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Astrocytes are intimately linked with brain vessels, a relationship that is critical for neuronal health and function. However, astroglial factors driving these physical and functional associations during postnatal brain development have yet to be identified. We characterized structural and transcriptional changes in mouse cortical astrocytes and microvessels during the first two postnatal weeks and found that high-mobility group box 1 (Hmgb1), normally upregulated with injury and involved in adult cerebrovascular repair, was highly expressed in astrocytes at birth and then decreased rapidly. Astrocyte-selective ablation ofHmgb1at birth affected astrocyte morphology and endfoot placement, altered distribution of endfoot proteins connexin43 and aquaporin-4, induced transcriptional changes in astrocytes related to cytoskeleton remodeling, and profoundly disrupted endothelial ultrastructure. While lack of astroglialHmgb1did not affect the blood-brain barrier or angiogenesis postnatally, it impaired neurovascular coupling and behavior in adult mice. These findings identify astroglialHmgb1as a key player in postnatal gliovascular maturation.

Summary Organoids (ORGs) are increasingly used as models of cerebral cortical development. Here, we compared transcriptome and cellular phenotypes between telencephalic ORGs and monolayers (MONs) generated in parallel from three biologically distinct induced pluripotent stem cell (iPSC) lines. Multiple readouts revealed increased proliferation in MONs, which was caused by increased integrin signaling. MONs also exhibited altered radial glia (RG) polarity and suppression of Notch signaling, as well as impaired generation of intermediate progenitors, outer RG, and cortical neurons, which were all partially reversed by reaggregation of dissociated cells. Network analyses revealed co-clustering of cell adhesion, Notch-related transcripts and their transcriptional regulators in a module strongly downregulated in MONs. The data suggest that ORGs, with respect to MONs, initiate more efficient Notch signaling in ventricular RG owing to preserved cell adhesion, resulting in subsequent generation of intermediate progenitors and outer RG, in a sequence that recapitulates the cortical ontogenetic process., Highlights • Organoid’s radial glia cells engage efficient Notch signaling • Monolayer hyperproliferation is due to increased integrin signaling • Neurogenesis' arrest by cell dissociation is partially reversed with reaggregation, In this research article, Scuderi and colleagues generated organoids and monolayers from three iPSC lines and characterized their transcriptomes, proteomes, and cellular architecture. Disorganized neural stem cell polarity in monolayers impaired Notch signaling and decreased a complex transcription factor cascade involved in the generation of intermediate neuronal progenitors and outer radial glia, disrupting cortical neuron differentiation.

Astroglial cells are key players in the development and maintenance of neurons and neuronal networks. Astroglia express steroid hormone receptors and show rapid responses to hormonal manipulations. However, despite important sex differences in the cortex and hippocampus, few studies have examined sex differences in astroglial cells in telencephalic development. To characterize the cortical astroglial translatome in male and female mice across postnatal development, we use translating ribosome affinity purification together with RNA sequencing and immunohistochemistry to phenotype astroglia at six developmental time points. Overall, we find two distinct astroglial phenotypes between early (P1-P7) and late development (P14-adult), independent of sex. We also find sex differences in gene expression patterns across development that peak at P7 and appear to result from males reaching a mature astroglial phenotype earlier than females. These developmental sex differences could have an impact on the construction of neuronal networks and windows of vulnerability to perturbations and disease.

