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Nature Communications Jun 2024To uncover molecular changes underlying blood-brain-barrier dysfunction in Alzheimer's disease, we performed single nucleus RNA sequencing in 24 Alzheimer's disease and...
To uncover molecular changes underlying blood-brain-barrier dysfunction in Alzheimer's disease, we performed single nucleus RNA sequencing in 24 Alzheimer's disease and control brains and focused on vascular and astrocyte clusters as main cell types of blood-brain-barrier gliovascular-unit. The majority of the vascular transcriptional changes were in pericytes. Of the vascular molecular targets predicted to interact with astrocytic ligands, SMAD3, upregulated in Alzheimer's disease pericytes, has the highest number of ligands including VEGFA, downregulated in Alzheimer's disease astrocytes. We validated these findings with external datasets comprising 4,730 pericyte and 150,664 astrocyte nuclei. Blood SMAD3 levels are associated with Alzheimer's disease-related neuroimaging outcomes. We determined inverse relationships between pericytic SMAD3 and astrocytic VEGFA in human iPSC and zebrafish models. Here, we detect vast transcriptome changes in Alzheimer's disease at the gliovascular-unit, prioritize perturbed pericytic SMAD3-astrocytic VEGFA interactions, and validate these in cross-species models to provide a molecular mechanism of blood-brain-barrier disintegrity in Alzheimer's disease.
Topics: Alzheimer Disease; Humans; Blood-Brain Barrier; Smad3 Protein; Zebrafish; Astrocytes; Vascular Endothelial Growth Factor A; Animals; Pericytes; Male; Induced Pluripotent Stem Cells; Female; Aged; Transcriptome; Brain; Aged, 80 and over; Disease Models, Animal
PubMed: 38902234
DOI: 10.1038/s41467-024-48926-6 -
Nature Communications Jun 2024Mutations in the FOXF1 gene, a key transcriptional regulator of pulmonary vascular development, cause Alveolar Capillary Dysplasia with Misalignment of Pulmonary Veins,...
Mutations in the FOXF1 gene, a key transcriptional regulator of pulmonary vascular development, cause Alveolar Capillary Dysplasia with Misalignment of Pulmonary Veins, a lethal lung disease affecting newborns and infants. Identification of new FOXF1 upstream regulatory elements is critical to explain why frequent non-coding FOXF1 deletions are linked to the disease. Herein, we use multiome single-nuclei RNA and ATAC sequencing of mouse and human patient lungs to identify four conserved endothelial and mesenchymal FOXF1 enhancers. We demonstrate that endothelial FOXF1 enhancers are autoactivated, whereas mesenchymal FOXF1 enhancers are regulated by EBF1 and GLI1. The cell-specificity of FOXF1 enhancers is validated by disrupting these enhancers in mouse embryonic stem cells using CRISPR/Cpf1 genome editing followed by lineage-tracing of mutant embryonic stem cells in mouse embryos using blastocyst complementation. This study resolves an important clinical question why frequent non-coding FOXF1 deletions that interfere with endothelial and mesenchymal enhancers can lead to the disease.
Topics: Forkhead Transcription Factors; Animals; Humans; Persistent Fetal Circulation Syndrome; Mice; Enhancer Elements, Genetic; Mesoderm; Lung; Endothelial Cells; Zinc Finger Protein GLI1; Embryonic Stem Cells; Pulmonary Alveoli
PubMed: 38898031
DOI: 10.1038/s41467-024-49477-6 -
BioRxiv : the Preprint Server For... Jun 2024The teleost consists of surface dwelling (surface fish) and cave dwelling (cavefish) forms. Cavefish have evolved in subterranean habitats characterized by reduced...
