The Experts below are selected from a list of 849 Experts worldwide ranked by ideXlab platform

Michael J Dunn - One of the best experts on this subject based on the ideXlab platform.

  • Preproendothelin 1 expression in human mesangial cells evidence for a p38 mitogen activated protein kinase protein kinases c dependent mechanism
    Journal of The American Society of Nephrology, 2001
    Co-Authors: Marco Foschi, Andrey Sorokin, Phillip F Pratt, Ann Mcginty, Giorgio La Villa, F Franchi, Michael J Dunn
    Abstract:

    : Endothelin-1 (ET-1) has been implicated in the pathogenesis of renal inflammation. This study investigated the mechanisms underlying the synergistic upregulation of preproET-1 gene expression in human mesangial cells after co-stimulation with thrombin and tumor necrosis factor alpha (TNFalpha). Whereas thrombin induced a moderate upregulation of preproET-1 mRNA, co-stimulation with TNFalpha resulted in a strong and protracted upregulation of this mRNA species. Thrombin+TNFalpha-induced upregulation of preproET-1 expression was found to require p38 mitogen-activated protein kinase and protein kinases C, whereas activation of extracellular signal-regulated kinase, c-Jun-N-terminal kinase, or intracellular Ca(2+) release were not required. Actinomycin D chase experiments suggested that enhanced stability of preproET-1 mRNA did not account for the increase in transcript levels. PreproET-1 promoter analysis demonstrated that the 5'-flanking region of preproET-1 encompassed positive regulatory elements engaged by thrombin. Negative modulation of thrombin-induced activation exerted by the distal 5' portion of preproET-1 promoter (-4.4 kbp to 204 bp) was overcome by co-stimulation with TNFalpha, providing a possible mechanism underlying the synergistic upregulation of preproET-1 expression by these two agonists. In conclusion, human mesangial cell expression of preproET-1 may be increased potently in the presence of two common proinflammatory mediators, thereby providing a potential mechanism for ET-1 production in inflammatory renal disease.

  • roles of calcium and kinases in regulation of thrombin stimulated Preproendothelin 1 transcription
    American Journal of Physiology-heart and Circulatory Physiology, 1996
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    The release of the vasoactive peptide endothelin-1 (ET-1) is Ca 2+ dependent after thrombin stimulation; however, little is known about the pathways involved. We studied the importance of Ca 2+ -dependent signal transduction pathways on preproET-1 mRNA induction in human endothelial cells. Thrombin-mediated preproET-1 mRNA induction was inhibited after clamping of cytosolic free Ca 2+ concentration ([Ca 2+ ] i ) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Chelation of extracellular Ca 2+ with ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid also had a significant inhibitory effect on the induction of preproET-1 mRNA. The Ca 2+ ionophore A23187 induced constitutive as well as thrombin-stimulated preproET-1 mRNAexpression. Mobilization of Ca 2+ stores into the cytosol by inhibition ofendoplasmic reticulum Ca 2+ -adenosinetriphosphatase with thapsigargin was effective also in inducing preproET-1 mRNA. Calmodulin antagonists W-7 and calmidazolium, as well as Ca 2+ /calmodulin-dependent kinase II inhibitor KN-62, significantly reduced thrombin-induced preproET-1 mRNA. Inhibition by cyclosporin A of the Ca 2+ -calmodulin-dependent phosphatase calcineurin potentiated constitutive preproET-1 mRNA. These data suggest that, in human endothelial cells, thrombin-mediated preproET-1 gene induction is regulated by a stimulatory Ca 2+ /calmodulin kinase II-dependent pathway.

  • Roles of calcium and kinases in regulation of thrombin-stimulated Preproendothelin-1 transcription.
    The American journal of physiology, 1996
    Co-Authors: Tobias A Marsen, M S Simonson, Michael J Dunn
    Abstract:

    The release of the vasoactive peptide endothelin-1 (ET-1) is Ca2+ dependent after thrombin stimulation; however, little is known about the pathways involved. We studied the importance of Ca(2+)-dependent signal transduction pathways on preproET-1 mRNA induction in human endothelial cells. Thrombin-mediated preproET-1 mRNA induction was inhibited after clamping of cytosolic free CA2+ concentration ([Ca2+]i) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Chelation of extracellular Ca2+ with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid also had a significant inhibitory effect on the induction of preproET-1 mRNA. The Ca2+ ionophore A23187 induced constitutive as well as thrombin-stimulated preproET-1 mRNA expression. Mobilization of Ca2+ stores into the cytosol by inhibition of endoplasmic reticulum Ca(2+)-adenosinetriphosphatase with thapsigargin was effective also in inducing preproET-1 mRNA. Calmodulin antagonists W-7 and calmidazolium, as well as Ca2+/calmodulin-dependent kinase II inhibitor KN-62, significantly reduced thrombin-induced preproET-1 mRNA. Inhibition by cyclosporin A of the Ca(2+)-calmodulin-dependent phosphatase calcineurin potentiated constitutive preproET-1 mRNA. These data suggest that, in human endothelial cells, thrombin-mediated preproET-1 gene induction is regulated by a stimulatory Ca2+/calmodulin kinase II-dependent pathway.

  • thrombin induces the Preproendothelin 1 gene in endothelial cells by a protein tyrosine kinase linked mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.

  • Thrombin Induces the Preproendothelin-1 Gene in Endothelial Cells by a Protein Tyrosine Kinase–Linked Mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.

Dror Harats - One of the best experts on this subject based on the ideXlab platform.

  • endothelin b receptor antagonist increases Preproendothelin 1 transcription in bovine aortic endothelial cells and in vivo
    Journal of Cardiovascular Pharmacology, 2006
    Co-Authors: Michael Peled, Aviv Shaish, Liron Frishman, Hofit Cohen, Dror Harats
    Abstract:

    Endothelin-1 (ET-1) receptor antagonists increase plasma immunoreactive ET-1 levels. However, their effect on Preproendothelin-1 (PPE-1) mRNA levels is still controversial. Few studies have found a decrease in PPE-1 mRNA levels in endothelial cells treated with the nonselective ET A/B receptor antagonist. and others demonstrated that an ET B blockade by the selective antagonist BQ788 increases PPE-1 mRNA levels. We studied the effect of ET A and ET B selective receptor antagonists on PPE-1 transcription, both in vitro and in vivo. Endothelial cells, transiently transfected with PPE-1 luciferase plasmid, were treated with ET-1 receptor antagonists. Bosentan, a dual ET A/B receptor antagonist, and BQ788 (ET B receptor antagonist) treatment resulted in a 1.6-fold and 1.3-fold increase, respectively in luciferase activity as compared with the untreated control. In contrast, the ET A receptor antagonist, BQ123, had no effect on luciferase activity. Transgenic mice that express the luciferase gene under the control of PPE-1 promoter were treated with Bosentan. Luciferase activity, PPE-1 mRNA levels, and plasma immunoreactive ET-1 levels were increased by 1.6-fold to 2.0-fold in the Bosentan-treated group compared with the untreated, control group. ET-1 receptor blockade increased PPE-1 transcription both in vitro and in vivo. The increased transcription can be attributed to ET B receptor blockade. because BQ-788, but not BQ-123, increased PPE-1 transcription.

  • inhibition of carcinogenesis in transgenic mouse models over expressing 15 lipoxygenase in the vascular wall under the control of murine Preproendothelin 1 promoter
    Cancer Letters, 2005
    Co-Authors: Dror Harats, Hofit Cohen, Dikla Benshushan, Ayelet Gonen, Iris Barshack, Iris Goldberg, Shoshana Greenberger, Israel Hodish, Ayelet Harari, Nira Vardabloom
    Abstract:

    Abstract Oxygenases are a family of enzymes that dioxygenate unsaturated fatty acids, thus initiating membrane oxidation and signaling molecule synthesis. The lipoxygenases (LOs), a family of lipid-peroxidizing enzymes that induce structural and metabolic changes in the cell in a number of pathophysiological conditions, belong to the oxygenases family. This class of enzymes has several subgroups, named 5-, 8-, 12- and 15-LOs, and these LO-isoforms are capable of oxygenating arachidonic and linoleic acid. 15-LOs were reported to play an inhibitory role in tumor angiogenesis and, consequently, they slow down carcinogenesis. It has been suggested that its anti-carcinogenic effect is conferred by promoting cell differentiation and apoptosis. Using transgenic mice that over-express 15-LO-1 in endothelial cells under the regulation of the murine Preproendothelin-1 promoter, we studied its effect on tumor and metastasis growth. We found that 15-LO-1 inhibited tumor and metastasis growth in the transgenic mice in two different models of cancer (mammary gland and Lewis lung carcinoma). This inhibition was concomitant with a higher number of apoptotic cells in the metastases of the transgenic mice and with a complicated network of multiple small blood vessels. This finding targets 15-LO as a new candidate in the treatment of carcinogenesis.

