The generation and properties of human cortical organoids as a disease model for malformations of cortical development

As three-dimensional“organ-like”aggregates,human cortical organoids have emerged as powerful models for studying human brain evolution and brain disorders with unique advantages of humanspecificity,fidelity and manipulation.Human cortical organoids derived from human pluripotent stem cells can elaborately replicate many of the key properties of human cortical development at the molecular,cellular,structural,and functional levels,including the anatomy,functional neural network,and interaction among different brain regions,thus facilitating the discovery of brain development and evolution.In addition to studying the neuro-electrophysiological features of brain cortex development,human cortical organoids have been widely used to mimic the pathophysiological features of cortical-related disease,especially in mimicking malformations of cortical development,thus revealing pathological mechanism and identifying effective drugs.In this review,we provide an overview of the generation of human cortical organoids and the properties of recapitulated cortical development and further outline their applications in modeling malformations of cortical development including pathological phenotype,underlying mechanisms and rescue strategies.

The molecular genetics of PI3K/PTEN/AKT/mTOR pathway in the malformations of cortical development

Malformations of cortical development(MCD)are a group of developmental disorders characterized by abnormal cortical structures caused by genetic or harmful environmental factors.Many kinds of MCD are caused by genetic variation.MCD is the common cause of intellectual disability and intractable epilepsy.With rapid advances in imaging and sequencing technologies,the diagnostic rate of MCD has been increasing,and many potential genes causing MCD have been successively identified.However,the high genetic heterogeneity of MCD makes it challenging to understand the molecular pathogenesis of MCD and to identify effective targeted drugs.Thus,in this review,we outline important events of cortical development.Then we illustrate the progress of molecular genetic studies about MCD focusing on the PI3K/PTEN/AKT/mTOR pathway.Finally,we briefly discuss the diagnostic methods,disease models,and therapeutic strategies for MCD.The information will facilitate further research on MCD.Understanding the role of the PI3K/PTEN/AKT/mTOR pathway in MCD could lead to a novel strategy for treating MCD-related diseases.

Establishment of human cerebral organoid systems to model early neural development and assess the central neurotoxicity of environmental toxins

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 matrigel for only 7 days during the rapid expansion phase of the neural epithelium and then removing them from the matrigel 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.

Electrical stimulation of cortical neurons promotes oligodendrocyte development and remyelination in the injured spinal cord

Background and early studies： Endogenous tri-potential neural stem cells （NSCs） exist in the adult mammalian central nervous system （CNS）. In the spinal cord, NSCs distribute throughout the entire cord, but exist predominately in white matter tracts. The phenotypic fate of these cells in white matter is glial, largely oligodendrocyte, but not neuronal.

Ultrastructure of focal cerebral cortex tissue from rats with focal cortical dysplasia

BACKGROUND： Developing a model of focal cortical dysplasia in microgyrus and observing the ultrastructure of focal tissue is of important significance for analyzing the pathology of cortical developmental disorder and the factors of structural changes. OBJECTIVE： This study was to observe the pathological characteristics of focal tissue around the microgyrus of rats with cortical developmental disorder using an electron microscope, so as to analyze the causes associated with cerebral cortical developmental disorder. DESIGN： A randomized controlled animal experiment. SETTING： The First Affiliated Hospital of Chongqing Medical University. MATERIALS： This study was carried out in the Chongqing Key Laboratory of Neurology, Room for Electron Microscope of Chongqing Medical University, and Laboratory Animal Center, Research Institute of Surgery, Daping Hospital, Third Military Medical University of Chinese PLA between January 2004 and August 2006. Eighteen healthy newborn male Wistar rats, weighing 3.0 - 6.0 g, provided by the Laboratory Animal Center, Daping Hospital, Third Military Medical University of Chinese PLA, were involved in this study. The protocol was carried out in accordance with animal ethics guidelines for the use and care of animals. Probes （Chongqing Wire ＆ Cable Factory, China） were made of copper core wire with diameter of 1mm. METHODS： The rats were randomly divided into 3 groups with 6 in each： normal control group, liquid nitrogen injured group and sham-operation group. （1）In the liquid nitrogen injured group, a blunt probe frozen by liquid nitrogen was placed on fronto-parietal crinial bone of rats for 8 s. A 3 - 5 cm of microgyrus was induced in the unilateral cerebral sensory cortical area. In the sham-operation group, probe was placed at the room temperature. In the normal control group, rats were untouched. （2） The conscious state and electrical activity of brain of rats in each group were observed. （3） 2 - 3 mm thickness of hippocampal tissue with coronary section was taken for observing its ultrastructure under a transmission electron microscope. MAIN OUTCOME MEASURES： （1） The ultrastructure of hippocampal tissue. （2）The conscious state and electrical activity of brain of rats. RESULTS： Eighteen rats were enrolled in the final analysis. （1） Observation of hippocampal ultrastructure： Electromicroscopic pathological findings showed that for each rat of the liquid nitrogen injured group, mitochondrium in the pyramidal neuron around the microgyrus was swelled, endoplasmic reticulum was expanded, glial cells were swelled, water gathered around the blood capillary, partial medullary sheath was degenerated, neuropilem was normal and no obviously abnormal synapse was found. （2） Changes in conscious state of rats： Rats in the normal control group and sham-operation group had no convulsive seizure, but those in the liquid nitrogen injured group had occasionally. Most of them showed increased activities, excitation and restlessness, scratching and frequent ＂ watching face-like activities＂. （3）Electrical activity of brain of rats： Electroencephalogram recording of liquid nitrogen injured group showed that small wave amplitude of rhythm took the main part. No typical sharp wave, V wave, sharp and slow wave, V and slow waves were discharged. CONCLUSION： Liquid nitrogen can lead to cerebral cortical developmental disorder. Pathological changes of ultrastructure of focal tissue around the microgyrus can provide pathological basis for epilepsy associated with focal cortical developmental disorder.

