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1
Logan, Sarah, Thiago Arzua, Yasheng Yan, Congshan Jiang, Xiaojie Liu, Lai-Kang Yu, Qing-Song Liu, and Xiaowen Bai. "Dynamic Characterization of Structural, Molecular, and Electrophysiological Phenotypes of Human-Induced Pluripotent Stem Cell-Derived Cerebral Organoids, and Comparison with Fetal and Adult Gene Profiles." Cells 9, no. 5 (May 23, 2020): 1301. http://dx.doi.org/10.3390/cells9051301.
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Background: The development of 3D cerebral organoid technology using human-induced pluripotent stem cells (iPSCs) provides a promising platform to study how brain diseases are appropriately modeled and treated. So far, understanding of the characteristics of organoids is still in its infancy. The current study profiled, for the first time, the electrophysiological properties of organoids at molecular and cellular levels and dissected the potential age equivalency of 2-month-old organoids to human ones by a comparison of gene expression profiles among cerebral organoids, human fetal and adult brains. Results: Cerebral organoids exhibit heterogeneous gene and protein markers of various brain cells, such as neurons, astrocytes, and vascular cells (endothelial cells and smooth muscle cells) at 2 months, and increases in neural, glial, vascular, and channel-related gene expression over a 2-month differentiation course. Two-month organoids exhibited action potentials, multiple channel activities, and functional electrophysiological responses to the anesthetic agent propofol. A bioinformatics analysis of 20,723 gene expression profiles showed the similar distance of gene profiles in cerebral organoids to fetal and adult brain tissues. The subsequent Ingenuity Pathway Analysis (IPA) of select canonical pathways related to neural development, network formation, and electrophysiological signaling, revealed that only calcium signaling, cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) signaling in neurons, glutamate receptor signaling, and synaptogenesis signaling were predicted to be downregulated in cerebral organoids relative to fetal samples. Nearly all cerebral organoid and fetal pathway phenotypes were predicted to be downregulated compared with adult tissue. Conclusions: This novel study highlights dynamic development, cellular heterogeneity and electrophysiological activity. In particular, for the first time, electrophysiological drug response recapitulates what occurs in vivo, and neural characteristics are predicted to be highly similar to the human brain, further supporting the promising application of the cerebral organoid system for the modeling of the human brain in health and disease. Additionally, the studies from these characterizations of cerebral organoids in multiple levels and the findings from gene comparisons between cerebral organoids and humans (fetuses and adults) help us better understand this cerebral organoid-based cutting-edge platform and its wide uses in modeling human brain in terms of health and disease, development, and testing drug efficacy and toxicity.
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Peng, Xiyao, Lei Wu, Qiushi Li, Yuqing Ge, Tiegang Xu, and Jianlong Zhao. "An Easy-to-Use Arrayed Brain–Heart Chip." Biosensors 14, no. 11 (October 22, 2024): 517. http://dx.doi.org/10.3390/bios14110517.
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Multi-organ chips are effective at emulating human tissue and organ functions and at replicating the interactions among tissues and organs. An arrayed brain–heart chip was introduced whose configuration comprises open culture chambers and closed biomimetic vascular channels distributed in a horizontal pattern, separated from each other by an endothelial barrier based on fibrin matrix. A 300 μm-high and 13.2 mm-long endothelial barrier surrounded each organoid culture chamber, thereby satisfying the material transport requirements. Numerical simulations were used to analyze the construction process of fibrin barriers in order to optimize the structural design and experimental manipulation, which exhibited a high degree of correlation with experiment results. In each interconnective unit, a cerebral organoid, a cardiac organoid, and endothelial cells were co-cultured stably for a minimum of one week. The permeability of the endothelial barrier and recirculating perfusion enabled cross talk between cerebral organoids and cardiac organoids, as well as between organoids and endothelial cells. This was corroborated by the presence of cardiac troponin I (cTnI) in the cerebral organoid culture chamber and the observation of cerebral organoid and endothelial cells invading the fibrin matrix after one week of co-culture. The arrayed chip was simple to manipulate, clearly visible under a microscope, and compatible with automated pipetting devices, and therefore had significant potential for application.
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Simsa, Robin, Theresa Rothenbücher, Hakan Gürbüz, Nidal Ghosheh, Jenny Emneus, Lachmi Jenndahl, David L. Kaplan, Niklas Bergh, Alberto Martinez Serrano, and Per Fogelstrand. "Brain organoid formation on decellularized porcine brain ECM hydrogels." PLOS ONE 16, no. 1 (January 28, 2021): e0245685. http://dx.doi.org/10.1371/journal.pone.0245685.
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Human brain tissue models such as cerebral organoids are essential tools for developmental and biomedical research. Current methods to generate cerebral organoids often utilize Matrigel as an external scaffold to provide structure and biologically relevant signals. Matrigel however is a nonspecific hydrogel of mouse tumor origin and does not represent the complexity of the brain protein environment. In this study, we investigated the application of a decellularized adult porcine brain extracellular matrix (B-ECM) which could be processed into a hydrogel (B-ECM hydrogel) to be used as a scaffold for human embryonic stem cell (hESC)-derived brain organoids. We decellularized pig brains with a novel detergent- and enzyme-based method and analyzed the biomaterial properties, including protein composition and content, DNA content, mechanical characteristics, surface structure, and antigen presence. Then, we compared the growth of human brain organoid models with the B-ECM hydrogel or Matrigel controls in vitro. We found that the native brain source material was successfully decellularized with little remaining DNA content, while Mass Spectrometry (MS) showed the loss of several brain-specific proteins, while mainly different collagen types remained in the B-ECM. Rheological results revealed stable hydrogel formation, starting from B-ECM hydrogel concentrations of 5 mg/mL. hESCs cultured in B-ECM hydrogels showed gene expression and differentiation outcomes similar to those grown in Matrigel. These results indicate that B-ECM hydrogels can be used as an alternative scaffold for human cerebral organoid formation, and may be further optimized for improved organoid growth by further improving protein retention other than collagen after decellularization.
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Bao, Zhongyuan, Kaiheng Fang, Zong Miao, Chong Li, Chaojuan Yang, Qiang Yu, Chen Zhang, Zengli Miao, Yan Liu, and Jing Ji. "Human Cerebral Organoid Implantation Alleviated the Neurological Deficits of Traumatic Brain Injury in Mice." Oxidative Medicine and Cellular Longevity 2021 (November 22, 2021): 1–16. http://dx.doi.org/10.1155/2021/6338722.
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Traumatic brain injury (TBI) causes a high rate of mortality and disability, and its treatment is still limited. Loss of neurons in damaged area is hardly rescued by relative molecular therapies. Based on its disease characteristics, we transplanted human embryonic stem cell- (hESC-) derived cerebral organoids in the brain lesions of controlled cortical impact- (CCI-) modeled severe combined immunodeficient (SCID) mice. Grafted organoids survived and differentiated in CCI-induced lesion pools in mouse cortical tissue. Implanted cerebral organoids differentiated into various types of neuronal cells, extended long projections, and showed spontaneous action, as indicated by electromyographic activity in the grafts. Induced vascularization and reduced glial scar were also found after organoid implantation, suggesting grafting could improve local situation and promote neural repair. More importantly, the CCI mice’s spatial learning and memory improved after organoid grafting. These findings suggest that cerebral organoid implanted in lesion sites differentiates into cortical neurons, forms long projections, and reverses deficits in spatial learning and memory, a potential therapeutic avenue for TBI.
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Harary, Paul M., Rachel Blue, Mackenzie Castellanos, Mehek Dedhia, Sarah Hamimi, Dennis Jgamadze, Benjamin Rees, et al. "Human brain organoid transplantation: ethical implications of enhancing specific cerebral functions in small-animal models." Molecular Psychology: Brain, Behavior, and Society 2 (June 6, 2023): 14. http://dx.doi.org/10.12688/molpsychol.17544.1.
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Brain organoids are self-organizing, three-dimensional tissues derived from pluripotent stem cells that recapitulate many aspects of the cellular diversity and architectural features of the developing brain. Recently, there has been growing interest in using human brain organoid transplantation in animal models as a means of addressing the limitations of in vitro culture, such as the lack of vascularization, and to explore the potential of organoids for neural repair. While there has been substantial debate on the ethical implications of brain organoid research, particularly the potential for organoids to exhibit higher-order brain functions such as consciousness, the impact of human organoid grafts on animal hosts has been less extensively discussed. Enhancement of host animal brain function may not be technically feasible at this time, but it is imperative to carefully consider the moral significance of these potential outcomes. Here, we discuss the ethical implications of enhancing somatosensation, motor processes, memory, and basic socialization in small-animal models. We consider the moral implications of such outcomes and if safeguards are needed to accommodate any increased moral status of animals transplanted with human brain organoids.
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Estridge, R. Chris, Jennifer E. O’Neill, and Albert J. Keung. "Matrigel Tunes H9 Stem Cell-Derived Human Cerebral Organoid Development." Organoids 2, no. 4 (October 5, 2023): 165–76. http://dx.doi.org/10.3390/organoids2040013.
Full textAbstract:
Human cerebral organoids are readily generated from human embryonic stem cells and human induced pluripotent stem cells and are useful in studying human neurodevelopment. Recent work with human cerebral organoids have explored the creation of different brain regions and the impacts of soluble and mechanical cues. Matrigel is a gelatinous, heterogenous mixture of extracellular matrix proteins, morphogens, and growth factors secreted by Engelbreth-Holm-Swarm mouse sarcoma cells. It is a core component of almost all cerebral organoid protocols, generally supporting neuroepithelial budding and tissue polarization; yet, its roles and effects beyond its general requirement in organoid protocols are not well understood, and its mode of delivery is variable, including the embedding of organoids within it or its delivery in soluble form. Given its widespread usage, we asked how H9 stem cell-derived hCO development and composition are affected by Matrigel dosage and delivery method. We found Matrigel exposure influences organoid size, morphology, and cell type composition. We also showed that greater amounts of Matrigel promote an increase in the number of choroid plexus (ChP) cells, and this increase is regulated by the BMP4 pathway. These results illuminate the effects of Matrigel on human cerebral organoid development and the importance of delivery mode and amount on organoid phenotype and composition.
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Shnaider, T. A. "Cerebral organoids: a promising model in cellular technologies." Vavilov Journal of Genetics and Breeding 22, no. 2 (April 8, 2018): 168–78. http://dx.doi.org/10.18699/vj18.344.
