CD46- and CD150-independent endothelial cell infection with wild-type measles viruses

The human cytomegalovirus (HCMV) genes UL128, UL130 and UL131A are essential for endothelial cell infection. Complementation of the defective UL131A gene of the non-endotheliotropic HCMV strain AD169 with wild-type UL131A in cis in an ectopic position restored endothelial cell tropism. The UL131A protein was found in virions in a complex with gH. Coinfection of fibroblasts with UL131A-negative and -positive viruses restored the endothelial cell tropism of UL131A-negative virions by complementing the virions with UL131A protein. Virus entry into endothelial cells, but not into fibroblasts, was blocked by an antipeptide antiserum to pUL131A. AD169, cis-complemented with wild-type UL131A, showed an impaired release of infectious particles from fibroblasts. A comparable defect in virus release was observed when UL131A was expressed ectopically in a virus background already expressing an intact copy of UL131A. In contrast, virus release from infected endothelial cells was not affected by UL131A. These data suggest a dual role for pUL131A in virus entry and virus exit from infected cells.

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Human Cytomegalovirus UL130 Protein Promotes Endothelial Cell Infection through a Producer Cell Modification of the Virion

Journal of Virology ◽

10.1128/jvi.79.13.8361-8373.2005 ◽

2005 ◽

Vol 79 (13) ◽

pp. 8361-8373 ◽

Cited By ~ 57

Author(s):

Marco Patrone ◽

Massimiliano Secchi ◽

Loretta Fiorina ◽

Mariagrazia Ierardi ◽

Gabriele Milanesi ◽

...

Keyword(s):

Endothelial Cell ◽

Human Cytomegalovirus ◽

Viral Infections ◽

Umbilical Vein ◽

Wild Type ◽

Cell Infection ◽

Virion Protein ◽

Human Embryonic Lung ◽

Virion Envelope ◽

Producer Cell

ABSTRACT Human cytomegalovirus (HCMV) growth in endothelial cells (EC) requires the expression of the UL131A-128 locus proteins. In this study, the UL130 protein (pUL130), the product of the largest gene of the locus, is shown to be a luminal glycoprotein that is inefficiently secreted from infected cells but is incorporated into the virion envelope as a Golgi-matured form. To investigate the mechanism of the UL130-mediated promotion of viral growth in EC, we performed a complementation analysis of a UL130 mutant strain. To provide UL130 in trans to viral infections, we constructed human embryonic lung fibroblast (HELF) and human umbilical vein endothelial cell (HUVEC) derivative cell lines that express UL130 via a retroviral vector. When the UL130-negative virus was grown in UL130-complementing HELF, the infectivity of progeny virions for HUVEC was restored to the wild-type level. In contrast, the infectivity of the UL130-negative virus for UL130-complementing HUVEC was low and similar to that of the same virus infecting control noncomplementing HUVEC. The UL130-negative virus, regardless of whether or not it had been complemented in the prior cycle, could form plaques only on UL130-complementing HUVEC, not control HUVEC. Because (i) both wild-type and UL130-transcomplemented virions maintained their infectivity for HUVEC after purification, (ii) UL130 failed to complement in trans the UL130-negative virus when it was synthesized in a cell separate from the one that produced the virions, and (iii) pUL130 is a virion protein, models are favored in which pUL130 acquisition in the producer cell renders HCMV virions competent for a subsequent infection of EC.

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Interaction of wild-type and catalytically inactive mutant forms of tissue-type plasminogen activator with human umbilical vein endothelial cell monolayers.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)39866-7 ◽

1990 ◽

Vol 265 (5) ◽

pp. 2755-2762

Author(s):

V Ramakrishnan ◽

D V Sinicropi ◽

R Dere ◽

W C Darbonne ◽

K B Bechtol ◽

...

Keyword(s):

Endothelial Cell ◽

Plasminogen Activator ◽

Umbilical Vein ◽

Tissue Type ◽

Wild Type ◽

Tissue Type Plasminogen Activator ◽

Human Umbilical Vein ◽

Cell Monolayers ◽

Type Plasminogen Activator ◽

Mutant Forms

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Mutational Mapping of UL130 of Human Cytomegalovirus Defines Peptide Motifs within the C-Terminal Third as Essential for Endothelial Cell Infection

Journal of Virology ◽

10.1128/jvi.00572-10 ◽

2010 ◽

Vol 84 (18) ◽

pp. 9019-9026 ◽

Cited By ~ 13

Author(s):

Andrea Schuessler ◽

Kerstin Laib Sampaio ◽

Laura Scrivano ◽

Christian Sinzger

Keyword(s):

