The role of brain organoids as model system for human disease

ACHAIKI IATRIKI | 2022; 41(2): 69–72


Christina Kyrousi1,2, Stavros Taraviras3

1First Department of Psychiatry, Medical School, National and Kapodistrian University of Athens, Eginition Hospital, Greece
2University Mental Health, Neurosciences and Precision Medicine Research Institute “Costas Stefanis”, Athens, Greece
3Department of Physiology, Medical School, University of Patras, Greece

Received: 21 Dec 2021; Accepted: 17 Feb 2022

Corresponding author: Christina Kyrousi, Assistant Professor, First Department of Psychiatry, Medical School, National and Kapodistrian University of Athens, Eginition Hospital, Greece and Affiliated Scientist, University Mental Health, Neurosciences and Precision Medicine Research Institute “Costas Stefanis”, Athens, Greece, E-mail:
Stavros Taraviras, Professor, Department of Physiology, Medical School, University of Patras, Greece, E-mail:

Key words: Brain organoids, human cortical development, brain disease modeling, cortical malformations


The human cerebral cortex represents the most highly developed part of the brain. It plays a vital role in processing and integrating information, modulating social and motor behaviors, planning and organization and thus it determines intelligence and personality in humans. Hence, the series of events resulting in its development must be tightly coordinated and regulated. Smaller or bigger changes or disruptions in the regulation of proliferation, differentiation, and migration of cells in the developing central nervous system may lead to malformations of the brain affecting its structure and function. This can cause a wide range of physiological and functional consequences, provoking brain-related diseases such as neurodevelopmental disorders. The main characteristics of such disorders are developmental delay, intellectual disability, and epilepsy. They can also be associated with psychiatric disorders affecting individuals from early postnatal life and throughout adulthood. Given the high societal and economic burden that such disorders impose, defining the pathophysiological mechanisms underlying their manifestation will help to better diagnose and will accelerate treatment. For this reason, over recent years scientists have made a significant effort to model brain diseases.

Mouse models revealed many aspects of the mechanism underlying proper cortical development, as well as the appearance of cortical malformations; however, their use is limited due to structural and functional differences between mice and humans. The latest advances in stem cell technology and the generation of induced pluripotent stem cells (iPSCs) offer a promising way to derive human cells of any tissue of interest from patients and control individuals to study the phenotype of patients affected by disease-causing mutations. The originally developed protocols yielded two-dimensional (2D) monolayer cultures of human neural progenitors and neurons, and were a big step forward in identifying human-specific molecular and cellular mechanisms related to brain development and disease. However, they did not allow insights into the effects of three-dimensional (3D) tissue context on cellular processes, a key feature that determines brain function, while the lack of cellular and molecular diversity was profound in such cultures. On the other hand, organoids offer a possibility to overcome these problems since they represent 3D, embryonic structures that reflect the 3D structure of organs. Brain organoids have been shown to reflect the 3D organization, cell-type composition, and transcriptional footprints of the developing human brain. For these reasons, in the past decade, such brain organoid protocols have been used to model many diseases and they are now representing a promising model system [1].

Brain organoids are characterized by high complexity in terms of cellular composition, as they consist of neural progenitors, neurons, astrocytes, and oligodendrocytes, by structural diversity, as they are organized in different cellular layers and by a higher degree of maturation than 2D cultures. All of these features allow regional interconnectivity and function similar to those observed in the human brain. Several different protocols for generating brain organoids have been published in the past decade. They are based either on intrinsic properties of neural progenitor cells to self-organize into 3D aggregates, or on guided differentiation programs engineering the external environment of the 3D aggregates which is achieved through the addition of morphogens mimicking endogenous patterning events. The first approach leads to the production of various cellular lineages yielding brain organoids composed of multiple regional identities of the brain within the same organoid. This allows the holistic modeling of brain structure, but it was reported to show increased variability between batches [2]. On the contrary, the second approach drives neural progenitors to acquire a specific brain region identity, which was proposed to reduce the variability between different organoid batches. However, they can be used only for specific applications because they lack complexity [2]. Over the last few years, different modifiers, namely small molecules, have been used to produce forebrain organoids (dorsal and ventral), midbrain organoids, hypothalamic or thalamic organoids, hippocampal organoids, spinal cord organoids, cerebellum organoids, and choroid plexus organoids, reviewed in [3]. In parallel, differently patterned organoids have been fused creating more complex models of the developing human nervous system leading to modeling interconnectivity in a tightly regulated approach [3] and they have been used as an alternative method of the intrinsic protocols. Nevertheless, these basic differences between these approaches need to be considered when choosing the appropriate 3D model to study different brain diseases.

