Evidence for telomerase involvement in the angiogenesis of astrocytic tumors: expression of human telomerase reverse transcriptase messenger RNA by vascular endothelial cells

2001 ◽  
Vol 94 (6) ◽  
pp. 961-971 ◽  
Author(s):  
Roberto Pallini ◽  
Francesco Pierconti ◽  
Maria Laura Falchetti ◽  
Daniela D'Arcangelo ◽  
Eduardo Fernandez ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Astrocytic Tumors ◽  
Htert Mrna ◽  
Content Type ◽  
Human Telomerase ◽  
Fine Print

Object. Evidence from recent in vitro studies indicates that reactivation of telomerase, the enzyme that synthesizes the telomere ends of chromosomes, is a crucial event in the unlimited clonal expansion of endothelial cells that precedes the neoplastic conversion of these cells. It is known that high-grade gliomas express telomerase and that, in these neoplasms, proliferating endothelial cells may undergo transformational changes with development of sarcomatous components within the primitive tumor. To assess whether telomerase is involved in the endothelial cell proliferation that characterizes brain tumor angiogenesis, the authors investigated at the single-cell level the expression of messenger (m)RNA for the human telomerase catalytic subunit human telomerase reverse transcriptase (hTERT) by vascular cells of astrocytic tumors. Methods. The in situ hybridization (ISH) method was performed by processing histological sections with specific riboprobes for hTERT and for c-myc, an oncogene that is known to upregulate hTERT. Results of the ISH studies were compared with proliferative activity, as estimated by Ki-67 immunostaining. The expression of hTERT mRNA by vascular endothelial cells was related to the histological grade of the tumor because it was detected in five (29%) of 17 low-grade astrocytomas, nine (56%) of 16 anaplastic astrocytomas, and 19 (100%) of 19 glioblastomas multiforme (GBMs). Expression of c-myc mRNA was strictly correlated with that of hTERT mRNA. In low-grade astrocytomas and anaplastic astrocytomas, a dissociation was noted between hTERT mRNA expression and the proliferation rate of endothelial cells. Conversely, GBMs displayed a significant correlation between the level of hTERT mRNA expression and endothelial cell proliferation. Data from an in vitro assay in which human umbilical vein endothelial cells were stimulated to proliferate by adding vascular endothelial growth factor and an ISH study of newly formed vessels surrounding brain infarcts confirmed that expression of hTERT mRNA does not merely reflect the proliferative status of endothelial cells but represents a specific feature of brain tumor neovascularization. Conclusions. The results of this study are consistent with a role of telomerase in the angiogenesis of astrocytic tumors. Expression of hTERT mRNA by tumor vascular cells is an early event during the progression of astrocytic tumors, which precedes endothelial cell proliferation and may represent a first sign of dedifferentiation. Other than elucidating the mechanisms of tumor angiogenesis, these results encourage research on antitelomerase drugs for the treatment of malignant gliomas.

scholarly journals Vascular Endothelial Growth Factor Mediates the Estrogen-Induced Breakdown of Tight Junctions between and Increase in Proliferation of Microvessel Endothelial Cells in the Baboon Endometrium

Endocrinology ◽  
2008 ◽  
Vol 149 (12) ◽  
pp. 6076-6083 ◽  
Author(s):  
Graham W. Aberdeen ◽  
Stanley J. Wiegand ◽  
Thomas W. Bonagura ◽  
Gerald J. Pepe ◽  
Eugene D. Albrecht
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Factor ◽  
Tight Junctions ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Endothelial Growth Factor ◽  
Vegf Trap

