Human Brain Development Research Articles

Brain organoids for hypoxic-ischemic studies: from bench to bedside

Our current knowledge regarding the development of the human brain mostly derives from experimental studies on non-human primates, sheep, and rodents. However, these studies may not completely simulate all the features of human brain development as a result of species differences and variations in pre- and postnatal brain maturation. Therefore, it is important to supplement the in vivo animal models to increase the possibility that preclinical studies have appropriate relevance for potential future human trials. Three-dimensional brain organoid culture technology could complement in vivo animal studies to enhance the translatability of the preclinical animal studies and the understanding of brain-related disorders. In this review, we focus on the development of a model of hypoxic-ischemic (HI) brain injury using human brain organoids to complement the translation from animal experiments to human pathophysiology. We also discuss how the development of these tools provides potential opportunities to study fundamental aspects of the pathophysiology of HI-related brain injury including differences in the responses between males and females.

Thyroid hormone action during GABAergic neuron maturation: The quest for mechanisms.

Thyroid hormone (TH) signaling plays a major role in mammalian brain development. Data obtained in the past years in animal models have pinpointed GABAergic neurons as a major target of TH signaling during development, which opens up new perspectives to further investigate the mechanisms by which TH affects brain development. The aim of the present review is to gather the available information about the involvement of TH in the maturation of GABAergic neurons. After giving an overview of the kinds of neurological disorders that may arise from disruption of TH signaling during brain development in humans, we will take a historical perspective to show how rodent models of hypothyroidism have gradually pointed to GABAergic neurons as a main target of TH signaling during brain development. The third part of this review underscores the challenges that are encountered when conducting gene expression studies to investigate the molecular mechanisms that are at play downstream of TH receptors during brain development. Unravelling the mechanisms of action of TH in the developing brain should help make progress in the prevention and treatment of several neurological disorders, including autism and epilepsy.

Tissue morphology influences the temporal program of human brain organoid development.

Progression through fate decisions determines cellular composition and tissue architecture, but how that same architecture may impact cell fate is less clear. We took advantage of organoids as a tractable model to interrogate this interaction of form and fate. Screening methodological variations revealed that common protocol adjustments impacted various aspects of morphology, from macrostructure to tissue architecture. We examined the impact of morphological perturbations on cell fate through integrated single nuclear RNA sequencing (snRNA-seq) and spatial transcriptomics. Regardless of the specific protocol, organoids with more complex morphology better mimicked invivo human fetal brain development. Organoids with perturbed tissue architecture displayed aberrant temporal progression, with cells being intermingled in both space and time. Finally, encapsulation to impart a simplified morphology led to disrupted tissue cytoarchitecture and a similar abnormal maturational timing. These data demonstrate that cells of the developing brain require proper spatial coordinates to undergo correct temporal progression.

A loss-of-function mutation in human Oxidation Resistance 1 disrupts the spatial–temporal regulation of histone arginine methylation in neurodevelopment

BackgroundOxidation Resistance 1 (OXR1) gene is a highly conserved gene of the TLDc domain-containing family. OXR1 is involved in fundamental biological and cellular processes, including DNA damage response, antioxidant pathways, cell cycle, neuronal protection, and arginine methylation. In 2019, five patients from three families carrying four biallelic loss-of-function variants in OXR1 were reported to be associated with cerebellar atrophy. However, the impact of OXR1 on cellular functions and molecular mechanisms in the human brain is largely unknown. Notably, no human disease models are available to explore the pathological impact of OXR1 deficiency.ResultsWe report a novel loss-of-function mutation in the TLDc domain of the human OXR1 gene, resulting in early-onset epilepsy, developmental delay, cognitive disabilities, and cerebellar atrophy. Patient lymphoblasts show impaired cell survival, proliferation, and hypersensitivity to oxidative stress. These phenotypes are rescued by TLDc domain replacement. We generate patient-derived induced pluripotent stem cells (iPSCs) revealing impaired neural differentiation along with dysregulation of genes essential for neurodevelopment. We identify that OXR1 influences histone arginine methylation by activating protein arginine methyltransferases (PRMTs), suggesting OXR1-dependent mechanisms regulating gene expression during neurodevelopment. We model the function of OXR1 in early human brain development using patient-derived brain organoids revealing that OXR1 contributes to the spatial–temporal regulation of histone arginine methylation in specific brain regions.ConclusionsThis study provides new insights into pathological features and molecular underpinnings associated with OXR1 deficiency in patients.

