PT  - JOURNAL ARTICLE
AU  - Barker, Stewart
AU  - Khan, Noorafza Q.
AU  - Wood, Elizabeth G.
AU  - Corder, Roger
TI  - 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
AID  - 10.1124/mol.59.2.163
DP  - 2001 Feb 01
TA  - Molecular Pharmacology
PG  - 163--169
VI  - 59
IP  - 2
4099  - http://molpharm.aspetjournals.org/content/59/2/163.short
4100  - http://molpharm.aspetjournals.org/content/59/2/163.full
SO  - Mol Pharmacol2001 Feb 01; 59
AB  - 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.