During blood vessel development, endothelial cells become specified toward arterial or venous fates to generate a circulatory network that provides nutrients and oxygen to, and removes metabolic ...waste from, all tissues. Arterial-venous specification occurs in conjunction with suppression of endothelial cell cycle progression; however, the mechanistic role of cell cycle state is unknown. Herein, using Cdh5-CreER
;R26FUCCI2aR reporter mice, we find that venous endothelial cells are enriched for the FUCCI-Negative state (early G1) and BMP signaling, while arterial endothelial cells are enriched for the FUCCI-Red state (late G1) and TGF-β signaling. Furthermore, early G1 state is essential for BMP4-induced venous gene expression, whereas late G1 state is essential for TGF-β1-induced arterial gene expression. Pharmacologically induced cell cycle arrest prevents arterial-venous specification defects in mice with endothelial hyperproliferation. Collectively, our results show that distinct endothelial cell cycle states provide distinct windows of opportunity for the molecular induction of arterial vs. venous fate.
Activin receptor-like kinase 1 (ALK1) is an endothelial serine-threonine kinase receptor for bone morphogenetic proteins (BMPs) 9 and 10. Inactivating mutations in the ALK1 gene cause hereditary ...haemorrhagic telangiectasia type 2 (HHT2), a disabling disease characterized by excessive angiogenesis with arteriovenous malformations (AVMs). Here we show that inducible, endothelial-specific homozygous Alk1 inactivation and BMP9/10 ligand blockade both lead to AVM formation in postnatal retinal vessels and internal organs including the gastrointestinal (GI) tract in mice. VEGF and PI3K/AKT signalling are increased on Alk1 deletion and BMP9/10 ligand blockade. Genetic deletion of the signal-transducing Vegfr2 receptor prevents excessive angiogenesis but does not fully revert AVM formation. In contrast, pharmacological PI3K inhibition efficiently prevents AVM formation and reverts established AVMs. Thus, Alk1 deletion leads to increased endothelial PI3K pathway activation that may be a novel target for the treatment of vascular lesions in HHT2.
The use of neural stem cell (NSC) therapy for the treatment of stroke patients is successfully paving its way into advanced phases of large-scale clinical trials. To understand how to optimize NSC ...therapeutic approaches, it is fundamental to decipher the crosstalk between NSC and other cells that comprise the NSC microenvironment (niche) and regulate their function,
; namely, the endothelial cells of the microvasculature. In this mini review, we first provide a concise summary of preclinical findings describing the signaling mechanisms between NSC and vascular endothelial cells and
. Second, we describe the progress made in the development of NSC therapy for the treatment of strokes.
Engineered tissues can be used to model human pathophysiology and test the efficacy and safety of drugs. Yet, to model whole-body physiology and systemic diseases, engineered tissues with preserved ...phenotypes need to physiologically communicate. Here we report the development and applicability of a tissue-chip system in which matured human heart, liver, bone and skin tissue niches are linked by recirculating vascular flow to allow for the recapitulation of interdependent organ functions. Each tissue is cultured in its own optimized environment and is separated from the common vascular flow by a selectively permeable endothelial barrier. The interlinked tissues maintained their molecular, structural and functional phenotypes over 4 weeks of culture, recapitulated the pharmacokinetic and pharmacodynamic profiles of doxorubicin in humans, allowed for the identification of early miRNA biomarkers of cardiotoxicity, and increased the predictive values of clinically observed miRNA responses relative to tissues cultured in isolation and to fluidically interlinked tissues in the absence of endothelial barriers. Vascularly linked and phenotypically stable matured human tissues may facilitate the clinical applicability of tissue chips.
Differentiation of hematopoietic stem cells into distinct cell types was thought to occur through a series of discrete, stable progenitor states. Work from Velten et al. (2017) now shows that ...hematopoietic cells differentiate via a mechanism of continuous lineage priming and thus represent a CLOUD-HSPC.
Differentiation of hematopoietic stem cells into distinct cell types was thought to occur through a series of discrete, stable progenitor states. Work from Velten et al. (2017) now shows that hematopoietic cells differentiate via a mechanism of continuous lineage priming and thus represent a CLOUD-HSPC.
The role of fluid shear stress in vasculature development and remodeling is well appreciated. However, the mechanisms regulating these effects remain elusive. We show that abnormal flow sensing in ...lymphatic endothelial cells (LECs) caused by Sdc4 or Pecam1 deletion in mice results in impaired lymphatic vessel remodeling, including abnormal valve morphogenesis. Ablation of either gene leads to the formation of irregular, enlarged and excessively branched lymphatic vessels. In both cases, lymphatic valve-forming endothelial cells are randomly oriented, resulting in the formation of abnormal valves. These abnormalities are much more pronounced in Sdc4
; Pecam1
double-knockout mice, which develop severe edema. In vitro, SDC4 knockdown human LECs fail to align under flow and exhibit high expression of the planar cell polarity protein VANGL2. Reducing VANGL2 levels in SDC4 knockdown LECs restores their alignment under flow, while VANGL2 overexpression in wild-type LECs mimics the flow alignment abnormalities seen in SDC4 knockdown LECs. SDC4 thus controls flow-induced LEC polarization via regulation of VANGL2 expression.