Modeling early human cortical development and evaluating neurotoxicity with a forebrain organoid system

The complexity and subtlety of brain development renders it challenging to examine effects of environmental toxicants on human fetal brain development. Advances in pluripotent cell-derived organoid systems open up novel avenues for human development, disease and toxicity modeling. Here, we have established a forebrain organoid system and recapitulated early human cortical development spatiotemporally including neuroepithelium induction, apical-basal axis formation, neural progenitor proliferation and maintenance, neuronal differentiation and layer/region patterning. To explore whether this forebrain organoid system is suitable for neurotoxicity modeling, we subjected the organoids to bisphenol A (BPA), a common environmental toxicant of global presence and high epidemic significance. BPA exposure caused substantial abnormalities in key cortical developmental events, inhibited progenitor cell proliferation and promoted precocious neuronal differentiation, leading premature progenitor cell depletion and aberrant cortical layer patterning and structural organization. Consistent with an antagonistic mechanism between thyroid hormone and BPA, T3 supplementation attenuated BPA-mediated cortical developmental abnormalities. Altogether, our in vitro recapitulation of cortical development with forebrain organoids provides a paradigm for efficient neural development and toxicity modeling and related remedy testing/screening.

The diploic venous system in Homo neanderthalensis and fossil Homo sapiens: A study using high-resolution computed tomography.

The diploic venous system has been hypothesized to be related to human brain evolution, though its evolutionary trajectory and physiological functions remain largely unclear. This study examines the characteristics of the diploic venous channels (DCs) in a selection of well-preserved Homo neanderthalensis and Upper Paleolithic Homo sapiens crania, searching for the differences between the two taxa and exploring the associations between brain anatomy and DCs. Five H. neanderthalensis and four H. sapiens fossil specimens from Western Europe were analyzed. Based on Micro-CT scanning and 3D reconstruction, the distribution pattern and draining orifices of the DCs were inspected qualitatively. The size of the DCs was quantified by volume calculation, and the degree of complexity was quantified by fractal analyses. High-resolution data show the details of the DC structures not documented in previous studies. H. neanderthalensis and H. sapiens specimens share substantial similarities in the DCs. The noticeable differences between the two samples manifest in the connecting points surrounding the frontal sinuses, parietal foramina, and asterional area. This study provides a better understanding of the anatomy of the DCs in H. neanderthalensis and H. sapiens. The connection patterns of the DCs have potential utility in distinguishing between the two taxa and in the phylogenetic and taxonomic discussion of the Neandertal-like specimens with controversial taxonomic status.

Multifield computational model for human brain development: Explicit numerical stabilization

AbstractThe morphological surface of the human brain is still puzzling to scientists. During the last few decades, studies have addressed different aspects of this phenomenon. The underlying cellular processes occurring at the microscopic scale are certainly important for the folding process. However, there is also an essential contribution from mechanical forces generated at the macroscopic scale during these processes. Many previous studies confirmed this fact, but still, most of them have not considered the link between cellular processes and mechanical forces. Recently, we have explained how the proliferation in different zones of the brain, cell migration, and neuronal connectivity affect the folding through a multifield computational model coupling an advection‐diffusion model with the theory of finite growth. We mathematically describe the cellular behavior of cells to control mechanical growth in both radial and circumferential directions. To deal with issues regarding numerical stability of the advection‐diffusion equation, which lead to temporal and spatial numerical oscillations, reduced efficiency of the numerical solver, and inaccurate results, we introduce an artificial diffusivity, which we apply only when the actual cell density does not satisfy the balance equation. The proposed method stabilizes the numerical solution and significantly improves the simulation results without additional numerical cost. It allows us to study brain development in 2D and 3D.

