Aneuploidy effects on human gene expression across three cell types.

Aneuploidy syndromes impact multiple organ systems but understanding of tissue-specific aneuploidy effects remains limited-especially for the comparison between peripheral tissues and relatively inaccessible tissues like brain. Here, we address this gap in knowledge by studying the transcriptomic effects of chromosome X, Y, and 21 aneuploidies in lymphoblastoid cell lines, fibroblasts and iPSC-derived neuronal cells (LCLs, FCL, and iNs, respectively). We root our analyses in sex chromosome aneuploidies, which offer a uniquely wide karyotype range for dosage effect analysis. We first harness a large LCL RNA-seq dataset from 197 individuals with one of 6 sex chromosome dosages (SCDs: XX, XXX, XY, XXY, XYY, and XXYY) to i) validate theoretical models of SCD sensitivity and ii) define an expanded set of 41 genes that show obligate dosage sensitivity to SCD and are all in cis (i.e., reside on the X or Y chromosome). We then use multiple complementary analyses to show that cis effects of SCD in LCLs are preserved in both FCLs (n = 32) and iNs (n = 24), whereas trans effects (i.e., those on autosomal gene expression) are mostly not preserved. Analysis of additional datasets confirms that the greater cross-cell type reproducibility of cis vs. trans effects is also seen in trisomy 21 cell lines. These findings i) expand our understanding of X, Y, and 21 chromosome dosage effects on human gene expression and ii) suggest that LCLs may provide a good model system for understanding cis effects of aneuploidy in harder-to-access cell types.

Fluoxetine Can Cause Epileptogenesis and Aberrant Neurogenesis in Male Wild Type Mice.

Antidepressants in general, and fluoxetine in particular, increase adult hippocampal neurogenesis (AHN) in mice. Here we asked how the antidepressant fluoxetine affects behavior and AHN in a corticosterone model of depression. In three groups of adult male C57BL/6j mice we administered either vehicle (VEH), corticosterone (CORT) treatment to induce a depression-like state or corticosterone plus a standard dose of fluoxetine (CORT+FLX). Following treatment, mice performed the open field test, the novelty suppressed feeding (NSF) test and the splash test. Neurogenesis was assessed by means of immunohistochemistry using BrdU and neuronal maturation markers. Unexpectedly, 42% of the CORT+FLX-treated mice exhibited severe weight loss, seizures and sudden death. As expected, the CORT treated group had altered behaviors compared to the VEH group, but the CORT+FLX mice that survived did not show any behavioral improvement compared to the CORT group. Antidepressants generally increase neurogenesis and here we also found that compared to CORT mice, CORT+FLX mice that survived had a significantly greater density of BrdU+, BrdU+DCX+ and BrdU+NeuN+ cells, suggesting increased neurogenesis. Moreover, the density of BrdU+NeuN+ cells was increased in an aberrant location, the hilus, of CORT+FLX mice, similar to previous studies describing aberrant neurogenesis following seizures. In conclusion, fluoxetine could induce considerable adverse effects in wild type mice, including seizure-like activity. Fluoxetine-induced neurogenesis increases could be related to this activity, therefore proneurogenic effects of fluoxetine and other antidepressants, especially in the absence of any behavioral therapeutic effects, should be interpreted with caution.

Intravital Imaging of the Murine Subventricular Zone with Three Photon Microscopy.

The mouse subventricular zone (SVZ) produces neurons throughout life. It is useful for mechanism discovery and is relevant for regeneration. However, the SVZ is deep, significantly restricting live imaging since current methods do not extend beyond a few hundred microns. We developed and adapted three-photon microscopy (3PM) for non-invasive deep brain imaging in live mice, but its utility in imaging the SVZ niche was unknown. Here, with fluorescent dyes and genetic labeling, we show successful 3PM imaging in the whole SVZ, extending to a maximum depth of 1.5 mm ventral to the dura mater. 3PM imaging distinguished multiple SVZ cell types in postnatal and juvenile mice. We also detected fine processes on neural stem cells interacting with the vasculature. Previous live imaging removed overlying cortical tissue or lowered lenses into the brain, which could cause inflammation and alter neurogenesis. We found that neither astrocytes nor microglia become activated in the SVZ, suggesting 3PM does not induce major damage in the niche. Thus, we show for the first time 3PM imaging of the SVZ in live mice. This strategy could be useful for intravital visualization of cell dynamics, molecular, and pathological perturbation and regenerative events.

