A Vascular-Centric Approach to Autism Spectrum Disorders.

Brain development and function are highly reliant on adequate establishment and maintenance of vascular networks. Early impairments in vascular health can impact brain maturation and energy metabolism, which may lead to neurodevelopmental anomalies. Our recent work not only provides novel insights into the development of cerebrovascular networks but also emphasizes the importance of their well-being for proper brain maturation. In particular, we have demonstrated that endothelial dysfunction in autism spectrum disorders (ASD) mouse models is causally related to altered behavior and brain metabolism. In the prenatal human brain, vascular cells change metabolic states in the second trimester. Such findings highlight the need to identify new cellular and molecular players in neurodevelopmental disorders, raising awareness about the importance of a healthy vasculature for brain development. It is thus essential to shift the mostly neuronal point of view in research on ASD and other neurodevelopmental disorders to also include vascular and metabolic features.

Choline metabolism underpins macrophage IL-4 polarization and RELMα up-regulation in helminth infection.

Type 2 cytokines like IL-4 are hallmarks of helminth infection and activate macrophages to limit immunopathology and mediate helminth clearance. In addition to cytokines, nutrients and metabolites critically influence macrophage polarization. Choline is an essential nutrient known to support normal macrophage responses to lipopolysaccharide; however, its function in macrophages polarized by type 2 cytokines is unknown. Using murine IL-4-polarized macrophages, targeted lipidomics revealed significantly elevated levels of phosphatidylcholine, with select changes to other choline-containing lipid species. These changes were supported by the coordinated up-regulation of choline transport compared to naïve macrophages. Pharmacological inhibition of choline metabolism significantly suppressed several mitochondrial transcripts and dramatically inhibited select IL-4-responsive transcripts, most notably, Retnla. We further confirmed that blocking choline metabolism diminished IL-4-induced RELMα (encoded by Retnla) protein content and secretion and caused a dramatic reprogramming toward glycolytic metabolism. To better understand the physiological implications of these observations, naïve or mice infected with the intestinal helminth Heligmosomoides polygyrus were treated with the choline kinase α inhibitor, RSM-932A, to limit choline metabolism in vivo. Pharmacological inhibition of choline metabolism lowered RELMα expression across cell-types and tissues and led to the disappearance of peritoneal macrophages and B-1 lymphocytes and an influx of infiltrating monocytes. The impaired macrophage activation was associated with some loss in optimal immunity to H. polygyrus, with increased egg burden. Together, these data demonstrate that choline metabolism is required for macrophage RELMα induction, metabolic programming, and peritoneal immune homeostasis, which could have important implications in the context of other models of infection or cancer immunity.

Astroglial Hmgb1 regulates postnatal astrocyte morphogenesis and cerebrovascular maturation.

Astrocytes are intimately linked with brain blood vessels, an essential relationship for neuronal function. However, astroglial factors driving these physical and functional associations during postnatal brain development have yet to be identified. By characterizing structural and transcriptional changes in mouse cortical astrocytes during the first two postnatal weeks, we find that high-mobility group box 1 (Hmgb1), normally upregulated with injury and involved in adult cerebrovascular repair, is highly expressed in astrocytes at birth and then decreases rapidly. Astrocyte-selective ablation of Hmgb1 at birth affects astrocyte morphology and endfoot placement, alters distribution of endfoot proteins connexin43 and aquaporin-4, induces transcriptional changes in astrocytes related to cytoskeleton remodeling, and profoundly disrupts endothelial ultrastructure. While lack of astroglial Hmgb1 does not affect the blood-brain barrier or angiogenesis postnatally, it impairs neurovascular coupling and behavior in adult mice. These findings identify astroglial Hmgb1 as an important player in postnatal gliovascular maturation.

16p11.2 haploinsufficiency reduces mitochondrial biogenesis in brain endothelial cells and alters brain metabolism in adult mice.

Neurovascular abnormalities in mouse models of 16p11.2 deletion autism syndrome are reminiscent of alterations reported in murine models of glucose transporter deficiency, including reduced brain angiogenesis and behavioral alterations. Yet, whether cerebrovascular alterations in 16p11.2df/+ mice affect brain metabolism is unknown. Here, we report that anesthetized 16p11.2df/+ mice display elevated brain glucose uptake, a phenomenon recapitulated in mice with endothelial-specific 16p11.2 haplodeficiency. Awake 16p11.2df/+ mice display attenuated relative fluctuations of extracellular brain glucose following systemic glucose administration. Targeted metabolomics on cerebral cortex extracts reveals enhanced metabolic responses to systemic glucose in 16p11.2df/+ mice that also display reduced mitochondria number in brain endothelial cells. This is not associated with changes in mitochondria fusion or fission proteins, but 16p11.2df/+ brain endothelial cells lack the splice variant NT-PGC-1α, suggesting defective mitochondrial biogenesis. We propose that altered brain metabolism in 16p11.2df/+ mice is compensatory to endothelial dysfunction, shedding light on previously unknown adaptative responses.