Endothelial Cell ◽

Human Cytomegalovirus ◽

Artificial Chromosome ◽

Protein Domain ◽

Cell Infection ◽

Peptide Motifs ◽

Charged Amino Acids ◽

C Terminus ◽

Charge Cluster ◽

A Charge

ABSTRACT The UL130 gene is one of the major determinants of endothelial cell (EC) tropism of human cytomegalovirus (HCMV). In order to define functionally important peptides within this protein, we have performed a charge-cluster-to-alanine (CCTA) mutational scanning of UL130 in the genetic background of a bacterial artificial chromosome-cloned endotheliotropic HCMV strain. A total of 10 charge clusters were defined, and in each of them two or three charged amino acids were replaced with alanines. While the six N-terminal clusters were phenotypically irrelevant, mutation of the four C-terminal clusters each caused a reduction of EC tropism. The importance of this protein domain was further emphasized by the fact that the C-terminal pentapeptide PNLIV was essential for infection of ECs, and the cell tropism could not be rescued by a scrambled version of this sequence. We conclude that the C terminus of the UL130 protein serves an important function for infection of ECs by HCMV. This makes UL130 a promising molecular target for antiviral strategies, e.g., the development of antiviral peptides.

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Abstract 3640: Mitochondrial Uncoupling Protein 2 Facilitates Endothelial Cell Growth and Angiogenesis via Reduction of Mitochondrial Superoxide

Circulation ◽

10.1161/circ.118.suppl_18.s_456-b ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Yukio Shimasaki ◽

Kai Chen ◽

John F Keaney

Keyword(s):

Endothelial Cells ◽

Endothelial Cell ◽

Cell Growth ◽

Mitochondrial Function ◽

Endothelial Cell Proliferation ◽

Wild Type ◽

Sprouting Angiogenesis ◽

Mitochondrial Superoxide ◽

Endothelial Cell Growth ◽

Aortic Explants

Background: Growing evidence suggests that mitochondrial function contributes to cell phenotype. One important component of mitochondrial function is the membrane potential that is controlled, in part, by uncoupling proteins (UCPs). Based on our previous data, the UCP2 is predominantly expressed in cultured endothelial cells. Therefore, we sought to examine the role of UCP2 in endothelial cell growth and angiogenesis. Methods and Results: Murine lung endothelial cells (MLECs) were isolated from UCP2-null and wild-type mice. UCP2-null cells were found less proliferative than wild-type cells (P<0.02, UCP2-null cells vs. wild-type cells, n=4). This defect of UCP2-null cells was rescued by UCP2 adenovirus transfection (19% increase, p<0.02 vs. LacZ adenovirus treated cells, n=3), and also rescued by transfection with manganese superoxide dismutase (MnSOD) adenovirus (53% increase, P<0.002 vs. LacZ adenovirus treated cells, n=3). We found a reciprocal relation such as no UCP2 expression and higher mitochondrial superoxide level in the MLECs (P<0.005, UCP2-null cells vs. wild-type cells, n=3), suggesting that mitochondrial superoxide may regulate endothelial cell growth. Then, we prepared murine aortic rings from UCP2-null and wild-type mice and embedded in rat tail collagen gel. The sprouting angiogenesis of UCP2-null explants was significantly less than wild-type explants (P<0.02, UCP2-null explants vs. wild-type explants, n=3– 4). Furthermore, MLECs from MnSOD-heterozygous mice showed less proliferation with lower expression of UCP2 protein and higher mitochondrial superoxide level compared to the MLECs from wild-type littermates (P<0.02, MnSOD-heterozygous cells vs. wild-type cells, n=4 – 8). We also observed less sprouting angiogenesis in MnSOD-heterozygous aortic explants than wild-type aortic explants (P<0.05, MnSOD-heterozygous explants vs. wild-type explants, n=3– 6). Conclusions: These data indicate that mitochondrial superoxide controls endothelial cell proliferation and angiogenesis, suggesting that mitochondrial metabolism modulates the endothelial cell growth and angiogenesis.

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Endothelial cell-specific reduction of heparan sulfate suppresses glioma growth in mice

Discover Oncology ◽

10.1007/s12672-021-00444-3 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Takamasa Kinoshita ◽

Hiroyuki Tomita ◽

Hideshi Okada ◽

Ayumi Niwa ◽

Fuminori Hyodo ◽

...