The long non-coding RNA Paupar promotes KAP1-dependent chromatin changes and regulates olfactory bulb neurogenesis.

Many long non-coding RNAs (lncRNAs) are expressed during central nervous system (CNS) development, yet their in vivo roles and mechanisms of action remain poorly understood. Paupar, a CNS-expressed lncRNA, controls neuroblastoma cell growth by binding and modulating the activity of transcriptional regulatory elements in a genome-wide manner. We show here that the Paupar lncRNA directly binds KAP1, an essential epigenetic regulatory protein, and thereby regulates the expression of shared target genes important for proliferation and neuronal differentiation. Paupar promotes KAP1 chromatin occupancy and H3K9me3 deposition at a subset of distal targets, through the formation of a ribonucleoprotein complex containing Paupar, KAP1 and the PAX6 transcription factor. Paupar-KAP1 genome-wide co-occupancy reveals a fourfold enrichment of overlap between Paupar and KAP1 bound sequences, the majority of which also appear to associate with PAX6. Furthermore, both Paupar and Kap1 loss-of-function in vivo disrupt olfactory bulb neurogenesis. These observations provide important conceptual insights into the trans-acting modes of lncRNA-mediated epigenetic regulation and the mechanisms of KAP1 genomic recruitment, and identify Paupar and Kap1 as regulators of neurogenesis in vivo.

The A30P α-synuclein mutation decreases subventricular zone proliferation.

Parkinson's disease (PD) is associated with olfactory defects in addition to dopaminergic degeneration. Dopaminergic signalling is necessary for subventricular zone (SVZ) proliferation and olfactory bulb (OB) neurogenesis. Alpha-synuclein (α-syn or Snca) modulates dopaminergic neurotransmission, and SNCA mutations cause familial PD, but how α-syn and its mutations affect adult neurogenesis is unclear. To address this, we studied a bacterial artificial chromosome transgenic mouse expressing the A30P SNCA familial PD point mutation on an Snca-/- background. We confirmed that the SNCA-A30P transgene recapitulates endogenous α-syn expression patterns and levels by immunohistochemical detection of endogenous α-syn in a wild-type mouse and transgenic SNCA-A30P α-syn protein in the forebrain. The number of SVZ stem cells (BrdU+GFAP+) was decreased in SNCA-A30P mice, whereas proliferating (phospho-histone 3+) cells were decreased in Snca-/- and even more so in SNCA-A30P mice. Similarly, SNCA-A30P mice had fewer Mash1+ transit-amplifying SVZ progenitor cells but Snca-/- mice did not. These data suggest the A30P mutation aggravates the effect of Snca loss in the SVZ. Interestingly, calbindin+ and calretinin (CalR)+ periglomerular neurons were decreased in both Snca-/-, and SNCA-A30P mice but tyrosine hydroxylase+ periglomerular OB neurons were only decreased in Snca-/- mice. Cell death decreased in the OB granule layer of Snca-/- and SNCA-A30P mice. In the same region, CalR+ numbers increased in Snca-/- and SNCA-A30P mice. Thus, α-syn loss and human A30P SNCA decrease SVZ proliferation, cell death in the OB and differentially alter interneuron numbers. Similar disruptions in human neurogenesis may contribute to the olfactory deficits, which are observed in PD.

Galectin-3 diminishes Wnt signaling in the postnatal subventricular zone.