[Description of a Pilot Project for Pediatric Occupational Therapy in Daycare and Community Settings]. / Description d'un projet pilote d'ergothérapie pédiatrique en milieux de garde et communautaires.

Background. Occupational therapy interventions that promote and prevent children's health and well-being aim to reduce health inequalities and foster protective factors. The purpose of this study is to describe a pilot community-based occupational therapy project for preschoolers in partnership with community organizations and childcare services. Method. A participatory action research approach was implemented with support from an advisory committee. An occupational therapist provided community-based occupational therapy services in a tiered organization model over one year. Findings. Services were offered in three tiers: 7 awareness workshops for parents and caregivers (Tier 1), 57 visits and 27 consultations in 8 community agencies (Tier 2), and 23 individual follow-ups (Tier 3). Implications. There is an opportunity to implement with community agencies and daycare settings an occupational therapy service based on community-based rehabilitation for children under 5 years of age.

Description. Les interventions ergothérapiques en promotion et prévention de la santé et du bien-être des enfants visent à réduire les inégalités de santé et favoriser les facteurs de protection. But. Cette étude vise à décrire un projet pilote de service d'ergothérapie à base communautaire, pour les enfants d'âge préscolaire, en partenariat avec les organismes communautaires et les milieux de garde. Méthodologie. Une approche de recherche-action participative avec soutien d'un comité aviseur a été mis en place. Une ergothérapeute a offert des services d'ergothérapie communautaire selon un modèle d'organisation par paliers durant un an. Résultats. Les services étaient offerts selon trois paliers : 7 ateliers de sensibilisation pour parents et intervenants (palier 1), 57 visites et 27 consultations dans 8 organismes communautaires (palier 2) et 23 suivis individuels (palier 3). Conséquences. Il est possible d'implanter un service d'ergothérapie s"inspirant de la réadaptation à base communautaire avec les organismes communautaires offrant des services aux enfants de 5 et moins et les milieux de garde.

Neonatal hyperoxia in mice triggers long-term cognitive deficits via impairments in cerebrovascular function and neurogenesis.

Preterm birth is the leading cause of death in children under 5 years of age. Premature infants who receive life-saving oxygen therapy often develop bronchopulmonary dysplasia (BPD), a chronic lung disease. Infants with BPD are at a high risk of abnormal neurodevelopment, including motor and cognitive difficulties. While neural progenitor cells (NPCs) are crucial for proper brain development, it is unclear whether they play a role in BPD-associated neurodevelopmental deficits. Here, we show that hyperoxia-induced experimental BPD in newborn mice led to lifelong impairments in cerebrovascular structure and function as well as impairments in NPC self-renewal and neurogenesis. A neurosphere assay utilizing nonhuman primate preterm baboon NPCs confirmed impairment in NPC function. Moreover, gene expression profiling revealed that genes involved in cell proliferation, angiogenesis, vascular autoregulation, neuronal formation, and neurotransmission were dysregulated following neonatal hyperoxia. These impairments were associated with motor and cognitive decline in aging hyperoxia-exposed mice, reminiscent of deficits observed in patients with BPD. Together, our findings establish a relationship between BPD and abnormal neurodevelopmental outcomes and identify molecular and cellular players of neonatal brain injury that persist throughout adulthood that may be targeted for early intervention to aid this vulnerable patient population.

An analysis of the influence of transfer learning when measuring the tortuosity of blood vessels.