Keyword(s):

Endothelial Cell ◽

Heparan Sulfate ◽

Vascular Endothelium ◽

Wild Type ◽

Brain Capillaries ◽

Gene Encoding ◽

Glioma Growth ◽

Underlying Mechanisms ◽

The Brain

Abstract Purpose Heparan sulfate (HS) is one of the factors that has been suggested to be associated with angiogenesis and invasion of glioblastoma (GBM), an aggressive and fast-growing brain tumor. However, it remains unclear how HS of endothelial cells is involved in angiogenesis in glioblastoma and its prognosis. Thus, we investigated the effect of endothelial cell HS on GBM development. Methods We generated endothelial cell-specific knockout of Ext1, a gene encoding a glycosyltransferase and essential for HS synthesis, and murine GL261 glioblastoma cells were orthotopically transplanted. Two weeks after transplantation, we examined the tumor progression and underlying mechanisms. Results The endothelial cell-specific Ext1 knockout (Ext1CKO) mice exhibited reduced HS expression specifically in the vascular endothelium of the brain capillaries compared with the control wild-type (WT) mice. GBM growth was significantly suppressed in Ext1CKO mice compared with that in WT mice. After GBM transplantation, the survival rate was significantly higher in Ext1CKO mice than in WT mice. We investigated how the effect of fibroblast growth factor 2 (FGF2), which is known as an angiogenesis-promoting factor, differs between Ext1CKO and WT mice by using an in vivo Matrigel assay and demonstrated that endothelial cell-specific HS reduction attenuated the effect of FGF2 on angiogenesis. Conclusions HS reduction in the vascular endothelium of the brain suppressed GBM growth and neovascularization in mice.

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Identification of an octamer element required for in vivo expression of the TIE1 gene in endothelial cells

Biochemical Journal ◽

10.1042/bj3600023 ◽

2001 ◽

Vol 360 (1) ◽

pp. 23-29 ◽

Cited By ~ 5

Author(s):

Stephane C. BOUTET ◽

Thomas QUERTERMOUS ◽

Bahaa M. FADEL

Keyword(s):

Endothelial Cells ◽

Endothelial Cell ◽

Molecular Mechanisms ◽

Vascular Development ◽

Common Mechanism ◽

Tyrosine Kinase Receptor ◽

Wild Type ◽

Binding Studies ◽

In Vivo Expression

TIE1, an endothelial-cell-specific tyrosine kinase receptor, is required for the survival and growth of microvascular endothelial cells during the capillary sprouting phase of vascular development. To investigate the molecular mechanisms that regulate the expression of TIE1 in the endothelium, we analysed transgenic mouse embryos carrying wild-type or mutant TIE1 promoter/LacZ constructs. Our data indicate that an upstream DNA octamer element (5′-ATGCAAAT-3′) is required for the in vivo expression of TIE1 in embryonic endothelial cells. Transgenic embryos carrying the wild-type TIE1 promoter (−466 to +78bp) fused to LacZ and spanning the octamer element demonstrate endothelial-cell-specific expression of the reporter transgene. Point mutations introduced within the octamer element result in a significant decrease of endothelial LacZ expression, suggesting that the octamer site functions as a positive regulator for TIE1 gene expression in endothelial cells. DNA–protein binding studies show that the octamer element exhibits an endothelial-cell-specific pattern of binding via interaction with endothelial-cell-restricted factor(s). Our findings suggest an important role for the octamer element in regulating the expression of the TIE1 receptor in the embryonic endothelium and suggest a common mechanism for the regulation of the angiogenic and cell-specific TIE1 and TIE2 genes during vascular development.

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Superoxide mediates endotoxin-induced platelet-endothelial cell adhesion in intestinal venules

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00311.2002 ◽

2003 ◽

Vol 284 (2) ◽

pp. H535-H541 ◽

Cited By ~ 46

Author(s):

Wolfgang H. Cerwinka ◽

Dianne Cooper ◽

Christian F. Krieglstein ◽

Chris R. Ross ◽

Joe M. McCord ◽

...

Keyword(s):

Endothelial Cell ◽

Vascular System ◽

Ischemia Reperfusion Injury ◽

Ischemia Reperfusion ◽

Fold Increase ◽

Induced Response ◽

Wild Type ◽

Binding Properties ◽

Platelet Endothelial Cell Adhesion ◽

Deficient Mice

Platelets have been implicated in the pathogenesis of different diseases of the vascular system, including atherosclerosis, sepsis, and ischemia-reperfusion injury; however, relatively little is known about the factors that regulate the interactions between circulating platelets and the vessel wall. The objective of this study was to define the contribution of superoxide to LPS-induced platelet-endothelial cell (P/E) adhesion in murine intestinal venules. The adhesion of rhodamine-6G-labeled murine platelets was monitored by intravital fluorescence microscopy. Four hours after LPS administration in control [wild-type (WT)] mice, an ∼10-fold increase in P/E adhesion was detected. This response did not result from LPS-induced platelet activation. The LPS-induced P/E adhesion was greatly attenuated in NAD(P)H oxidase-deficient mice and in WT mice rendered neutropenic with anti-neutrophil serum, whereas the response was unchanged in WT mice receiving a CD18 blocking MAb or in CD18-deficient mice. A chimeric form of MnSOD that exhibits the binding properties of extracellular SOD also attenuated the LPS-induced response in WT mice. These findings indicate that neutrophil-derived superoxide plays a major role in the modulation of endotoxin-induced P/E adhesion.