Postnatal subventricular zone (pSVZ) stem and progenitor cell proliferation is regulated by several developmental signaling pathways such as Wnt/ß-catenin. However, the molecular regulation of Wnt function in the pSVZ is poorly understood. We previously showed that Wnt signaling is upregulated in an SVZ gliomagenesis in vivo model. As well, the pro-inflammatory molecule Galectin-3 (Gal-3) increases Wnt signaling in cancer cells and is expressed in the SVZ. Therefore, we asked if Gal-3 has a similar function on Wnt signaling in the pSVZ. We interrogated Wnt signaling using a signaling reporter as well as immunohistochemistry and showed that Wnt signaling predominates upstream in the pSVZ lineage but is downregulated in migrating neuroblasts. Biochemical analysis of SVZ cells, in vivo and in neurosphere stem/progenitor cells, showed that Gal-3 physically interacts with multiple forms of ß-catenin, which is a major downstream regulator of Wnt signaling. Functional analyses demonstrated, in vitro and in vivo, that Gal-3 knockdown increases Wnt signaling and conversely that Gal-3 OE inhibits Wnt/ß-catenin signaling in the pSVZ. This latter result suggested that Gal-3, which is consistently increased in brain injury, may decrease pSVZ proliferation. We showed that Gal-3 OE decreased proliferation without altering cell cycle re-entry and that it increased p27Kip1, a molecule which induces cell cycle exit. Our data uncover a novel regulator of Wnt signaling in the SVZ, Gal-3, which does so in a manner opposite to cancer.

Galectin-3 modulates postnatal subventricular zone gliogenesis.

Postnatal subventricular zone (SVZ) neural stem cells generate forebrain glia, namely astrocytes and oligodendrocytes. The cues necessary for this process are unclear, despite this phase of brain development being pivotal in forebrain gliogenesis. Galectin-3 (Gal-3) is increased in multiple brain pathologies and thereby regulates astrocyte proliferation and inflammation in injury. To study the function of Gal-3 in inflammation and gliogenesis, we carried out functional studies in mouse. We overexpressed Gal-3 with electroporation and using immunohistochemistry surprisingly found no inflammation in the healthy postnatal SVZ. This allowed investigation of inflammation-independent effects of Gal-3 on gliogenesis. Loss of Gal-3 function via knockdown or conditional knockout reduced gliogenesis, whereas Gal-3 overexpression increased it. Gal-3 overexpression also increased the percentage of striatal astrocytes generated by the SVZ but decreased the percentage of oligodendrocytes. These novel findings were further elaborated with multiple analyses demonstrating that Gal-3 binds to the bone morphogenetic protein receptor one alpha (BMPR1α) and increases bone morphogenetic protein (BMP) signaling. Conditional knockout of BMPR1α abolished the effect of Gal-3 overexpression on gliogenesis. Gain-of-function of Gal-3 is relevant in pathological conditions involving the human forebrain, which is particularly vulnerable to hypoxia/ischemia during perinatal gliogenesis. Hypoxic/ischemic injury induces astrogliosis, inflammation and cell death. We show that Gal-3 immunoreactivity was increased in the perinatal human SVZ and striatum after hypoxia/ischemia. Our findings thus show a novel inflammation-independent function for Gal-3; it is necessary for gliogenesis and when increased in expression can induce astrogenesis via BMP signaling.

Polycomb Protein Eed is Required for Neurogenesis and Cortical Injury Activation in the Subventricular Zone.

The postnatal subventricular zone (SVZ) harbors neural stem cells (NSCs) that exhibit robust neurogenesis. However, the epigenetic mechanisms that maintain NSCs and regulate neurogenesis remain unclear. We report that label-retaining SVZ NSCs express Eed, the core component of Polycomb repressive complex 2. In vivo and in vitro conditional knockout and knockdown show Eed is necessary for maintaining NSC proliferation, neurogenesis and neurosphere formation. We discovered that Eed functions to maintain p21 protein levels in NSCs by repressing Gata6 transcription. Both Gata6 overexpression and p21 knockdown reduced neurogenesis, while Gata6 knockdown or p21 overexpression partially rescued neurogenesis after Eed loss. Furthermore, genetic deletion of Eed impaired injury induced SVZ proliferation and emigration. These data reveal a novel epigenetic regulated pathway and suggest an essential role for Eed in SVZ homeostasis and injury.

Calretinin interneuron density in the caudate nucleus is lower in autism spectrum disorder.