To assess whether there is a link between estrogen, vascular endothelial growth factor (VEGF), and early aspects of uterine angiogenesis, an acute temporal study was conducted in which ovariectomized baboons were pretreated with VEGF Trap, which sequesters endogenous VEGF, and administered estradiol at time 0 h. Serum estradiol levels approximated 500 pg/ml 4–6 h after estradiol administration. VEGF mRNA levels in endometrial glandular epithelial and stromal cells were increased to values 6 h after estradiol that were 3.74 ± 0.99-fold (mean ± se) and 5.70 ± 1.60-fold greater (P < 0.05), respectively, than at 0 h. Microvessel interendothelial cell tight junctions, which control paracellular permeability, were present in the endometrium at time 0 h, but not evident 6 h after estradiol administration. Thus, microvessel paracellular cleft width increased (P < 0.01, ANOVA) from 5.03 ± 0.22 nm at 0 h to 7.27 ± 0.48 nm 6 h after estrogen. In contrast, tight junctions remained intact, and paracellular cleft widths were unaltered in estradiol/VEGF Trap and vehicle-treated animals. Endometrial microvessel endothelial cell mitosis, i.e. percent Ki67+/Ki67− immunolabeled endothelial cells, increased (P < 0.05) from 2.9 ± 0.3% at 0 h to 21.4 ± 7.0% 6 h after estrogen treatment but was unchanged in estradiol/VEGF Trap and vehicle-treated animals. In summary, the estrogen-induced disruption of endometrial microvessel endothelial tight junctions and increase in endothelial cell proliferation were prevented by VEGF Trap. Therefore, we propose that VEGF mediates the estrogen-induced increase in microvessel permeability and endothelial cell proliferation as early steps in angiogenesis in the primate endometrium.

scholarly journals Sonic hedgehog selectively promotes lymphangiogenesis after kidney injury through noncanonical pathway

2019 ◽  
Vol 317 (4) ◽  
pp. F1022-F1033 ◽  
Author(s):  
Hui Zhuo ◽  
Dong Zhou ◽  
Yuanyuan Wang ◽  
Hongyan Mo ◽  
Ying Yu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Sonic Hedgehog ◽  
Lymphatic Vessels ◽  
Kidney Injury ◽  
Lymphatic Endothelial Cell ◽  
Cell Counting ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial

Kidney fibrosis is associated with an increased lymphangiogenesis, characterized by the formation and expansion of new lymphatic vessels. However, the trigger and underlying mechanism responsible for the growth of lymphatic vessels in diseased kidney remain poorly defined. Here, we report that tubule-derived sonic hedgehog (Shh) ligand is a novel lymphangiogenic factor that plays a crucial role in mediating lymphatic endothelial cell proliferation and expansion. Shh was induced in renal tubular epithelium in various models of fibrotic chronic kidney disease, and this was accompanied by an expansion of lymphatic vessels in adjacent areas. In vitro, Shh selectively promoted the proliferation of human dermal lymphatic endothelial cells (HDLECs) but not human umbilical vein endothelial cells, as assessed by cell counting, MTT assay, and bromodeoxyuridine incorporation. Shh also induced the expression of vascular endothelial growth factor receptor-3, cyclin D1, and proliferating cell nuclear antigen in HDLECs. Shh did not affect the expression of Gli1, the downstream target and readout of canonical hedgehog signaling, but activated ERK-1/2 in HDLECs. Inhibition of Smoothened with small-molecule inhibitor or blockade of ERK-1/2 activation abolished the lymphatic endothelial cell proliferation induced by Shh. In vivo, inhibition of Smoothened also repressed lymphangiogenesis and attenuated renal fibrosis. This study identifies Shh as a novel mitogen that selectively promotes lymphatic, but not vascular, endothelial cell proliferation and suggests that tubule-derived Shh plays an essential role in mediating lymphangiogenesis after kidney injury.

scholarly journals Serine 26 in Early Growth Response‐1 Is Critical for Endothelial Proliferation, Migration, and Network Formation

2021 ◽  
Author(s):  
Fernando S. Santiago ◽  
Yue Li ◽  
Levon M. Khachigian
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Response ◽  
Network Formation ◽  
Early Growth ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Early Growth Response ◽  
Early Growth Response 1