Decreased frontal lobe complexity in left-behind children during joint attention: a fNIRS study with multivariable and multiscale sample entropy analysis.

Human brain development is shaped by experiences, especially during preschool, the critical period for cognitive and socioemotional development. This study employed the functional Near-Infrared Spectroscopy technique to explore the neural differences between left-behind children (LBC) and non-left-behind children (NLBC) on joint attention. Through collecting brain image data of 50 children (26 boys, aged 65.08 ± 6.28months) and conducting multivariable and multiscale sample entropy (MMSE) analysis, the present study found that: (i) LBC showed lower brain complexity than NLBC in right prefrontal cortex; (ii) all participants demonstrated higher brain complexity in responding to joint attention conditions, compared to initiating joint attention ones; (iii) their brain complexity during joint attention was negatively associated with their emotional abilities. The findings advance our understanding of early brain development in LBC by providing evidence for the neural process characteristics of joint attention. Implications for early intervention to promote their brain development are also addressed.

Expanding the Genetic and Mutation Spectrum of ASPM-associated Neurodevelopmental Disorders

Neurodevelopmental disorders (NDDs) are a class of disorders affecting brain development and function; they are characterized by extensive genetic and clinical variability. We performed clinical, genetic, biochemical, and molecular analyses on two consanguineous families with microcephaly exhibiting an NDD. Detailed clinical investigation and molecular diagnosis were performed using whole-exome sequencing (WES), followed by Sanger sequencing for the affected families. WES revealed disease-causing homozygous variants in two families associated with microcephaly and NDDs. In family A and family B, we identified two previously reported homozygous variants (c.3978G>A; Trp1326* and c.4309C>A; p.Arg1437Ser) in the ASPM gene. Both the variants were further confirmed using bi-directional Sanger sequencing. In the present study, we presented literature review regarding the NDDs and microcephaly associated with ASPM pathogenesis. These findings contribute to studies of genotype–phenotype correlation, genetic counseling of the families, inclusion of ASPM in newborn screening, and further understanding of human brain function and development.

Hedonic eating, obesity, and addiction result from increased neuropeptide Y in the nucleus accumbens during human brain evolution

The nucleus accumbens (NAc) is central to motivation and action, exhibiting one of the highest densities of neuropeptide Y (NPY) in the brain. Within the NAc, NPY plays a role in reward and is involved in emotional behavior and in increasing alcohol and drug addiction and fat intake. Here, we examined NPY innervation and neurons of the NAc in humans and other anthropoid primates in order to determine whether there are differences among these various species that would correspond to behavioral or life history variables. We quantified NPY-immunoreactive axons and neurons in the NAc of 13 primate species, including humans, great apes, and monkeys. Our data show that the human brain is unique among primates in having denser NPY innervation within the NAc, as measured by axon length density to neuron density, even after accounting for brain size. Combined with our previous finding of increased dopaminergic innervation in the same region, our results suggest that the neurochemical profile of the human NAc appears to have rendered our species uniquely susceptible to neurophysiological conditions such as addiction. The increase in NPY specific to the NAc may represent an adaptation that favors fat intake and contributes to an increased vulnerability to eating disorders, obesity, as well as alcohol and drug dependence. Along with our findings for dopamine, these deeply rooted structural attributes of the human brain are likely to have emerged early in the human clade, laying the groundwork for later brain expansion and the development of cognitive and behavioral specializations.

