SummaryGrowing evidence implicates the importance of glia, particularly astrocytes, in neurological and psychiatric diseases. Here, we describe a rapid and robust method for the differentiation of highly pure populations of replicative astrocytes from human induced pluripotent stem cells (hiPSCs), via a neural progenitor cell (NPC) intermediate. We evaluated this protocol across 42 NPC lines (derived from 30 individuals). Transcriptomic analysis demonstrated that hiPSC-astrocytes from four individuals are highly similar to primary human fetal astrocytes and characteristic of a non-reactive state. hiPSC-astrocytes respond to inflammatory stimulants, display phagocytic capacity, and enhance microglial phagocytosis. hiPSC-astrocytes also possess spontaneous calcium transient activity. Our protocol is a reproducible, straightforward (single medium), and rapid (<30 days) method to generate populations of hiPSC-astrocytes that can be used for neuron-astrocyte and microglia-astrocyte co-cultures for the study of neuropsychiatric disorders.

Summary Zika virus (ZIKV) infects fetal and adult human brain, and is associated with serious neurological complications. To date, no therapeutic treatment is available to treat ZIKV infected patients. We performed a high content chemical screen using human embryonic stem cell derived cortical neuron progenitor cells (hNPCs) and found that hippeastrine hydrobromide (HH) and amodiaquine dihydrochloride dihydrate (AQ), can inhibit ZIKV infection in hNPCs. Further validation showed that HH also rescues ZIKV-induced growth and differentiation defects in hNPCs and human fetal-like forebrain organoids. Finally, HH and AQ inhibit ZIKV infection in adult mouse brain in vivo. Strikingly, HH suppresses viral propagation when administered to adult mice with active ZIKV infection, highlighting its therapeutic potential. Our approach highlights the power of stem cell-based screens and validation in human forebrain organoids and mouse models in identifying drug candidates for treating ZIKV infection and related neurological complications in fetal and adult patients.

We report locus-specific disintegration of megabase-scale chromosomal conformations in brain after neuronal ablation of Kmt1e/Setdb1 histone H3-lysine 9 methyltransferase, including a large topologically associated 1.2Mb domain conserved in human and mouse and encompassing >70 genes at the clustered Protocadherin (cPcdh) locus. TADcPcdh in mutant neurons showed abnormal accumulations of CTCF transcriptional regulator and 3D genome organizer at cryptic binding sites, converted into permissive state with DNA cytosine hypomethylation and histone hyperacetylation. Broadly upregulated expression across cPcdh included defective S-type Protocadherin single-cell stochastic constraint. Setdb1-dependent loop formations, bypassing 0.2–1Mb of linear genome, radiated from TADPcdh fringes towards cPcdh cis-regulatory sequences, counterbalanced shorter-range facilitative promoter-enhancer contacts and carried loop-bound polymorphisms associated with genetic risk for schizophrenia. We show that KRAB-zinc finger Setdb1 repressor complex, shielding neuronal 3D genomes from excess CTCF binding, is critically required for structural maintenance of TADcPcdh.

The power of human induced pluripotent stem cell (hiPSC)-based studies to resolve the smaller effects of common variants within the size of cohorts that can be realistically assembled remains uncertain. We identified and accounted for a variety of technical and biological sources of variation in a large case/control schizophrenia (SZ) hiPSC-derived cohort of neural progenitor cells and neurons. Reducing the stochastic effects of the differentiation process by correcting for cell type composition boosted the SZ signal and increased the concordance with post-mortem data sets. We predict a growing convergence between hiPSC and post-mortem studies as both approaches expand to larger cohort sizes. For studies of complex genetic disorders, to maximize the power of hiPSC cohorts currently feasible, in most cases and whenever possible, we recommend expanding the number of individuals even at the expense of the number of replicate hiPSC clones.