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The development of the vascular system is crucial in supporting the growth and health of all other organs in the body, and vascular system dysfunction is the major cause of human morbidity and mortality. This chapter discusses three successive processes that govern vascular system development, starting with the differentiation of the primitive vascular system in early embryonic development, followed by its remodeling into a functional circulatory system composed of arteries and veins, and its final maturation and acquisition of an organ specific semi-permeable barrier that controls nutrient uptake into tissues and hence controls organ physiology. Along these steps, endothelial cells forming the inner lining of all blood vessels acquire extensive heterogeneity in terms of gene expression patterns and function, that we are only beginning to understand. These advances contribute to overall knowledge of vascular biology and are predicted to unlock the unprecedented therapeutic potential of the endothelium as an avenue for treatment of diseases associated with dysfunctional vasculature.

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BACKGROUND: Acute kidney injury (AKI) in sepsis patients increases patient mortality. Endothelial cells are important players in the pathophysiology of sepsis-associated AKI (SA-AKI), yet knowledge regarding their spatiotemporal involvement in coagulation disbalance and leukocyte recruitment is lacking. This study investigated the identity and kinetics of responses of different microvascular compartments in kidney cortex in response to SA-AKI. METHODS: Laser microdissected arterioles, glomeruli, peritubular capillaries, and postcapillary venules from kidneys of mice subjected to cecal ligation and puncture (CLP) were analyzed using RNA sequencing. Differential expression and pathway enrichment analyses identified genes involved in coagulation and inflammation. A selection of these genes was evaluated by RT-qPCR in microvascular compartments of renal biopsies from patients with SA-AKI. The role of two identified genes in lipopolysaccharide-induced endothelial coagulation and inflammatory activation were determined in vitro in HUVEC using siRNA-based gene silencing. RESULTS: CLP-sepsis in mice induced altered expression of approximately 400 genes in the renal microvasculature, with microvascular compartments exhibiting unique spatiotemporal responses. In mice, changes in gene expression related to coagulation and inflammation were most extensive in glomeruli at early and intermediate time points, with high induction of Plat, Serpine1, Thbd, Icam1, Stat3, and Ifitm3. In human SA-AKI, PROCR and STAT3 were induced in postcapillary venules, while SERPINE1 expression was diminished. IFITM3 was increased in arterioles and glomeruli. In vitro studies revealed that STAT3 and IFITM3 partly control endothelial coagulation and inflammatory activation. CONCLUSION: Renal microvascular compartments in mice and humans exhibited heterogeneous changes in coagulation- and inflammation-related gene expression in response to SA-AKI. Additional research should aim at understanding the functional consequences of the here described heterogeneous microvascular responses to establish the usefulness of identified genes as therapeutic targets in SA-AKI.

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Choroidal neovascularization (CNV) is a hallmark of neovascular age-related macular degeneration (nAMD) and a major contributor to vision loss in nAMD cases. However, the identification of specific cell types associated with nAMD remains challenging. Herein, we performed single-cell sequencing to comprehensively explore the cellular diversity and understand the foundational components of the retinal pigment epithelium (RPE)/choroid complex. We unveiled 10 distinct cell types within the RPE/choroid complex. Notably, we observed significant heterogeneity within endothelial cells (ECs), fibroblasts, and macrophages, underscoring the intricate nature of the cellular composition in the RPE/choroid complex. Within the EC category, four distinct clusters were identified and EC cluster 0 was tightly associated with choroidal neovascularization. We identified five clusters of fibroblasts actively involved in the pathogenesis of nAMD, influencing fibrotic responses, angiogenic effects, and photoreceptor function. Additionally, three clusters of macrophages were identified, suggesting their potential roles in regulating the progression of nAMD through immunomodulation and inflammation regulation. Through CellChat analysis, we constructed a complex cell-cell communication network, revealing the role of EC clusters in interacting with fibroblasts and macrophages in the context of nAMD. These interactions were found to govern angiogenic effects, fibrotic responses, and inflammatory processes. In summary, this study reveals noteworthy cellular heterogeneity in the RPE/choroid complex and provides valuable insights into the pathogenesis of CNV. These findings will open up potential avenues for deep understanding and targeted therapeutic interventions in nAMD.

