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Today, it is recognized that medicines will eventually be needed during pregnancy, either due to gestation-related medical conditions or pre-existing diseases. Also, the rate of drug prescription to pregnant women has increased over the past few years, in accordance with the increasing trend to postpone childbirth to a later age. However, information regarding teratogenic risk in humans is often missing for most of the purchased drugs. Inter-species differences have limited the suitability of animal models to predict human-specific outcomes, contributing to misidentified human teratogenicity. Therefore, the development of physiologically relevant in vitro humanized models can be the key to surpassing this limitation. In this context, this review describes the pathway towards the introduction of human pluripotent stem cell-derived models in developmental toxicity studies.

The embryonic development of the human heart under healthy and pathological conditions is still very poorly understood. In a recently published Nature Communications paper, a iBB team led by Margarida Diogo, in collaboration with Perpetua Pinto do Ó at i3S, engineered a unique organ-like model from stem cells that recreates several stages of embryonic development of the human heart. This humanized model opens now the path to study, in a tissue engineering lab, the development of congenital heart diseases, such as coronary vascular diseases and myocardium non-compaction cardiomyopathies, and can be further applied in drug screening and toxicology assays during embryonic heart development. This last topic is particularly relevant since pregnant women normally do not take part in clinical trials, making this human cell-based in vitro platform the only viable alternative to assess developmental cardiotoxicity. The work was a part of the PhD thesis in Bioengineering – MIT Portugal program of Mariana Branco, the first author of the paper, and was funded by the FCT project “Mini-Hearts - Organoid Engineering for Production of 3D Cardiovascular Microtissues from Human Induced Pluripotent Stem Cells for Cardiotoxicity”.  

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 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.

Human adult stem cells and patient-derived induced pluripotent stem cells represent promising tools to understand human biology, development, and disease. Under a permissive environment, stem cell derivatives can self-organize and reconstruct their native environment, resulting in the creation of organ-like entities known as organoids. Although organoids represent a breakthrough in the stem cell field, there are still considerable shortcomings preventing their widespread use, namely their variability, limited function, and reductionist size. In this recently published review article in Trends in Biotechnology, Miguel Tenreiro (Ph.D. student in Bioengineering), Prof. Margarida Diogo, and co-authors from iBB highlight emerging technologies to fill in the gaps in organoid design and provide insights on how they can be best utilized to fulfill the potential of organoids.

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.

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.