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The development of the human brain is a complex multi-stage process including the formation of various types of neural cells and their interactions. Many fundamental mechanisms of neurogenesis have been established due to the studying of model animals. However, significant differences in the brain structure compared to other animals do not allow considering all aspects of the human brain formation, which could play the main role in the development of unique cognitive abilities for human. Four years ago, Lancaster’s group elaborated human pluripotent stem cell-derived three-dimensional cerebral organoid technology, which opened a unique opportunity for researchers to model early stages of human neurogenesis in vitro. Cerebral organoids closely remodel many endogenous brain regions with specific cell composition like ventricular zone with radial glia, choroid plexus, and cortical plate with upper and deeper-layer neurons. Moreover, human brain development includes interactions between different brain regions. Generation of hybrid three-dimensional cerebral organoids with different brain region identity allows remodeling some of them, including long-distance neuronal migration or formation of major axonal tracts. In this review, we consider the technology of obtaining human pluripotent stem cell-derived three-dimensional cerebral organoids with different modifications and with different brain region identity. In addition, we discuss successful implementation of this technology in fundamental and applied research like modeling of different neurodevelopmental disorders and drug screening. Finally, we regard existing problems and prospects for development of human pluripotent stem cell-derived threedimensional cerebral organoid technology.
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Roosen, Mieke, Chris Meulenbroeks, Phylicia Stathi, Joris Maas, Julie Morscio, Jens Bunt, and Marcel Kool. "BIOL-11. PRECLINICAL MODELLING OF PEDIATRIC BRAIN TUMORS USING ORGANOID TECHNOLOGY." Neuro-Oncology 25, Supplement_1 (June 1, 2023): i8. http://dx.doi.org/10.1093/neuonc/noad073.030.
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Abstract Molecular characterization has resulted in improved classification of pediatric brain tumors, leading to many novel (sub)types with distinct oncodriving events. To study tumor biology and to perform translational research on each of these tumors, preclinical models are essential. However, we are currently lacking sufficient models, especially in vitro, to represent each (sub)type and their heterogeneity. To generate large series of preclinical in vitro models for pediatric brain tumors, we are using organoid technology. Cells from patient samples and patient-derived xenograft samples have been taken into culture to establish 3D organoids using tumor type specific culture conditions. These organoid lines retain the molecular characteristics of the original tumor tissue. They can be used to perform high-throughput drug screens, genetic manipulations, and co-cultures with, for instance, immune cells. Viable tissue is not always available for all tumor (sub)types and specific oncodrivers. To circumvent this lack of tissue, we can also induce tumors in vitro. Therefore, we generate cerebral and cerebellar brain organoids from human pluripotent stem cells. These organoids mimic human developing brain cells and can be genetically manipulated to model different brain tumor types. These genetically engineered brain tumor models allow us to study the cellular origins of pediatric brain tumors and the different tumor driving mechanisms. Tumors induced in the brain organoids histologically and molecularly resemble human patient samples based on (single cell) transcriptomic analyses. Moreover, the tumor cells are able to establish xenografts in mouse brains. In summary, organoid technology provides a novel avenue to establish in vitro models for pediatric brain tumors. At the meeting we will present data for various new ependymoma, medulloblastoma and embryonal brain tumor organoid models.
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Delepine, Chloe, Vincent A. Pham, Hayley W. S. Tsang, and Mriganka Sur. "GSK3ß inhibitor CHIR 99021 modulates cerebral organoid development through dose-dependent regulation of apoptosis, proliferation, differentiation and migration." PLOS ONE 16, no. 5 (May 5, 2021): e0251173. http://dx.doi.org/10.1371/journal.pone.0251173.
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Cerebral organoids generated from human pluripotent stem cells (hiPSCs) are unique in their ability to recapitulate human-specific neurodevelopmental events. They are capable of modeling the human brain and its cell composition, including human-specific progenitor cell types; ordered laminar compartments; and both cell-specific transcriptional signatures and the broader telencephalic transcriptional landscape. The serine/threonine kinase, GSK3β, plays a critical role in neurodevelopment, controlling processes as varied as neurogenesis, morphological changes, polarization, and migration. In the generation of cerebral organoids, inhibition of GSK3β at low doses has been used to increase organoid size and decrease necrotic core. However, little is known of the effects of GSK3β inhibition on organoid development. Here, we demonstrate that while low dose of GSK3β inhibitor CHIR 99021 increases organoid size, higher dose actually reduces organoid size; with the highest dose arresting organoid growth. To examine the mechanisms that may contribute to the phenotypic size differences observed in these treatment groups, we show that low dose of CHIR 99021 increases cell survival, neural progenitor cell proliferation and neuronal migration. A higher dose, however, decreases not only apoptosis but also proliferation, and arrests neural differentiation, enriching the pool of neuroepithelial cells, and decreasing the pools of early neuronal progenitors and neurons. These results reveal new mechanisms of the pleiotropic effects of GSK3β during organoid development, providing essential information for the improvement of organoid production and ultimately shedding light on the mechanisms of embryonic brain development.
10
Yakoub, Abraam M., and Mark Sadek. "Development and Characterization of Human Cerebral Organoids." Cell Transplantation 27, no. 3 (March 2018): 393–406. http://dx.doi.org/10.1177/0963689717752946.
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Studies of human neurodevelopmental disorders and stem cell–based regenerative transplants have been hampered by the lack of a model of the developing human brain. Stem cell–derived neurons suffer major limitations, including the ability to recapitulate the 3-dimensional architecture of a brain tissue and the representation of multiple layers and cell types that contribute to the overall brain functions in vivo. Recently, cerebral organoid technology was introduced; however, such technology is still in its infancy, and its low reproducibility and limitations significantly reduce the reliability of such a model as it currently exists, especially considering the complexity of cerebral-organoid protocols. Here we have tested and compared multiple protocols and conditions for growth of organoids, and we describe an optimized methodology, and define the necessary and sufficient factors that support the development of optimal organoids. Our optimization criteria included organoids’ overall growth and size, stratification and representation of the various cell types, inter-batch variability, analysis of neuronal maturation, and even the cost of the procedure. Importantly, this protocol encompasses a plethora of technical tips that allow researchers to easily reproduce it and obtain reliable organoids with the least variability, and showcases a robust array of approaches to characterize successful organoids. This optimized protocol provides a reliable system for genetic or pharmacological (drug development) screens and may enhance understanding and therapy of human neurodevelopmental disorders, including harnessing the therapeutic potential of stem cell–derived transplants.
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Tanaka, Yoshiaki, and In-Hyun Park. "Regional specification and complementation with non-neuroectodermal cells in human brain organoids." Journal of Molecular Medicine 99, no. 4 (March 2, 2021): 489–500. http://dx.doi.org/10.1007/s00109-021-02051-9.
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AbstractAlong with emergence of the organoids, their application in biomedical research has been currently one of the most fascinating themes. For the past few years, scientists have made significant contributions to deriving organoids representing the whole brain and specific brain regions. Coupled with somatic cell reprogramming and CRISPR/Cas9 editing, the organoid technologies were applied for disease modeling and drug screening. The methods to develop organoids further improved for rapid and efficient generation of cerebral organoids. Additionally, refining the methods to develop the regionally specified brain organoids enabled the investigation of development and interaction of the specific brain regions. Recent studies started resolving the issue in the lack of non-neuroectodermal cells in brain organoids, including vascular endothelial cells and microglia, which play fundamental roles in neurodevelopment and are involved in the pathophysiology of acute and chronic neural disorders. In this review, we highlight recent advances of neuronal organoid technologies, focusing on the region-specific brain organoids and complementation with endothelial cells and microglia, and discuss their potential applications to neuronal diseases.
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da Silva, Bárbara, Ryan K. Mathew, Euan S. Polson, Jennifer Williams, and Heiko Wurdak. "Spontaneous Glioblastoma Spheroid Infiltration of Early-Stage Cerebral Organoids Models Brain Tumor Invasion." SLAS DISCOVERY: Advancing the Science of Drug Discovery 23, no. 8 (March 15, 2018): 862–68. http://dx.doi.org/10.1177/2472555218764623.
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Organoid methodology provides a platform for the ex vivo investigation of the cellular and molecular mechanisms underlying brain development and disease. The high-grade brain tumor glioblastoma multiforme (GBM) is considered a cancer of unmet clinical need, in part due to GBM cell infiltration into healthy brain parenchyma, making complete surgical resection improbable. Modeling the process of GBM invasion in real time is challenging as it requires both tumor and neural tissue compartments. Here, we demonstrate that human GBM spheroids possess the ability to spontaneously infiltrate early-stage cerebral organoids (eCOs). The resulting formation of hybrid organoids demonstrated an invasive tumor phenotype that was distinct from noncancerous adult neural progenitor (NP) spheroid incorporation into eCOs. These findings provide a basis for the modeling and quantification of the GBM infiltration process using a stem-cell-based organoid approach, and may be used for the identification of anti-GBM invasion strategies.
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Li, Xiaodong, Abdullah Shopit, and Jingmin Wang. "A Comprehensive Update of Cerebral Organoids between Applications and Challenges." Oxidative Medicine and Cellular Longevity 2022 (December 5, 2022): 1–10. http://dx.doi.org/10.1155/2022/7264649.
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The basic technology of stem cells has been developed and created organoids, which have established a strong interest in regenerative medicine. Different cell types have been used to generate cerebral organoids, which include interneurons and oligodendrocytes (OLs). OLs are fundamental for brain development. Abundant studies have displayed that brain organoids can recapitulate fundamental and vital features of the human brain, such as cellular regulation and distribution, neuronal networks, electrical activities, and physiological structure. The organoids contain essential ventral brain domains and functional cortical interneurons, which are similar to the developing cortex and medial ganglionic eminence (MGE). So, brain organoids have provided a singular model to study and investigate neurological disorder mechanisms and therapeutics. Furthermore, the blood brain barrier (BBB) organoids modeling contributes to accelerate therapeutic discovery for the treatment of several neuropathologies. In this review, we summarized the advances of the brain organoids applications to investigate neurological disorder mechanisms such as neurodevelopmental and neurodegenerative disorders, mental disorders, brain cancer, and cerebral viral infections. We discussed brain organoids’ therapeutic application as a potential therapeutic unique method and highlighted in detail the challenges and hurdles of organoid models.