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Major adverse cardiovascular events occurring upon coronary artery bypass graft surgery are typically accompanied by endothelial dysfunction. Total arterial revascularisation, which employs both left and right internal thoracic arteries instead of the saphenous vein to create a bypass, is associated with better mid- and long-term outcomes. We suggested that molecular profiles of human coronary artery endothelial cells (HCAECs) and human internal mammary artery endothelial cells (HITAECs) are coherent in terms of transcriptomic and proteomic signatures, which were then investigated by RNA sequencing and ultra-high performance liquid chromatography-mass spectrometry, respectively. Both HCAECs and HITAECs overexpressed molecules responsible for the synthesis of extracellular matrix (ECM) components, basement membrane assembly, cell-ECM adhesion, organisation of intercellular junctions, and secretion of extracellular vesicles. HCAECs were characterised by higher enrichment with molecular signatures of basement membrane construction, collagen biosynthesis and folding, and formation of intercellular junctions, whilst HITAECs were notable for augmented pro-inflammatory signaling, intensive synthesis of proteins and nitrogen compounds, and enhanced ribosome biogenesis. Despite HCAECs and HITAECs showing a certain degree of molecular heterogeneity, no specific markers at the protein level have been identified. Coherence of differentially expressed molecular categories in HCAECs and HITAECs suggests synergistic interactions between these ECs in a bypass surgery scenario.

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The use of cardiovascular implants is commonplace in clinical practice. However, reproducing the key bioactive and adaptive properties of native cardiovascular tissues with an artificial replacement is highly challenging. Exciting new treatment strategies are under development to regenerate (parts of) cardiovascular tissues directly in situ using immunomodulatory biomaterials. Direct exposure to the bloodstream and hemodynamic loads is a particular challenge, given the risk of thrombosis and adverse remodeling that it brings. However, the blood is also a source of (immune) cells and proteins that dominantly contribute to functional tissue regeneration. This review explores the potential of the blood as a source for the complete or partial in situ regeneration of cardiovascular tissues, with a particular focus on the endothelium, being the natural blood-tissue barrier. We pinpoint the current scientific challenges to enable rational engineering and testing of blood-contacting implants to leverage the regenerative potential of the blood.

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Endothelial cells (ECs) in the microvasculature in organs are active participants in the pathophysiology of sepsis. Tyrosine protein kinase receptor Tie2 (Tek; Tunica interna Endothelial cell Kinase) is thought to play a role in their inflammatory response, yet data are inconclusive. We investigated acute endotoxemia-induced changes in the expression of Tie2 and inflammation-associated endothelial adhesion molecules E-selectin and VCAM-1 (vascular cell adhesion molecule-1) in kidneys and lungs in inducible, EC-specific Tie2 knockout mice. The extent of Tie2 knockout in healthy mice differed between microvascular beds, with low to absent expression in arterioles in kidneys and in capillaries in lungs. In kidneys, Tie2 mRNA dropped more than 70% upon challenge with lipopolysaccharide (LPS) in both genotypes, with no change in protein. In renal arterioles, tamoxifen-induced Tie2 knockout was associated with higher VCAM-1 protein expression in healthy conditions. This did not increase further upon challenge of mice with LPS, in contrast to the increased expression occurring in control mice. Also, in lungs, Tie2 mRNA levels dropped within 4 h after LPS challenge in both genotypes, while Tie2 protein levels did not change. In alveolar capillaries, where tamoxifen-induced Tie2 knockout did not affect the basal expression of either adhesion molecule, a 4-fold higher E-selectin protein expression was observed after exposure to LPS compared to controls. The here-revealed heterogeneous effects of absence of Tie2 in ECs in kidney and lung microvasculature in health and in response to acute inflammatory activation calls for further in vivo investigations into the role of Tie2 in EC behavior.

