Single cell analysis of dup15q syndrome reveals developmental and postnatal molecular changes in autism.

Duplication 15q (dup15q) syndrome is the most common genetic cause of autism spectrum disorder (ASD). Due to a higher genetic and phenotypic homogeneity compared to idiopathic autism, dup15q syndrome provides a well-defined setting to investigate ASD mechanisms. Previous bulk gene expression studies identified shared molecular changes in ASD. However, how cell type specific changes compare across different autism subtypes and how they change during development is largely unknown. In this study, we used single cell and single nucleus mRNA sequencing of dup15q cortical organoids from patient iPSCs, as well as post-mortem patient brain samples. We find cell-type specific dysregulated programs that underlie dup15q pathogenesis, which we validate by spatial resolved transcriptomics using brain tissue samples. We find degraded identity and vulnerability of deep-layer neurons in fetal stage organoids and highlight increased molecular burden of postmortem upper-layer neurons implicated in synaptic signaling, a finding shared between idiopathic ASD and dup15q syndrome. Gene co-expression network analysis of organoid and postmortem excitatory neurons uncovers modules enriched with autism risk genes. Organoid developmental modules were involved in transcription regulation via chromatin remodeling, while postmortem modules were associated with synaptic transmission and plasticity. The findings reveal a shifting landscape of ASD cellular vulnerability during brain development.

LIF signaling regulates outer radial glial to interneuron fate during human cortical development.

Radial glial (RG) development is essential for cerebral cortex growth and organization. In humans, the outer radial glia (oRG) subtype is expanded and gives rise to diverse neurons and glia. However, the mechanisms regulating oRG differentiation are unclear. oRG cells express leukemia-inhibitory factor (LIF) receptors during neurogenesis, and consistent with a role in stem cell self-renewal, LIF perturbation impacts oRG proliferation in cortical tissue and organoids. Surprisingly, LIF treatment also increases the production of inhibitory interneurons (INs) in cortical cultures. Comparative transcriptomic analysis identifies that the enhanced IN population resembles INs produced in the caudal ganglionic eminence. To evaluate whether INs could arise from oRGs, we isolated primary oRG cells and cultured them with LIF. We observed the production of INs from oRG cells and an increase in IN abundance following LIF treatment. Our observations suggest that LIF signaling regulates the capacity of oRG cells to generate INs.

Tropism of SARS-CoV-2 for human cortical astrocytes.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) readily infects a variety of cell types impacting the function of vital organ systems, with particularly severe impact on respiratory function. Neurological symptoms, which range in severity, accompany as many as one-third of COVID-19 cases, indicating a potential vulnerability of neural cell types. To assess whether human cortical cells can be directly infected by SARS-CoV-2, we utilized stem-cell-derived cortical organoids as well as primary human cortical tissue, both from developmental and adult stages. We find significant and predominant infection in cortical astrocytes in both primary tissue and organoid cultures, with minimal infection of other cortical populations. Infected and bystander astrocytes have a corresponding increase in inflammatory gene expression, reactivity characteristics, increased cytokine and growth factor signaling, and cellular stress. Although human cortical cells, particularly astrocytes, have no observable ACE2 expression, we find high levels of coronavirus coreceptors in infected astrocytes, including CD147 and DPP4. Decreasing coreceptor abundance and activity reduces overall infection rate, and increasing expression is sufficient to promote infection. Thus, we find tropism of SARS-CoV-2 for human astrocytes resulting in inflammatory gliosis-type injury that is dependent on coronavirus coreceptors.

Arsenite interrupts neurodevelopmental processes of human and rat neural progenitor cells: The role of reactive oxygen species and species-specific antioxidative defense.

Arsenic exposure disturbs brain development in humans. Although developmental neurotoxicity (DNT) of arsenic has been studied in vivo and in vitro, its mode-of-action (MoA) is not completely understood. Here, we characterize the adverse neurodevelopmental effects of sodium arsenite on developing human and rat neural progenitor cells (hNPC, rNPC). Moreover, we analyze the involvement of reactive oxygen species (ROS) and the role of the glutathione (GSH)-dependent antioxidative defense for arsenite-induced DNT in a species-specific manner. We determined IC50 values for sodium arsenite-dependent (0.1-10 µM) inhibition of hNPC and rNPC migration (6.0 µM; >10 µM), neuronal (2.7 µM; 4.4 µM) and oligodendrocyte (1.1 µM; 2.0 µM) differentiation. ROS involvement was studied by quantifying the expression of ROS-regulated genes, measuring glutathione (GSH) levels, inhibiting GSH synthesis and co-exposing cells to the antioxidant N-acetylcysteine. Arsenite reduces NPC migration, neurogenesis and oligodendrogenesis of differentiating hNPC and rNPC at sub-cytotoxic concentrations. Species-specific arsenite cytotoxicity and induction of antioxidative gene expression is inversely related to GSH levels with rNPC possessing >3-fold the amount of GSH than hNPC. Inhibition of GSH synthesis increased the sensitivity towards arsenite in rNPC > hNPC. N-acetylcysteine antagonized arsenite-mediated induction of HMOX1 expression as well as reduction of neuronal and oligodendrocyte differentiation in hNPC suggesting involvement of oxidative stress in arsenite DNT. hNPC are more sensitive towards arsenite-induced neurodevelopmental toxicity than rNPC, probably due to their lower antioxidative defense capacities. This species-specific MoA data might be useful for adverse outcome pathway generation and future integrated risk assessment strategies concerning DNT.

A transcriptome comparison of time-matched developing human, mouse and rat neural progenitor cells reveals human uniqueness.

It is widely accepted that human brain development has unique features that cannot be represented by rodents. Obvious reasons are the evolutionary distance and divergent physiology. This might lead to false predictions when rodents are used for safety or pharmacological efficacy studies. For a better translation of animal-based research to the human situation, human in vitro systems might be useful. In this study, we characterize developing neural progenitor cells from prenatal human and time-matched rat and mouse brains by analyzing the changes in their transcriptome profile during neural differentiation. Moreover, we identify hub molecules that regulate neurodevelopmental processes like migration and differentiation. Consequences of modulation of three of those hubs on these processes were studied in a species-specific context. We found that although the gene expression profiles of the three species largely differ qualitatively and quantitatively, they cluster in similar GO terms like cell migration, gliogenesis, neurogenesis or development of multicellular organism. Pharmacological modulation of the identified hub molecules triggered species-specific cellular responses. This study underlines the importance of understanding species differences on the molecular level and advocates the use of human based in vitro models for pharmacological and toxicological research.