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Bock, Minsung, Sung Jun Hong, Songzi Zhang, Yerin Yu, Somin Lee, Haeeun Shin, Byung Hyune Choi, and Inbo Han. "Morphogenetic Designs, and Disease Models in Central Nervous System Organoids." International Journal of Molecular Sciences 25, no. 14 (July 15, 2024): 7750. http://dx.doi.org/10.3390/ijms25147750.
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Since the emergence of the first cerebral organoid (CO) in 2013, advancements have transformed central nervous system (CNS) research. Initial efforts focused on studying the morphogenesis of COs and creating reproducible models. Numerous methodologies have been proposed, enabling the design of the brain organoid to represent specific regions and spinal cord structures. CNS organoids now facilitate the study of a wide range of CNS diseases, from infections to tumors, which were previously difficult to investigate. We summarize the major advancements in CNS organoids, concerning morphogenetic designs and disease models. We examine the development of fabrication procedures and how these advancements have enabled the generation of region-specific brain organoids and spinal cord models. We highlight the application of these organoids in studying various CNS diseases, demonstrating the versatility and potential of organoid models in advancing our understanding of complex conditions. We discuss the current challenges in the field, including issues related to reproducibility, scalability, and the accurate recapitulation of the in vivo environment. We provide an outlook on prospective studies and future directions. This review aims to provide a comprehensive overview of the state-of-the-art CNS organoid research, highlighting key developments, current challenges, and prospects in the field.
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Wong, HakKei. "The importance of cerebral organoid technology in medicine." Highlights in Science, Engineering and Technology 2 (June 22, 2022): 179–85. http://dx.doi.org/10.54097/hset.v2i.572.
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Because of the intricate nature of the nervous systems, neurological diseases have always been one of the least studied areas of pathology and medicine. Currently, there is no cure for these kinds of diseases but only medications or therapies that relieve symptoms and minimise suffering. Thus, cerebral organoids derived from human pluripotent stem cells are produced in order to study the development and pathology of the human brain, especially the embryonic stage, and to model neurological diseases. In this dissertation, I will make a judgement on the appropriate usage of cerebral organoid in investigating neurological disease through exploring and assessing the effectiveness of the cerebral organoids modeling Zika Virus and Alzheimer’s disease and examining the ethical issues arising from this practice.
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Berdenis van Berlekom, Amber, Raphael Kübler, Jeske W. Hoogeboom, Daniëlle Vonk, Jacqueline A. Sluijs, R. Jeroen Pasterkamp, Jinte Middeldorp, et al. "Exposure to the Amino Acids Histidine, Lysine, and Threonine Reduces mTOR Activity and Affects Neurodevelopment in a Human Cerebral Organoid Model." Nutrients 14, no. 10 (May 23, 2022): 2175. http://dx.doi.org/10.3390/nu14102175.
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Evidence of the impact of nutrition on human brain development is compelling. Previous in vitro and in vivo results show that three specific amino acids, histidine, lysine, and threonine, synergistically inhibit mTOR activity and behavior. Therefore, the prenatal availability of these amino acids could be important for human neurodevelopment. However, methods to study the underlying mechanisms in a human model of neurodevelopment are limited. Here, we pioneer the use of human cerebral organoids to investigate the impact of amino acid supplementation on neurodevelopment. In this study, cerebral organoids were exposed to 10 mM and 50 mM of the amino acids threonine, histidine, and lysine. The impact was determined by measuring mTOR activity using Western blots, general cerebral organoid size, and gene expression by RNA sequencing. Exposure to threonine, histidine, and lysine led to decreased mTOR activity and markedly reduced organoid size, supporting findings in rodent studies. RNA sequencing identified comprehensive changes in gene expression, with enrichment in genes related to specific biological processes (among which are mTOR signaling and immune function) and to specific cell types, including proliferative precursor cells, microglia, and astrocytes. Altogether, cerebral organoids are responsive to nutritional exposure by increasing specific amino acid concentrations and reflect findings from previous rodent studies. Threonine, histidine, and lysine exposure impacts the early development of human cerebral organoids, illustrated by the inhibition of mTOR activity, reduced size, and altered gene expression.
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Sapir, Gal, Daniel J. Steinberg, Rami I. Aqeilan, and Rachel Katz-Brull. "Real-Time Non-Invasive and Direct Determination of Lactate Dehydrogenase Activity in Cerebral Organoids—A New Method to Characterize the Metabolism of Brain Organoids?" Pharmaceuticals 14, no. 9 (August 30, 2021): 878. http://dx.doi.org/10.3390/ph14090878.
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Organoids are a powerful tool in the quest to understand human diseases. As the developing brain is extremely inaccessible in mammals, cerebral organoids (COs) provide a unique way to investigate neural development and related disorders. The aim of this study was to utilize hyperpolarized 13C NMR to investigate the metabolism of COs in real-time, in a non-destructive manner. The enzymatic activity of lactate dehydrogenase (LDH) was determined by quantifying the rate of [1-13C]lactate production from hyperpolarized [1-13C]pyruvate. Organoid development was assessed by immunofluorescence imaging. Organoid viability was confirmed using 31P NMR spectroscopy. A total of 15 organoids collated into 3 groups with a group total weight of 20–77 mg were used in this study. Two groups were at the age of 10 weeks and one was at the age of 33 weeks. The feasibility of this approach was demonstrated in both age groups, and the LDH activity rate was found to be 1.32 ± 0.75 nmol/s (n = 3 organoid batches). These results suggest that hyperpolarized NMR can be used to characterize the metabolism of brain organoids with a total tissue wet weight of as low as 20 mg (<3 mm3) and a diameter ranging from 3 to 6 mm.
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Li, Chong, Jonas Simon Fleck, Catarina Martins-Costa, Thomas R. Burkard, Jan Themann, Marlene Stuempflen, Angela Maria Peer, et al. "Single-cell brain organoid screening identifies developmental defects in autism." Nature 621, no. 7978 (September 13, 2023): 373–80. http://dx.doi.org/10.1038/s41586-023-06473-y.
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AbstractThe development of the human brain involves unique processes (not observed in many other species) that can contribute to neurodevelopmental disorders1–4. Cerebral organoids enable the study of neurodevelopmental disorders in a human context. We have developed the CRISPR–human organoids–single-cell RNA sequencing (CHOOSE) system, which uses verified pairs of guide RNAs, inducible CRISPR–Cas9-based genetic disruption and single-cell transcriptomics for pooled loss-of-function screening in mosaic organoids. Here we show that perturbation of 36 high-risk autism spectrum disorder genes related to transcriptional regulation uncovers their effects on cell fate determination. We find that dorsal intermediate progenitors, ventral progenitors and upper-layer excitatory neurons are among the most vulnerable cell types. We construct a developmental gene regulatory network of cerebral organoids from single-cell transcriptomes and chromatin modalities and identify autism spectrum disorder-associated and perturbation-enriched regulatory modules. Perturbing members of the BRG1/BRM-associated factor (BAF) chromatin remodelling complex leads to enrichment of ventral telencephalon progenitors. Specifically, mutating the BAF subunit ARID1B affects the fate transition of progenitors to oligodendrocyte and interneuron precursor cells, a phenotype that we confirmed in patient-specific induced pluripotent stem cell-derived organoids. Our study paves the way for high-throughput phenotypic characterization of disease susceptibility genes in organoid models with cell state, molecular pathway and gene regulatory network readouts.
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Batara, Don Carlo Ramos, Shuchang Zhou, Moon-Chang Choi, and Sung-Hak Kim. "Glioblastoma organoid technology: an emerging preclinical models for drug discovery." Organoid 2 (February 25, 2022): e7. http://dx.doi.org/10.51335/organoid.2022.2.e7.
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Glioblastoma multiforme (GBM) is the most prevalent type of primary brain tumor among adults, and it has a median overall survival of 12 to 15 months upon diagnosis. Despite significant improvements in GBM research, therapeutic options are still limited and survival rates have not significantly improved. Accordingly, clinical and translational studies are hampered due to the lack of suitable preclinical models that accurately reflect the brain tumor architecture and its microenvironment. Scientists have recently developed cerebral organoids, which are artificial 3-dimensional brain-like tissue. Organoid technology provides new cancer modeling options, which could help us better understand GBM pathogenesis and design personalized treatments. In this review, we summarize recent developments in organoid GBM models, highlighting their advantages in cancer modeling, as well as their challenges and limitations and potential future directions in GBM therapy.
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Sivitilli, Adam A., Jessica T. Gosio, Bibaswan Ghoshal, Alesya Evstratova, Daniel Trcka, Parisa Ghiasi, J. Javier Hernandez, Jean Martin Beaulieu, Jeffrey L. Wrana, and Liliana Attisano. "Robust production of uniform human cerebral organoids from pluripotent stem cells." Life Science Alliance 3, no. 5 (April 17, 2020): e202000707. http://dx.doi.org/10.26508/lsa.202000707.
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Human cerebral organoid (hCO) models offer the opportunity to understand fundamental processes underlying human-specific cortical development and pathophysiology in an experimentally tractable system. Although diverse methods to generate brain organoids have been developed, a major challenge has been the production of organoids with reproducible cell type heterogeneity and macroscopic morphology. Here, we have directly addressed this problem by establishing a robust production pipeline to generate morphologically consistent hCOs and achieve a success rate of >80%. These hCOs include both a radial glial stem cell compartment and electrophysiologically competent mature neurons. Moreover, we show using immunofluorescence microscopy and single-cell profiling that individual organoids display reproducible cell type compositions that are conserved upon extended culture. We expect that application of this method will provide new insights into brain development and disease processes.
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Bunt, Jens, Mieke Roosen, Evie Egelmeers, Joris Maas, Zelda Ode, and Marcel Kool. "TMOD-02. GEBTO: GENETICALLY ENGINEERED BRAIN TUMOR ORGANOIDS AS A NOVEL PRECLINICAL MODEL." Neuro-Oncology 23, Supplement_1 (June 1, 2021): i35—i36. http://dx.doi.org/10.1093/neuonc/noab090.143.