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The hemostatic system involves an array of circulating coagulation factors that work in concert with platelets and the vascular endothelium to promote clotting in a space- and time-defined manner. Despite equal systemic exposure to circulating factors, bleeding and thrombotic diseases tend to prefer specific sites, suggesting an important role for local factors. This may be provided by endothelial heterogeneity. Endothelial cells differ not only between arteries, veins, and capillaries but also between microvascular beds from different organs, which present unique organotypic morphology and functional and molecular profiles. Accordingly, regulators of hemostasis are not uniformly distributed in the vasculature. The establishment and maintenance of endothelial diversity are orchestrated at the transcriptional level. Recent transcriptomic and epigenomic studies have provided a global picture of endothelial cell heterogeneity. In this review, we discuss the organotypic differences in the hemostatic profile of endothelial cells; we focus on 2 major endothelial regulators of hemostasis, namely von Willebrand factor and thrombomodulin, to provide examples of transcriptional mechanisms that control heterogeneity; finally, we consider some of the methodological challenges and opportunities for future studies.

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Diabetic retinopathy (DR), a leading cause of vision loss in working-age adults, induces mosaic patterns of vasculopathy that may be associated with spatial heterogeneity of intraretinal endothelial cells. We recently reported that secretogranin III (Scg3), a neuron-derived angiogenic and vascular leakage factor, selectively binds retinal vessels of diabetic but not healthy mice. Here, we investigated endothelial heterogeneity of three retinal vascular plexuses in DR pathogenesis and the therapeutic implications. Our unique in vivo ligand binding assay detected a 22.7-fold increase in Scg3 binding to retinal vessels of diabetic mice relative to healthy mice. Functional immunohistochemistry revealed that Scg3 predominantly binds to the DR-stressed CD31- deep retinal vascular plexus but not to the relatively healthy CD31+ superficial and intermediate plexuses within the same diabetic retina. In contrast, VEGF bound to healthy and diabetic retinal vessels indiscriminately with low activity. FITC-dextran assays indicated that selectively increased retinal vascular leakage coincides with Scg3 binding in diabetic mice that was independent of VEGF, whereas VEGF-induced leakage did not distinguish between diabetic and healthy mice. Dose-response curves showed that the anti-Scg3 humanized antibody (hAb) and anti-VEGF aflibercept alleviated DR leakage with equivalent efficacies, and that the combination acted synergistically. These findings suggest: (i) the deep plexus is highly sensitive to DR; (ii) Scg3 binding to the DR deep plexus coincides with the loss of CD31 and compromised endothelial junctions; (iii) anti-Scg3 hAb alleviates vascular leakage by selectively targeting the DR-stressed deep plexus within the same diabetic retina; (iv) combined anti-Scg3 and anti-VEGF treatments synergistically ameliorate DR through distinct mechanisms.

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Lungs are constantly exposed to environmental perturbations and therefore have remarkable capacity to regenerate in response to injury. Sustained lung injuries, aging, and increased genomic instability, however, make lungs particularly susceptible to disrepair and fibrosis. Pulmonary fibrosis constitutes a major cause of morbidity and is often relentlessly progressive, leading to death from respiratory failure. The pulmonary vasculature, which is critical for gas exchanges and plays a key role during lung development, repair, and regeneration, becomes aberrantly remodeled in patients with progressive pulmonary fibrosis. Although capillary rarefaction and increased vascular permeability are recognized as distinctive features of fibrotic lungs, the role of vasculature dysfunction in the pathogenesis of pulmonary fibrosis has only recently emerged as an important contributor to the progression of this disease. This review summarizes current findings related to lung vascular repair and regeneration and provides recent insights into the vascular abnormalities associated with the development of persistent lung fibrosis.