Autism spectrum disorder is a debilitating condition with possible neurodevelopmental origins but unknown neuroanatomical correlates. Whereas investigators have paid much attention to the cerebral cortex, few studies have detailed the basal ganglia in autism. The caudate nucleus may be involved in the repetitive movements and limbic changes of autism. We used immunohistochemistry for calretinin and neuropeptide Y in 24 age- and gender-matched patients with autism spectrum disorder and control subjects ranging in age from 13 to 69 years. Patients with autism had a 35% lower density of calretinin+ interneurons in the caudate that was driven by loss of small calretinin+ neurons. This was not caused by altered size of the caudate, as its cross-sectional surface areas were similar between diagnostic groups. Controls exhibited an age-dependent increase in the density of medium and large calretinin+ neurons, whereas subjects with autism did not. Diagnostic groups did not differ regarding ionized calcium-binding adapter molecule 1+ immunoreactivity for microglia, suggesting chronic inflammation did not cause the decreased calretinin+ density. There was no statistically significant difference in the density of neuropeptide Y+ neurons between subjects with autism and controls. The decreased calretinin+ density may disrupt the excitation/inhibition balance in the caudate leading to dysfunctional corticostriatal circuits. The description of such changes in autism spectrum disorder may clarify pathomechanisms and thereby help identify targets for drug intervention and novel therapeutic strategies.

STAT1-induced ASPP2 transcription identifies a link between neuroinflammation, cell polarity, and tumor suppression.

Inflammation and loss of cell polarity play pivotal roles in neurodegeneration and cancer. A central question in both diseases is how the loss of cell polarity is sensed by cell death machinery. Here, we identify apoptosis-stimulating protein of p53 with signature sequences of ankyrin repeat-, SH3 domain-, and proline-rich region-containing protein 2 (ASPP2), a haploinsufficient tumor suppressor, activator of p53, and regulator of cell polarity, as a transcriptional target of signal transducer and activator of transcription 1 (STAT1). LPS induces ASPP2 expression in murine macrophage and microglial cell lines, a human monocyte cell line, and primary human astrocytes in vitro. LPS and IFNs induce ASPP2 transcription through an NF-κB RELA/p65-independent but STAT1-dependent pathway. In an LPS-induced maternal inflammation mouse model, LPS induces nuclear ASPP2 in vivo at the blood-cerebral spinal fluid barrier (the brain's barrier to inflammation), and ASPP2 mediates LPS-induced apoptosis. Consistent with the role of ASPP2 as a gatekeeper to inflammation, ASPP2-deficient brains possess enhanced neuroinflammation. Elevated ASPP2 expression is also observed in mouse models and human neuroinflammatory disease tissue, where ASPP2 was detected in GFAP-expressing reactive astrocytes that coexpress STAT1. Because the ability of ASPP2 to maintain cellular polarity is vital to CNS development, our findings suggest that the identified STAT1/ASPP2 pathway may connect tumor suppression and cell polarity to neuroinflammation.

Loss of galectin-3 decreases the number of immune cells in the subventricular zone and restores proliferation in a viral model of multiple sclerosis.

Multiple sclerosis (MS) frequently starts near the lateral ventricles, which are lined by subventricular zone (SVZ) progenitor cells that can migrate to lesions and contribute to repair. Because MS-induced inflammation may decrease SVZ proliferation and thus limit repair, we studied the role of galectin-3 (Gal-3), a proinflammatory protein. Gal-3 expression was increased in periventricular regions of human MS in post-mortem brain samples and was also upregulated in periventricular regions in a murine MS model, Theiler's murine encephalomyelitis virus (TMEV) infection. Whereas TMEV increased SVZ chemokine (CCL2, CCL5, CCL, and CXCL10) expression in wild type (WT) mice, this was inhibited in Gal-3(-/-) mice. Though numerous CD45+ immune cells entered the SVZ of WT mice after TMEV infection, their numbers were significantly diminished in Gal-3(-/-) mice. TMEV also reduced neuroblast and proliferative SVZ cell numbers in WT mice but this was restored in Gal-3(-/-) mice and was correlated with increased numbers of doublecortin+ neuroblasts in the corpus callosum. In summary, our data showed that loss of Gal-3 blocked chemokine increases after TMEV, reduced immune cell migration into the SVZ, reestablished SVZ proliferation and increased the number of progenitors in the corpus callosum. These results suggest Gal-3 plays a central role in modulating the SVZ neurogenic niche's response to this model of MS.

Cuprizone demyelination induces a unique inflammatory response in the subventricular zone.

