ABSTRACT
Owing to recent medical and technological advances in neonatal care, infants born extremely premature have increased survival rates1,2. After birth, these infants are at high risk of hypoxic episodes because of lung immaturity, hypotension and lack of cerebral-flow regulation, and can develop a severe condition called encephalopathy of prematurity3. Over 80% of infants born before post-conception week 25 have moderate-to-severe long-term neurodevelopmental impairments4. The susceptible cell types in the cerebral cortex and the molecular mechanisms underlying associated gray-matter defects in premature infants remain unknown. Here we used human three-dimensional brain-region-specific organoids to study the effect of oxygen deprivation on corticogenesis. We identified specific defects in intermediate progenitors, a cortical cell type associated with the expansion of the human cerebral cortex, and showed that these are related to the unfolded protein response and changes. Moreover, we verified these findings in human primary cortical tissue and demonstrated that a small-molecule modulator of the unfolded protein response pathway can prevent the reduction in intermediate progenitors following hypoxia. We anticipate that this human cellular platform will be valuable for studying the environmental and genetic factors underlying injury in the developing human brain.
Subject(s)
Brain Injuries/etiology , Hypoxia, Brain/etiology , Models, Neurological , Brain Injuries/metabolism , Brain Injuries/pathology , Cell Hypoxia/genetics , Cell Hypoxia/physiology , Cerebral Cortex/metabolism , Cerebral Cortex/pathology , Humans , Hypoxia, Brain/metabolism , Hypoxia, Brain/pathology , Infant, Extremely Premature , Infant, Newborn , Neural Stem Cells/metabolism , Neural Stem Cells/pathology , Neurogenesis/genetics , Neurogenesis/physiology , Organoids/metabolism , Organoids/pathology , T-Box Domain Proteins/metabolism , Unfolded Protein ResponseABSTRACT
An understanding of how heterozygous loss-of-function mutations in autism spectrum disorder (ASD) risk genes, such as TBR1, contribute to ASD remains elusive. Conditional Tbr1 deletion during late mouse gestation in cortical layer 6 neurons (Tbr1layer6 mutants) provides novel insights into its function, including dendritic patterning, synaptogenesis, and cell-intrinsic physiology. These phenotypes occur in heterozygotes, providing insights into mechanisms that may underlie ASD pathophysiology. Restoring expression of Wnt7b largely rescues the synaptic deficit in Tbr1layer6 mutant neurons. Furthermore, Tbr1layer6 heterozygotes have increased anxiety-like behavior, a phenotype seen ASD. Integrating TBR1 chromatin immunoprecipitation sequencing (ChIP-seq) and RNA sequencing (RNA-seq) data from layer 6 neurons and activity of TBR1-bound candidate enhancers provides evidence for how TBR1 regulates layer 6 properties. Moreover, several putative TBR1 targets are ASD risk genes, placing TBR1 in a central position both for ASD risk and for regulating transcriptional circuits that control multiple steps in layer 6 development essential for the assembly of neural circuits.
Subject(s)
DNA-Binding Proteins/genetics , Gene Dosage/physiology , Neocortex/cytology , Neocortex/physiology , Nerve Net/cytology , Nerve Net/physiology , Animals , Animals, Newborn , Cells, Cultured , DNA-Binding Proteins/biosynthesis , Maze Learning/physiology , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neocortex/chemistry , Nerve Net/chemistry , T-Box Domain ProteinsABSTRACT
SUMMARY: Non-negative matrix factorization (NMF) is an increasingly used algorithm for the analysis of complex high-dimensional data. BRB-ArrayTools is a widely used software system for the analysis of gene expression data with almost 9000 registered users in over 65 countries. We have developed a NMF analysis plug-in in BRB-ArrayTools for unsupervised sample clustering of microarray gene expression data. Our analysis tool also incorporates an algorithm for Semi-NMF which can handle both positive and negative elements for log-ratio data. Output includes a heat map of sample clusters and differentially expressed genes with extensive biological annotation. For comparison, output also includes the results of K-means clustering. AVAILABILITY: The NMF analysis plug-in is freely available in BRB-ArrayTools for non-commercial users. BRB-ArrayTools can be downloaded at http://linus.nci.nih.gov/BRB-ArrayTools.html. The algorithms used for NMF and Semi-NMF are available at ftp://linus.nci.nih.gov/pub/NMF.