BACKGROUND AND OBJECTIVE: Convolutional Neural Networks (CNNs) can provide excellent results regarding the segmentation of blood vessels. One important aspect of CNNs is that they can be trained on large amounts of data and then be made available, for instance, in image processing software. The pre-trained CNNs can then be easily applied in downstream blood vessel characterization tasks, such as the calculation of the length, tortuosity, or caliber of the blood vessels. Yet, it is still unclear if pre-trained CNNs can provide robust, unbiased, results in downstream tasks involving the morphological analysis of blood vessels. Here, we focus on measuring the tortuosity of blood vessels and investigate to which extent CNNs may provide biased tortuosity values even after fine-tuning the network to a new dataset under study. METHODS: We develop a procedure for quantifying the influence of CNN pre-training in downstream analyses involving the measurement of morphological properties of blood vessels. Using the methodology, we compare the performance of CNNs that were trained on images containing blood vessels having high tortuosity with CNNs that were trained on blood vessels with low tortuosity and fine-tuned on blood vessels with high tortuosity. The opposite situation is also investigated. RESULTS: We show that the tortuosity values obtained by a CNN trained from scratch on a dataset may not agree with those obtained by a fine-tuned network that was pre-trained on a dataset having different tortuosity statistics. In addition, we show that improving the segmentation accuracy does not necessarily lead to better tortuosity estimation. To mitigate the aforementioned issues, we propose the application of data augmentation techniques even in situations where they do not improve segmentation performance. For instance, we found that the application of elastic transformations was enough to prevent an underestimation of 8% of blood vessel tortuosity when applying CNNs to different datasets. CONCLUSIONS: The results highlight the importance of developing new methodologies for training CNNs with the specific goal of reducing the error of morphological measurements, as opposed to the traditional approach of using segmentation accuracy as a proxy metric for performance evaluation.

Isolation and functional characterization of primary endothelial cells from mouse cerebral cortex.

Endothelial cells (ECs) lining blood vessels are implicated in organ development, function, and maintenance. We present a detailed protocol enabling isolation and characterization of primary mouse brain ECs, including quality controls and functional assays. These procedures promote survival of primary brain ECs for the assessment of endothelial health. Since alterations in brain ECs are involved in the onset and progression of neurological disorders, this protocol represents a valuable tool to better understand the roles of ECs in brain health. For complete details on the use and execution of this profile, please refer to Ouellette et al. (2020).

From Neurodevelopmental to Neurodegenerative Disorders: The Vascular Continuum.

Structural and functional integrity of the cerebral vasculature ensures proper brain development and function, as well as healthy aging. The inability of the brain to store energy makes it exceptionally dependent on an adequate supply of oxygen and nutrients from the blood stream for matching colossal demands of neural and glial cells. Key vascular features including a dense vasculature, a tightly controlled environment, and the regulation of cerebral blood flow (CBF) all take part in brain health throughout life. As such, healthy brain development and aging are both ensured by the anatomical and functional interaction between the vascular and nervous systems that are established during brain development and maintained throughout the lifespan. During critical periods of brain development, vascular networks remodel until they can actively respond to increases in neural activity through neurovascular coupling, which makes the brain particularly vulnerable to neurovascular alterations. The brain vasculature has been strongly associated with the onset and/or progression of conditions associated with aging, and more recently with neurodevelopmental disorders. Our understanding of cerebrovascular contributions to neurological disorders is rapidly evolving, and increasing evidence shows that deficits in angiogenesis, CBF and the blood-brain barrier (BBB) are causally linked to cognitive impairment. Moreover, it is of utmost curiosity that although neurodevelopmental and neurodegenerative disorders express different clinical features at different stages of life, they share similar vascular abnormalities. In this review, we present an overview of vascular dysfunctions associated with neurodevelopmental (autism spectrum disorders, schizophrenia, Down Syndrome) and neurodegenerative (multiple sclerosis, Huntington's, Parkinson's, and Alzheimer's diseases) disorders, with a focus on impairments in angiogenesis, CBF and the BBB. Finally, we discuss the impact of early vascular impairments on the expression of neurodegenerative diseases.

Vascular contributions to 16p11.2 deletion autism syndrome modeled in mice.

While the neuronal underpinnings of autism spectrum disorder (ASD) are being unraveled, vascular contributions to ASD remain elusive. Here, we investigated postnatal cerebrovascular development in the 16p11.2df/+ mouse model of 16p11.2 deletion ASD syndrome. We discover that 16p11.2 hemizygosity leads to male-specific, endothelium-dependent structural and functional neurovascular abnormalities. In 16p11.2df/+ mice, endothelial dysfunction results in impaired cerebral angiogenesis at postnatal day 14, and in altered neurovascular coupling and cerebrovascular reactivity at postnatal day 50. Moreover, we show that there is defective angiogenesis in primary 16p11.2df/+ mouse brain endothelial cells and in induced-pluripotent-stem-cell-derived endothelial cells from human carriers of the 16p11.2 deletion. Finally, we find that mice with an endothelium-specific 16p11.2 deletion (16p11.2ΔEC) partially recapitulate some of the behavioral changes seen in 16p11.2 syndrome, specifically hyperactivity and impaired motor learning. By showing that developmental 16p11.2 haploinsufficiency from endothelial cells results in neurovascular and behavioral changes in adults, our results point to a potential role for endothelial impairment in ASD.