BACKGROUND: Cuprizone leads to demyelination of the corpus callosum (CC) and activates progenitor cells in the adjacent subventricular zone (SVZ), a stem cell niche which contributes to remyelination. The healthy SVZ contains semi-activated microglia and constitutively expresses the pro-inflammatory molecule galectin-3 (Gal-3) suggesting the niche uniquely regulates inflammation. METHODS: We studied the inflammatory response to cuprizone in the SVZ and CC in Gal-3 knockout mice using immunohistochemistry and with the in vitro neurosphere assay. RESULTS: Cuprizone caused loss of myelin basic protein (MBP) immunofluorescence in the CC suggesting demyelination. Cuprizone increased the density of CD45+/Iba1+ microglial cells and also increased Gal-3 expression in the CC. Surprisingly, the number of Gal-3+ and CD45+ cells decreased in the SVZ after cuprizone, suggesting inflammation was selectively reduced therein. Inflammation can regulate SVZ proliferation and indeed the number of phosphohistone H3+ (PHi3+) cells decreased in the SVZ but increased in the CC in both genotypes after cuprizone treatment. BrdU+ SVZ cell numbers also decreased in the SVZ after cuprizone, and this effect was significantly greater at 3 weeks in Gal-3 (-/-) mice compared to WT, suggesting Gal-3 normally limits SVZ cell emigration following cuprizone treatment. CONCLUSIONS: This study reveals a uniquely regulated inflammatory response in the SVZ and shows that Gal-3 participates in remyelination in the cuprizone model. This contrasts with more severe models of demyelination which induce SVZ inflammation and suggests the extent of demyelination affects the SVZ neurogenic response.

Disruption of Visc-2, a Brain-Expressed Conserved Long Noncoding RNA, Does Not Elicit an Overt Anatomical or Behavioral Phenotype.

Although long noncoding RNAs (lncRNAs) are proposed to play essential roles in mammalian neurodevelopment, we know little of their functions from their disruption in vivo. Combining evidence for evolutionary constraint and conserved expression data, we previously identified candidate lncRNAs that might play important and conserved roles in brain function. Here, we demonstrate that the sequence and neuronal transcription of lncRNAs transcribed from the previously uncharacterized Visc locus are conserved across diverse mammals. Consequently, one of these lncRNAs, Visc-2, was selected for targeted deletion in the mouse, and knockout animals were subjected to an extremely detailed anatomical and behavioral characterization. Despite a neurodevelopmental expression pattern of Visc-2 that is highly localized to the cortex and sites of neurogenesis, anomalies in neither cytoarchitecture nor neuroproliferation were identified in knockout mice. In addition, no abnormal motor, sensory, anxiety, or cognitive behavioral phenotypes were observed. These results are important because they contribute to a growing body of evidence that lncRNA loci contribute on average far less to brain and biological functions than protein-coding loci. A high-throughput knockout program focussing on lncRNAs, similar to that currently underway for protein-coding genes, will be required to establish the distribution of their organismal functions.

Blocked angiogenesis in Galectin-3 null mice does not alter cellular and behavioral recovery after middle cerebral artery occlusion stroke.

Angiogenesis is thought to decrease stroke size and improve behavioral outcomes and therefore several clinical trials are seeking to augment it. Galectin-3 (Gal-3) expression increases after middle cerebral artery occlusion (MCAO) and has been proposed to limit damage 3days after stroke. We carried out mild MCAO that damages the striatum but spares the cerebral cortex and SVZ. Gal-3 gene deletion prevented vascular endothelial growth factor (VEGF) upregulation after MCAO. This inhibited post-MCAO increases in endothelial proliferation and angiogenesis in the striatum allowing us to uniquely address the function of angiogenesis in this model of stroke. Apoptosis and infarct size were unchanged in Gal-3(-/-) mice 7 and 14 days after MCAO, suggesting that angiogenesis does not affect lesion size. Microglial and astrocyte activation/proliferation after MCAO was similar in wild type and Gal-3(-/-) mice. In addition, openfield activity, motor hemiparesis, proprioception, reflex, tremors and grooming behaviors were essentially identical between WT and Gal-3(-/-) mice at 1, 3, 7, 10 and 14 days after MCAO, suggesting that penumbral angiogenesis has limited impact on behavioral recovery. In addition to angiogenesis, increased adult subventricular zone (SVZ) neurogenesis is thought to provide neuroprotection after stroke in animal models. SVZ neurogenesis and migration to lesion were overall unaffected by the loss of Gal-3, suggesting no compensation for the lack of angiogenesis in Gal-3(-/-) mice. Because angiogenesis and neurogenesis are usually coordinately regulated, identifying their individual effects on stroke has hitherto been difficult. These results show that Gal-3 is necessary for angiogenesis in stroke in a VEGF-dependant manner, but suggest that angiogenesis may be dispensable for post-stroke endogenous repair, therefore drawing into question the clinical utility of augmenting angiogenesis.