  • targeting gene expression to the vascular wall in transgenic mice using the murine Preproendothelin 1 promoter
    Journal of Clinical Investigation, 1995
    Co-Authors: Dror Harats, Hiroki Kurihara, Yukiko Kurihara, Paula Nanette Belloni, Heather Oakley, Amy Ziober, Don Ackley, Gary Cain, Richard Lawn, Elliott Sigal
    Abstract:

    To develop a system for overexpressing genes in the vascular wall, we created transgenic mice using the reporter gene luciferase and the murine Preproendothelin-1 promoter. In vitro analysis suggested that the murine 5'-flanking region contained endothelial-specific elements in a 5.9-kb fragment. Five transgenic mice colonies established from independent founders all exhibited the highest level of luciferase activity in the aorta with up to 8,540 light units per microgram of protein. Immunohistochemistry with anti-luciferase antisera revealed high levels of expression in the endothelial cells of both large and small arteries and lower levels of expression in veins and capillaries. Significant expression was also seen in arterial smooth muscle cells and in select epithelial surfaces which is consistent with the known distribution of endothelin-1 in mammals. To further demonstrate the targeting capability of this system, we overexpressed the lipid-peroxidating enzyme, human 15-lipoxygenase, in the vessel wall of transgenic mice. As with luciferase, expression of active enzyme and immunohistochemical localization in vascular cells were documented in transgenic animals. Hence, this new system can be used to direct expression of molecules to the vascular wall for the purpose of examining the biological significance of either overexpression or inhibition of select proteins. (J. Clin. Invest. 1995. 95:1335-1344.)

Michael S Simonson - One of the best experts on this subject based on the ideXlab platform.

  • pneumoperitoneum upregulates Preproendothelin 1 messenger rna
    Surgical Endoscopy and Other Interventional Techniques, 2001
    Co-Authors: J A Ambrose, Michael S Simonson, Raymond P Onders, Nicholas T Stowe, Ann V Robinson, Scott M Wilhelm, James A Schulak
    Abstract:

    Background Laparoscopic pneumoperitoneum has been shown to decrease glomerular filtration rate (GFR) and urine volume (UV). Endothelin-1 (ET-1), a potent renal vasoconstrictor, has been implicated. The purpose of this study was to determine renal function, ET-1 gene expression, and peptide localization in kidneys subjected to CO2 pneumoperitoneum.

  • specific repression of the Preproendothelin 1 gene in intracranial arteriovenous malformations
    Journal of Neurosurgery, 1997
    Co-Authors: R Patrick L Rhoten, Youssef G Comair, Daniel Shedid, Douglas Chyatte, Michael S Simonson
    Abstract:

    ✓ Cerebrovascular arteriovenous malformations (AVMs) display abnormal vascular development and dysautoregulation of blood flow. Genetic mechanisms that contribute to the pathogenesis and phenotype of cerebral AVMs are unknown. As a first step in understanding the pathophysiology of AVMs, the authors investigated the hypothesis that endothelial dysfunction—specifically, deregulation of endothelin-1 (ET-1) secretion—contributes to the abnormal vascular phenotype and the lack of hemodynamic autoregulation elaborated by these lesions. Endothelin-1 peptide and Preproendothelin-1 (ppET1) messenger RNA were not detected in the intranidal vasculature of all 17 patients with AVMs studied, but were prominently expressed in human control subjects with normal cerebrovasculature (p < 0.01). Although AVM vasculature lacked ET-1, its expression was prominent in vasculature distant from these lesions, suggesting local repression of the ppET-1 gene. Local repression of ET-1 was specific to AVMs; ET-1 in vascular malformat...