Recent studies have suggested that cortical astroglia play an important role in depressive-like behaviors. Potential astroglial contributions have been proposed based on their known neuroplastic functions, such as glutamate recycling and synaptic plasticity. However, the specific mechanisms by which astroglial cells may contribute or protect against a depressive phenotype remain unknown. To delineate astroglial changes that accompany depressive-like behavior, we used astroglial-specific bacTRAP mice exposed to chronic variable stress (CVS) and profiled the astroglial translatome using translating ribosome affinity purification (TRAP) in conjunction with RNAseq. As expected, CVS significantly increased anxiety- and depressive-like behaviors and corticosterone levels and decreased GFAP expression in astroglia, although this did not reflect a change in the total number of astroglial cells. TRAPseq results showed that CVS decreased genes associated with astroglial plasticity: RhoGTPases, growth factor signaling, and transcription regulation, and increased genes associated with the formation of extracellular matrices such as perineuronal nets (PNNs). PNNs inhibit neuroplasticity and astroglia contribute to the formation, organization, and maintenance of PNNs. To validate our TRAPseq findings, we showed an increase in PNNs following CVS. Degradation of PNNs in the prefrontal cortex of mice exposed to CVS reversed the CVS-induced behavioral phenotype in the forced swim test. These data lend further support to the neuroplasticity hypothesis of depressive behaviors and, in particular, extend this hypothesis beyond neuronal plasticity to include an overall decrease in genes associated with cortical astroglial plasticity following CVS. Further studies will be needed to assess the antidepressant potential of directly targeting astroglial cell function in models of depression.

Astroglial cells are emerging as key players in the development and homeostatic maintenance of neurons and neuronal networks. Astroglial cell functions are critical to neuronal migration and maturation, myelination, and synapse dynamics, however little is known about astroglial phenotypic changes over development. Furthermore, astroglial cells express steroid hormone receptors and show rapid responses to hormonal manipulations, however, despite important sex differences in telencephalic regions such as the cortex and hippocampus few studies have examined sex differences in astroglial cells in development and outside of the hypothalamus and amygdala. To phenotype cortical astroglial cells across postnatal development while considering potential sex differences, we used translating ribosome affinity purification together with RNA sequencing (TRAPseq) and immunohistochemistry to phenotype the entire astroglial translatome in males and females at key developmental time points: P1, P4, P7, P14, P35 and in adulthood. Overall, we found two distinct astroglial phenotypes between early (P1-P7) and late development (P14-Adult). We also noted that although astroglia show few basal sex differences in adulthood, they show significant sex differences in developmental gene expression patterns, with peak sex differences observed at P7. At least part of the sex differences observed at P7 appear to be due to males reaching a more mature astroglial phenotype earlier than females. Together, these data clearly delineate and phenotype astroglia across development and identify sex differences in astroglial developmental programs. Importantly, these developmental sex differences could have an impact on the construction and maintenance of neuronal networks and potential developmental windows of vulnerability to neurologic and psychiatric disease.

Human induced pluripotent stem cells (hiPSCs) are emerging as a useful tool for modelling in vitro early brain development and neurological disorders. Molecular mechanisms and cell interactions that regulate the neurodevelopment at early stages remain unclear because of human brain’s complexity and limitations of functional studies. Two major culture methodologies are used to differentiate in vitro hiPSCs into neurons: monolayer (2D) and organoid (3D) cultures. Here we investigate the effect of cell dissociation and the loss of 3D organization during the early differentiation process of neuronal progenitors. Using the same culture media, we first differentiated hiPSCs into neural progenitor cells (NPCs) and then induced their differentiation into neurons in 3 different modalities: 3D undissociated organoids, dissociated NPCs followed by immediate re-aggregation into an organoid, and dissociated NPCs cultured as monolayer. We assessed neuronal differentiation efficiency of each method by immunocytochemistry, qPCR, western blot, and RNA-Seq analysis over a time course. Our data revealed substantial differences in gene and protein expression among the three systems, including genes of the Notch pathway (e.g. NEUROD1, NEUROG2), earliest determinants of cortical region differentiation (e.g. SOX1, FEZF1) as well as later transcriptional regulators that specify cortical neuron subtypes (e.g. TBR1, CTIP2), which were all downregulated in monolayer. Moreover, we found that genes and pathways mediating cell-to-cell interactions (e.g. CNTNs, CAMs) were mostly upregulated in the 3D culture systems, whereas cell-extracellular matrix interaction molecules (e.g. ITG, LAM) were mostly upregulated in 2D, indicating that cell surface molecules may be involved in specification of neuronal cell types. Our results address the methodological question of the appropriateness of a differentiation method for a particular experimental goal, and, beyond that, reveal important early determinants that exert a decisive influence on neuronal differentiation and regional specification of human neural stem cells., Italian Journal of Anatomy and Embryology, Vol. 122, No. 1 (Supplement) 2017

