iBB

Our researchers were involved in many different ways at this year's edition of FIC.A – Festival internacional de Ciência 2022. Vasco Bonifácio was involved in the activities "The Perfume Laboratory", "The Air We Live In", "Black Ink Secrets", and also assisted the "Virtual Tour To The International Space Station (ISS)" using VR glasses. Tiago Fernandes talked to young audiences about "Challenges and opportunities in Regenerative Medicine and Stem Cells". Our student Diana Marques talked to kids about "The Future of food and 3D printing", an activity developed under the umbrella of the initiative "Soapbox Science Lisbon" that promotes the visibility of women in Science. Finally, we were also represented in the Técnico stand with the hands-on activity "Bioreactors for Cell Expansion".

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.

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.

Human morphogenesis is a complex process involving distinct microenvironmental and physical signals that are manipulated in space and time to give rise to complex tissues and organs. The development of organoids represents a novel way to modeling such complexity. Advances in the bioengineering field have allowed the manipulation of different components, including cellular and noncellular factors, to better mimic the natural microenvironment and generate better organoid models of human morphogenesis. In a paper published in Stem Cells International, a team of researchers from the Stem Cell Engineering Research Group (SCERG) at iBB in collaboration with the Institute of Molecular Medicine reviewed the bioengineering strategies used to control the initial state and spatiotemporal positioning of cells within organoids and, lastly, the growth and remodeling of multicellular aggregates to achieve mini organ-like structures.

Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) have an enormous potential for in vitro modeling of cardiac diseases and for testing the effect and toxicity of new drugs. However, when compared to adult CMs, hiPSC-CMs are very immature, exhibiting low structural development and functionality. SCERG-iBB researchers developed a novel methodological framework to quantify structural aspects of hiPSC-CMs during long-term culture. hiPSC-CMs showed significant progression in several structural characteristics namely cardiomyocyte fiber density and length. Importantly, this methodology contributes to set new metrics to develop applications for drug screening and disease modeling for hiPSC-CMs. The research has been published on Biochemical and Biophysical Research Communications.

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.

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.

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.

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.

The ability to differentiate neural progenitors (NP) from human induced pluripotent stem cells (hiPSCs) provides an opportunity to develop new applications for cellular therapy, disease modelling and drug screening. SCERG-iBB researchers developed a platform that can be applied towards the study of the effect of neurotoxic molecules that impair normal embryonic development, such as the antiepileptic drug valproic acid (VPA). It was verified that exposure to VPA led to a prevalence of NP structures over neuronal differentiation, confirmed by analysis of the expression of neural cell adhesion molecule, and neural rosette number and morphology. This methodology can potentially complement current toxicity tests for the detection of teratogenic compounds that can interfere with normal embryonic development. The work was published in Toxicology Letters.