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Tumor vasculature abnormality creates a microenvironment that is not suitable for anti-tumor immune response and thereby induces resistance to immunotherapy. Remodeling of dysfunctional tumor blood vessels by anti-angiogenic approaches, known as vascular normalization, reshapes the tumor microenvironment toward an immune-favorable one and improves the effectiveness of immunotherapy. The tumor vasculature serves as a potential pharmacological target with the capacity of promoting an anti-tumor immune response. In this review, the molecular mechanisms involved in tumor vascular microenvironment-modulated immune reactions are summarized. In addition, the evidence of pre-clinical and clinical studies for the combined targeting of pro-angiogenic signaling and immune checkpoint molecules with therapeutic potential are highlighted. The heterogeneity of endothelial cells in tumors that regulate tissue-specific immune responses is also discussed. The crosstalk between tumor endothelial cells and immune cells in individual tissues is postulated to have a unique molecular signature and may be considered as a potential target for the development of new immunotherapeutic approaches.

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Current techniques for the detection of vasa vasorum (VV) in vascular pathology include staining for endothelial cell (EC) markers such as CD31 or VE-cadherin. However, this approach does not permit an objective assessment of vascular geometry upon vasospasm and the clinical relevance of endothelial specification markers found in developmental biology studies remains unclear. Here, we performed a combined immunostaining of rat abdominal aorta (rAA) and human saphenous vein (hSV) for various EC or vascular smooth muscle cell (VSMC) markers and found that the latter (e.g., alpha smooth muscle actin (α-SMA) or smooth muscle myosin heavy chain (SM-MHC)) ensure a several-fold higher signal-to-noise ratio irrespective of the primary antibody origin, fluorophore, or VV type (arterioles, venules, or capillaries). Further, α-SMA or SM-MHC staining allowed unbiased evaluation of the VV area under vasospasm. Screening of the molecular markers of endothelial heterogeneity (mechanosensitive transcription factors KLF2 and KLF4, arterial transcription factors HES1, HEY1, and ERG, venous transcription factor NR2F2, and venous/lymphatic markers PROX1, LYVE1, VEGFR3, and NRP2) have not revealed specific markers of any lineage in hSV (although KLF2 and PROX1 were restricted to venous endothelium in rAA), suggesting the need in high-throughput searches for the clinically relevant signatures of arterial, venous, lymphatic, or capillary differentiation.

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The blood microvascular endothelium consists of a heterogeneous population of cells with regionally distinct morphologies and transcriptional signatures in different tissues and organs. In addition to providing an anti-thrombogenic surface for blood flow, endothelial cells perform a multitude of additional regulatory tasks involving organogenesis, metabolism, angiogenesis, inflammation, repair and organ homeostasis. To communicate with surrounding cells and accomplish their many functions, endothelial cells secrete angiocrine factors including growth factors, chemokines, cytokines, extracellular matrix components, and proteolytic enzymes. Nonendothelial parenchymal and stromal cells in turn regulate endothelial growth, differentiation and survival during embryonal development and in the adult by paracrine mechanisms. Driven by advances in molecular biology, animal genetics, single cell transcriptomics and microscopic imaging, knowledge of organotypic vasculatures has expanded rapidly in recent years. The kidney vasculature, in particular, has been the focus of intensive investigation and represents a primary example of how endothelial heterogeneity and crosstalk with nonendothelial cells contribute to the development and function of a vital organ. In this paper, we review the morphology, function, and development of the kidney vasculature, with an emphasis on blood microvascular endothelial heterogeneity, and provide examples of endothelial and nonendothelial-derived factors that are critically involved in kidney development, growth, response to injury, and homeostasis.