Ectonucleotide phosphatase/phosphodiesterase 1 (ENPP1) deficiency results in either lethal arterial calcifications (‘Generalized Arterial Calcification of Infancy’ – GACI), phosphate wasting rickets (‘Autosomal Recessive Hypophosphatemic Rickets type 2’ – ARHR2), early onset osteoporosis, or progressive spinal rigidity (‘Ossification of the Posterior Longitudinal Ligament’ – OPLL). As ENPP1 generates a strong endogenous mineralization inhibitor – extracellular pyrophosphate (PPi) – ENPP1 deficiency should not result in reduced bone volume, and therefore the mechanism ENPP1 associated osteoporosis is not apparent given current understanding of the enzyme’s function. To investigate genetic pathways driving the skeletal phenotype of ENPP1 deficiency we compared gene expression in Enpp1(asj/asj) mice and WT sibling pairs by RNAseq and qPCR in whole bones, and in the liver and kidney by qPCR, directly correlating gene expression with measures of bone microarchitectural and biomechanical phenotypes. Unbiased analysis of the differentially expressed genes compared to relevant human disease phenotypes revealed that Enpp1(asj/asj) mice exhibit strong signatures of osteoporosis, ARHR2 and OPLL. We found that ENPP1 deficient mice exhibited reduced gene transcription of Wnt ligands in whole bone and increased transcription of soluble Wnt inhibitors in the liver and kidney, suggestive of multiorgan inhibition of Wnt activity. Consistent with Wnt suppression in bone, Collagen gene pathways in bone were significantly decreased and Fgf23 was significantly increased, all of which directly correlated with bone microarchitectural defects and fracture risk in Enpp1(asj/asj) mice. Moreover, the bone findings in 10-week old mice correlated with Enpp1 transcript counts but not plasma [PPi], suggesting that the skeletal phenotype at 10 weeks is driven by catalytically independent ENPP1 function. In contrast, the bone findings in 23-week Enpp1(asj/asj) mice strongly correlated with plasma PPi, suggesting that chronically low PPi drives the skeletal phenotype in older mice. Finally, correlation between Enpp1 and Fgf23 transcription suggested ENPP1 regulation of Fgf23, which we confirmed by dosing Enpp1(asj/asj) mice with soluble ENPP1-Fc and observing suppression of intact plasma FGF23 and ALP. In summary, our findings suggest that osteoporosis associated with ENPP1 deficiency involves the suppression of Wnt via catalytically independent Enpp1 pathways, and validates Enpp1(asj/asj) mice as tools to better understand OPLL and Paradoxical Mineralization Disorders.

Supplemental material, Supplementary_Data_File_3 for A Further Analysis and Commentary on: Profiling Changes in Cortical Astroglial Cells Following Chronic Stress by Gianfilippo Coppola, Gareth M Rurak, Stephanie Simard and Natalina Salmaso in Journal of Experimental Neuroscience