Background Vascular endothelial cell proliferation, migration, and network formation are key proangiogenic processes involving the prototypic immediate early gene product, Egr‐1 (early growth response‐1). Egr‐1 undergoes phosphorylation at a conserved Ser26 but its function is completely unknown in endothelial cells or any other cell type. Methods and Results A CRISPR/Cas9 strategy was used to introduce a homozygous Ser26>Ala mutation into endogenous Egr‐1 in human microvascular endothelial cells. In the course of generating mutant cells, we produced cells with homozygous deletion in Egr ‐1 caused by frameshift and premature termination. We found that Ser26 mutation in Egr‐1, or Egr‐1 deletion, perturbed endothelial cell proliferation in models of cell counting or real‐time growth using the xCELLigence System. We found that Ser26 mutation or Egr‐1 deletion ameliorated endothelial cell migration toward VEGF‐A 165 (vascular endothelial growth factor‐A) in a dual‐chamber model. On solubilized basement membrane preparations, Ser26 mutation or Egr‐1 deletion prevented endothelial network (or tubule) formation, an in vitro model of angiogenesis. Flow cytometry further revealed that Ser26 mutation or Egr‐1 deletion elevated early and late apoptosis. Finally, we demonstrated that Ser26 mutation or Egr‐1 deletion increased VE‐cadherin (vascular endothelial cadherin) expression, a regulator of endothelial adhesion and signaling, permeability, and angiogenesis. Conclusions These findings not only indicate that Egr‐1 is essential for endothelial cell proliferation, migration, and network formation, but also show that point mutation in Ser26 is sufficient to impair each of these processes and trigger apoptosis as effectively as the absence of Egr‐1. This highlights the importance of Ser26 in Egr‐1 for a range of proangiogenic processes.

scholarly journals The role of progesterone in endometrial angiogenesis in pregnant and ovariectomised mice

Reproduction ◽  
2005 ◽  
Vol 129 (6) ◽  
pp. 765-777 ◽  
Author(s):  
Lisa M Walter ◽  
Peter A W Rogers ◽  
Jane E Girling
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Plasma Progesterone ◽  
Ovariectomized Mice ◽  
Pregnant Mice ◽  
Endothelial Growth Factor

The role of progesterone (and oestrogen) in endometrial angiogenesis remains controversial. The aims of this study were to quantify endometrial angiogenesis in pregnant mice and to investigate the role of progesterone in promoting endothelial cell proliferation in ovariectomized mice. Uteri were collected on days 1 to 4 of pregnancy when circulating progesterone concentrations were increasing, prior to implantation. Before dissection, mice were injected with bromodeoxyuridine (BrdU) enabling proliferating endothelial cells to be quantified with CD31/BrdU double-immunohistochemistry. There was a significant increase in proliferating endothelial cells on day 3 of pregnancy when plasma progesterone also increased. To determine if this endothelial cell proliferation was due to progesterone, an experiment was performed on ovariectomised mice. One group was treated with a single oestradiol injection on day 8 after ovariectomy, followed by a no-treatment day and three consecutive daily injections of progesterone. Other groups were treated with either the vehicle, oestradiol or progesterone injections only; all were dissected on day 13 following ovariectomy. Unexpectedly, mice treated with progesterone-only had the highest amount of endothelial cell proliferation and oestrogen priming was found to significantly reduce this progesterone-induced endothelial cell proliferation. To determine if this proliferation is mediated by vascular endothelial growth factor (VEGF), a further experiment in which VEGF anti-serum was administered concurrently with the progesterone injections was performed. Endothelial cell proliferation was reduced but not abolished suggesting progesterone-induced endometrial angiogenesis is only partly mediated by VEGF. Results indicate that oestrogen priming is not required for progesterone to stimulate endometrial endothelial cell proliferation and that oestrogen inhibits progesterone-induced angiogenesis in ovariectomised mice.

scholarly journals Regulation of Endothelial Cell Proliferation and Vascular Assembly through Distinct mTORC2 Signaling Pathways

2015 ◽  
Vol 35 (7) ◽  
pp. 1299-1313 ◽  
Author(s):  
Shan Wang ◽  
Katherine R. Amato ◽  
Wenqiang Song ◽  
Victoria Youngblood ◽  
Keunwook Lee ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Vascular Endothelial Cells ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Relative Contribution ◽  
Protein Kinase Cα