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Inhibition of antigen-receptor signaling by Platelet Endothelial Cell Adhesion Molecule-1 (CD31) requires functional ITIMs, SHP-2, and p56lck

Blood ◽

10.1182/blood.v97.8.2351 ◽

2001 ◽

Vol 97 (8) ◽

pp. 2351-2357 ◽

Cited By ~ 97

Author(s):

Debra K. Newman ◽

Christin Hamilton ◽

Peter J. Newman

Keyword(s):

T Cells ◽

Endothelial Cell ◽

Cell Adhesion Molecule ◽

Antigen Receptor ◽

Inhibitory Effect ◽

Wild Type ◽

Antigen Receptor Signaling ◽

Platelet Endothelial Cell Adhesion ◽

Cell Adhesion Molecule 1 ◽

Endothelial Cell Adhesion

Abstract Platelet Endothelial Cell Adhesion Molecule-1 (PECAM-1, CD31) is a 130-kd member of the immunoglobulin gene superfamily that is expressed on the surface of platelets, endothelial cells, myeloid cells, and certain lymphocyte subsets. PECAM-1 has recently been shown to contain functional immunoreceptor tyrosine-based inhibitory motifs (ITIMs) within its cytoplasmic domain, and co-ligation of PECAM-1 with the T-cell antigen receptor (TCR) results in tyrosine phosphorylation of PECAM-1, recruitment of Src homology 2 domain-containing protein tyrosine phosphatase-2 (SHP-2), and attenuation of TCR-mediated cellular signaling. To determine the molecular basis of PECAM-1 inhibitory signaling in lymphocytes, the study sought to (1) establish the importance of the PECAM-1 ITIMs for its inhibitory activity, (2) determine the relative importance of SHP-2 versus SHP-1 in mediating the inhibitory effect of PECAM-1, and (3) identify the protein tyrosine kinases required for PECAM-1 tyrosine phosphorylation in T cells. Co-ligation of wild-type PECAM-1 with the B-cell antigen receptor expressed on chicken DT40 B cells resulted in a marked reduction of calcium mobilization—similar to previous observations in T cells. In contrast, co-ligation of an ITIM-less form of PECAM-1 had no inhibitory effect. Furthermore, wild-type PECAM-1 was unable to attenuate calcium mobilization in SHP-2–deficient DT40 variants despite abundant levels of SHP-1 in these cells. Finally, PECAM-1 failed to become tyrosine phosphorylated in p56lck-deficient Jurkat T cells. Together, these data provide important insights into the molecular requirements for PECAM-1 regulation of antigen receptor signaling.

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Activation of endothelial NADPH oxidase during normoxic lung ischemia is KATP channel dependent

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00210.2005 ◽

2005 ◽

Vol 289 (6) ◽

pp. L954-L961 ◽

Cited By ~ 51

Author(s):

Qunwei Zhang ◽

Ikuo Matsuzaki ◽

Shampa Chatterjee ◽

Aron B. Fisher

Keyword(s):

Endothelial Cells ◽

Membrane Potential ◽

Endothelial Cell ◽

Cell Membrane ◽

Nadph Oxidase ◽

Membrane Depolarization ◽

Fluorescence Probes ◽

Ros Generation ◽

Wild Type ◽

Channel Dependent

Previous studies have shown endothelial cell membrane depolarization and generation of reactive oxygen species (ROS) in endothelial cells with abrupt reduction in shear stress (ischemia). This study evaluated the role of ATP-sensitive potassium (KATP) channels and NADPH oxidase in the ischemic response by using Kir6.2−/− and gp91phox−/− mice. To evaluate ROS generation, we subjected isolated perfused mouse lungs labeled with 2′,7′-dichlorodihydrofluorescein (DCF), hydroethidine (HE), or diphenyl-1-pyrenylphosphine (DPPP) to control perfusion followed by global ischemia. In wild-type C57BL/6J mice, imaging of subpleural endothelial cells showed a time-dependent increase in intensity for all three fluorescence probes with ischemia, which was blocked by preperfusion with cromakalim (a KATP channel agonist) or diphenyleneiodonium (DPI, a flavoprotein inhibitor). Endothelial cell fluorescence with bis-oxonol, a membrane potential probe, increased during lung ischemia indicating cell membrane depolarization. The change in membrane potential with ischemia in lungs of gp91phox−/− mice was similar to wild type, but ROS generation did not occur. Lungs from Kir6.2−/− showed marked attenuation of the change in both membrane potential and ROS production. Thus membrane depolarization during lung ischemia requires the presence of a KATP channel and is required for activation of NADPH oxidase and endothelial ROS generation.

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