INTRODUCTION The human cerebral cortex has undergone an extraordinary increase in size and complexity during mammalian evolution. Cortical cell lineages are specified in the embryo, and genetic and epidemiological evidence implicates early cortical development in the etiology of neuropsychiatric disorders such as autism spectrum disorder (ASD), intellectual disabilities, and schizophrenia. Most of the disease-implicated genomic variants are located outside of genes, and the interpretation of noncoding mutations is lagging behind owing to limited annotation of functional elements in the noncoding genome. RATIONALE We set out to discover gene-regulatory elements and chart their dynamic activity during prenatal human cortical development, focusing on enhancers, which carry most of the weight upon regulation of gene expression. We longitudinally modeled human brain development using human induced pluripotent stem cell (hiPSC)–derived cortical organoids and compared organoids to isogenic fetal brain tissue. RESULTS Fetal fibroblast–derived hiPSC lines were used to generate cortically patterned organoids and to compare oganoids’ epigenome and transcriptome to that of isogenic fetal brains and external datasets. Organoids model cortical development between 5 and 16 postconception weeks, thus enabling us to study transitions from cortical stem cells to progenitors to early neurons. The greatest changes occur at the transition from stem cells to progenitors. The regulatory landscape encompasses a total set of 96,375 enhancers linked to target genes, with 49,640 enhancers being active in organoids but not in mid-fetal brain, suggesting major roles in cortical neuron specification. Enhancers that gained activity in the human lineage are active in the earliest stages of organoid development, when they target genes that regulate the growth of radial glial cells. Parallel weighted gene coexpression network analysis (WGCNA) of transcriptome and enhancer activities defined a number of modules of coexpressed genes and coactive enhancers, following just six and four global temporal patterns that we refer to as supermodules, likely reflecting fundamental programs in embryonic and fetal brain. Correlations between gene expression and enhancer activity allowed stratifying enhancers into two categories: activating regulators (A-regs) and repressive regulators (R-regs). Several enhancer modules converged with gene modules, suggesting that coexpressed genes are regulated by enhancers with correlated patterns of activity. Furthermore, enhancers active in organoids and fetal brains were enriched for ASD de novo variants that disrupt binding sites of homeodomain, Hes1, NR4A2, Sox3, and NFIX transcription factors. CONCLUSION We validated hiPSC-derived cortical organoids as a suitable model system for studying gene regulation in human embryonic brain development, evolution, and disease. Our results suggest that organoids may reveal how noncoding mutations contribute to ASD etiology. Summary of the study, analyses, and main results. Data were generated for iPSC-derived human telencephalic organoids and isogenic fetal cortex. Organoids modeled embryonic and early fetal cortex and show a larger repertoire of enhancers. Enhancers could be divided into activators and repressors of gene expression. We derived networks of modules and supermodules with correlated gene and enhancer activities, some of which were implicated in autism spectrum disorders (ASD).

SummaryAutism spectrum disorder (ASD) is a disorder of brain development. Most cases lack a clear etiology or genetic basis, and the difficulty of re-enacting human brain development has precluded understanding of ASD pathophysiology. Here we use three-dimensional neural cultures (organoids) derived from induced pluripotent stem cells (iPSCs) to investigate neurodevelopmental alterations in individuals with severe idiopathic ASD. While no known underlying genomic mutation could be identified, transcriptome and gene network analyses revealed upregulation of genes involved in cell proliferation, neuronal differentiation, and synaptic assembly. ASD-derived organoids exhibit an accelerated cell cycle and overproduction of GABAergic inhibitory neurons. Using RNA interference, we show that overexpression of the transcription factor FOXG1 is responsible for the overproduction of GABAergic neurons. Altered expression of gene network modules and FOXG1 are positively correlated with symptom severity. Our data suggest that a shift toward GABAergic neuron fate caused by FOXG1 is a developmental precursor of ASD.

Autism spectrum disorder (ASD) affects as many as 1 in 68 children and is said to be the fastest-growing serious developmental disability in the United States. There is currently no medical cure or diagnostic test for ASD. Furthermore, the U.S. Food and Drug Administration has yet to approve a single drug for the treatment of autism's core symptoms. Despite numerous genome studies and the identification of hundreds of genes that may cause or predispose children to ASD, the pathways underlying the pathogenesis of idiopathic ASD still remain elusive. Post-mortem brain samples, apart from being difficult to obtain, offer little insight into a disorder that arises through the course of development. Furthermore, ASD is a disorder of highly complex, human-specific behaviors, making it difficult to model in animals. Stem cell models of ASD can be generated by performing skin biopsies of ASD patients and then dedifferentiating these fibroblasts into human-induced pluripotent stem cells (hiPSCs). iPSCs closely resemble embryonic stem cells and retain the unique genetic signature of the ASD patient from whom they were originally derived. Differentiation of these iPSCs into neurons essentially recapitulates the ASD patient's neuronal development in a dish, allowing for a patient-specific model of ASD. Here we review our current understanding of the underlying neurobiology of ASD and how the use of stem cells can advance this understanding, possibly leading to new therapeutic avenues.

