) and improve the biological performance of cells both in vitro and in vivo. This Special Issue welcomes full research papers, communications and reviews on recent exciting developments of polymeric scaffolds for tissue engineering and regenerative medicine applications.

The project "UROPRINT: Urinary bladder bioprinting for fully autologous transplantation" has been recommended for funding by the EU. The goal of UROPRINT is to laser print fully functional immunocompatible urothelial tissue ex vivo and in vivo for bladder augmentation and replacement. The project is led by the Institute of Biomedical Research of the Academy of Athens and involves iBB-SCERG, Asphalion SL, Institute of Comunication and Computer Systems, Optics11 BV and METATISSUE/JFCC-Biosolutions, LDA.

Photo details: National Institute of Standards and Technology, Public domain, via Wikimedia Commons.

The project "SMART:New technologies and strategies for Stem cell Manufacturing bioprocess monitoring and ARTificial intelligence-based modeling" has been recommended for funding by FCT (2020 Call for SR&TD Project Grants). The goal of SMART is to develop new monitoring and modeling strategies envisaging more efficient bioreactor-based processes for the manufacturing of human induced Pluripotent Stem Cell-derived cardiomyocytes for biomedical applications, using artificial intelligence to integrate bioprocess monitoring data with biological data. The project, which falls within the scientific area of Chemical Engineering, is headed by Carlos Rodrigues from SCERG-iBB.

Umbilical cord blood (UCB) is an accepted and appealing alternative source of hematopoietic stem/progenitor cells (HSPC) for hematopoietic cell transplants. However, low UCB volume recovered from births results in an unsatisfactory cell dose for transplants in adults, having initially limited transplants of a single UCB unit to pediatric patients. Ex vivo expansion of HSPC based on the addition of exogenous cytokines.has been pursued to address this problem. Notably, selection of individual cytokines and their concentrations for an expansion cocktail has differed between existing strategies. To improve the effectiveness of these platforms, namely targeting clinical approval, iBB researchers optimized the cytokine cocktails in two clinically relevant expansion platforms for HSPC, a liquid suspension culture system (CS_HSPC) and a co-culture system with bone marrow derived mesenchymal stromal cells (CS_HSPC/MSC).. The tailored and novel optimized cocktails determined made it possible to individually maximize cytokine contribution in both studied platforms, leading to an increase in the expansion platform performance, while allowing a rational side-by-side comparison between them and enhancing our knowledge on the impact of cytokine supplementation on the HSPC expansion process. The results achieved were published in Frontiers in Bioengineering and Biotechnology, Stem Cell Systems Bioengineering section.

Cell-based therapies can enhance the specificity of anti-cancer therapeutic agents. In this context, human mesenchymal stromal cells (MSC) hold a promising future as cell delivery systems for anti-cancer proteins due to their innate tropism for tumors. iBB researchers Marília Silva, Gabriel Monteiro, Arsénio Fialho, Nuno Bernardes and Cláudia Lobato da Silva, engineered human MSC through non-viral methods to secrete a human codon-optimized version of azurin (hazu), a bacterial protein with demonstrated anti-cancer activity towards different cancer models in vitro and in vivo. Upon treatment with conditioned media (CM) from these engineered cells, a decrease in cancer cell proliferation, migration and invasion was seen, and an increase in cell death was observed for breast and lung cancer cell models. The results achieved by SCERG- and BSRG-iBB researchers were published in Frontiers in Cell and Developmental Biology, Stem Cell Research section.

Evguenia Bekman and Tiago Fernandes from SCERG-iBB together with Simão Teixeira da Rocha from IMM-FMUL have been awarded by the Amélia de Mello Foundation (FAM) with the Pedro Maria José de Mello Costa Duarte Scholarship to carry out the project “Exploring new therapeutic strategies for Angelman syndrome: A disease modeling approach based on 3D cerebellar organoids derived from patient iPSCs”. The Angelman Syndrome is a genetic-neurological disease discovered in the 60s of the last century by the English pediatrician Harry Angelman, which is estimated to affect 1 in every 15,000 live births and which is characterized by developmental delay, difficulty in speech, sleep disorders, convulsions, disconnected movements and a frequent smile.

Tiago G. Fernandes from SCERG-iBB is guest-editing a special topic for the open-access journal "Frontiers in Bioengineering and Biotechnology" entitled "Stem Cell Systems Bioengineering". Advances in the bioengineering field have allowed the manipulation of singular aspects of the cellular microenvironment, and this Research Topic aims to focus on outstanding examples of bioengineering approaches used to promote the self-organization of human cells and the production of tissue-like structure formation.