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Abstract Background One of the bottlenecks in basic and translational research on pediatric brain tumors, is the lack of suitable and representative preclinical models to study tumor biology and drug sensitivity. Over the last decades, extensive molecular characterization has uncovered many entities and subgroups with their unique oncodriving events. However, this heterogeneity is currently not reflected in the models available, especially not for in vitro models. Objectives We aim to generate genetically engineered brain tumor organoids (GEBTO) to represent the molecular variety of embryonal brain tumors and ependymomas. Method Human brain organoids derived from embryonic stem cells are generated to represent the region of tumor origin. To mimic oncodriving events, DNA plasmids are introduced via electroporation in the organoid cells to knockout tumor suppressor genes or overexpress oncogenes. Results Cerebellar and cerebral forebrain organoids were generated as the tissue of origin for medulloblastoma and supratentorial ependymoma (ST-EPN), respectively. Based on the detection of GFP protein encoded by DNA plasmids, the organoid cells can be manipulated within a wide developmental window, which corresponds with the presence of the proposed cells of origin. Different oncodrivers and combinations thereof are now being tested to see whether they result in ectopic growth in cerebral or cerebellar organoids. When successful, the GEBTOs are histologically and molecularly characterized using (single cell) transcriptomic and epigenomic analyses to see how well they resemble human tumors. Discussion Although further development is required, GEBTOs provide a novel avenue to model especially rare pediatric brain tumors, for which tissue and therefore patient-derived models are limited. It also allows for in-depth analyses of the potential cells of origin and the contribution of different mutations to tumor biology.
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Sukhinich, K. K., K. M. Shakirova, E. B. Dashinimaev, and M. A. Aleksandrova. "Development of 3D Cerebral Aggregates in the Brain Ventricles of Adult Mice." Russian Journal of Developmental Biology 52, no. 3 (May 2021): 164–75. http://dx.doi.org/10.1134/s1062360421030061.
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Abstract The cerebral organoids are three-dimensional cell cultures formed from brain-specific cell types arising from embryonic or pluripotent stem cells. Organoids provide an opportunity to study the early stages of brain development and diseases of the central nervous system. However, the modeling of organoids is associated with a number of unsolved problems. Organoid production techniques involve a complex cell culture process that requires special media, growth factors, and often the use of a bioreactor. Even under standardized conditions, structures of different morphology are formed: from disorganized cell aggregates to structured minibrains, which are selected for study. For natural reasons, organoids grown in vitro do not have a blood supply, which limits their development. We tried to obtain cerebral aggregates similar to organoids in an in vivo model, where vascular growth and tissue blood supply are provided, for which we transplanted a cell suspension from the mouse embryonic neocortex into the lateral ventricles of the brain of adult mice. Therefore, the medium for cultivation was the cerebrospinal fluid, and the lateral ventricles of the brain, where it circulates, served as a bioreactor. The results showed that the neocortex from E14.5 is a suitable source of stem/progenitor cells that self-assemble into three-dimensional aggregates and vascularized in vivo. The aggregates consisted of a central layer of mature neurons, the marginal zone free of cells and a glia limitans, which resembled cerebral organoids. Thus, the lateral ventricles of the adult mouse brain can be used to obtain vascularized cell aggregates resembling cerebral organoids.
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Robles, Denise, Andrew Boreland, Zhiping Pang, and Jeffrey Zahn. "A Cerebral Organoid Connectivity Apparatus to Model Neuronal Tract Circuitry." Micromachines 12, no. 12 (December 17, 2021): 1574. http://dx.doi.org/10.3390/mi12121574.
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Mental disorders have high prevalence, but the efficacy of existing therapeutics is limited, in part, because the pathogenic mechanisms remain enigmatic. Current models of neural circuitry include animal models and post-mortem brain tissue, which have allowed enormous progress in understanding the pathophysiology of mental disorders. However, these models limit the ability to assess the functional alterations in short-range and long-range network connectivity between brain regions that are implicated in many mental disorders, e.g., schizophrenia and autism spectrum disorders. This work addresses these limitations by developing an in vitro model of the human brain that models the in vivo cerebral tract environment. In this study, microfabrication and stem cell differentiation techniques were combined to develop an in vitro cerebral tract model that anchors human induced pluripotent stem cell-derived cerebral organoids (COs) and provides a scaffold to promote the formation of a functional connecting neuronal tract. Two designs of a Cerebral Organoid Connectivity Apparatus (COCA) were fabricated using SU-8 photoresist. The first design contains a series of spikes which anchor the CO to the COCA (spiked design), whereas the second design contains flat supporting structures with open holes in a grid pattern to anchor the organoids (grid design); both designs allow effective media exchange. Morphological and functional analyses reveal the expression of key neuronal markers as well as functional activity and signal propagation along cerebral tracts connecting CO pairs. The reported in vitro models enable the investigation of critical neural circuitry involved in neurodevelopmental processes and has the potential to help devise personalized and targeted therapeutic strategies.
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Krieger, Teresa G., Stephan M. Tirier, Jeongbin Park, Katharina Jechow, Tanja Eisemann, Heike Peterziel, Peter Angel, Roland Eils, and Christian Conrad. "Modeling glioblastoma invasion using human brain organoids and single-cell transcriptomics." Neuro-Oncology 22, no. 8 (April 16, 2020): 1138–49. http://dx.doi.org/10.1093/neuonc/noaa091.
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Abstract Background Glioblastoma (GBM) consists of devastating neoplasms with high invasive capacity, which have been difficult to study in vitro in a human-derived model system. Therapeutic progress is also limited by cellular heterogeneity within and between tumors, among other factors such as therapy resistance. To address these challenges, we present an experimental model using human cerebral organoids as a scaffold for patient-derived GBM cell invasion. Methods This study combined tissue clearing and confocal microscopy with single-cell RNA sequencing of GBM cells before and after co-culture with organoid cells. Results We show that tumor cells within organoids extend a network of long microtubes, recapitulating the in vivo behavior of GBM. Transcriptional changes implicated in the invasion process are coherent across patient samples, indicating that GBM cells reactively upregulate genes required for their dispersion. Potential interactions between GBM and organoid cells identified by an in silico receptor–ligand pairing screen suggest functional therapeutic targets. Conclusions Taken together, our model has proven useful for studying GBM invasion and transcriptional heterogeneity in vitro, with applications for both pharmacological screens and patient-specific treatment selection on a time scale amenable to clinical practice.
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Tongkrajang, Nongnat, Porntida Kobpornchai, Pratima Dubey, Urai Chaisri, and Kasem Kulkeaw. "Modelling amoebic brain infection caused by Balamuthia mandrillaris using a human cerebral organoid." PLOS Neglected Tropical Diseases 18, no. 6 (June 20, 2024): e0012274. http://dx.doi.org/10.1371/journal.pntd.0012274.
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The lack of disease models adequately resembling human tissue has hindered our understanding of amoebic brain infection. Three-dimensional structured organoids provide a microenvironment similar to human tissue. This study demonstrates the use of cerebral organoids to model a rare brain infection caused by the highly lethal amoeba Balamuthia mandrillaris. Cerebral organoids were generated from human pluripotent stem cells and infected with clinically isolated B. mandrillaris trophozoites. Histological examination showed amoebic invasion and neuron damage following coculture with the trophozoites. The transcript profile suggested an alteration in neuron growth and a proinflammatory response. The release of intracellular proteins specific to neuronal bodies and astrocytes was detected at higher levels postinfection. The amoebicidal effect of the repurposed drug nitroxoline was examined using the human cerebral organoids. Overall, the use of human cerebral organoids was important for understanding the mechanism of amoeba pathogenicity, identify biomarkers for brain injury, and in the testing of a potential amoebicidal drug in a context similar to the human brain.
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Ferreira, Rodolfo Sanches, Bruno H. S. Araujo, and Oswaldo Okamoto. "MODL-06. ASSESSMENT OF ONCOLYTIC VIRUS SPECIFICITY AND CYTOTOXICITY IN A HYBRID GLIOBLASTOMA-CEREBRAL ORGANOID MODEL." Neuro-Oncology 24, Supplement_7 (November 1, 2022): vii292. http://dx.doi.org/10.1093/neuonc/noac209.1134.
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Abstract Recent studies have demonstrated potent oncolytic effects of wild-type Zika virus (ZIKV) against primary central nervous system (CNS) tumors, including Medulloblastoma, Atypical Teratoid/Rhabdoid Tumor (AT/RT), and Glioblastoma (GBM). However, the neurotropism of ZIKV urges further evaluation of specific tumor-targeting properties and comparative toxicity to non-neoplastic neural cells in order to address its therapeutic potential. We have developed a hybrid organoid model by co-culturing GBM cells with mature human cerebral organoids and assessed cytotoxicity of the Brazilian ZIKV isolate (ZIKVBR) towards normal brain organoid cells and GBM cells. Human induced pluripotent stem cells (hiPSC)-derived cerebral organoids were co-cultured with 105 GFP-expressing cells of three different GBM cell lines. A total of 20.000 PFU of ZIKVBR or mock condition was added per co-cultured model. Viable cells were analyzed by flow cytometry (FC) and fluorescence microscopy at different time points. ZIKVBR presence was assessed by PFU assay. Although ZIKVBR infection did not cause a pronounced reduction of GFP+ cell proportion over time in all GBM cell lines, cell viability analysis showed a greater amount of non-viable GFP+ cells over GFP- cells in all ZIKVBR-treated groups, compared to their corresponding mock groups. PFU assays of cell culture supernatants confirmed the presence of infectious viral particles in treated groups and absence in mock groups. Our data reveal that ZIKVBR has a potential oncolytic effect characterized by preferential killing of GBM tumor cells over normal cerebral cells, in a hybrid organoid model. These findings support the development of an oncolytic virus therapy platform based on ZIKV.
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Tomasso, Federica, Wai Chin Chong, Shivaprasad Bhuvanendran, Sridevi Yadavilli, Roger Packer, and Javad Nazarian. "NFS-22. INVESTIGATING MALIGNANT TRANSFORMATION IN NF1 PEDIATRIC GLIOMAS USING AN IPSCS-DERIVED CEREBRAL ORGANOID MODEL." Neuro-Oncology 26, Supplement_4 (June 18, 2024): 0. http://dx.doi.org/10.1093/neuonc/noae064.580.