  • roles of calcium and kinases in regulation of thrombin stimulated Preproendothelin 1 transcription
    American Journal of Physiology-heart and Circulatory Physiology, 1996
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    The release of the vasoactive peptide endothelin-1 (ET-1) is Ca 2+ dependent after thrombin stimulation; however, little is known about the pathways involved. We studied the importance of Ca 2+ -dependent signal transduction pathways on preproET-1 mRNA induction in human endothelial cells. Thrombin-mediated preproET-1 mRNA induction was inhibited after clamping of cytosolic free Ca 2+ concentration ([Ca 2+ ] i ) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Chelation of extracellular Ca 2+ with ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid also had a significant inhibitory effect on the induction of preproET-1 mRNA. The Ca 2+ ionophore A23187 induced constitutive as well as thrombin-stimulated preproET-1 mRNAexpression. Mobilization of Ca 2+ stores into the cytosol by inhibition ofendoplasmic reticulum Ca 2+ -adenosinetriphosphatase with thapsigargin was effective also in inducing preproET-1 mRNA. Calmodulin antagonists W-7 and calmidazolium, as well as Ca 2+ /calmodulin-dependent kinase II inhibitor KN-62, significantly reduced thrombin-induced preproET-1 mRNA. Inhibition by cyclosporin A of the Ca 2+ -calmodulin-dependent phosphatase calcineurin potentiated constitutive preproET-1 mRNA. These data suggest that, in human endothelial cells, thrombin-mediated preproET-1 gene induction is regulated by a stimulatory Ca 2+ /calmodulin kinase II-dependent pathway.

  • thrombin induces the Preproendothelin 1 gene in endothelial cells by a protein tyrosine kinase linked mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.

  • Thrombin Induces the Preproendothelin-1 Gene in Endothelial Cells by a Protein Tyrosine Kinase–Linked Mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.

Elliott Sigal - One of the best experts on this subject based on the ideXlab platform.

  • targeting gene expression to the vascular wall in transgenic mice using the murine Preproendothelin 1 promoter
    Journal of Clinical Investigation, 1995
    Co-Authors: Dror Harats, Hiroki Kurihara, Yukiko Kurihara, Paula Nanette Belloni, Heather Oakley, Amy Ziober, Don Ackley, Gary Cain, Richard Lawn, Elliott Sigal
    Abstract:

    To develop a system for overexpressing genes in the vascular wall, we created transgenic mice using the reporter gene luciferase and the murine Preproendothelin-1 promoter. In vitro analysis suggested that the murine 5'-flanking region contained endothelial-specific elements in a 5.9-kb fragment. Five transgenic mice colonies established from independent founders all exhibited the highest level of luciferase activity in the aorta with up to 8,540 light units per microgram of protein. Immunohistochemistry with anti-luciferase antisera revealed high levels of expression in the endothelial cells of both large and small arteries and lower levels of expression in veins and capillaries. Significant expression was also seen in arterial smooth muscle cells and in select epithelial surfaces which is consistent with the known distribution of endothelin-1 in mammals. To further demonstrate the targeting capability of this system, we overexpressed the lipid-peroxidating enzyme, human 15-lipoxygenase, in the vessel wall of transgenic mice. As with luciferase, expression of active enzyme and immunohistochemical localization in vascular cells were documented in transgenic animals. Hence, this new system can be used to direct expression of molecules to the vascular wall for the purpose of examining the biological significance of either overexpression or inhibition of select proteins. (J. Clin. Invest. 1995. 95:1335-1344.)

Tobias A Marsen - One of the best experts on this subject based on the ideXlab platform.

  • roles of calcium and kinases in regulation of thrombin stimulated Preproendothelin 1 transcription
    American Journal of Physiology-heart and Circulatory Physiology, 1996
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    The release of the vasoactive peptide endothelin-1 (ET-1) is Ca 2+ dependent after thrombin stimulation; however, little is known about the pathways involved. We studied the importance of Ca 2+ -dependent signal transduction pathways on preproET-1 mRNA induction in human endothelial cells. Thrombin-mediated preproET-1 mRNA induction was inhibited after clamping of cytosolic free Ca 2+ concentration ([Ca 2+ ] i ) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Chelation of extracellular Ca 2+ with ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid also had a significant inhibitory effect on the induction of preproET-1 mRNA. The Ca 2+ ionophore A23187 induced constitutive as well as thrombin-stimulated preproET-1 mRNAexpression. Mobilization of Ca 2+ stores into the cytosol by inhibition ofendoplasmic reticulum Ca 2+ -adenosinetriphosphatase with thapsigargin was effective also in inducing preproET-1 mRNA. Calmodulin antagonists W-7 and calmidazolium, as well as Ca 2+ /calmodulin-dependent kinase II inhibitor KN-62, significantly reduced thrombin-induced preproET-1 mRNA. Inhibition by cyclosporin A of the Ca 2+ -calmodulin-dependent phosphatase calcineurin potentiated constitutive preproET-1 mRNA. These data suggest that, in human endothelial cells, thrombin-mediated preproET-1 gene induction is regulated by a stimulatory Ca 2+ /calmodulin kinase II-dependent pathway.