Markers of Neuroontogenesis in the Stress-Hyporesponsive Period: Comparison of Laboratory Rodents and Humans

Nowadays there is a number of neuropsychiatric diseases that are considered to be associated with early life stress. Various models are used on laboratory rodents to elucidate the mechanisms of the pathogenesis of psychopathologies that cannot be studied in humans. For successful translation of data, it is necessary to compare the processes of neuroontogenesis at the moment of exposure and subsequent periods. There are many comparative studies concerning the development of neurons and neuronal networks, as well as changes in the hypothalamic-pituitary-adrenal axis. In recent years, it has been reliably shown that glial cells are an important participant in both brain development and its response to stress. The opinion that it is microglia and astrocytes that represent the most promising targets for therapeutic intervention in stress-related diseases is supported. However, there are still no comparative analytical studies covering both stress-realizing systems and neuronal and glial markers of development. This review fills this gap. Here we provide a new perspective for considering the problems of modeling childhood stress and translating the data obtained. The presented analysis, on the one hand, supplements the existing understanding of the correspondence between the stages of brain development in laboratory rodents and humans, and, on the other hand, marks points of growth and raises new questions for researchers of stress in early ontogenesis.

Application of Human Brain Organoids-Opportunities and Challenges in Modeling Human Brain Development and Neurodevelopmental Diseases.

Brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that reflect early brain organization. These organoids contain different cell types, including neurons and glia, similar to those found in the human brain. Human brain organoids provide unique opportunities to model features of human brain development that are not well-reflected in animal models. Compared with traditional cell cultures and animal models, brain organoids offer a more accurate representation of human brain development and function, rendering them suitable models for neurodevelopmental diseases. In particular, brain organoids derived from patients' cells have enabled researchers to study diseases at different stages and gain a better understanding of disease mechanisms. Multi-brain regional assembloids allow for the investigation of interactions between distinct brain regions while achieving a higher level of consistency in molecular and functional characterization. Although organoids possess promising features, their usefulness is limited by several unresolved constraints, including cellular stress, hypoxia, necrosis, a lack of high-fidelity cell types, limited maturation, and circuit formation. In this review, we discuss studies to overcome the natural limitations of brain organoids, emphasizing the importance of combinations of all neural cell types, such as glia (astrocyte, oligodendrocytes, and microglia) and vascular cells. Additionally, considering the similarity of organoids to the developing brain, regionally patterned brain organoid-derived neural stem cells (NSCs) could serve as a scalable source for cell replacement therapy. We highlight the potential application of brain organoid-derived cells in disease cell therapy within this field.

Individual-level brain morphological similarity networks: Current methodologies and applications.

The human brain is an extremely complex system in which neurons, clusters of neurons, or regions are connected to form a complex network. With the development of neuroimaging techniques, magnetic resonance imaging (MRI)-based brain networks play a key role in our understanding of the intricate architecture of human brain. Among them, the structural MRI-based brain morphological network approach has attracted increasing attention due to the advantages in data acquisition, image quality, and in revealing the structural organizing principles intrinsic to the brain. This review is to summarize the methodology and related applications of individual-level morphological networks. There have been a growing number of studies related to brain morphological similarity networks. Conventional morphological networks are intersubject covariance networks constructed using a certain morphological indicator of a group of subjects; individual-level morphological networks, on the other hand, measure the morphological similarity between brain regions for individual brains and can reflect the morphological information of single subjects. In recent years, individual morphological networks have demonstrated significant worth in exploring the topological changes of the human brain under both normal and disease conditions. Such studies provided novel perspectives for understanding human brain development and exploring the pathological mechanisms of neuropsychiatric disorders. This paper mainly focuses on the studies of brain morphological networks at the individual level, introduces several ways for network construction, reviews representative work in this field, and finally points out current problems and future directions.

Genetics of human brain development.

Brain development in humans is achieved through precise spatiotemporal genetic control, the mechanisms of which remain largely elusive. Recently, integration of technological advances in human stem cell-based modelling with genome editing has emerged as a powerful platform to establish causative links between genotypes and phenotypes directly in the human system. Here, we review our current knowledge of complex genetic regulation of each key step of human brain development through the lens of evolutionary specialization and neurodevelopmental disorders and highlight the use of human stem cell-derived 2D cultures and 3D brain organoids to investigate human-enriched features and disease mechanisms. We also discuss opportunities and challenges of integrating new technologies to reveal the genetic architecture of human brain development and disorders.