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Abstract BACKGROUND Neurofibromatosis Type 1 (NF1) is an autosomal dominant tumor predisposition syndrome affecting 1 in 3000 children around the world. Although NF1 inactivation is recognized as the cause of low-grade glioma (LGG) development, little is known about subsequent genetic alterations that lead to malignant transformation. One of the limitations in understanding the molecular events underlying the malignant transformation of NF1 gliomas is the lack of appropriate NF1 preclinical models for its comprehensive profiling. OBJECTIVE Brain organoids have emerged as powerful in vitro models for studying the interaction between tumor cells and the tumor microenvironment. In this study, we present a novel cerebral organoid model, developed to investigate the molecular events leading to malignant transformation in NF1 pediatric gliomas. METHODS Induced pluripotent stem cells (iPSCs)-derived cerebral organoids were generated using NF1wt and NF1mut iPSCs, the latter reprogrammed from patients with plexiform neurofibromas (Carlos III Health Institute, Spain). Immunofluorescence analysis was performed at four different developmental stages to confirm their correct cytoarchitectural development and differentiation into heterogeneous cell populations. Furthermore, four mCherry-labelled isogenic glioma cells lines, RES 186 wild type, NF1-/-, CDKN2a-/-,NF1-/-/CDKN2a-/- were co-cultured with the developed cerebral organoids and their migration potential and invasiveness were compared using two-photon microscopy. RESULTS/DISCUSSION NF1mut cerebral organoids display ventricular-like zone at early stages of development (d15-d30), characterized by luminal Sox2+ neural stem cells surrounded by Tuj1+ immature neuron. These organized structures disappear at later stages, while progenitor cells start to differentiate into fully mature neurons and glial cells (d45-d60). Additionally, after 10 days of co-culture, all four RES186 isogenic cell lines infiltrated the organoid structure with different invasion patterns. The established organoid model will provide a platform for evaluating the impact of sequential mutational events on LGG transformation, taking into consideration the role exerted by the tumor microenvironment in supporting tumor cell invasion and migration.
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Roosen, Mieke, Julie Morscio, Phylicia Stathi, Norman Mack, Benjamin Schwalm, Panagiotis A. Polychronopoulos, Mariëtte E. G. Kranendonk, Eelco Hoving, Jens Bunt, and Marcel Kool. "EPEN-17.IN VITRO MODELLING OF PEDIATRIC SUPRATENTORIAL EPENDYMOMAS USING CEREBRAL ORGANOIDS." Neuro-Oncology 26, Supplement_4 (June 18, 2024): 0. http://dx.doi.org/10.1093/neuonc/noae064.219.
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Abstract BACKGROUND Ependymomas (EPN) are glial tumors of the central nervous system occurring in children and adults. ZFTA-fusion positive and YAP1-fusion positive EPN are the main supratentorial (ST) subgroups in children. While most ST-EPN-YAP1 patients survive, only 50% of ZFTA positive patients survive longer than 5 years. Improving survival of ST-EPN patients requires a better understanding of tumor biology. Human embryonal stem cell-derived (hESC) brain organoids provide a novel opportunity to model ST-EPN and study the impact of oncogenic fusions on tumor development within a healthy brain environment. METHODS hESC-derived cerebral organoids were genetically modified with YAP1 or ZFTA fusion genes and histologically and molecularly analyzed using antibody stainings, bulk and single cell RNA sequencing. RESULTS scRNA-seq analyses showed that our cerebral organoids mimic embryonal brain development and that radial glia, the presumed cell-of-origin of ST-EPN are abundant in 11-day old organoids. At this timepoint, electroporation of oncogenic ZFTA-RELA or YAP1-MAMLD1/YAP1-FAM118B fusions led to ectopic tumor outgrowth. Histological and molecular analyses showed that ZFTA and YAP1 EPN tumor organoids displayed different phenotypes and fusion-specific gene signatures, closely resembling human ZFTA and YAP1 EPN patient samples. ScRNA-seq data of organoid tumors showed a skewed differentiation compared to normal development with ZFTA tumors being more neuronal and YAP1 tumors having a more extracellular matrix-like phenotype. Analysis of the healthy compartment showed that YAP1 tumors influence the differentiation of the healthy cells and in both subtypes a new cell cluster was identified with high expression of tumor associated markers. Intercellular interactions revealed potential targetable tumor(specific) interactions such as YBX1-NOTCH1 in ZFTA or GNAI2-CAV1 in YAP1 tumor organoid models, as well as CD99-CD81 in both. CONCLUSIONS These models contribute to a better molecular and biological understanding of ependymomas and can be used to identify targeted therapies, especially those targeting the tumor microenvironment.
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Santos, Alexandra C., George Nader, Dana El Soufi El Sabbagh, Karolina Urban, Liliana Attisano, and Peter L. Carlen. "Treating Hyperexcitability in Human Cerebral Organoids Resulting from Oxygen-Glucose Deprivation." Cells 12, no. 15 (July 27, 2023): 1949. http://dx.doi.org/10.3390/cells12151949.
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Human cerebral organoids resemble the 3D complexity of the human brain and have the potential to augment current drug development pipelines for neurological disease. Epilepsy is a complex neurological condition characterized by recurrent seizures. A third of people with epilepsy do not respond to currently available pharmaceutical drugs, and there is not one drug that treats all subtypes; thus, better models of epilepsy are needed for drug development. Cerebral organoids may be used to address this unmet need. In the present work, human cerebral organoids are used along with electrophysiological methods to explore oxygen-glucose deprivation as a hyperexcitability agent. This activity is investigated in its response to current antiseizure drugs. Furthermore, the mechanism of action of the drug candidates is probed with qPCR and immunofluorescence. The findings demonstrate OGD-induced hyperexcitable changes in the cerebral organoid tissue, which is treated with cannabidiol and bumetanide. There is evidence for NKCC1 and KCC2 gene expression, as well as other genes and proteins involved in the complex development of GABAergic signaling. This study supports the use of organoids as a platform for modelling cerebral cortical hyperexcitability that could be extended to modelling epilepsy and used for drug discovery.
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Schneider, Eric, Leigh Ann Samsa, and Veljko Dubljević. "Political and ethical landscape of brain organoid research." Molecular Psychology: Brain, Behavior, and Society 2 (April 19, 2023): 3. http://dx.doi.org/10.12688/molpsychol.17521.1.
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Human cerebral organoids (hCOs), produced in labs through directed cell culture of embryonic or induced pluripotent stem cells, closely replicate the 3-dimensional architecture of the human brain on a micro scale. This technology has been used to model neurological disease and shows promise to complement or supplant animal subjects in preclinical therapeutic investigation. However, attention must be paid by researchers and institutions to the various ethical concerns associated with hCO development. Human-animal chimeras produced through the grafting of hCOs have shown integration of neurological function, calling into question the moral status of both the animal chimeras and the organoid itself. Sensationalist reporting on such acts may also prompt public backlash, potentially jeopardizing hCO research and the promised benefits thereof. Moreover, concerns arise over privacy and consent for past and prospective donors of stem cells used to produce organoids. Genetic information may be considered privileged to the public domain and disrupted trust can reduce the supply of willing donors. Though hCOs are believed thus far to lack the capacity for consciousness and cognitive function, consideration must be given to their potential status as moral agents with further development or enhancement. Boundaries concerning organoids adhered to by researchers have been largely voluntary and informal to this point. By edict or by the power of the purse, governmental regulatory agencies ought to formalize necessary guidelines to ensure compliance with ethical principles and the adequate representation of all affected stakeholders in future decisions.
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Yin, He. "Human brain organoids combined with CRISPR technology to gain insight into neurological diseases." Highlights in Science, Engineering and Technology 102 (July 11, 2024): 75–79. http://dx.doi.org/10.54097/m3grdg15.
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As the most complex organ of human beings, the brain's functions cover various high-level intellectual activities such as cognition, language and thinking. These functions are closely related to the developmental processes in the cerebral cortex granted the composition structure of the nervous system in the cortex. Therefore, scientists have long been exploring the developmental processes of the cerebral cortex and the constituent structures of the nervous system, the purpose of the system is to deepen comprehend how the brain works. The study of neurological diseases by improving three-dimensional brain models is a hot issue at present, because the brain is very complex and fragile, and many therapies are weak and still theoretical. Brain organoids are a kind of three-dimensional tissue culture derived from human pluripotent stem cells (hPSCs), which can simulate the type composition, spatial structure, physiological function and other characteristics of human brain cells by culture in vitro. The use of brain organoid technology can simulate the development process of the brain in vitro, providing an unprecedented opportunity to study the occurrence of brain function and nervous system diseases, and combining CRISPR technology for screening, so that it can be further applied efficiently. This research will summarize the development process, progress and deficiency of brain organoids and the research situation of neurological diseases, and make a summary of the current development and prospect of the future development.
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Schneider, Eric, Leigh Ann Samsa, and Veljko Dubljević. "A Political and ethical landscape of brain organoid research." Molecular Psychology: Brain, Behavior, and Society 2 (March 15, 2024): 3. http://dx.doi.org/10.12688/molpsychol.17521.2.
Full textAbstract:
Human cerebral organoids (hCOs), produced in labs through directed cell culture of embryonic or induced pluripotent stem cells, closely replicate the 3-dimensional architecture of the human brain on a micro scale. This technology has been used to model neurological disease and shows promise to complement or supplant animal subjects in preclinical therapeutic investigation. However, attention must be paid by researchers and institutions to the various ethical concerns associated with hCO development. Human-animal chimeras produced through the grafting of hCOs have shown integration of neurological function, calling into question the moral status of both the animal chimeras and the organoid itself. Sensationalist reporting on such acts may also prompt public backlash, potentially jeopardizing hCO research and the promised benefits thereof. Moreover, concerns arise over privacy and consent for past and prospective donors of stem cells used to produce organoids. Genetic information may be considered privileged to the public domain and disrupted trust can reduce the supply of willing donors. Though hCOs are believed thus far to lack the capacity for consciousness and cognitive function, consideration must be given to their potential status as moral agents with further development or enhancement. Boundaries concerning organoids adhered to by researchers have been largely voluntary and informal to this point. By edict or by the power of the purse, governmental regulatory agencies ought to formalize necessary guidelines to ensure compliance with ethical principles and the adequate representation of all affected stakeholders in future decisions.
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Weth, Freya R., Lifeng Peng, Erin Paterson, Swee T. Tan, and Clint Gray. "Utility of the Cerebral Organoid Glioma ‘GLICO’ Model for Screening Applications." Cells 12, no. 1 (December 30, 2022): 153. http://dx.doi.org/10.3390/cells12010153.