Pure Salivatory Seizures Secondary to a Subtle Malformation of the Right Parietal Cortex

Background: Salivatory seizures are a singularly rare condition, which can occur both in idiopathic and symptomatic epilepsies. Objectives: To describe and discuss the case of an adolescent patient with sleep-triggered “pure” salivatory seizures associated with a subtle cortical malformation of the right parietal cortex. Case report: Herein, we report a 17-year-old female who started to present salivatory paroxysms, which occasionally secondarily generalized, shortly after falling asleep, at the age of eight years. Video-electroencephalographic monitoring with scalp electrodes failed to show any epileptiform activity during the several recorded clinical events. Brain MRI and curvilinear reconstruction revealed, in the three orthogonal planes, a subtle cortical thickening, limited to a single gyrus in the right parietal cortex, suggestive of a focal cortical malformation. After antiepileptic drug therapy was optimized, the patient became seizure-free. Conclusion: An epilepsy diagnosis should be pursued in patients presenting isolated, paroxysmal hypersalivation, despite possible negative scalp EEG studies.

Ultrasonographic Characteristics of Cortical Sulcus Development in the Human Fetus between 18 and 41 Weeks of Gestation

Background： Fetal brain development is a complicated process that continues throughout pregnancy. Fetal sulcus development has typical morphological features. Assessment of fetal sulcus development to understand the cortical maturation and development by prenatal ultrasound has become widespread. This study aimed to explore a reliable method to assess cortical sulcus and to describe the normal sonographic features of cortical sulcus development in the human fetus between 18 and 41 weeks of gestation. Methods： A cross-sectional study was designed to examine the fetal cortical sulcus development at 18-41 weeks of gestation. Ultrasound was used to examine the insula, sylvian fissure （SF）, parieto-occipital fissure （POF）, and calcarine fissure （CF）. Bland-Altman plots were used for assessing the concordance, and the intraclass correlation coefficient was used for assessing the reliability. Results： SF images were successfully obtained in 100% of participants at 22 weeks of gestation, while the POF images and CF images could be obtained in 100% at 23 weeks of gestation and 24 weeks of gestation, respectively. The SF width, temporal lobe depth, POF depth, and the CF depth increased with the developed gestation. The width of uncovered insula and the POF angle decreased with the developed gestation. By 23 weeks of gestation, the insula was beginning to be covered. Moreover, it completed at 35 weeks of gestation. The intra- and inter-observer agreements showed consistent reproducibility. Conclusions： This study defined standard views of the fetal sulcus as well as the normal reference ranges of these sulcus measurements between 18 and 41 weeks of gestation. Such ultrasonographic measurements could be used to identil fetuses at risk of fetal neurological structural disorders.

Transcriptome Analysis Identifies SenZfp536,a Sense LncRNA that Suppresses Self-renewal of Cortical Neural Progenitors

Long non-coding RNAs(lncRNAs)regulate transcription to control development and homeostasis in a variety of tissues and organs.However,their roles in the development of the cerebral cortex have not been well elucidated.Here,a bioinformatics pipeline was applied to delineate the dynamic expression and potential cis-regulating effects of mouse lncRNAs using transcriptome data from 8 embryonic time points and sub-regions of the developing cerebral cortex.We further characterized a sense lncRNA,SenZfp536,which is transcribed downstream of and partially overlaps with the protein-coding gene Zfp536.Both SenZfp536 and Zfp536 were predominantly expressed in the proliferative zone of the developing cortex.Zfp536 was cis-regulated by SenZfp536,which facilitates looping between the promoter of Zfp536 and the genomic region that transcribes SenZfp536.Surprisingly,knocking down or activating the expression of SenZfp536 increased or compromised the proliferation of cortical neural progenitor cells(NPCs),respectively.Finally,overexpressing Zfp536 in cortical NPCs reversed the enhanced proliferation of cortical NPCs caused by SenZfp536 knockdown.The study deepens our understanding of how lncRNAs regulate the propagation of cortical NPCs through cis-regulatory mechanisms.