Dynamic cytoskeletal regulation of cell shape supports resilience of lymphatic endothelium.

Lymphatic capillaries continuously take up interstitial fluid and adapt to resulting changes in vessel calibre. The mechanisms by which the permeable monolayer of loosely connected lymphatic endothelial cells (LECs) maintains mechanical stability remain elusive. Here we identify dynamic cytoskeletal regulation of LEC shape, induced by isotropic stretch, as crucial for the integrity and function of dermal lymphatic capillaries.

Cdc42 is crucial for mural cell migration, proliferation and patterning of the retinal vasculature.

Aims: Mural cells constitute the outer lining of blood vessels and are essential for vascular development and function. Mural cell loss or malfunction has been associated with numerous diseases including diabetic retinopathy, stroke and amyotrophic lateral sclerosis. In this work, we investigate the role of CDC42 in mural cells in vivo, using the developing mouse retina as a model.

Intravital Imaging of Disease Mechanisms in a Mouse Model of CCM Skin Lesions-Brief Report.

Background: Cerebral cavernous malformation (CCM) is a disease characterized by vascular malformations that primarily develop in the brain. These malformations are prone to leak, and their rupture or thrombotic closure can cause life-threatening hemorrhages and strokes. Mouse models have been instrumental to study the disease, but most cause premature lethality, precluding the investigation of disease mechanisms through intravital microscopy.

Mosaic deletion of claudin-5 reveals rapid non-cell-autonomous consequences of blood-brain barrier leakage.

Claudin-5 (CLDN5) is an endothelial tight junction protein essential for blood-brain barrier (BBB) formation. Abnormal CLDN5 expression is common in brain disease, and knockdown of Cldn5 at the BBB has been proposed to facilitate drug delivery to the brain. To study the consequences of CLDN5 loss in the mature brain, we induced mosaic endothelial-specific Cldn5 gene ablation in adult mice (Cldn5).

Incongruence between transcriptional and vascular pathophysiological cell states.

The Notch pathway is a major regulator of endothelial transcriptional specification. Targeting the Notch receptors or Delta-like ligand 4 (Dll4) dysregulates angiogenesis. Here, by analyzing single and compound genetic mutants for all Notch signaling members, we find significant differences in the way ligands and receptors regulate liver vascular homeostasis.

Mural Cell SRF Controls Pericyte Migration, Vessel Patterning and Blood Flow.

Background: Pericytes and vascular smooth muscle cells, collectively known as mural cells, are recruited through PDGFB (platelet-derived growth factor B)-PDGFRB (platelet-derived growth factor receptor beta) signaling. MCs are essential for vascular integrity, and their loss has been associated with numerous diseases. Most of this knowledge is based on studies in which MCs are insufficiently recruited or fully absent upon inducible ablation.

Sphingosine 1-Phosphate Receptor Signaling Establishes AP-1 Gradients to Allow for Retinal Endothelial Cell Specialization.

Transcriptional mechanisms that drive angiogenesis and organotypic vascular endothelial cell specialization are poorly understood. Here, we show that retinal endothelial sphingosine 1-phosphate receptors (S1PRs), which restrain vascular endothelial growth factor (VEGF)-induced angiogenesis, spatially restrict expression of JunB, a member of the activator protein 1 (AP-1) family of transcription factors (TFs). Mechanistically, VEGF induces JunB expression at the sprouting vascular front while S1PR-dependent vascular endothelial (VE)-cadherin assembly suppresses JunB expression in the nascent vascular network, thus creating a gradient of this TF.

CDC42 Deletion Elicits Cerebral Vascular Malformations via Increased MEKK3-Dependent KLF4 Expression.

Rationale: Aberrant formation of blood vessels precedes a broad spectrum of vascular complications; however, the cellular and molecular events governing vascular malformations are not yet fully understood.

Objective: Here, we investigated the role of CDC42 (cell division cycle 42) during vascular morphogenesis and its relative importance for the development of cerebrovascular malformations.

Methods And Results: To avoid secondary systemic effects often associated with embryonic gene deletion, we generated an endothelial-specific and inducible knockout approach to study postnatal vascularization of the mouse brain.

Defective endothelial cell migration in the absence of Cdc42 leads to capillary-venous malformations.

Formation and homeostasis of the vascular system requires several coordinated cellular functions, but their precise interplay during development and their relative importance for vascular pathologies remain poorly understood. Here, we investigated the endothelial functions regulated by Cdc42 and their relevance during angiogenic sprouting and vascular morphogenesis in the postnatal mouse retina. We found that Cdc42 is required for endothelial tip cell selection, directed cell migration and filopodia formation, but dispensable for cell proliferation or apoptosis.

Prickle is phosphorylated by Nemo and targeted for degradation to maintain Prickle/Spiny-legs isoform balance during planar cell polarity establishment.

Planar cell polarity (PCP) instructs tissue patterning in a wide range of organisms from fruit flies to humans. PCP signaling coordinates cell behavior across tissues and is integrated by cells to couple cell fate identity with position in a developing tissue. In the fly eye, PCP signaling is required for the specification of R3 and R4 photoreceptors based upon their positioning relative to the dorso-ventral axis.

Visualization of vascular mural cells in developing brain using genetically labeled transgenic reporter mice.

The establishment of a fully functional blood vascular system requires elaborate angiogenic and vascular maturation events in order to fulfill organ-specific anatomical and physiological needs. Although vascular mural cells, i.e.