Ependymal ciliary dysfunction and reactive astrocytosis in a reorganized subventricular zone after stroke.

Subventricular zone (SVZ) astrocytes and ependymal cells are both derived from radial glia and may have similar gliotic reactions after stroke. Diminishing SVZ neurogenesis worsens outcomes in mice, yet the effects of stroke on SVZ astrocytes and ependymal cells are poorly understood. We used mouse experimental stroke to determine if SVZ astrocytes and ependymal cells assume similar phenotypes and if stroke impacts their functions. Using lateral ventricular wall whole mount preparations, we show that stroke caused SVZ reactive astrocytosis, disrupting the neuroblast migratory scaffold. Also, SVZ vascular density and neural proliferation increased but apoptosis did not. In contrast to other reports, ependymal denudation and cell division was never observed. Remarkably, however, ependymal cells assumed features of reactive astrocytes post stroke, robustly expressing de novo glial fibrillary acidic protein, enlargening and extending long processes. Unexpectedly, stroke disrupted motile cilia planar cell polarity in ependymal cells. This suggested ciliary function was affected and indeed ventricular surface flow was slower and more turbulent post stroke. Together, these results demonstrate that in response to stroke there is significant SVZ reorganization with implications for both pathophysiology and therapeutic strategies.

Rosemary extract activates oligodendrogenesis genes in mouse brain and improves learning and memory ability.

Rosemary (Rosmarinus officinalis L.) is a rich source of dietary bioactive compounds such as rosmarinic acid and carnosol with a large repertoire of pharmacological properties, including anti-inflammatory and neuroprotective activities. In the present study, we investigated rosemary as a potential new therapeutic agent for cognitive function and other symptoms of aging. In this present study, we have aimed to investigate the effects of oral administration of rosemary extract (RME) on learning and memory in the context of other biomarkers-related cognitive function and neurotransmitter levels in senescent accelerated prone 8 (SAMP8) mouse, a model of accelerating aging and Alzheimer's disease. The Morris water maze (MWM) test showed improved spatial learning and memory behavior in RME treated SAMP8 mouse. Moreover, RME decreased Aß42 and inflammatory cytokine levels and increased BDNF, Sirt1, and neurotransmitter levels in SAMP8 mouse. Whole-genome microarray analysis revealed that RME significantly increased gene expression related to oligodendrocyte differentiation, myelination, and ATP production in the hippocampus and decreased gene expression related to stress, neuroinflammation, and apoptosis. Also, in the SAMP8 hippocampus, RME significantly increased Olig1 and Olig2 expression. Altogether, our study is the first to report improvement of spatial learning and memory of RME, modulation of genes important for oligodendrogenesis, and Anti-neuroinflammatory effect by suppressing Aß42 levels in mouse brain and thus highlights the prospects of RME in the treatment of cognitive dysfunction and aging.

Galectin-3 maintains cell motility from the subventricular zone to the olfactory bulb.

The adult brain subventricular zone (SVZ) produces neuroblasts that migrate through the rostral migratory stream (RMS) to the olfactory bulb (OB) in a specialized niche. Galectin-3 (Gal-3) regulates proliferation and migration in cancer and is expressed by activated macrophages after brain injury. The function of Gal-3 in the normal brain is unknown, but we serendipitously found that it was expressed by ependymal cells and SVZ astrocytes in uninjured mice. Ependymal cilia establish chemotactic gradients and astrocytes form glial tubes, which combine to aid neuroblast migration. Whole-mount preparations and electron microscopy revealed that both ependymal cilia and SVZ astrocytes were disrupted in Gal3(-/-) mice. Interestingly, far fewer new BrdU(+) neurons were found in the OB of Gal3(-/-) mice, than in wild-type mice 2 weeks after labeling. However, SVZ proliferation and cell death, as well as OB differentiation rates were unaltered. This suggested that decreased migration in vivo was sufficient to decrease the number of new OB neurons. Two-photon time-lapse microscopy in forebrain slices confirmed decreased migration; cells were slower and more exploratory in Gal3(-/-) mice. Gal-3 blocking antibodies decreased migration and dissociated neuroblast cell-cell contacts, whereas recombinant Gal-3 increased migration from explants. Finally, we showed that expression of phosphorylated epidermal growth factor receptor (EGFR) was increased in Gal3(-/-) mice. These results suggest that Gal-3 is important in SVZ neuroblast migration, possibly through an EGFR-based mechanism, and reveals a role for this lectin in the uninjured brain.