Currently, we have a poor understanding of the pathogenesis of neurodevelopmental disorders, owing to the fact that post-mortem and imaging studies are only capable of measuring the postnatal status quo and offer little insight into the processes that give rise to the observed outcomes. Human induced pluripotent stem cells (hiPSCs) in principle should prove powerful in elucidating the pathways that give rise to neurodevelopmental disorders. HiPSCs are embryonic stem cell-like cells that can be derived from somatic cells. They retain the unique genetic signature of the individual from whom they were derived from, and thus allow researchers to recapitulate that individual’s idiosyncratic neural development in a dish. In the case of diseased individuals, we can reenact the disease-altered trajectory of brain development and examine how and why phenotypic and molecular abnormalities arise in these diseased brains. In this paper, we review various aspects of hiPSC biology and experimental design as well as discuss existing hiPSC models of neurodevelopmental disorders. As already shown by some studies discussed in this review, our hope is that iPSCs will illuminate the pathophysiology of developmental disorders of the CNS and lead to therapeutic avenues for the millions that today suffer from neurodevelopmental disorders.

High-resolution molecular phenotyping of human IPSC organoids using CLARITY and 2-photon microscopy Simone Tomasi, Soraya Scuderi, Anahita Amiri, Giovanna G. Altobelli, Jessica Mariani, Cheryl Dambrot, Gianfilippo Coppola, Flora M. Vaccarino To understand the role of gene regulation in human brain development and neuropsychiatric disorders, it is essential to develop cellular models of the human brain. Induced pluripotent stem cells (iPSC)-derived brain organoids can be used to investigate the role of gene regulatory elements, noncoding RNA, and in general, noncoding disease associated gene variants in brain development and function. Organoids enable gradients of morphogenes and other extracellular cues to build up in the intercellular milieu and to interact with the genetic and epigenetic background of a given progenitor cell during the course of brain development. We have developed an iPSC-derived organoid model of the early human forebrain, where differentiation of cortical excitatory and inhibitory neurons can be studied in a reproducible fashion, enabling a more precise identification of molecular events crucially involved in the specification of distinct neuronal subtypes. However, a precise assessment of protein and RNA expression in intact organoids is hampered by the limited penetration of molecular probes, therefore requiring the preparation of thin sections and greatly limiting the capacity of exploring molecular and cellular features in a 3D environment. Here, we labeled telencephalic excitatory and inhibitory lineages in using pLenti-CAMKII-GFP and pLenti-DlxI12b-BG-DsRed vectors, then used two-photon microscopy to image the genetically-encoded fluorescence at higher resolution in live forebrain organoids. Next, we used CLARITY to clear the organoids and perform immunostainings on the intact cell aggregates. Our current protocol enables a 3D reconstruction of GFP/TdTomato filled cells allowing the analysis of axonal and dendritic arborization, dendrite length, synapse and spine distribution, as well as stereological counts of structures labeled by specific markers. Using these combined approaches, we aim at comparing intra-organoid layer cytoarchitecture and its emerging connectivity with parallel data from RNA-seq and ChIP-seq experiments, and develop new tools for linking the molecular and cellular features of organoids derived from different individuals.

Few studies have been conducted to understand post-zygotic accumulation of mutations in cells of the healthy human body. We reprogrammed 32 skin fibroblast cells from families of donors into human induced pluripotent stem cell (hiPSC) lines. The clonal nature of hiPSC lines allows a high-resolution analysis of the genomes of the founder fibroblast cells without being confounded by the artifacts of single-cell whole-genome amplification. We estimate that on average a fibroblast cell in children has 1035 mostly benign mosaic SNVs. On average, 235 SNVs could be directly confirmed in the original fibroblast population by ultradeep sequencing, down to an allele frequency (AF) of 0.1%. More sensitive droplet digital PCR experiments confirmed more SNVs as mosaic with AF as low as 0.01%, suggesting that 1035 mosaic SNVs per fibroblast cell is the true average. Similar analyses in adults revealed no significant increase in the number of SNVs per cell, suggesting that a major fraction of mosaic SNVs in fibroblasts arises during development. Mosaic SNVs were distributed uniformly across the genome and were enriched in a mutational signature previously observed in cancers and in de novo variants and which, we hypothesize, is a hallmark of normal cell proliferation. Finally, AF distribution of mosaic SNVs had distinct narrow peaks, which could be a characteristic of clonal cell selection, clonal expansion, or both. These findings reveal a large degree of somatic mosaicism in healthy human tissues, link de novo and cancer mutations to somatic mosaicism, and couple somatic mosaicism with cell proliferation.