A functional network of blood vessels is essential for organ growth and homeostasis, yet how the vasculature matures and maintains homeostasis remains elusive in live mice. By longitudinally tracking ...the same neonatal endothelial cells (ECs) over days to weeks, we found that capillary plexus expansion is driven by vessel regression to optimize network perfusion. Neonatal ECs rearrange positions to evenly distribute throughout the developing plexus and become positionally stable in adulthood. Upon local ablation, adult ECs survive through a plasmalemmal self-repair response, while neonatal ECs are predisposed to die. Furthermore, adult ECs reactivate migration to assist vessel repair. Global ablation reveals coordinated maintenance of the adult vascular architecture that allows for eventual network recovery. Lastly, neonatal remodeling and adult maintenance of the skin vascular plexus are orchestrated by temporally restricted, neonatal VEGFR2 signaling. Our work sheds light on fundamental mechanisms that underlie both vascular maturation and adult homeostasis in vivo.
Display omitted
•Plexus maturation is mediated by coordinated regression and rearrangement of ECs•Adult ECs are positionally stable but reactivate migration following local ablation•Global ablation reveals resilience of network architecture to large-scale cell loss•Adult but not neonatal ECs are competent for plasmalemmal self-repair
Maturation and expansion of skin vasculature is driven by vessel regression, with endothelial cells rearranging to distribute and optimize network perfusion. During adulthood, positionally stable cells reactivate migration to assist vessel repair. Following membrane injury, adult endothelial cells survive through a self-repair response, while neonatal cells are predisposed to die, leading to vessel regression.
Development of blood-forming (hemogenic) endothelial cells that give rise to hematopoietic stem and progenitor cells (HSPCs) is critical during embryogenesis to generate the embryonic and postnatal ...hematopoietic system. We previously demonstrated that the specification of murine hemogenic endothelial cells is promoted by retinoic acid (RA) signaling and requires downstream endothelial cell cycle control. Whether this mechanism is conserved in human hemogenic endothelial cell specification is unknown. Here, we present a protocol to derive primordial endothelial cells from human embryonic stem cells and promote their specification toward hemogenic endothelial cells. Furthermore, we demonstrate that RA treatment significantly increases human hemogenic endothelial cell specification. That is, RA promotes endothelial cell cycle arrest to enable RA-induced instructive signals to upregulate the genes needed for hematopoietic transition. These insights provide guidance for the ex vivo generation of autologous human hemogenic endothelial cells that are needed to produce human HSPCs for regenerative medicine applications.
Display omitted
•A simple and efficient protocol to derive human hemogenic endothelial cells is described•Functional human hemogenic endothelial cells are characterized on a clonal level•Retinoic acid signaling is required for human hemogenic endothelial specification•Cell-cycle state is a critical determinant of human endothelial cell specification
Retinoic acid signaling and cell-cycle control are known to promote mouse hemogenic endothelial cell specification. Qiu et al. present a protocol to derive endothelial cells from human stem cells and promote their hemogenic specification. These insights provide guidance for generating human hemogenic endothelial cells for regenerative medicine applications.
Atherosclerosis, the major cause of myocardial infarction and stroke, results from converging inflammatory, metabolic, and biomechanical factors. Arterial lesions form at sites of low and disturbed ...blood flow but are suppressed by high laminar shear stress (LSS) mainly via transcriptional induction of the anti-inflammatory transcription factor, Kruppel-like factor 2 (Klf2). We therefore performed a whole genome CRISPR-Cas9 screen to identify genes required for LSS induction of Klf2. Subsequent mechanistic investigation revealed that LSS induces Klf2 via activation of both a MEKK2/3-MEK5-ERK5 kinase module and mitochondrial metabolism. Mitochondrial calcium and ROS signaling regulate assembly of a mitophagy- and p62-dependent scaffolding complex that amplifies MEKK-MEK5-ERK5 signaling. Blocking the mitochondrial pathway in vivo reduces expression of KLF2-dependent genes such as eNOS and inhibits vascular remodeling. Failure to activate the mitochondrial pathway limits Klf2 expression in regions of disturbed flow. This work thus defines a connection between metabolism and vascular inflammation that provides a new framework for understanding and developing treatments for vascular disease.
Urodeles and fetal mammals are capable of impressive epimorphic regeneration in a variety of tissues, whereas the typical default response to injury in adult mammals consists of inflammation and scar ...tissue formation. One component of epimorphic regeneration is the recruitment of resident progenitor and stem cells to a site of injury. Bioactive molecules resulting from degradation of extracellular matrix (ECM) have been shown to recruit a variety of progenitor and stem cells in vitro in adult mammals. The ability to recruit multipotential cells to the site of injury by in vivo administration of chemotactic ECM degradation products in a mammalian model of digit amputation was investigated in the present study. Adult, 6- to 8-week-old C57/BL6 mice were subjected to midsecond phalanx amputation of the third digit of the right hind foot and either treated with chemotactic ECM degradation products or left untreated. At 14 days after amputation, mice treated with ECM degradation products showed an accumulation of heterogeneous cells that expressed markers of multipotency, including Sox2, Sca1, and Rex1 (Zfp42). Cells isolated from the site of amputation were capable of differentiation along neuroectodermal and mesodermal lineages, whereas cells isolated from control mice were capable of differentiation along only mesodermal lineages. The present findings demonstrate the recruitment of endogenous stem cells to a site of injury, and/or their generation/proliferation therein, in response to ECM degradation products.
PDF