Integrative omics reveals rapidly evolving regulatory sequences driving primate brain evolution.

Although continual expansion of the brain during primate evolution accounts for our enhanced cognitive capabilities, the drivers of brain evolution have scarcely been explored in these ancestral nodes. Here we performed large-scale comparative genomic, transcriptomic and epigenomic analyses to investigate the evolutionary alterations acquired by brain genes, and provide comprehensive listings of innovatory genetic elements along the evolutionary path from ancestral primates to human. The regulatory sequences associated with brain-expressed genes experienced rapid change, particularly in the ancestor of the Simiiformes. Extensive comparisons of single cell and bulk transcriptomic data between primate and non-primate brains revealed that these regulatory sequences may drive the high expression of certain genes in primate brains. Employing in utero electroporation into mouse embryonic cortex, we show that the primate-specific brain-biased gene BMP7 was recruited, probably in the ancestor of the Simiiformes, to regulate neuronal proliferation in the primate ventricular zone. Our study provides a comprehensive listing of genes and regulatory changes along the brain evolution lineage of ancestral primates leading to human. These data should be invaluable for future functional studies that will deepen our understanding not only of the genetic basis of human brain evolution but also of inherited disease.

Molecular features driving cellular complexity of human brain evolution.

Human-specific genomic changes contribute to the unique functionalities of the human brain1-5. The cellular heterogeneity of the human brain6,7 and the complex regulation of gene expression highlight the need to characterize human-specific molecular features at cellular resolution. Here we analysed single-nucleus RNA-sequencing and single-nucleus assay for transposase-accessible chromatin with sequencing datasets for human, chimpanzee and rhesus macaque brain tissue from posterior cingulate cortex. We show a human-specific increase of oligodendrocyte progenitor cells and a decrease of mature oligodendrocytes across cortical tissues. Human-specific regulatory changes were accelerated in oligodendrocyte progenitor cells, and we highlight key biological pathways that may be associated with the proportional changes. We also identify human-specific regulatory changes in neuronal subtypes, which reveal human-specific upregulation of FOXP2 in only two of the neuronal subtypes. We additionally identify hundreds of new human accelerated genomic regions associated with human-specific chromatin accessibility changes. Our data also reveal that FOS::JUN and FOX motifs are enriched in the human-specifically accessible chromatin regions of excitatory neuronal subtypes. Together, our results reveal several new mechanisms underlying the evolutionary innovation of human brain at cell-type resolution.

Semi-Automatic GUI Platform to Characterize Brain Development in Preterm Children Using Ultrasound Images.

The third trimester of pregnancy is the most critical period for human brain development, during which significant changes occur in the morphology of the brain. The development of sulci and gyri allows for a considerable increase in the brain surface. In preterm newborns, these changes occur in an extrauterine environment that may cause a disruption of the normal brain maturation process. We hypothesize that a normalized atlas of brain maturation with cerebral ultrasound images from birth to term equivalent age will help clinicians assess these changes. This work proposes a semi-automatic Graphical User Interface (GUI) platform for segmenting the main cerebral sulci in the clinical setting from ultrasound images. This platform has been obtained from images of a cerebral ultrasound neonatal database images provided by two clinical researchers from the Hospital Sant Joan de Déu in Barcelona, Spain. The primary objective is to provide a user-friendly design platform for clinicians for running and visualizing an atlas of images validated by medical experts. This GUI offers different segmentation approaches and pre-processing tools and is user-friendly and designed for running, visualizing images, and segmenting the principal sulci. The presented results are discussed in detail in this paper, providing an exhaustive analysis of the proposed approach's effectiveness.