Experiments, simulations, and numerical bifurcation analysis are used to study the incompressible flow between two opposed tubes with disks mounted at their exits. The experiments in this axisymmetric geometry show that for low and equal Reynolds numbers, Re, at both nozzles, the flow remains symmetric about the plane halfway through the nozzle exits and the stagnation plane is located halfway between the two jets. When Re is increased past a critical value, asymmetric flow fields are obtained even when the momentum fluxes of the two opposed streams are equal. For unequal Re at the jet exits, when the fixed velocity (and the corresponding Reynolds number, Re 1 ) of one stream is low, the stagnation plane location, SPL, changes smoothly with the Re 2 . For high enough Re 1 , a hysteretic jump of SPL is observed. Particle Image Velocimetry and flow visualization demonstrate that within the hysteretic range, the two stable flow fields are anti-symmetric. The experimental setup is also studied with transient incompressible flow simulations using a spectral element solver. It is found that to accurately model the flow, we either need to extend the domain into the nozzles, or impose experimental velocity profiles at the nozzle exits. As in the experiments asymmetric flows are obtained past a critical Re. Finally, bifurcation analysis using a Newton–Picard method shows that the transition from symmetric to asymmetric flows results from the loss of stability of the symmetric flows at a pitchfork bifurcation.

An experimental study was conducted to develop and characterize systematically a new turbulence generator system to yield large turbulent Reynolds numbers in a compact configuration. The effect of the geometric parameters of two families of high-blockage plates on the resulting turbulent flow field was systematically studied: one series of plates was characterized by the number and distribution of circular openings; a second series had non-circular opening(s) with different shapes, distribution and position of the opening(s). The plates were placed upstream of a contoured contraction and the near field at the centerline of the resulting turbulent free jet was characterized by hot-wire anemometry in terms of mean axial velocity, turbulence intensity, turbulence length scales and corresponding Reynolds numbers. The plate with a central, non-circular opening produced the best compromise of highest turbulence levels along with excellent uniformity in average velocity and turbulence intensity, as evidenced by scan in the transverse direction. It appears to be the most promising one. By comparison with more traditional approaches to turbulence generation, we increased the turbulent Reynolds numbers based on the integral length scale to values on the order of 1000, which was one of the design objectives. Other plate geometries also yielded intense turbulence, but, in some cases, exhibited spurious frequency peaks in their power spectrum. The turbulent generation approach is to be adapted to combustion studies to reproduce conditions typical of practical system in relatively small experimental set-ups that are well-suited for bench-top experiments.

We propose a highly turbulent counterflow flame as a very useful benchmark of complexity intermediate between laminar flames and practical systems. By operating in a turbulent Reynolds number regime of relevance to practical systems such as gas turbines and internal combustion engines, it retains the interaction of turbulence and chemistry of such environments, but offers several advantages including: (a) the achievement of high Reynolds numbers without pilot flames, which is particularly advantageous from a modeling standpoint; (b) control of the transition from stable flames to local extinction/reignition conditions; (c) compactness of the domain by comparison with jet flames, with obvious advantages from both a diagnostic and, especially, a computational viewpoint; and (d) the reduction or, altogether, elimination of soot formation, thanks to the high strain rates and low residence times of such a system, and the establishment of conditions of large stoichiometric mixture fraction, as required for robust flame stabilization. We demonstrate the phenomenology of such highly strained turbulent flames under conditions spanning unpremixed, partially premixed and premixed regimes. The system lends itself to the validation of DNS and other computational models. It is also well-suited for the examination of practical fuel blends – a need that is becoming more and more pressing in view of the anticipated diversification of the future fossil fuel supply.