These protocols were used to model early human CNS development, neuronal survival and maturation, human brain evolution, and human brain diseases [1]. Indeed, since the publication of the first intrinsic brain organoid protocol [4], numerous studies have been published modeling a great variety of different brain-related diseases. Amongst the first diseases that have been modeled were the malformations of cortical development (MCDs), such as microcephaly, macrocephaly, cortical heterotopias, and lissencephaly. Interestingly, structural defects of the developing cortex are among the main clinical phenotypes in the previously mentioned diseases. Using mainly the intrinsic protocols for generating brain organoids, a human-specific mechanism involving the proper regulation of the mitotic spindle orientation in the transition from apical radial glial cells to basal radial glial cells, the novel neural progenitors responsible for the neuronal expansion observed in the human cortex, was described in patients with microcephaly following mutations in genes such as CDK5RAP2 [4] and ASPM [5] or after zika virus infection [6]. Besides, macrocephaly was also modeled using brain organoids as well as disorders implicating alterations in the gyrification index of the brain scrutinizing the human-specific function of genes including PTEN, LIS1 and YWHAE [7-9]. Finally, neuronal heterotopias were also extensively studied using brain organoids contributing to our limited knowledge of the involvement of intrinsic and extrinsic signaling on neural progenitors’ function, and neuronal migration profile in the formation of the human cortex. These studies have shown that the morphology, position, and function of neural progenitors, as well as the migration behavior of human neurons during cortical development, are regulated amongst others by DCHS1, FAT4, and LGALS3BP [10,11] contributing to the establishment of human cortical complexity.

Besides cortical malformations, other brain-related neurodevelopmental disorders have been modeled including αutism spectrum disorder (ASD) [12,13], Rett syndrome [14], and Timothy syndrome [15]. Using brain organoids, the hypothesis of the excitatory/inhibitory imbalance in autistic brains has been tested and the involvement of genes such as FOGX1 and CHD8 has been shown. Additionally, using assembloids, interneuron migration defects were suggested as one of the causing mechanisms of Timothy syndrome, while brain organoids harboring mutations in the gene MECP2 showed defects such as increased proliferation and decreased differentiation potentials of neural progenitors suggesting a novel mechanism for Rett Syndrome. Lastly, although brain organoids were shown to recapitulate early steps of brain development, modified protocols have been used for modeling neurodegenerative disorders including Alzheimer’s disease (AD) [16], Amyotrophic lateral sclerosis (ALS) [17], Parkinson’s disease (PD) [18] schizophrenia [19] and others. This enabled the modeling of these genetic neurodegenerative diseases in a human cellular context, highlighting for example i) the cellular mechanisms involved in the accumulation of amyloidogenic Aβ peptides in AD, ii) the impaired motor features upon neuronal degradation in ALS and iii) the decreased neurite length of dopaminergic neurons in LRRK2 mutant (PD) organoids. Of note, these cellular systems have highlighted potential developmental deficits underlined classical neurodegenerative disorders. For instance, in schizophrenia cellular mechanisms, such as cell cycle control dysregulation of the key neural progenitor type, the radial glial cells, were described as a consequence of the disruption of DISC1 and NDEL1 interaction. This could shape a novel understanding of the causes of neurodegenerative disorders that may contribute to changes in diagnosis and therapeutic strategies in the future.

From all the above, it is clear that with the use of brain organoids we were able to describe human-specific mechanisms that upon disruption lead to MCDs, neurodevelopmental or neurodegenerative disorders. Nevertheless, it is not always easy to estimate where the limitations of the organoids lie. Even though many studies have shown remarkable similarities between brain organoids and the fetal brain, it is also clear that not all aspects of brain development are accurately reflected in these in vitro cultures. For instance, it has been reported that the proportion of several cell types is altered in organoids compared to the primary tissue. Indeed, single-cell-RNA-sequencing, immunofluorescence, and FACS analysis from several labs have shown that the proportion of glial cells – astrocytes and oligodendrocytes – is lower, same as the ratio of apical and basal progenitors in the neurogenic zones. Furthermore, endothelial cells are missing and white matter regions are also very much underrepresented within the organoids. In addition, due to the absence of vascularization, the organoid’s nutrient and oxygen supply are suboptimal and thus their size and stochasticity of developmental fate choices remain limited. These limitations are the driving force for future improvements of organoid cultures. Attempts for such improvement are already in line, such as the addition of microfilaments and scaffolding components or the generation of the “organoids on a chip”, micro-engineered promising models that will allow the control and manipulation of fluid flow with a high degree of accuracy [20]. These will assist the improvement of the morphology and nutrient support of organoids, which ultimately will ameliorate organoid cultures and will ensure the high quality of this model. Regardless of the remaining limitations from the use of organoids, it is widely accepted that they represent one of the best models so far for studying human-specific mechanisms of brain development and disease and constitute an evolutionary approach which opens new avenues in the diagnosis and treatment of brain-related disorders.

