Endothelin-1 Attenuates ω3 Fatty Acid–Induced Apoptosis by Inhibition of Caspase 3

• Published in: Hypertension (Dallas, Tex. 1979) Tex. 1979), 2000-01, Vol.35 (1, Part 2 Suppl), p.287-287
• Main Authors: Diep, Quy N, Intengan, Hope D, Schiffrin, Ernesto L
• Format: Article
• Language: English
• ISSN: 0194-911X; 1524-4563
• DOI: 10.1161/01.hyp.35.1.287

ABSTRACTEndothelin-1 (ET-1) may be involved in the induction of vascular hypertrophy in hypertension. ET-1 may also modulate vascular growth through the exertion of antiapoptotic effects. The ω3 fatty acids (ω3 FAs), which have antiproliferative effects in various cell types, may have a beneficial role in hypertension. We tested the hypothesis that ET-1 could act as a survival factor against ω3 FA–induced apoptosis and attempted to elucidate possible molecular mechanisms underlying the protective action of ET-1 on docosahexaenoic acid (DHA)-induced apoptosis. Mesenteric vascular smooth muscle cells were stimulated with DHA, a representative ω3 FA. Dose-response curves of DHA at different apoptotic stages were assessed with the use of flow cytometry(1) very earlyplasma membrane phosphatidylserine (PS) translocation; (2) earlychange in mitochondrial transmembrane potential (ΔΨm); and (3) latecell cycle analysis. Expression of the proapoptotic protein bax and the antiapoptotic protein bcl-2 was determined with Western blot assay. The activity and the expression of caspase 3, which is a critical proteolytic enzyme involved in the death-signaling pathway, were evaluated with a fluorometric immunosorbent enzyme assay and Western blot analysis, respectively. Apoptosis, which was detected with PS translocation, ΔΨm disruption, and cell cycle analysis, was increased dose dependently by DHA. DHA-induced apoptosis was attenuated through exposure to ET-1 for 1 hour before DHA in cell cycle analysis. The interference of ET-1 with DHA-induced apoptosis, as detected with cell cycle analysis, was not apparent at the membrane (PS translocation) or the mitochondrial (ΔΨm) level. The increase in bax/bcl-2 ratio in DHA-stimulated cells was not affected by ET-1. However, DHA increased both caspase 3 activity and the active forms of caspase 3 (20 and 17 kDa), resulting in enhanced DNA fragmentation as shown through Hoechst staining and fluorescence microscopy, which were attenuated by ET-1 pretreatment. In conclusion, DHA, an ω3 FA, induced apoptosis in vascular smooth muscle cells in a dose-dependent manner. ET-1 exerted important protective effects through the attenuation of DHA-induced caspase 3 activation and subsequent DNA fragmentation in the late stages of apoptosis.