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AIMS: Coronary vasculature formation is a critical event during cardiac development, essential for heart function throughout perinatal and adult life. However, current understanding of coronary vascular development has largely been derived from transgenic mouse models. The aim of this study was to characterize the transcriptome of the human foetal cardiac endothelium using single-cell RNA sequencing (scRNA-seq) to provide critical new insights into the cellular heterogeneity and transcriptional dynamics that underpin endothelial specification within the vasculature of the developing heart. METHODS AND RESULTS: We acquired scRNA-seq data of over 10 000 foetal cardiac endothelial cells (ECs), revealing divergent EC subtypes including endocardial, capillary, venous, arterial, and lymphatic populations. Gene regulatory network analyses predicted roles for SMAD1 and MECOM in determining the identity of capillary and arterial populations, respectively. Trajectory inference analysis suggested an endocardial contribution to the coronary vasculature and subsequent arterialization of capillary endothelium accompanied by increasing MECOM expression. Comparative analysis of equivalent data from murine cardiac development demonstrated that transcriptional signatures defining endothelial subpopulations are largely conserved between human and mouse. Comprehensive characterization of the transcriptional response to MECOM knockdown in human embryonic stem cell-derived EC (hESC-EC) demonstrated an increase in the expression of non-arterial markers, including those enriched in venous EC. CONCLUSIONS: scRNA-seq of the human foetal cardiac endothelium identified distinct EC populations. A predicted endocardial contribution to the developing coronary vasculature was identified, as well as subsequent arterial specification of capillary EC. Loss of MECOM in hESC-EC increased expression of non-arterial markers, suggesting a role in maintaining arterial EC identity.

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Forming the inner layer of the vascular system, endothelial cells (ECs) facilitate a multitude of crucial physiological processes throughout the body. Vascular ECs enable the vessel wall passage of nutrients and diffusion of oxygen from the blood into adjacent cellular structures. ECs regulate vascular tone and blood coagulation as well as adhesion and transmigration of circulating cells. The multitude of EC functions is reflected by tremendous cellular diversity. Vascular ECs can form extremely tight barriers, thereby restricting the passage of xenobiotics or immune cell invasion, whereas, in other organ systems, the endothelial layer is fenestrated (e.g., glomeruli in the kidney), or discontinuous (e.g., liver sinusoids) and less dense to allow for rapid molecular exchange. ECs not only differ between organs or vascular systems, they also change along the vascular tree and specialized subpopulations of ECs can be found within the capillaries of a single organ. Molecular tools that enable selective vascular targeting are helpful to experimentally dissect the role of distinct EC populations, to improve molecular imaging and pave the way for novel treatment options for vascular diseases. This review provides an overview of endothelial diversity and highlights the most successful methods for selective targeting of distinct EC subpopulations.

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Exercise is a powerful driver of physiological angiogenesis during adulthood, but the mechanisms of exercise-induced vascular expansion are poorly understood. We explored endothelial heterogeneity in skeletal muscle and identified two capillary muscle endothelial cell (mEC) populations that are characterized by differential expression of ATF3/4. Spatial mapping showed that ATF3/4+ mECs are enriched in red oxidative muscle areas while ATF3/4low ECs lie adjacent to white glycolytic fibers. In vitro and in vivo experiments revealed that red ATF3/4+ mECs are more angiogenic when compared with white ATF3/4low mECs. Mechanistically, ATF3/4 in mECs control genes involved in amino acid uptake and metabolism and metabolically prime red (ATF3/4+) mECs for angiogenesis. As a consequence, supplementation of non-essential amino acids and overexpression of ATF4 increased proliferation of white mECs. Finally, deleting Atf4 in ECs impaired exercise-induced angiogenesis. Our findings illustrate that spatial metabolic angiodiversity determines the angiogenic potential of muscle ECs.

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Endothelial cells display heterogeneous properties based on location and function. How this heterogeneity influences endothelial barrier stability both between and within vessel subtypes is unexplored. In this study, we find that endothelial cells exhibit heterogeneous barrier properties on inter-organ and intra-vessel levels. Using intravital microscopy and sequential stimulation of the ear dermis with vascular endothelial growth factor-A (VEGFA) and/or histamine, we observe distinct, reappearing sites, common for both agonists, where leakage preferentially takes place. Through repetitive stimulation of the diaphragm and trachea, we find inter-organ conservation of such predetermined leakage sites. Qualitatively, predetermined sites display distinct leakage properties and enhanced barrier breakdown compared to less susceptible regions. Mechanistically, laminin α5 is reduced at predetermined sites, which is linked to reduced junctional vascular endothelial (VE)-cadherin and enhanced VEGFA-induced VE-cadherin phosphorylation. These data highlight functional intra-vessel heterogeneity that defines predetermined sites with distinct leakage properties and that may disproportionately impact pathological vascular leakage.