  • Roles of calcium and kinases in regulation of thrombin-stimulated Preproendothelin-1 transcription.
    The American journal of physiology, 1996
    Co-Authors: Tobias A Marsen, M S Simonson, Michael J Dunn
    Abstract:

    The release of the vasoactive peptide endothelin-1 (ET-1) is Ca2+ dependent after thrombin stimulation; however, little is known about the pathways involved. We studied the importance of Ca(2+)-dependent signal transduction pathways on preproET-1 mRNA induction in human endothelial cells. Thrombin-mediated preproET-1 mRNA induction was inhibited after clamping of cytosolic free CA2+ concentration ([Ca2+]i) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Chelation of extracellular Ca2+ with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid also had a significant inhibitory effect on the induction of preproET-1 mRNA. The Ca2+ ionophore A23187 induced constitutive as well as thrombin-stimulated preproET-1 mRNA expression. Mobilization of Ca2+ stores into the cytosol by inhibition of endoplasmic reticulum Ca(2+)-adenosinetriphosphatase with thapsigargin was effective also in inducing preproET-1 mRNA. Calmodulin antagonists W-7 and calmidazolium, as well as Ca2+/calmodulin-dependent kinase II inhibitor KN-62, significantly reduced thrombin-induced preproET-1 mRNA. Inhibition by cyclosporin A of the Ca(2+)-calmodulin-dependent phosphatase calcineurin potentiated constitutive preproET-1 mRNA. These data suggest that, in human endothelial cells, thrombin-mediated preproET-1 gene induction is regulated by a stimulatory Ca2+/calmodulin kinase II-dependent pathway.

  • thrombin induces the Preproendothelin 1 gene in endothelial cells by a protein tyrosine kinase linked mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.

  • Thrombin Induces the Preproendothelin-1 Gene in Endothelial Cells by a Protein Tyrosine Kinase–Linked Mechanism
    Circulation Research, 1995
    Co-Authors: Tobias A Marsen, Michael S Simonson, Michael J Dunn
    Abstract:

    Abstract Thrombin stimulates synthesis and secretion of endothelin-1 (ET-1), a vasoactive peptide that triggers responses in the vascular endothelium and smooth muscle. We investigated the signal transduction pathways by which thrombin stimulates preproET-1 gene expression and ET-1 peptide secretion in macrovascular cells (human umbilical vein endothelial cells [HUVECs] and bovine pulmonary artery endothelial cells [BPAECs]) and microvascular cells (human microvascular endothelial cell line [HMEC-1]). Thrombin (4 U/mL) stimulated maximal induction of ET-1 peptide secretion and preproET-1 mRNA after 2 hours in HUVECs and BPAECs and after 1 hour in HMEC-1. A synthetic thrombin receptor activator peptide confirmed ligand-specific receptor actions to induce preproET-1 mRNA. Protein kinase C (PKC) activation by phorbol ester transiently induced preproET-1 mRNA but had no effect on ET-1 peptide synthesis. PKC inhibitors sangivamycin and calphostin C and PKC depletion failed to suppress thrombin-stimulated preproET-1 mRNA. Adenylate cyclase and cAMP-dependent protein kinase did not participate in thrombin-induced preproET-1 gene activation. Thrombin stimulated a rapid increase in phosphotyrosine-containing proteins, suggesting a role for tyrosine phosphorylation in thrombin signaling. These data demonstrate that thrombin induces the preproET-1 gene and ET-1 peptide synthesis by a PKC-independent PTK-dependent pathway in macrovascular and microvascular endothelial cells. Protein tyrosine kinase inhibitors herbimycin A and genistein blocked thrombin-stimulated preproET-1 mRNA and peptide secretion, whereas daidzein, which lacks inhibitory activity, did not suppress thrombin-induced ET-1.