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Glioblastoma, a grade IV astrocytoma, is regarded as the most aggressive primary brain tumour with an overall median survival of 16.0 months following the standard treatment regimen of surgical resection, followed by radiotherapy and chemotherapy with temozolomide. Despite such intensive treatment, the tumour almost invariably recurs. This poor prognosis has most commonly been attributed to the initiation, propagation, and differentiation of cancer stem cells. Despite the unprecedented advances in biomedical research over the last decade, the current in vitro models are limited at preserving the inter- and intra-tumoural heterogeneity of primary tumours. The ability to understand and manipulate complex cancers such as glioblastoma requires disease models to be clinically and translationally relevant and encompass the cellular heterogeneity of such cancers. Therefore, brain cancer research models need to aim to recapitulate glioblastoma stem cell function, whilst remaining amenable for analysis. Fortunately, the recent development of 3D cultures has overcome some of these challenges, and cerebral organoids are emerging as cutting-edge tools in glioblastoma research. The opportunity to generate cerebral organoids via induced pluripotent stem cells, and to perform co-cultures with patient-derived cancer stem cells (GLICO model), has enabled the analysis of cancer development in a context that better mimics brain tissue architecture. In this article, we review the recent literature on the use of patient-derived glioblastoma organoid models and their applicability for drug screening, as well as provide a potential workflow for screening using the GLICO model. The proposed workflow is practical for use in most laboratories with accessible materials and equipment, a good first pass, and no animal work required. This workflow is also amenable for analysis, with separate measures of invasion, growth, and viability.
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Shakya, Sajina, Christopher G. Hubert, and Justin D. Lathia. "A Material Transfer Agreement between Glioblastoma and Normal Brain Cells." Cancer Discovery 15, no. 2 (February 7, 2025): 261–63. https://doi.org/10.1158/2159-8290.cd-24-1661.
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Summary: Tumor cells communicate with normal cells in various ways, typically leading to the exploitation of resources of the normal cells by tumor cells for their benefit. In this issue, Mangena and colleagues use three-dimensional organoid models to show the transfer of GFP and mRNA from malignant glioblastoma to nonmalignant cells in cerebral organoid models; this transfer is facilitated by extracellular vesicles and possibly tunneling nanotubes, demonstrating how nonmalignant cells in the tumor microenvironment can be exploited by neighboring malignant cells. See related article by Mangena et al., p. 299
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Andrews, Madeline G., and Tomasz J. Nowakowski. "Human brain development through the lens of cerebral organoid models." Brain Research 1725 (December 2019): 146470. http://dx.doi.org/10.1016/j.brainres.2019.146470.
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Gumbs, Stephanie B. H., Amber Berdenis van Berlekom, Raphael Kübler, Pauline J. Schipper, Lavina Gharu, Marco P. Boks, Paul R. Ormel, Annemarie M. J. Wensing, Lot D. de Witte, and Monique Nijhuis. "Characterization of HIV-1 Infection in Microglia-Containing Human Cerebral Organoids." Viruses 14, no. 4 (April 16, 2022): 829. http://dx.doi.org/10.3390/v14040829.
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The achievement of an HIV cure is dependent on the eradication or permanent silencing of HIV-latent viral reservoirs, including the understudied central nervous system (CNS) reservoir. This requires a deep understanding of the molecular mechanisms of HIV’s entry into the CNS, latency establishment, persistence, and reversal. Therefore, representative CNS culture models that reflect the intercellular dynamics and pathophysiology of the human brain are urgently needed in order to study the CNS viral reservoir and HIV-induced neuropathogenesis. In this study, we characterized a human cerebral organoid model in which microglia grow intrinsically as a CNS culture model to study HIV infection in the CNS. We demonstrated that both cerebral organoids and isolated organoid-derived microglia (oMG), infected with replication-competent HIVbal reporter viruses, support productive HIV infection via the CCR5 co-receptor. Productive HIV infection was only observed in microglial cells. Fluorescence analysis revealed microglia as the only HIV target cell. Susceptibility to HIV infection was dependent on the co-expression of microglia-specific markers and the CD4 and CCR5 HIV receptors. Altogether, this model will be a valuable tool within the HIV research community to study HIV–CNS interactions, the underlying mechanisms of HIV-associated neurological disorders (HAND), and the efficacy of new therapeutic and curative strategies on the CNS viral reservoir.
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Wei, NaiLi, ZiFang Quan, Hailiang Tang, and JianHong Zhu. "Three-Dimensional Organoid System Transplantation Technologies in Future Treatment of Central Nervous System Diseases." Stem Cells International 2017 (2017): 1–14. http://dx.doi.org/10.1155/2017/5682354.
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In recent years, scientists have made great achievements in understanding the development of human brain and elucidating critical elements of stepwise spatiotemporal control strategies in neural stem cell specification lineage, which facilitates successful induction of neural organoid in vitro including the cerebral cortex, cerebellar, neural tube, hippocampus cortex, pituitary, and optic cup. Besides, emerging researches on neural organogenesis promote the application of 3D organoid system transplantation in treating central nervous system (CNS) diseases. Present review will categorize current researches on organogenesis into three approaches: (a) stepwise, direct organization of region-specific or population-enriched neural organoid; (b) assemble and direct distinct organ-specific progenitor cells or stem cells to form specific morphogenesis organoid; and (c) assemble embryoid bodies for induction of multilayer organoid. However, the majority of these researches focus on elucidating cellular and molecular mechanisms involving in brain organogenesis or disease development and only a few of them conducted for treating diseases. In this work, we will compare three approaches and also analyze their possible indications for diseases in future treatment on the basis of their distinct characteristics.
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Choe, Mu Seog, and Min Young Lee. "A brain metastasis model for breast cancer using human embryonic stem cell-derived cerebral organoids." Organoid 2 (August 25, 2022): e25. http://dx.doi.org/10.51335/organoid.2022.2.e25.
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Background: Breast cancer is a common cause of brain metastasis. Although breast cancer has relatively high survival rates, its survival rate after metastasis to the brain is lower. Conventional two-dimensional cell culture models and animal models are widely used in metastatic cancer research, and these models have tremendously contributed to the understanding of this disease. However, these models have some limitations, such as different physiological features and genetic backgrounds.Methods: We established a simple metastatic breast cancer model using human pluripotent stem cell-derived cerebral organoids (COs)—in this case, breast cancer cerebral organoids (BC-COs).Results: Using the BC-CO model, we induced the metastasis of MDA-MB-231 cells into COs by co-culture of cells with COs and compared the differences between adapted cancer cells in BC-COs and non-adapted cells. Our results showed that the proliferative capacity increased in adapted cells. Additionally, the expression levels of endothelial-mesenchymal transition markers and cancer stem cells were significantly higher in adapted cancer cells.Conclusion: We conclude that metastasis promotes the metastatic capacity of breast cancer cells. Our results also showed that the BC-CO model could be a novel tool for research on brain metastasis in breast cancer.
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Amiri, Anahita, Gianfilippo Coppola, Soraya Scuderi, Feinan Wu, Tanmoy Roychowdhury, Fuchen Liu, Sirisha Pochareddy, et al. "Transcriptome and epigenome landscape of human cortical development modeled in organoids." Science 362, no. 6420 (December 13, 2018): eaat6720. http://dx.doi.org/10.1126/science.aat6720.
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Genes implicated in neuropsychiatric disorders are active in human fetal brain, yet difficult to study in a longitudinal fashion. We demonstrate that organoids from human pluripotent cells model cerebral cortical development on the molecular level before 16 weeks postconception. A multiomics analysis revealed differentially active genes and enhancers, with the greatest changes occurring at the transition from stem cells to progenitors. Networks of converging gene and enhancer modules were assembled into six and four global patterns of expression and activity across time. A pattern with progressive down-regulation was enriched with human-gained enhancers, suggesting their importance in early human brain development. A few convergent gene and enhancer modules were enriched in autism-associated genes and genomic variants in autistic children. The organoid model helps identify functional elements that may drive disease onset.
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Bessler, N., C. Ruiz Moreno, A. Wezenaar, N. Dommann, F. Keramati, H. C. R. Ariese, C. Honhoff, et al. "P07.07.B INDUCTION AND TARGETING OF DIFFUSE MIDLINE GLIOMA IN A NOVEL HUMAN PONTINE ORGANOID MODEL." Neuro-Oncology 25, Supplement_2 (September 1, 2023): ii52. http://dx.doi.org/10.1093/neuonc/noad137.167.
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Abstract BACKGROUND Diffuse Midline Glioma (DMG) is a rare and aggressive pediatric brain cancer with no chance of survival, highlighting a critical need for therapy development. Disease representative models can aid the search for effective treatments, but human preclinical models that reflect the unique developmental features and anatomical environment of DMG, are so far lacking. Latest research on DMG suggest a fetal neurodevelopmental origin with a stem cell-like cellular profile. Here, we developed a novel pontine hindbrain model to create de novo H3K27M DMG and applied it to study early tumorigenesis and immunotherapy response. MATERIAL AND METHODS Timely sequenced morphogens were applied to developing cerebral organoids, to create a new pontine patterned brain organoid model. Bulk sequencing and 3D imaging over a period of 16 weeks were used to determine ideal patterning conditions, confirm the correct brain regionality and reconstruct cellular developmental dynamics. Introducing H3K27M with common pontine mutations P53 and PDGFRA-D842V via electroporation resulted in de novo H3K27M DMG, which subsequently was characterized based on WHO-histopathological criteria and single cell sequencing. Exploiting this new human DMG model, we applied barcode-based genetic lineage tracing paired with single cell sequencing, to delineate tumorigenesis, and anti-GD2 chimeric antigen receptor (CAR) T cell therapy to investigate CAR T cell responses. RESULTS Our novel pontine hindbrain organoid model inherently gives rise to all relevant macroglia and pons-specific neurons, resembling the same developmental dynamics as seen in humans. Moreover, de novo DMG introduced in these pontine organoids strongly resembles patient cancer, including its cellular human-specific heterogeneity and invasive nature, outperforming existing gold standard PDX and cell line models. As a first model for investigating DMG, the applied barcoded lineage-tracing delineates cancerous transforming precursor states and how they contribute to the diverse lineage among known DMG cancer cell populations. Finally, administration of CAR T cells recreates treatment outcomes observed in patients and demonstrates a high level of CAR T cell heterogeneity. CONCLUSION We generated a bona fide DMG pontine hindbrain organoid model, the first human in vitro model for this specific region of the brain. The matching resemblance to patient tumor paired with the relevant healthy brain environment gives this model the potential for new insights into DMG early tumorigenesis and microenvironmental impact, as well as next generation therapy development.