UNLABELLED
The teleost consists of surface dwelling (surface fish) and cave dwelling (cavefish) forms. Cavefish have evolved in subterranean habitats characterized by reduced oxygen levels (hypoxia) and show constructive and regressive phenotypic traits controlled by increased Sonic hedgehog (Shh) signaling along the embryonic midline. The enhancement of primitive hematopoietic domains, which are formed bilaterally in the anterior and posterior lateral plate mesoderm, are responsible for the development of more larval erythrocytes in cavefish relative to surface fish. In this study, we determine the role of hypoxia and Shh signaling in the development and evolution of primitive hematopoiesis in cavefish. We show that hypoxia treatment during embryogenesis increases primitive hematopoiesis and erythrocyte development in surface fish. We also demonstrate that upregulation of Shh midline signaling by treatment with the Smoothened agonist SAG increases primitive hematopoiesis and erythrocyte development in surface fish, whereas Shh downregulation via treatment with the Smoothened inhibitor cyclopamine decreases these traits in cavefish. Together these results suggest that hematopoietic enhancement is regulated by hypoxia and the Shh signaling system. Lastly, we demonstrate that hypoxia treatment enhances expression of Shh signaling along the midline of surface fish embryos. Thus, we conclude that a hypoxia-Shh axis may drive the adaptive evolution of primitive hematopoiesis and erythrocyte development in cavefish.
HIGHLIGHTS
Hypoxia increases hematopoiesis and erythrocytes in surface fishShh upregulation increases hematopoiesis and erythrocytes in surface fishShh inhibition decreases hematopoiesis and erythrocytes in cavefishHypoxia upregulates Shh along the embryonic midline in surface fish.
PubMed: 38895301
DOI: 10.1101/2024.06.09.598120 -
International Journal of Molecular... May 2024Derived from axial structures, Sonic Hedgehog (Shh) is secreted into the paraxial mesoderm, where it plays crucial roles in sclerotome induction and myotome...
Derived from axial structures, Sonic Hedgehog (Shh) is secreted into the paraxial mesoderm, where it plays crucial roles in sclerotome induction and myotome differentiation. Through conditional loss-of-function in quail embryos, we investigate the timing and impact of Shh activity during early formation of sclerotome-derived vertebrae and ribs, and of lateral mesoderm-derived sternum. To this end, Hedgehog interacting protein (Hhip) was electroporated at various times between days 2 and 5. While the vertebral body and rib primordium showed consistent size reduction, rib expansion into the somatopleura remained unaffected, and the sternal bud developed normally. Additionally, we compared these effects with those of locally inhibiting BMP activity. Transfection of Noggin in the lateral mesoderm hindered sternal bud formation. Unlike Hhip, BMP inhibition via Noggin or Smad6 induced myogenic differentiation of the lateral dermomyotome lip, while impeding the growth of the myotome/rib complex into the somatic mesoderm, thus affirming the role of the lateral dermomyotome epithelium in rib guidance. Overall, these findings underscore the continuous requirement for opposing gradients of Shh and BMP activity in the morphogenesis of proximal and distal flank skeletal structures, respectively. Future research should address the implications of these early interactions to the later morphogenesis and function of the musculo-skeletal system and of possible associated malformations.
Topics: Animals; Hedgehog Proteins; Ribs; Spine; Gene Expression Regulation, Developmental; Mesoderm; Quail; Somites; Bone Morphogenetic Proteins; Carrier Proteins
PubMed: 38891790
DOI: 10.3390/ijms25115602 -
Nature Communications Jun 2024Cell-fate decisions during mammalian gastrulation are poorly understood outside of rodent embryos. The embryonic disc of pig embryos mirrors humans, making them a useful... (Comparative Study)
Comparative Study
Cell-fate decisions during mammalian gastrulation are poorly understood outside of rodent embryos. The embryonic disc of pig embryos mirrors humans, making them a useful proxy for studying gastrulation. Here we present a single-cell transcriptomic atlas of pig gastrulation, revealing cell-fate emergence dynamics, as well as conserved and divergent gene programs governing early porcine, primate, and murine development. We highlight heterochronicity in extraembryonic cell-types, despite the broad conservation of cell-type-specific transcriptional programs. We apply these findings in combination with functional investigations, to outline conserved spatial, molecular, and temporal events during definitive endoderm specification. We find early FOXA2 + /TBXT- embryonic disc cells directly form definitive endoderm, contrasting later-emerging FOXA2/TBXT+ node/notochord progenitors. Unlike mesoderm, none of these progenitors undergo epithelial-to-mesenchymal transition. Endoderm/Node fate hinges on balanced WNT and hypoblast-derived NODAL, which is extinguished upon endodermal differentiation. These findings emphasise the interplay between temporal and topological signalling in fate determination during gastrulation.