Genes associated with cognitive ability and HAR show overlapping expression patterns in human cortical neuron types

GWAS have identified numerous genes associated with human cognition but their cell type expression profiles in the human brain are unknown. These genes overlap with human accelerated regions (HARs) implicated in human brain evolution and might act on the same biological processes. Here, we investigated whether these gene sets are expressed in adult human cortical neurons, and how their expression relates to neuronal function and structure. We find that these gene sets are preferentially expressed in L3 pyramidal neurons in middle temporal gyrus (MTG). Furthermore, neurons with higher expression had larger total dendritic length (TDL) and faster action potential (AP) kinetics, properties previously linked to intelligence. We identify a subset of genes associated with TDL or AP kinetics with predominantly synaptic functions and high abundance of HARs.

Novel effects of Ras-MAPK pathogenic variants on the developing human brain and their link to gene expression and inhibition abilities

The RASopathies are genetic syndromes associated with pathogenic variants causing dysregulation of the Ras/mitogen-activated protein kinase (Ras-MAPK) pathway, essential for brain development, and increased risk for neurodevelopmental disorders. Yet, the effects of most pathogenic variants on the human brain are unknown. We examined: (1) How Ras-MAPK activating variants of PTPN11/SOS1 protein-coding genes affect brain anatomy. (2) The relationship between PTPN11 gene expression levels and brain anatomy, and (3) The relevance of subcortical anatomy to attention and memory skills affected in the RASopathies. We collected structural brain MRI and cognitive-behavioral data from 40 pre-pubertal children with Noonan syndrome (NS), caused by PTPN11 (n = 30) or SOS1 (n = 10) variants (age 8.53 ± 2.15, 25 females), and compared them to 40 age- and sex-matched typically developing controls (9.24 ± 1.62, 27 females). We identified widespread effects of NS on cortical and subcortical volumes and on determinants of cortical gray matter volume, surface area (SA), and cortical thickness (CT). In NS, we observed smaller volumes of bilateral striatum, precentral gyri, and primary visual area (d’s < −0.8), and extensive effects on SA (d’s > |0.8|) and CT (d’s > |0.5|) relative to controls. Further, SA effects were associated with increasing PTPN11 gene expression, most prominently in the temporal lobe. Lastly, PTPN11 variants disrupted normative relationships between the striatum and inhibition functioning. We provide evidence for the effects of Ras-MAPK pathogenic variants on striatal and cortical anatomy as well as links between PTPN11 gene expression and cortical SA increases, and striatal volume and inhibition skills. These findings provide essential translational information on the Ras-MAPK pathway’s effect on human brain development and function.

Cerebral-Organoid-Derived Exosomes Alleviate Oxidative Stress and Promote LMX1A-Dependent Dopaminergic Differentiation.

The remarkable advancements related to cerebral organoids have provided unprecedented opportunities to model human brain development and diseases. However, despite their potential significance in neurodegenerative diseases such as Parkinson's disease (PD), the role of exosomes from cerebral organoids (OExo) has been largely unknown. In this study, we compared the effects of OExo to those of mesenchymal stem cell (MSC)-derived exosomes (CExo) and found that OExo shared similar neuroprotective effects to CExo. Our findings showed that OExo mitigated H2O2-induced oxidative stress and apoptosis in rat midbrain astrocytes by reducing excess ROS production, antioxidant depletion, lipid peroxidation, mitochondrial dysfunction, and the expression of pro-apoptotic genes. Notably, OExo demonstrated superiority over CExo in promoting the differentiation of human-induced pluripotent stem cells (iPSCs) into dopaminergic (DA) neurons. This was attributed to the higher abundance of neurotrophic factors, including neurotrophin-4 (NT-4) and glial-cell-derived neurotrophic factor (GDNF), in OExo, which facilitated the iPSCs' differentiation into DA neurons in an LIM homeobox transcription factor 1 alpha (LMX1A)-dependent manner. Our study provides novel insight into the biological properties of cerebral organoids and highlights the potential of OExo in the treatment of neurodegenerative diseases such as PD.