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Gebing, Philip, Stefanos Loizou, Sebastian Hänsch, Julian Schliehe-Diecks, Lea Spory, Pawel Stachura, Aleksandra Pandyra, et al. "CNS Invasion of TCF3::PBX1+ Leukemia Cells Requires Upregulation of AP-1 Signaling As Revealed By Brain Organoid Model." Blood 142, Supplement 1 (November 28, 2023): 1407. http://dx.doi.org/10.1182/blood-2023-178613.
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Introduction: Involvement of the central nervous system (CNS) remains a challenge in childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Currently, the investigation of CNS leukemia mechanisms relies heavily on 2D cell culture and mouse models. However, given the differences between human and murine CNS in cellular identity and architecture, it becomes crucial to explore alternative models to study CNS leukemia. Moreover, novel targets for diagnosis and treatment of CNS ALL are critically needed. Methods: In this study, we established a pioneering 3D co-culture model that combines human induced pluripotent stem cell (iPSC)-derived cerebral organoids with BCP-ALL cells. We developed new methods to extract data from our invasion assays by enhancing the visualization of 3D organoid images, enabling us to accurately measure the invasion depth of leukemia cells compared to healthy controls within the organoids relative to their surface. To gain further insights on leukemia cells invading the organoids compared to those in the non-invaded fraction, we conducted RNA sequencing and immunofluorescence staining. Subsequently, we validated these results in a BCP-ALL in vivo mouse model and in patients initially diagnosed with CNS-positive BCP-ALL compared to CNS-negative cases within a cohort of 100 BCP-ALL patients. Results: Our experiments demonstrated robust and deep engraftment of TCF3::PBX1+ leukemia cell lines and patient-derived xenograft (PDX) cells into cerebral organoids (within a 14-day of co-culture). In contrast, the engraftment of healthy human CD34+ hematopoietic stem and progenitor cells (HSPCs) was limited ( p < 0.05) as compared to leukemia cells. Utilizing the co-culture model, we successfully validated the targeting of CNS leukemia-relevant pathways, such as CD79a/Igα or CXCR4-SDF1, via genetic knockdown or blocking experiments using inhibitor, respectively, which reduced the invasion of BCP-ALL cells into the organoids. Of note, RNA sequencing and immunofluorescence staining analysis revealed a significant upregulation of members of the AP-1 transcription factor complex, namely FOS, FOSB, and JUN, in the organoid-invading cells. Furthermore, we found an enrichment of AP-1 pathway genes in PDX cells recovered from the CNS compared to spleen blasts of mice transplanted with TCF3::PBX1+ PDX BCP-ALL cells (n = 5), thereby supporting the critical role of AP-1 signaling in CNS disease (Figure 1A). In line with these findings, we observed significantly higher levels of the AP-1 gene JUN in patients initially diagnosed with CNS-positive BCP-ALL compared to CNS-negative cases (mean JUN expression: 633.4 ± 78.51 in CNS-negative vs 1142 ± 296.5 in CNS-positive) within a cohort of 100 BCP-ALL patients (Figure 1B). Summary: In summary, we present ALL co-culture systems with iPSC-derived cerebral organoids as a promising complementary model to investigate CNS involvement in BCP-ALL including therapeutic targeting approaches, and identified the AP-1 pathway as a marker of CNS disease in TCF3::PBX1+ BCP-ALL.
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Gallego Villarejo, Lucia, Wanda M. Gerding, Lisa Bachmann, Luzie H. I. Hardt, Stefan Bormann, Huu Phuc Nguyen, and Thorsten Müller. "Optical Genome Mapping Reveals Genomic Alterations upon Gene Editing in hiPSCs: Implications for Neural Tissue Differentiation and Brain Organoid Research." Cells 13, no. 6 (March 14, 2024): 507. http://dx.doi.org/10.3390/cells13060507.
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Genome editing, notably CRISPR (cluster regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9), has revolutionized genetic engineering allowing for precise targeted modifications. This technique’s combination with human induced pluripotent stem cells (hiPSCs) is a particularly valuable tool in cerebral organoid (CO) research. In this study, CRISPR/Cas9-generated fluorescently labeled hiPSCs exhibited no significant morphological or growth rate differences compared with unedited controls. However, genomic aberrations during gene editing necessitate efficient genome integrity assessment methods. Optical genome mapping, a high-resolution genome-wide technique, revealed genomic alterations, including chromosomal copy number gain and losses affecting numerous genes. Despite these genomic alterations, hiPSCs retain their pluripotency and capacity to generate COs without major phenotypic changes but one edited cell line showed potential neuroectodermal differentiation impairment. Thus, this study highlights optical genome mapping in assessing genome integrity in CRISPR/Cas9-edited hiPSCs emphasizing the need for comprehensive integration of genomic and morphological analysis to ensure the robustness of hiPSC-based models in cerebral organoid research.
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Damodharan, Sudarshawn, Peter Favreau, Connie Lebakken, and Mahua Dey. "BIOL-19. DIFFUSE MIDLINE GLIOMA CEREBRAL ORGANOID MODEL AND MULTIOMICS CHARACTERIZATION." Neuro-Oncology 25, Supplement_1 (June 1, 2023): i10. http://dx.doi.org/10.1093/neuonc/noad073.038.
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Abstract Diffuse midline glioma (DMG) are highly aggressive malignancies of the central nervous system that primarily affect the pediatric population. These tumors are historically universally fatal with no curative treatment options available. There is a need to identify more targeted and optimal treatments for these patients. Current approaches to pre-clinical therapeutic testing have been limited by many obstacles to effectively translate theses to patients. It is known that the interactions between tumors and the other components of the tumor microenvironment (TME) can change the response to therapeutic interventions. This especially holds true for brain tumors and the complex neural network encompassed within their TME. Given this, it is crucial to develop more realistic DMG models that integrate this to conduct therapeutic testing rather than relying upon conventional cell culture models. The goal of our study was to develop a three-dimensional DMG cerebral organoid model derived from human induced pluripotent stem cells (iPSCs) co-cultured with three different DMG patient-derived xenograft (PDX) cell lines to better mimic the TME for therapeutic testing. We were able to successfully integrate our three cell lines into the cerebral organoids, capturing TME interactions along with performing multiomic profiling for better characterization. We next plan to perform therapeutic testing to further validate the model and improve preclinical drug screening for DMG.
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Singh, Sanjay, Maxime Munyeshyaka, Joy Gumin, Jing Yang, Daniel Ledbetter, Anwar Hossain, Brittany Parker Kerrigan, and Frederick Lang. "TAMI-24. BEHAVIOR OF GLIOBLASTOMA STEM-LIKE CELLS WITH KNOWN IDH1 STATUS IN CEREBRAL ORGANOIDS." Neuro-Oncology 22, Supplement_2 (November 2020): ii218. http://dx.doi.org/10.1093/neuonc/noaa215.913.
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Abstract Glioblastomas (GBM) exhibit high proliferative index, areas of necrosis, high vascularization, and are highly invasive to normal brain tissues. The most common and lethal form of GBMs are primary GBMs, with no prior clinical history. Whereas, secondary GBMs arise from low-grade gliomas and are associated with IDH1 mutation. Pre-clinical studies of GBM largely depend on patient-derived GBM stem-like cells (GSCs) in vitro and in vivo as orthotopic xenografts. Cerebral organoids (COs) derived from induced pluripotent stem cells can serve as allogenic in vitro model systems to study interactions between normal brain and GSCs. COs have been shown to harbor neural stem cells and their differentiated progenies as well as microglia within distinct niches. Here, we co-cultured 45 day-old COs and MDA-GSCs lines representing mesenchymal sub-group (M-MDA-GSC), classical sub-group (C-MDA-GSC), and IDH1 mutant (IDH1R132H-MDA-GSC). MDA-GSCs stably express fluorescent proteins and is used to track GSCs within COs. These GSC bearing COs were fixed, embedded, sectioned, immuno-stained, and imaged by confocal microscope. There was a positive correlation between GSC numbers in allografted niche and invasion into COs as measured from the edge of organoid, M-MDA-GSC (R2=0.99; 0.89μm/cell), C-MDA-GSC (R2=0.92; 0.66μm/cell), and IDH1R132H-MDA-GSC (R2=0.89; 0.5μm/cell). Additionally, M-MDA-GSCs had significantly high percentage of Ki67+ve invasive cells (24%) in comparison to C-MDA-GSCs (5.1%; p=0.0057). As a measure of interaction of MDA-GSC with normal cells, we assessed proximity of IBA1+ve microglia in GSC niche within organoids and show that M-MDA-GSC and IDH1R132H-MDA-GSC highly co-localized with IBA1+ve microglia on day12 of co-culture. In conclusion, our cerebral organoid-based allograft study shows that mesenchymal GSCs (M-MDA-GSC) are most invasive whereas IDH1 mutant GSCs (IDH1R132H-MDA-GSC) are least invasive. C-MDA-GSCs are least proliferative while invading into normal COs. Uniqueness of CO based allograft system is highlighted by observed similarity between M-MDA-GSC and IDH1R132H-MDA-GSC for their potential to attract IBA1+ve microglia.
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Acharya, Prabha, Pranav Joshi, Sunil Shrestha, Na Young Choi, Sehoon Jeong, and Moo-Yeal Lee. "Uniform cerebral organoid culture on a pillar plate by simple and reproducible spheroid transfer from an ultralow attachment well plate." Biofabrication, January 4, 2024. http://dx.doi.org/10.1088/1758-5090/ad1b1e.
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Abstract Human induced pluripotent stem cell (iPSCs)-derived brain organoids have potential to recapitulate the earliest stages of brain development, serving as an effective in vitro model for studying both normal brain development and disorders. However, current brain organoid culture methods face several challenges, including low throughput, high variability in organoid generation, and time-consuming, multiple transfer and encapsulation of cells in hydrogels throughout the culture. These limitations hinder the widespread application of brain organoids including high-throughput assessment of compounds in clinical and industrial lab settings. In this study, we demonstrate a straightforward approach of generating multiple cerebral organoids from iPSCs on a pillar plate platform, eliminating the need for labor-intensive, multiple transfer and encapsulation steps to ensure the reproducible generation of cerebral organoids. We formed embryoid bodies (EBs) in an ultra-low attachment (ULA) 384-well plate and subsequently transferred them to the pillar plate containing Matrigel, using a straightforward sandwiching and inverting method. Each pillar on the pillar plate contains a single spheroid, and the success rate of spheroid transfer was in a range of 95 - 100%. Using this approach, we robustly generated cerebral organoids on the pillar plate and demonstrated an intra-batch coefficient of variation (CV) below 9 - 19% based on ATP-based cell viability and compound treatment. Notably, our spheroid transfer method in combination with the pillar plate allows miniaturized culture of cerebral organoids, alleviates the issue of organoid variability, and has potential to significantly enhance assay throughput by allowing in situ organoid assessment as compared to conventional organoid culture in 6-/24-well plates, petri dishes, and spinner flasks.