Mammaliantargetofrapamycin (mTOR) is a serine/threonine kinase that regulates a diverse array of cellular processes, including cell growth, survival, metabolism, and cytoskeleton dynamics. mTOR functions in two distinct complexes, mTORC1 and mTORC2, whose activities and substrate specificities are regulated by complex specific cofactors, including Raptor and Rictor, respectively. Little is known regarding the relative contribution of mTORC1 versus mTORC2 in vascular endothelial cells. Using mouse models of Raptor or Rictor gene targeting, we discovered that Rictor ablation inhibited vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation and assemblyin vitroand angiogenesisin vivo, whereas the loss of Raptor had only a modest effect on endothelial cells (ECs). Mechanistically, the loss of Rictor reduced the phosphorylation of AKT, protein kinase Cα (PKCα), and NDRG1 without affecting the mTORC1 pathway. In contrast, the loss of Raptor increased the phosphorylation of AKT despite inhibiting the phosphorylation of S6K1, a direct target of mTORC1. Reconstitution of Rictor-null cells with myristoylated AKT (Myr-AKT) rescued vascular assembly in Rictor-deficient endothelial cells, whereas PKCα rescued proliferation defects. Furthermore, tumor neovascularizationin vivowas significantly decreased upon EC-specific Rictor deletion in mice. These data indicate that mTORC2 is a critical signaling node required for VEGF-mediated angiogenesis through the regulation of AKT and PKCα in vascular endothelial cells.

scholarly journals Mid-luteal angiogenesis and function in the primate is dependent on vascular endothelial growth factor

2001 ◽  
Vol 168 (3) ◽  
pp. 409-416 ◽  
Author(s):  
SE Dickson ◽  
R Bicknell ◽  
HM Fraser
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Factor ◽  
Luteal Phase ◽  
Smooth Muscle Actin ◽  
Proliferation Index ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Endothelial Growth Factor

Vascular endothelial growth factor (VEGF) is essential for the angiogenesis required for the formation of the corpus luteum; however, its role in ongoing luteal angiogenesis and in the maintenance of the established vascular network is unknown. The aim of this study was to determine whether VEGF inhibition could intervene in ongoing luteal angiogenesis using immunoneutralisation of VEGF starting in the mid-luteal phase. In addition, the effects on endothelial cell survival and the recruitment of periendothelial support cells were examined. Treatment with a monoclonal antibody to VEGF, or mouse gamma globulin for control animals, commenced on day 7 after ovulation and continued for 3 days. Bromodeoxyuridine (BrdU), used to label proliferating cells to obtain a proliferation index, was administered one hour before collecting ovaries from control and treated animals. Ovarian sections were stained using antibodies to BrdU, the endothelial cell marker, CD31, the pericyte marker, alpha-smooth muscle actin, and 3' end DNA fragments as a marker for apoptosis. VEGF immunoneutralisation significantly suppressed endothelial cell proliferation and the area occupied by endothelial cells while increasing pericyte coverage and the incidence of endothelial cell apoptosis. Luteal function was markedly compromised by anti-VEGF treatment as judged by a 50% reduction in plasma progesterone concentration. It is concluded that ongoing angiogenesis in the mid-luteal phase is primarily driven by VEGF, and that a proportion of endothelial cells of the mid-luteal phase vasculature are dependent on VEGF support.

scholarly journals Transactivation of Fetal Liver Kinase-1/Kinase-Insert Domain-Containing Receptor by Lysophosphatidylcholine Induces Vascular Endothelial Cell Proliferation

Endocrinology ◽  
2006 ◽  
Vol 147 (3) ◽  
pp. 1377-1385 ◽  
Author(s):  
Yoshiko Fujita ◽  
Masanori Yoshizumi ◽  
Yuki Izawa ◽  
Nermin Ali ◽  
Hideki Ohnishi ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cell ◽  
Fetal Liver ◽  
Vascular Endothelial Cell ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial

Abstract 117: RhoC Regulates VEGF-induced Signaling in Endothelial Cells

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Luke Hoeppner ◽  
Sutapa Sinha ◽  
Ying Wang ◽  
Resham Bhattacharya ◽  
Shamit Dutta ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Vascular Permeability ◽  
Rho Gtpase ◽  
Endothelial Cell Migration ◽  
Calcium Flux ◽  
Endothelial Cell Proliferation ◽  
Vegf Receptor ◽  
Vegf Signaling