We have studied numerically steady and unsteady flow in a straight and a helically stented common carotid artery, in order to model porcine experimental results that show reduced intimal hyperplasia (IH) in the helical case. The combination of flow pulsatility and three-dimensionality generates a sweeping motion of the Dean vortices, which overall reduced extremes of both oxygen flux to the vessel wall and wall shear stress (WSS). Since IH and atherosclerosis affect preferentially low WSS regions, these findings imply that vessel three-dimensionality and flow pulsatility can play important protective roles in respect of these diseases. The amplitude and frequency of the velocity waveform are important parameters of the system. Increase in amplitude increases WSS and oxygen flux to the vessel wall. Increase in frequency has a small effect; it increases WSS but has no effect on the oxygen flux to the vessel wall.

Arterial geometry is commonly non-planar and associated with swirling blood flow. In this study, we examine the effect of arterial three-dimensionality on the distribution of wall shear stress (WSS) and the mass transfer of oxygen from the blood to the vessel wall in a U-bend, by modelling the blood vessels as either cylindrical or helical conduits. The results show that under physiological flow conditions, three-dimensionality can reduce both the range and extent of low WSS regions and substantially increase oxygen flux through the walls. The Sherwood number and WSS distributions between the three-dimensional helical model and a human coronary artery show remarkable qualitative agreement, implying that coronary arteries may potentially be described with a relatively simple idealized three-dimensional model, characterized by a small number of well-defined geometric parameters. The flow pattern downstream of a planar bend results in separation of the Sh number and WSS effects, a finding that implies means of investigating them individually.

Recent research on disparate psychiatric disorders has implicated rare variants in genes involved in global gene regulation and chromatin modification, as well as many common variants located primarily in regulatory regions of the genome. Understanding precisely how these variants contribute to disease will require a deeper appreciation for the mechanisms of gene regulation in the developing and adult human brain. The PsychENCODE project aims to produce a public resource of multidimensional genomic data using tissue- and cell type–specific samples from approximately 1,000 phenotypically well-characterized, high-quality healthy and disease-affected human post-mortem brains, as well as functionally characterize disease-associated regulatory elements and variants in model systems. We are beginning with a focus on autism spectrum disorder, bipolar disorder and schizophrenia, and expect that this knowledge will apply to a wide variety of psychiatric disorders. This paper outlines the motivation and design of PsychENCODE.

Tourette syndrome (TS) is a developmental neuropsychiatric disorder that is characterized by vocal and motor tics and is estimated to affect 0.2–1 % of the population. Despite evidence for a substantial genetic contribution toward disease risk, identification and replication of associations between genetic variants and TS have been challenging. Rare mutations in several genes have been identified, yet it is unclear whether these genes or their biological pathways play a role in the majority of TS cases. Similar to other complex neuropsychiatric disorders, it is likely that multiple variations in multiple genes, both within the individual and the population, may act together with environmental factors to confer risk. Centralized consortia are collecting larger patient cohorts required to identify candidate genes and biological pathways for TS. Recent profiling of gene expression in the striatum, a brain region highly implicated in TS, indicates downregulation of GABAergic and cholinergic interneurons and shows overlap with gene networks implicated by rare structural genetic variants. Together, integrated analyses of common and rare sequence and structural variation, transcriptomic variation, and bioinformatic analyses of convergent gene networks and pathways may elucidate the genetic etiology of TS.

A simple model that simulates a single biomass particle devolatilization is described. The model takes into account the main physical and chemical factors influencing the phenomenon at high temperatures (>700 K), where the production of gaseous components far outweighs that of liquids. The predictions of the model are shown to be in good agreement with published data. The model is then applied to the devolatilization of biomass in a fluidized bed, in which attention is focused on heat transfer, particle mixing and elutriation, and gas production. Predictions on the overall devolatilization time for a biomass particle are compared with experimental results obtained in a fluidized bed reactor in which the process was monitored by continuous measurement of the bed pressure. Good correspondence of predicted with calculated values was obtained, supporting the validity of the many approximations made in the derivation of the governing relationships for the pyrolysis process. On decrit un modele simple simulant une devolatilisation de particules de biomasse. Le modele prend en consideration les principaux facteurs physiques et chimiques qui influencent le phenomene a des temperatures elevees (> 700 K), alors que la production de composantes gazeuses surpasse de loin celle des liquides. On montre que les predictions du modele montre un bon accord avec des donnees publiees. Le modele est ensuite applique a la devolatilisation de biomasse dans un lit fiuidise, ou l'accent est mis sur le transfert de chaleur, le melange et la decantation des particules et la production de gaz. Des predictions du temps global de devolatilisation pour une particule de biomasse sont comparees a des resultats experimentaux obtenus dans un reacteur a lit fiuidise dans lequel le procede a ete surveille par des mesures continues de la pression du lit. On a trouve un bon accord entre les valeurs predites et les valeurs calculees, ce qui corrobore la validite des nombreuses approximations realisees dans l'estimation des relations gouvernantes le procede de pyrolyse.