Topics: Animals; Gastrulation; Endoderm; Swine; Single-Cell Analysis; Gene Expression Regulation, Developmental; Mice; Embryo, Mammalian; Cell Differentiation; Mesoderm; Transcriptome; Hepatocyte Nuclear Factor 3-beta; Cell Lineage; T-Box Domain Proteins; Epithelial-Mesenchymal Transition
PubMed: 38890321
DOI: 10.1038/s41467-024-49407-6 -
Biology Open Jun 2024Regular spatial patterns are ubiquitous forms of organization in nature. In animals, regular patterns can be found from the cellular scale to the tissue scale, and from...
Regular spatial patterns are ubiquitous forms of organization in nature. In animals, regular patterns can be found from the cellular scale to the tissue scale, and from early stages of development to adulthood. To understand the formation of these patterns, how they assemble and mature, and how they are affected by perturbations, a precise quantitative description of the patterns is essential. However, accessible tools that offer in-depth analysis without the need for computational skills are lacking for biologists. Here, we present PatternJ, a novel toolset to analyze regular one-dimensional patterns precisely and automatically. This toolset, to be used with the popular imaging processing program ImageJ/Fiji, facilitates the extraction of key geometric features within and between pattern repeats in static images and time-lapse series. We validate PatternJ with simulated data and test it on images of sarcomeres from insect muscles and contracting cardiomyocytes, actin rings in neurons, and somites from zebrafish embryos obtained using confocal fluorescence microscopy, STORM, electron microscopy, and brightfield imaging. We show that the toolset delivers subpixel feature extraction reliably even with images of low signal-to-noise ratio. PatternJ's straightforward use and functionalities make it valuable for various scientific fields requiring quantitative one-dimensional pattern analysis, including the sarcomere biology of muscles or the patterning of mammalian axons, speeding up discoveries with the bonus of high reproducibility.
Topics: Animals; Axons; Image Processing, Computer-Assisted; Zebrafish; Sarcomeres; Somites; Software; Algorithms
PubMed: 38887972
DOI: 10.1242/bio.060548 -
Frontiers in Cell and Developmental... 2024Several differentiation protocols have enabled the generation of intermediate mesoderm (IM)-derived cells from human pluripotent stem cells (hPSC). However, the...
Several differentiation protocols have enabled the generation of intermediate mesoderm (IM)-derived cells from human pluripotent stem cells (hPSC). However, the substantial variability between existing protocols for generating IM cells compromises their efficiency, reproducibility, and overall success, potentially hindering the utility of urogenital system organoids. Here, we examined the role of high levels of Nodal signaling and BMP activity, as well as WNT signaling in the specification of IM cells derived from a UCSD167i-99-1 human induced pluripotent stem cells (hiPSC) line. We demonstrate that precise modulation of WNT and BMP signaling significantly enhances IM differentiation efficiency. Treatment of hPSC with 3 μM CHIR99021 induced TBXT+/MIXL1+ mesoderm progenitor (MP) cells after 48 h of differentiation. Further treatment with a combination of 3 μM CHIR99021 and 4 ng/mL BMP4 resulted in the generation of OSR1+/GATA3+/PAX2+ IM cells within a subsequent 48 h period. Molecular characterization of differentiated cells was confirmed through immunofluorescence staining and RTqPCR. Hence, this study establishes a consistent and reproducible protocol for differentiating hiPSC into IM cells that faithfully recapitulates the molecular signatures of IM development. This protocol holds promise for improving the success of protocols designed to generate urogenital system organoids , with potential applications in regenerative medicine, drug discovery, and disease modeling.
PubMed: 38887514
DOI: 10.3389/fcell.2024.1395723 -
Brain Research Bulletin Nov 2023This study was designed to investigate the role of pericytes in the pathogenesis of perioperative neurocognitive disorder (PND).
AIMS
This study was designed to investigate the role of pericytes in the pathogenesis of perioperative neurocognitive disorder (PND).