Vascular permeability factor/vascular endothelial growth factor A (VEGF) is a central regulator of angiogenesis and potently promotes vascular permeability. VEGF plays a key role in the pathologies of heart disease, stroke, and cancer. Therefore, understanding the molecular regulation of VEGF signaling is an important pursuit. Rho GTPase proteins play various roles in vasculogenesis and angiogenesis. While the functions of RhoA and RhoB in these processes have been well defined, little is known about the role of RhoC in VEGF-mediated signaling in endothelial cells and vascular development. Here, we describe how RhoC modulates VEGF signaling to regulate endothelial cell proliferation, migration and permeability. We found VEGF stimulation activates RhoC in human umbilical vein endothelial cells (HUVECs), which was completely blocked after VEGF receptor 2 (VEGFR-2) knockdown indicating that VEGF activates RhoC through VEGFR-2 signaling. Interestingly, RhoC knockdown delayed the degradation of VEGFR-2 compared to control siRNA treated HUVECs, thus implicating RhoC in VEGFR-2 trafficking. In light of our results suggesting VEGF activates RhoC through VEGFR-2, we sought to determine whether RhoC regulates vascular permeability through the VEGFR-2/phospholipase Cγ (PLCγ) /Ca 2+ /eNOS cascade. We found RhoC knockdown in VEGF-stimulated HUVECs significantly increased PLC-γ1 phosphorylation at tyrosine 783, promoted basal and VEGF-stimulated eNOS phophorylation at serine 1177, and increased calcium flux compared with control siRNA transfected HUVECs. Taken together, our findings suggest RhoC negatively regulates VEGF-induced vascular permeability. We confirmed this finding through a VEGF-inducible zebrafish model of vascular permeability by observing significantly greater vascular permeability in RhoC morpholino (MO)-injected zebrafish than control MO-injected zebrafish. Furthermore, we showed that RhoC promotes endothelial cell proliferation and negatively regulates endothelial cell migration. Our data suggests a scenario in which RhoC promotes proliferation by upregulating -catenin in a Wnt signaling-independent manner, which in turn, promotes Cyclin D1 expression and subsequently drives cell cycle progression.

The role of matrix metalloproteinase activity in the maturation of human capillary endothelial cells in vitro

1999 ◽  
Vol 112 (10) ◽  
pp. 1599-1609 ◽  
Author(s):  
B.M. Kraling ◽  
D.G. Wiederschain ◽  
T. Boehm ◽  
M. Rehn ◽  
J.B. Mulliken ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Endothelial Cell Proliferation ◽  
Protein Levels ◽  
Matrix Deposition ◽  
Capillary Endothelial Cells ◽  
Vessel Maturation

Vessel maturation during angiogenesis (the formation of new blood vessels) is characterized by the deposition of new basement membrane and the downregulation of endothelial cell proliferation in the new vessels. Matrix remodeling plays a crucial, but still poorly understood role, in angiogenesis regulation. We present here a novel assay system with which to study the maturation of human capillary endothelial cells in vitro. When human dermal microvascular endothelial cells (HDMEC) were cultured in the presence of dibutyryl cAMP (Bt2) and hydrocortisone (HC), the deposition of a fibrous lattice of matrix molecules consisting of collagens type IV, type XVIII, laminin and thrombospondin was induced. In basal medium (without Bt2 and HC), HDMEC released active matrix metalloproteinases (MMPs) into the culture medium. However, MMP protein levels were significantly reduced by treatment with Bt2 and HC, while protein levels and activity of endogenous tissue inhibitor of MMPs (TIMP) increased. This shift in the proteolytic balance and matrix deposition was inhibited by the specific protein kinase A inhibitors RpcAMP and KT5720 or by substituting analogues without reported glucocorticoid activity for HC. The addition of MMP inhibitors human recombinant TIMP-1 or 1,10-phenanthroline to cultures under basal conditions induced matrix deposition in a dose-dependent manner, which was not observed with the serine protease inhibitor epsilon-amino-n-caproic acid (ACA). The deposited basement membrane-type of matrix reproducibly suppressed HDMEC proliferation and increased HDMEC adhesion to the substratum. These processes of matrix deposition and downregulation of endothelial cell proliferation, hallmarks of differentiating new capillaries in the end of angiogenesis, were recapitulated in our cell culture system by decreasing the matrix-degrading activity. These data suggest that our cell culture assay provides a simple and feasible model system for the study of capillary endothelial cell differentiation and vessel maturation in vitro.

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