Background Genome-wide association studies have not revealed any risk-conferring common genetic variants in Tourette syndrome (TS), requiring the adoption of alternative approaches to investigate the pathophysiology of this disorder. Methods We obtained the basal ganglia transcriptome by RNA sequencing in the caudate and putamen of nine TS and nine matched normal control subjects. Results We found 309 downregulated and 822 upregulated genes in the caudate and putamen (striatum) of TS individuals. Using data-driven gene network analysis, we identified 17 gene coexpression modules associated with TS. The top-scoring downregulated module in TS was enriched in striatal interneuron transcripts, which was confirmed by decreased numbers of cholinergic and gamma-aminobutyric acidergic interneurons by immunohistochemistry in the same regions. The top-scoring upregulated module was enriched in immune-related genes, consistent with activation of microglia in patients' striatum. Genes implicated by copy number variants in TS were enriched in the interneuron module, as well as in a protocadherin module. Module clustering revealed that the interneuron module was correlated with a neuronal metabolism module. Conclusions Convergence of differential expression, network analyses, and module clustering, together with copy number variants implicated in TS, strongly implicates disrupted interneuron signaling in the pathophysiology of severe TS and suggests that metabolic alterations may be linked to their death or dysfunction.

Intimal hyperplasia (IH) is a leading cause of obstruction of vascular interventions, including arterial stents, bypass grafts and arteriovenous grafts and fistulae. Proposals to account for arterial stent-associated IH include wall damage, low wall shear stress (WSS), disturbed flow and, although not widely recognized, wall hypoxia. The common non-planarity of arterial geometry and flow, led us to develop a bare-metal, nitinol, self-expanding stent with three-dimensional helical-centreline geometry. This was deployed in one common carotid artery of healthy pigs, with a straight-centreline, but otherwise identical (conventional) stent deployed contralaterally. Both stent types deformed the arteries, but the helical-centreline device additionally deformed them helically and caused swirling of intraluminal flow. At sacrifice, one month post stent deployment, histology revealed significantly less IH in the helical-centreline than straight-centreline stented vessels. Medial cross-sectional area was not significantly different in helical-centreline than straight-centreline stented vessels. By contrast, luminal cross-sectional area was significantly larger in helical-centreline than straight-centreline stented vessels. Mechanisms considered to account for those results include enhanced intraluminal WSS and enhanced intraluminal blood–vessel wall mass transport, including of oxygen, in the helical-centreline stented vessels. Consistent with the latter proposal, adventitial microvessel density was lower in the helical-centreline stented than straight-centreline stented vessels.

The problem of calculating particle trajectories on unstructured meshes using a high-order polynomial approximation of the velocity field is addressed. The calculation of the particle trajectory is based on a Runge–Kutta integration in time. A convenient way of implementing high-order approximations is to employ an auxiliary mapping that transforms a finite element into a topologically equivalent parent element within a normalized parametric space. This presents two possible choices of space in which to perform the time integration of the particle position: the physical space or the parametric space. We present algorithms for implementing both particle tracking strategies using high-order elements and discuss their merits. The main drawback of both methods is their reliance on nonlinear procedures to calculate the particle trajectory. A novel alternative hybrid approach that advances a particle in both the physical and the parametric space without requiring nonlinear iterations is proposed. The error introduced by the alternative linearized procedures and their effect in the rate of convergence of the time integration is discussed. Finally, the performance of the different algorithms is compared using a set of analytical and computational, linear and high-order, velocity fields.