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Tissue-engineered vascular grafts (TEVGs) have enormous potential for vascular replacement therapy. However, thrombosis and intimal hyperplasia are important problems associated with TEVGs especially small diameter TEVGs (<6 mm) after transplantation. Endothelialization of TEVGs is a key point to prevent thrombosis. Here, we discuss different types of endothelialization and different seed cells of tissue-engineered vascular grafts. Meanwhile, endothelial heterogeneity is also discussed. Based on it, we provide a new perspective for selecting suitable types of endothelialization and suitable seed cells to improve the long-term patency rate of tissue-engineered vascular grafts with different diameters and lengths.

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Transient receptor potential vanilloid 4 (TRPV4) ion channels on the endothelial cell membrane are widely regarded as a crucial Ca2+ influx pathway that promotes endothelium-dependent vasodilation. The downstream vasodilatory targets of endothelial TRPV4 channels vary among different vascular beds, potentially contributing to endothelial cell heterogeneity. Although numerous studies have examined the role of endothelial TRPV4 channels using specific pharmacological tools over the past decade, their physiological significance remains unclear, mainly due to a lack of endothelium-specific knockouts. Moreover, the loss of endothelium-dependent vasodilation is a significant contributor to vascular dysfunction in cardiovascular disease. The activity of endothelial TRPV4 channels is impaired in cardiovascular disease; therefore, strategies targeting the mechanisms that reduce endothelial TRPV4 channel activity may restore vascular function and provide therapeutic benefit. In this chapter, we discuss endothelial TRPV4 channel-dependent signaling mechanisms, the heterogeneity in endogenous activators and targets of endothelial TRPV4 channels, and the role of endothelial TRPV4 channels in the pathogenesis of cardiovascular diseases. We also discuss potentially interesting future research directions that may provide novel insights into the physiological and pathological roles of endothelial TRPV4 channels.

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Liver sinusoidal endothelial cells (LSECs) are the first liver cells to encounter waste macromolecules, pathogens, and toxins in blood. LSECs are highly specialized to mediate the clearance of these substances via endocytic scavenger receptors and are equipped with fenestrae that mediate the passage of macromolecules toward hepatocytes. Although some transcription factors (TFs) are known to play a role in LSEC specialization, information about the specialized LSEC signature and its transcriptional determinants remains incomplete.Based on a comparison of liver, heart, and brain endothelial cells (ECs), we established a 30-gene LSEC signature comprising both established and newly identified markers, including 7 genes encoding TFs. To evaluate the LSEC TF regulatory network, we artificially increased the expression of the 7 LSEC-specific TFs in human umbilical vein ECs. Although Zinc finger E-box-binding protein 2, homeobox B5, Cut-like homolog 2, and transcription factor EC (TCFEC) had limited contributions, musculoaponeurotic fibrosarcoma (C-MAF), GATA binding protein 4 (GATA4), and MEIS homeobox 2 (MEIS2) emerged as stronger inducers of LSEC marker expression. Furthermore, a combination of C-MAF, GATA4, and MEIS2 showed a synergistic effect on the increase of LSEC signature genes, including liver/lymph node-specific ICAM-3 grabbing non-integrin (L-SIGN) (or C-type lectin domain family member M (CLEC4M)), mannose receptor C-Type 1 (MRC1), legumain (LGMN), G protein-coupled receptor 182 (GPR182), Plexin C1 (PLXNC1), and solute carrier organic anion transporter family member 2A1 (SLCO2A1). Accordingly, L-SIGN, MRC1, pro-LGMN, GPR182, PLXNC1, and SLCO2A1 protein levels were elevated by this combined overexpression. Although receptor-mediated endocytosis was not significantly induced by the triple TF combination, it enhanced binding to E2, the hepatitis C virus host-binding protein. We conclude that C-MAF, GATA4, and MEIS2 are important transcriptional regulators of the unique LSEC fingerprint and LSEC interaction with viruses. Additional factors are however required to fully recapitulate the molecular, morphological, and functional LSEC fingerprint.NEW & NOTEWORTHY Liver sinusoidal endothelial cells (LSECs) are the first liver cells to encounter waste macromolecules, pathogens, and toxins in the blood and are highly specialized. Although some transcription factors are known to play a role in LSEC specialization, information about the specialized LSEC signature and its transcriptional determinants remains incomplete. Here, we show that Musculoaponeurotic Fibrosarcoma (C-MAF), GATA binding protein 4 (GATA4), and Meis homeobox 2 (MEIS2) are important transcriptional regulators of the unique LSEC signature and that they affect the interaction of LSECs with viruses.