Conflict of interest disclosure

None to declare

Declaration of funding sources

None to declare

Author contribution

CK, ST: conceptualization, writing original draft, writing review and editing manuscript


1. Kyrousi C, Cappello S. Using brain organoids to study human neurodevelopment, evolution and disease. Wiley Interdiscip Rev Dev Biol. 2020;9(1):e347.
2. Yoon SJ, Elahi LS, Pașca AM, Marton RM, Gordon A, Revah O, et al. Reliability of human cortical organoid generation. Nat Methods. 2019;16(1):75-8.
3. Bhattacharya A, Choi WWY, Muffat J, Li Y. Modeling developmental brain diseases using human pluripotent stem cell-derived brain organoids – progress and perspective. J Mol Biol. 2022;434(3):167386.
4. Lancaster MA, Renner M, Martin CA, Wenzel D, Bicknell LS, Hurles ME, et al. Cerebral organoids model human brain development and microcephaly. Nature. 2013;501(7467):373-9.
5. Li R, Sun L, Fang A, Li P, Wu Q, Wang X. Recapitulating cortical development with organoid culture in vitro and modeling abnormal spindle-like (ASPM related primary) microcephaly disease. Protein Cell. 2017;8(11):823-33.
6. Qian X, Nguyen HN, Jacob F, Song H, Ming GL. Using brain organoids to understand Zika virus-induced microcephaly. Development. 2017;144(6):952-7.
7. Li Y, Muffat J, Omer A, Bosch I, Lancaster MA, Sur M, et al. Induction of Expansion and Folding in Human Cerebral Organoids. Cell Stem Cell. 2017;20(3):385-96.e3.
8. Bershteyn M, Nowakowski TJ, Pollen AA, Di Lullo E, Nene A, Wynshaw-Boris A, et al. Human iPSC-Derived Cerebral Organoids Model Cellular Features of Lissencephaly and Reveal Prolonged Mitosis of Outer Radial Glia. Cell Stem Cell. 2017;20(4):435-49.e4.
9. Iefremova V, Manikakis G, Krefft O, Jabali A, Weynans K, Wilkens R, et al. An Organoid-Based Model of Cortical Development Identifies Non-Cell-Autonomous Defects in Wnt Signaling Contributing to Miller-Dieker Syndrome. Cell Rep. 2017;19(1):50-9.
10. Klaus J, Kanton S, Kyrousi C, Ayo-Martin AC, Di Giaimo R, Riesenberget S, al. Altered neuronal migratory trajectories in human cerebral organoids derived from individuals with neuronal heterotopia. Nat Med. 2019;25(4):561-8.
11. Kyrousi C, O’Neill AC, Brazovskaja A, He Z, Kielkowski P, Coquand L, et al. Extracellular LGALS3BP regulates neural progenitor position and relates to human cortical complexity. Nat Commun. 2021;12(1):6298.
12. Mariani J, Coppola G, Zhang P, Abyzov A, Provini L, Tomasini L, et al. FOXG1-Dependent Dysregulation of GABA/Glutamate Neuron Differentiation in Autism Spectrum Disorders. Cell. 2015;162(2):375-90.
13. Wang P, Mokhtari R, Pedrosa E, Kirschenbaum M, Bayrak C, Zheng D, et al. CRISPR/Cas9-mediated heterozygous knockout of the autism gene CHD8 and characterization of its transcriptional networks in cerebral organoids derived from iPS cells. Mol Autism. 2017;8:11.
14. Mellios N, Feldman D, Sheridan S, Ip JPK, Kwok S, Amoah SK, et al. Human cerebral organoids reveal deficits in neurogenesis and neuronal migration in MeCP2-deficient neural progenitors. Mol Psychiatry. 2018;23:791.
15. Birey F, Andersen J, Makinson CD, Islam S, Wei W, Huberet N, et al. Assembly of functionally integrated human forebrain spheroids. Nature. 2017;545(7652):54-9.
16. Gonzalez C, Armijo E, Bravo-Alegria J, Becerra-Calixto A, Mays CE, Soto C. Modeling amyloid beta and tau pathology in human cerebral organoids. Mol Psychiatry. 2018;23(12):2363-74.
17. Osaki T, Uzel SGM, Kamm RD. Microphysiological 3D model of amyotrophic lateral sclerosis (ALS) from human iPS-derived muscle cells and optogenetic motor neurons. Sci Adv. 2018;4(10):eaat5847.
18. Kim H, Park HJ, Choi H, Chang Y, Park H, Shin J, et al. Modeling G2019S-LRRK2 Sporadic Parkinson’s Disease in 3D Midbrain Organoids. Stem Cell Reports. 2019;12(3):518-31.
19. Ye F, Kang E, Yu C, Qian X, Jacob F, Yu C, et al. DISC1 Regulates Neurogenesis via Modulating Kinetochore Attachment of Ndel1/Nde1 during Mitosis. Neuron. 2017;96(5):1204.
20. Qiao H, Zhang YS, Chen P. Commentary: Human brain organoid-on-a-chip to model prenatal nicotine exposure. Front Bioeng Biotechnol. 2018 ;6:138.