VEGF Receptor Tyrosine Kinases: Key Regulators of Vascular Function.

Vascular endothelial growth factor receptor (VEGFR) tyrosine kinases are key regulators of vascular development in vertebrates. Their activation is regulated through a family of secreted glycoproteins, the vascular endothelial growth factors (VEGFs). Expression, proteolytic processing, and diffusion range of VEGF proteins need to be tightly regulated, due to their crucial roles in development.

Gpr116 Receptor Regulates Distinctive Functions in Pneumocytes and Vascular Endothelium.

Despite its known expression in both the vascular endothelium and the lung epithelium, until recently the physiological role of the adhesion receptor Gpr116/ADGRF5 has remained elusive. We generated a new mouse model of constitutive Gpr116 inactivation, with a large genetic deletion encompassing exon 4 to exon 21 of the Gpr116 gene. This model allowed us to confirm recent results defining Gpr116 as necessary regulator of surfactant homeostasis.

New imaging methods and tools to study vascular biology.

Purpose Of Review: Throughout history, development of novel microscopy techniques has been of fundamental importance to advance the vascular biology field.This review offers a concise summary of the most recently developed imaging techniques and discusses how they can be applied to vascular biology. In addition, we reflect upon the most important fluorescent reporters for vascular research that are currently available.

Excessive vascular sprouting underlies cerebral hemorrhage in mice lacking αVβ8-TGFβ signaling in the brain.

Vascular development of the central nervous system and blood-brain barrier (BBB) induction are closely linked processes. The role of factors that promote endothelial sprouting and vascular leak, such as vascular endothelial growth factor A, are well described, but the factors that suppress angiogenic sprouting and their impact on the BBB are poorly understood. Here, we show that integrin αVβ8 activates angiosuppressive TGFβ gradients in the brain, which inhibit endothelial cell sprouting.

Endocytosis regulates VEGF signalling during angiogenesis.

Endocytosis has proved to be a versatile mechanism regulating diverse cellular processes, ranging from nutrient uptake to intracellular signal transduction. New work reinforces the importance of endocytosis for VEGF receptor signalling and angiogenesis in the developing eye, and describes a mechanism for its differential regulation in angiogenic versus quiescent endothelial cells.

The sphingosine-1-phosphate receptor S1PR1 restricts sprouting angiogenesis by regulating the interplay between VE-cadherin and VEGFR2.

Angiogenesis, the process by which new blood vessels arise from preexisting ones, is critical for embryonic development and is an integral part of many disease processes. Recent studies have provided detailed information on how angiogenic sprouts initiate, elongate, and branch, but less is known about how these processes cease. Here, we show that S1PR1, a receptor for the blood-borne bioactive lipid sphingosine-1-phosphate (S1P), is critical for inhibition of angiogenesis and acquisition of vascular stability.

Nemo kinase phosphorylates β-catenin to promote ommatidial rotation and connects core PCP factors to E-cadherin-β-catenin.

Frizzled planar cell polarity (PCP) signaling regulates cell motility in several tissues, including ommatidial rotation in Drosophila melanogaster. The Nemo kinase (Nlk in vertebrates) has also been linked to cell-motility regulation and ommatidial rotation but its mechanistic role(s) during rotation remain obscure. We show that nemo functions throughout the entire rotation movement, increasing the rotation rate.

Endothelial-mural cell signaling in vascular development and angiogenesis.

Mural cells are essential components of blood vessels and are necessary for normal development, homeostasis, and organ function. Alterations in mural cell density or the stable attachment of mural cells to the endothelium is associated with several human diseases such as diabetic retinopathy, venous malformation, and hereditary stroke. In addition mural cells are implicated in regulating tumor growth and have thus been suggested as potential antiangiogenic targets in tumor therapy.

Microscopic analysis of the adult Drosophila retina using semithin plastic sections.

The regular appearance and the repetitive nature of the Drosophila eye, consisting of several hundred identical multicellular units, the ommatidia, has long served as an invaluable experimental system to study cell-cell interactions, inductive signaling events, cell proliferation, programmed cell death, cell differentiation, cell organization, and planar cell polarity among others. Importantly, the eye is dispensable for viability and fertility of the fly and thus, it can easily be manipulated, making it an ideal target for genetic screens. This chapter described an essential technique in the analysis of different genotypes in the adult fly eye, and allows detailed analyses with single cell resolution.

Transgenic Drosophila models of Noonan syndrome causing PTPN11 gain-of-function mutations.

Mutations in the PTPN11 gene, which encodes the protein tyrosine phosphatase SHP-2, causes Noonan syndrome (NS), an autosomal dominant disorder with pleomorphic developmental abnormalities. Certain germline and somatic PTPN11 mutations cause leukemias. Mutations have gain-of-function (GOF) effects with the commonest NS allele, N308D, being weaker than the leukemia-causing mutations.

Egfr signaling regulates ommatidial rotation and cell motility in the Drosophila eye via MAPK/Pnt signaling and the Ras effector Canoe/AF6.

Epidermal Growth Factor-receptor (Egfr) signaling is evolutionarily conserved and controls a variety of different cellular processes. In Drosophila these include proliferation, patterning, cell-fate determination, migration and survival. Here we provide evidence for a new role of Egfr signaling in controlling ommatidial rotation during planar cell polarity (PCP) establishment in the Drosophila eye.