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Blood vessels are lined by endothelial cells engaged in distinct organ-specific functions but little is known about their characteristic gene expression profiles. RNA-Sequencing of the brain, lung, and heart endothelial translatome identified specific pathways, transporters and cell-surface markers expressed in the endothelium of each organ, which can be visualized at http://www.rehmanlab.org/ribo. We found that endothelial cells express genes typically found in the surrounding tissues such as synaptic vesicle genes in the brain endothelium and cardiac contractile genes in the heart endothelium. Complementary analysis of endothelial single cell RNA-Seq data identified the molecular signatures shared across the endothelial translatome and single cell transcriptomes. The tissue-specific heterogeneity of the endothelium is maintained during systemic in vivo inflammatory injury as evidenced by the distinct responses to inflammatory stimulation. Our study defines endothelial heterogeneity and plasticity and provides a molecular framework to understand organ-specific vascular disease mechanisms and therapeutic targeting of individual vascular beds.

Blood vessels supply nutrients, oxygen and other key molecules to all of the organs in the body. Cells lining the blood vessels, called endothelial cells, regulate which molecules pass from the blood to the organs they supply. For example, brain endothelial cells prevent toxic molecules from getting into the brain, and lung endothelial cells allow immune cells into the lungs to fight off bacteria or viruses.Determining which genes are switched on in the endothelial cells of major organs might allow scientists to determine what endothelial cells do in the brain, heart, and lung, and how they differ; or help scientists deliver drugs to a particular organ. If endothelial cells from different organs switch on different groups of genes, each of these groups of genes can be thought of as a 'genetic signature' that identifies endothelial cells from a specific organ.Now, Jambusaria et al. show that brain, heart, and lung endothelial cells have distinct genetic signatures. The experiments used mice that had been genetically modified to have tags on their endothelial cells. These tags made it possible to isolate RNA ­ a molecule similar to DNA that contains the information about which genes are active ­ from endothelial cells without separating the cells from their tissue of origin. Next, RNA from endothelial cells in the heart, brain and lung was sequenced and analyzed.The results show that each endothelial cell type has a distinct genetic signature under normal conditions and infection-like conditions. Unexpectedly, the experiments also showed that genes that were thought to only be switched on in the cells of specific tissues are also on in the endothelial cells lining the blood vessels of the tissue. For example, genes switched on in brain cells are also active in brain endothelial cells, and genes allowing heart muscle cells to pump are also on in the endothelial cells of the heart blood vessels.The endothelial cell genetic signatures identified by Jambusaria et al. can be used as "postal codes" to target drugs to a specific organ via the endothelial cells that feed it. It might also be possible to use these genetic signatures to build organ-specific blood vessels from stem cells in the laboratory. Future work will try to answer why endothelial cells serving the heart and brain use genes from these organs.

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