What has single-cell transcriptomics taught us about long non-coding RNAs in the ventricular-subventricular zone?

Long non-coding RNA (lncRNA) function is mediated by the process of transcription or through transcript-dependent associations with proteins or nucleic acids to control gene regulatory networks. Many lncRNAs are transcribed in the ventricular-subventricular zone (V-SVZ), a postnatal neural stem cell niche. lncRNAs in the V-SVZ are implicated in neurodevelopmental disorders, cancer, and brain disease, but their functions are poorly understood. V-SVZ neurogenesis capacity declines with age due to stem cell depletion and resistance to neural stem cell activation. Here we analyzed V-SVZ transcriptomics by pooling current single-cell RNA-seq data. They showed consistent lncRNA expression during stem cell activation, lineage progression, and aging. In conjunction with epigenetic and genetic data, we predicted V-SVZ lncRNAs that regulate stem cell activation and differentiation. Some of the lncRNAs validate known epigenetic mechanisms, but most remain uninvestigated. Our analysis points to several lncRNAs that likely participate in key aspects of V-SVZ stem cell activation and neurogenesis in health and disease.

Regulation of immunological tolerance by the p53-inhibitor iASPP.

Maintenance of immunological homeostasis between tolerance and autoimmunity is essential for the prevention of human diseases ranging from autoimmune disease to cancer. Accumulating evidence suggests that p53 can mitigate phagocytosis-induced adjuvanticity thereby promoting immunological tolerance following programmed cell death. Here we identify Inhibitor of Apoptosis Stimulating p53 Protein (iASPP), a negative regulator of p53 transcriptional activity, as a regulator of immunological tolerance. iASPP-deficiency promoted lung adenocarcinoma and pancreatic cancer tumorigenesis, while iASPP-deficient mice were less susceptible to autoimmune disease. Immune responses to iASPP-deficient tumors exhibited hallmarks of immunosuppression, including activated regulatory T cells and exhausted CD8+ T cells. Interestingly, iASPP-deficient tumor cells and tumor-infiltrating myeloid cells, CD4+, and γδ T cells expressed elevated levels of PD-1H, a recently identified transcriptional target of p53 that promotes tolerogenic phagocytosis. Identification of an iASPP/p53 axis of immune homeostasis provides a therapeutic opportunity for both autoimmune disease and cancer.

Integration of 3D-printed cerebral cortical tissue into an ex vivo lesioned brain slice.

Engineering human tissue with diverse cell types and architectures remains challenging. The cerebral cortex, which has a layered cellular architecture composed of layer-specific neurons organised into vertical columns, delivers higher cognition through intricately wired neural circuits. However, current tissue engineering approaches cannot produce such structures. Here, we use a droplet printing technique to fabricate tissues comprising simplified cerebral cortical columns. Human induced pluripotent stem cells are differentiated into upper- and deep-layer neural progenitors, which are then printed to form cerebral cortical tissues with a two-layer organization. The tissues show layer-specific biomarker expression and develop a structurally integrated network of processes. Implantation of the printed cortical tissues into ex vivo mouse brain explants results in substantial structural implant-host integration across the tissue boundaries as demonstrated by the projection of processes and the migration of neurons, and leads to the appearance of correlated Ca2+ oscillations across the interface. The presented approach might be used for the evaluation of drugs and nutrients that promote tissue integration. Importantly, our methodology offers a technical reservoir for future personalized implantation treatments that use 3D tissues derived from a patient's own induced pluripotent stem cells.