Endothelin-1 expression in vascular adventitial fibroblasts.
Endothelial cells are a major source of endothelin (ET)-1, but the possibility that vascular adventitial fibroblasts generate ET-1 has not been explored. We hypothesized that aortic adventitial fibroblasts have the ability to produce ET-1, which may contribute to extracellular matrix synthesis. Vascular adventitial fibroblasts were isolated from mouse aorta and incubated with various concentrations of angiotensin II (ANG II). mRNA levels of preproET-1 and type I procollagen were detected with relative RT-PCR. ET-1 levels in culture medium were measured with ELISA. Protein levels of procollagen were detected with Western blotting. ANG II (10 and 100 nM, 1 microM) induced a time- and concentration-dependent increase in preproET-1 mRNA levels (P < 0.05). Induction of preproET-1 mRNA was accompanied by release of immunoreactive peptide ET-1 (P < 0.05). ANG II-evoked increases in preproET-1 mRNA expression and ET-1 release were blocked by losartan (100 microM), an AT1 receptor antagonist, but not PD-123319 (100 microM), an AT2 receptor antagonist. To further confirm our findings, we cloned and then sequenced vascular fibroblast preproET-1 bidirectionally with T7 and M13 reverse sequencing primers. Their nucleotide sequences were identical to preproET-1 cDNA from mouse vascular endothelial cells (accession no. AB081657). Moreover, ANG II-induced type I procollagen mRNA and protein expression were inhibited by BQ-123 (10 microM), an ET(A) receptor inhibitor, but not BQ-788 (10 microM), an ET(B) receptor inhibitor, suggesting a significant role of adventitial ET-1 in regulation of extracellular matrix synthesis. The results demonstrate that vascular adventitial fibroblasts are able to synthesize and release ET-1 in response to ANG II.
An
Sheng Jun
SJ
Dept. of Community Health Sciences, Faculty of Applied Health Sciences, Brock Univ., St. Catharines, ON, Canada L2S 3Y6.
Boyd
Ryan
R
Wang
Ying
Y
Qiu
Xiaofan
X
Wang
Hui Di
HD
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001013:Aorta, Thoracic / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000378:metabolism; D003238:Connective Tissue / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D019332:Endothelin-1 / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism; D044022:Receptor, Endothelin A / Q000378:metabolism; D044023:Receptor, Endothelin B / Q000378:metabolism
D024042:Collagen Type I; D018076:DNA, Complementary; D019332:Endothelin-1; D012333:RNA, Messenger; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D000804:Angiotensin II