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Sozzi, Edoardo, Janko Kajtez, Andreas Bruzelius, Milan Finn Wesseler, Fredrik Nilsson, Marcella Birtele, Niels B. Larsen, et al. "Silk scaffolding drives self-assembly of functional and mature human brain organoids." Frontiers in Cell and Developmental Biology 10 (October 14, 2022). http://dx.doi.org/10.3389/fcell.2022.1023279.
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Human pluripotent stem cells (hPSCs) are intrinsically able to self-organize into cerebral organoids that mimic features of developing human brain tissue. These three-dimensional structures provide a unique opportunity to generate cytoarchitecture and cell-cell interactions reminiscent of human brain complexity in a dish. However, current in vitro brain organoid methodologies often result in intra-organoid variability, limiting their use in recapitulating later developmental stages as well as in disease modeling and drug discovery. In addition, cell stress and hypoxia resulting from long-term culture lead to incomplete maturation and cell death within the inner core. Here, we used a recombinant silk microfiber network as a scaffold to drive hPSCs to self-arrange into engineered cerebral organoids. Silk scaffolding promoted neuroectoderm formation and reduced heterogeneity of cellular organization within individual organoids. Bulk and single cell transcriptomics confirmed that silk cerebral organoids display more homogeneous and functionally mature neuronal properties than organoids grown in the absence of silk scaffold. Furthermore, oxygen sensing analysis showed that silk scaffolds create more favorable growth and differentiation conditions by facilitating the delivery of oxygen and nutrients. The silk scaffolding strategy appears to reduce intra-organoid variability and enhances self-organization into functionally mature human brain organoids.
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Barnhart, Andrew J., and Kris Dierickx. "Cultures and cures: neurodiversity and brain organoids." BMC Medical Ethics 22, no. 1 (May 17, 2021). http://dx.doi.org/10.1186/s12910-021-00627-1.
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Abstract Background Research with cerebral organoids is beginning to make significant progress in understanding the etiology of autism spectrum disorder (ASD). Brain organoid models can be grown from the cells of donors with ASD. Researchers can explore the genetic, developmental, and other factors that may give rise to the varieties of autism. Researchers could study all of these factors together with brain organoids grown from cells originating from ASD individuals. This makes brain organoids unique from other forms of ASD research. They are like a multi-tool, one with significant versatility for the scope of ASD research and clinical applications. There is hope that brain organoids could one day be used for precision medicine, like developing tailored ASD drug treatments. Main body Brain organoid researchers often incorporate the medical model of disability when researching the origins of ASD, especially when the research has the specific aim of potentially finding tailored clinical treatments for ASD individuals. The neurodiversity movement—a developmental disability movement and paradigm that understands autism as a form of natural human diversity—will potentially disagree with approaches or aims of cerebral organoid research on ASD. Neurodiversity advocates incorporate a social model of disability into their movement, which focuses more on the social, attitudinal, and environmental barriers rather than biophysical or psychological deficits. Therefore, a potential conflict may arise between these perspectives on how to proceed with cerebral organoid research regarding neurodevelopmental conditions, especially ASD. Conclusions Here, we present these perspectives and give at least three initial recommendations to achieve a more holistic and inclusive approach to cerebral organoid research on ASD. These three initial starting points can build bridges between researchers and the neurodiversity movement. First, neurodiverse individuals should be included as co-creators in both the scientific process and research communication. Second, clinicians and neurodiverse communities should have open and respectful communication. Finally, we suggest a continual reconceptualization of illness, impairment, disability, behavior, and person.
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Hu, Daiyu, Yuanqing Cao, Chenglin Cai, Guangming Wang, Min Zhou, Luying Peng, Yantao Fan, Qiong Lai, and Zhengliang Gao. "Establishment of human cerebral organoid systems to model early neural development and assess the central neurotoxicity of environmental toxins." Neural Regeneration Research, January 31, 2024. http://dx.doi.org/10.4103/nrr.nrr-d-23-00928.
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Abstract Human brain development is a complex process, and animal models often have significant limitations. To address this, researchers have developed pluripotent stem cell-derived three-dimensional structures, known as brain-like organoids, to more accurately model early human brain development and disease. To enable more consistent and intuitive reproduction of early brain development, in this study, we incorporated forebrain organoid culture technology into the traditional unguided method of brain organoid culture. This involved embedding organoids in matrix glue for only 7 days during the rapid expansion phase of the neural epithelium and then removing them from the matrix glue for further cultivation, resulting in a new type of human brain organoid system. This cerebral organoid system replicated the temporospatial characteristics of early human brain development, including neuroepithelium derivation, neural progenitor cell production and maintenance, neuron differentiation and migration, and cortical layer patterning and formation, providing more consistent and reproducible organoids for developmental modeling and toxicology testing. As a proof of concept, we applied the heavy metal cadmium to this newly improved organoid system to test whether it could be used to evaluate the neurotoxicity of environmental toxins. Brain organoids exposed to cadmium for 7 or 14 days manifested severe damage and abnormalities in their neurodevelopmental patterns, including bursts of cortical cell death and premature differentiation. Cadmium exposure caused progressive depletion of neural progenitor cells and loss of organoid integrity, accompanied by compensatory cell proliferation at ectopic locations. The convenience, flexibility, and controllability of this newly developed organoid platform make it a powerful and affordable alternative to animal models for use in neurodevelopmental, neurological, and neurotoxicological studies.
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Xue, Jun, Youjun Chu, Yanwang Huang, Ming Chen, Meng Sun, Zhiqin Fan, Yonghe Wu, and Liang Chen. "A tumorigenicity evaluation platform for cell therapies based on brain organoids." Translational Neurodegeneration 13, no. 1 (October 29, 2024). http://dx.doi.org/10.1186/s40035-024-00446-5.
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Abstract Background Tumorigenicity represents a critical challenge in stem cell-based therapies requiring rigorous monitoring. Conventional approaches for tumorigenicity evaluation are based on animal models and have numerous limitations. Brain organoids, which recapitulate the structural and functional complexity of the human brain, have been widely used in neuroscience research. However, the capacity of brain organoids for tumorigenicity evaluation needs to be further elucidated. Methods A cerebral organoid model produced from human pluripotent stem cells (hPSCs) was employed. Meanwhile, to enhance the detection sensitivity for potential tumorigenic cells, we created a glioblastoma-like organoid (GBM organoid) model from TP53−/−/PTEN−/− hPSCs to provide a tumor microenvironment for injected cells. Midbrain dopamine (mDA) cells from human embryonic stem cells were utilized as a cell therapy product. mDA cells, hPSCs, mDA cells spiked with hPSCs, and immature mDA cells were then injected into the brain organoids and NOD SCID mice. The injected cells within the brain organoids were characterized, and compared with those injected in vivo to evaluate the capability of the brain organoids for tumorigenicity evaluation. Single-cell RNA sequencing was performed to identify the differential gene expression between the cerebral organoids and the GBM organoids. Results Both cerebral organoids and GBM organoids supported maturation of the injected mDA cells. The hPSCs and immature mDA cells injected in the GBM organoids showed a significantly higher proliferative capacity than those injected in the cerebral organoids and in NOD SCID mice. Furthermore, the spiked hPSCs were detectable in both the cerebral organoids and the GBM organoids. Notably, the GBM organoids demonstrated a superior capacity to enhance proliferation and pluripotency of spiked hPSCs compared to the cerebral organoids and the mouse model. Kyoto Encyclopedia of Genes and Genomes analysis revealed upregulation of tumor-related metabolic pathways and cytokines in the GBM organoids, suggesting that these factors underlie the high detection sensitivity for tumorigenicity evaluation. Conclusions Our findings suggest that brain organoids could represent a novel and effective platform for evaluating the tumorigenic risk in stem cell-based therapies. Notably, the GBM organoids offer a superior platform that could complement or potentially replace traditional animal-based models for tumorigenicity evaluation.
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Acharya, Prabha, Sunil Shrestha, Pranav Joshi, Na Young Choi, Vinod Kumar Reddy Lekkala, Soo-Yeon Kang, Gabriel Ni, and Moo-Yeal Lee. "Dynamic culture of cerebral organoids using a pillar/perfusion plate for the assessment of developmental neurotoxicity." Biofabrication, October 14, 2024. http://dx.doi.org/10.1088/1758-5090/ad867e.
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Abstract Despite the potential toxicity of commercial chemicals to the development of the nervous system (known as developmental neurotoxicity or DNT), conventional in vitro cell models have primarily been employed for the assessment of acute neuronal toxicity. On the other hand, animal models used for the assessment of DNT are not physiologically relevant due to the heterogenic difference between humans and animals. In addition, animal models are low-throughput, time-consuming, expensive, and ethically questionable. Recently, human brain organoids have emerged as a promising alternative to assess the detrimental effects of chemicals on the developing brain. However, conventional organoid culture systems have several technical limitations including low throughput, lack of reproducibility, insufficient maturity of organoids, and the formation of the necrotic core due to limited diffusion of nutrients and oxygen. To address these issues and establish predictive DNT models, cerebral organoids were differentiated in a dynamic condition in a unique pillar/perfusion plate, which were exposed to test compounds to evaluate DNT potential. The pillar/perfusion plate facilitated uniform, dynamic culture of cerebral organoids with improved proliferation and maturity by rapid, bidirectional flow generated on a digital rocker. Day 9 cerebral organoids in the pillar/perfusion plate were exposed to ascorbic acid (DNT negative) and methylmercury (DNT positive) in a dynamic condition for 1 and 3 weeks, and changes in organoid morphology and neural gene expression were measured to determine DNT potential. As expected, ascorbic acid didn’t induce any changes in organoid morphology and neural gene expression. However, exposure of day 9 cerebral organoids to methylmercury resulted in significant changes in organoid morphology and neural gene expression. Interestingly, methylmercury did not induce adverse changes in cerebral organoids in a static condition, thus highlighting the importance of dynamic organoid culture in DNT assessment.