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Molecular Pharmacology

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Rapid CommunicationAccelerated Communication

Effect of an Antisense Oligodeoxynucleotide to Endothelin-Converting Enzyme-1c (ECE-1c) on ECE-1c mRNA, ECE-1 Protein and Endothelin-1 Synthesis in Bovine Pulmonary Artery Smooth Muscle Cells

Stewart Barker, Noorafza Q. Khan, Elizabeth G. Wood and Roger Corder
Molecular Pharmacology February 2001, 59 (2) 163-169; DOI: https://doi.org/10.1124/mol.59.2.163
Stewart Barker
Department of Experimental Therapeutics, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, Charterhouse Square Campus, London, EC1M 6BQ, UK
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Noorafza Q. Khan
Department of Experimental Therapeutics, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, Charterhouse Square Campus, London, EC1M 6BQ, UK
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Elizabeth G. Wood
Department of Experimental Therapeutics, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, Charterhouse Square Campus, London, EC1M 6BQ, UK
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Roger Corder
Department of Experimental Therapeutics, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, Charterhouse Square Campus, London, EC1M 6BQ, UK
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Abstract

Endothelin-1 (ET-1) is secreted from endothelial and vascular smooth muscle cells (VSMC) after intracellular hydrolysis of big ET-1 by endothelin converting enzyme (ECE). The metallopeptidase called ECE-1 is widely thought to be the physiological ECE, but unequivocal evidence of this role has yet to be provided. Endothelial cells express four isoforms of ECE-1 (ECE-1a, ECE-1b, ECE-1c, and ECE-1d), but the identity of ECE-1 isoforms expressed in VSMC is less clear. Here, we describe the characterization of ECE-1 isoforms in bovine pulmonary artery smooth muscle cells (BPASMC) and the effect on ET-1 synthesis of an antisense oligodeoxynucleotide (ODN) to ECE-1c. Reverse transcriptase-polymerase chain reaction (RT-PCR) evaluation of total RNA from BPASMC showed that ECE-1a and ECE-1d were not expressed. Sequencing of cloned ECE-1 cDNA products and semiquantitative RT-PCR demonstrated that ECE-1b and ECE-1c were expressed in BPASMC, with ECE-1c being the predominant isoform. Basal release of ET-1 from BPASMC was low. Treatment for 24 h with tumor necrosis factor-α (TNFα) stimulated ET-1 production by up to 10-fold with parallel increases in levels of preproET-1 mRNA. Levels of ECE-1c mRNA were also raised after TNFα, whereas amounts of ECE-1b mRNA were decreased significantly. Treatment of BPASMC with a phosphorothioate antisense ODN to ECE-1c caused a marked reduction in ECE-1c mRNA levels and ECE-1 protein levels. However, basal and TNFα-stimulated ET-1 release were largely unaffected by the ECE-1c antisense ODN despite the inhibition of ECE-1c synthesis. Hence, an endopeptidase distinct from ECE-1 is mainly responsible big ET-1 processing in BPASMC.

Footnotes

    • Received August 22, 2000.
    • Accepted October 18, 2000.
  • Send reprint requests to: Prof. Roger Corder, Department of Experimental Therapeutics, William Harvey Research Institute, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, Charterhouse Square Campus, London, EC1M 6BQ, UK. E-mail: r.corder{at}mds.qmw.ac.uk

  • This work was supported by the Institut de Recherches Internationales Servier and The William Harvey Research Foundation.

  • The American Society for Pharmacology and Experimental Therapeutics
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Molecular Pharmacology: 59 (2)
Molecular Pharmacology
Vol. 59, Issue 2
1 Feb 2001
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Effect of an Antisense Oligodeoxynucleotide to Endothelin-Converting Enzyme-1c (ECE-1c) on ECE-1c mRNA, ECE-1 Protein and Endothelin-1 Synthesis in Bovine Pulmonary Artery Smooth Muscle Cells
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Rapid CommunicationAccelerated Communication

Effect of an Antisense Oligodeoxynucleotide to Endothelin-Converting Enzyme-1c (ECE-1c) on ECE-1c mRNA, ECE-1 Protein and Endothelin-1 Synthesis in Bovine Pulmonary Artery Smooth Muscle Cells

Stewart Barker, Noorafza Q. Khan, Elizabeth G. Wood and Roger Corder
Molecular Pharmacology February 1, 2001, 59 (2) 163-169; DOI: https://doi.org/10.1124/mol.59.2.163

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Rapid CommunicationAccelerated Communication

Effect of an Antisense Oligodeoxynucleotide to Endothelin-Converting Enzyme-1c (ECE-1c) on ECE-1c mRNA, ECE-1 Protein and Endothelin-1 Synthesis in Bovine Pulmonary Artery Smooth Muscle Cells

Stewart Barker, Noorafza Q. Khan, Elizabeth G. Wood and Roger Corder
Molecular Pharmacology February 1, 2001, 59 (2) 163-169; DOI: https://doi.org/10.1124/mol.59.2.163
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