METHODS
In this study, we established a PND model via sevoflurane anesthesia and tibial fracture surgery in 2-month-old and 16-month-old male C57BL/6 mice. On the third postoperative day, the mice were subjected to behavioral testing or sacrificed to collect brain tissue. The progression of hippocampal blood-brain barrier (BBB) disruption and neuroinflammation was detected using transmission electron microscope and immunofluorescence. We also used western blotting to measure the levels of plasma-derived protein immunoglobulin G (IgG) and albumin in the hippocampus to assess the leakage of the BBB.
RESULTS
Aged mice did not experience age-related cognitive decline and BBB disruption compared with younger mice but only increased glial cell activity. Anesthesia/Surgery damaged cognitive function, reduced pericyte coverage, decreased the length of capillaries and levels of occludin and claudin-5, destroyed the structure of the BBB, exacerbated IgG and albumin accumulation in the hippocampus, and enhanced the activation of microglia and astrocytes in the hippocampus of aged mice. However, these negative effects did not occur in young mice.
CONCLUSION
Our study showed that the loss of pericytes led to increased BBB permeability and neuroinflammation after anesthesia/surgery in aged mice, ultimately resulting in cognitive dysfunction.
Topics: Animals; Blood-Brain Barrier; Pericytes; Male; Mice; Mice, Inbred C57BL; Hippocampus; Cognitive Dysfunction; Cognition; Aging; Sevoflurane; Anesthesia
PubMed: 38867419
DOI: 10.1016/j.brainresbull.2023.110799 -
Frontiers in Cell and Developmental... 2024Vertebrate body axis formation initiates during gastrulation and continues within the tail bud at the posterior end of the embryo. Major structures in the trunk are...
Vertebrate body axis formation initiates during gastrulation and continues within the tail bud at the posterior end of the embryo. Major structures in the trunk are paired somites, which generate the musculoskeletal system, the spinal cord-forming part of the central nervous system, and the notochord, with important patterning functions. The specification of these different cell lineages by key signalling pathways and transcription factors is essential, however, a global map of cell types and expressed genes in the avian trunk is missing. Here we use high-throughput sequencing approaches to generate a molecular map of the emerging trunk and tailbud in the chick embryo. Single cell RNA-sequencing (scRNA-seq) identifies discrete cell lineages including somites, neural tube, neural crest, lateral plate mesoderm, ectoderm, endothelial and blood progenitors. In addition, RNA-seq of sequential tissue sections (RNA-tomography) provides a spatially resolved, genome-wide expression dataset for the avian tailbud and emerging body, comparable to other model systems. Combining the single cell and RNA-tomography datasets, we identify spatially restricted genes, focusing on somites and early myoblasts. Thus, this high-resolution transcriptome map incorporating cell types in the embryonic trunk can expose molecular pathways involved in body axis development.
PubMed: 38863942
DOI: 10.3389/fcell.2024.1382960 -
Science Progress 2024Pericytes (PCs) are versatile cells integral to the microcirculation wall, exhibiting specific stem cell traits. They are essential in modulating blood flow, ensuring... (Review)
Review
Pericytes (PCs) are versatile cells integral to the microcirculation wall, exhibiting specific stem cell traits. They are essential in modulating blood flow, ensuring vascular permeability, maintaining homeostasis, and aiding tissue repair process. Given their involvement in numerous disease-related pathological and physiological processes, the regulation of PCs has emerged as a focal point of research. Adenomyosis is characterized by the presence of active endometrial glands and stroma encased by an enlarged and proliferative myometrial layer, further accompanied by fibrosis and new blood vessel formation. This distinct pathological condition might be intricately linked with PCs. This article comprehensively reviews the markers associated with PCs, their contributions to angiogenesis, blood flow modulation, and fibrotic processes. Moreover, it provides a comprehensive overview of the current research on adenomyosis pathophysiology, emphasizing the potential correlation and future implications regarding PCs and the development of adenomyosis.
Topics: Adenomyosis; Pericytes; Humans; Female; Neovascularization, Pathologic; Animals; Fibrosis; Endometrium; Myometrium; Biomarkers
PubMed: 38863331
DOI: 10.1177/00368504241257126