Ethanol inhibits γ-secretase proteolytic activity in vascular smooth muscle cells.
Ethanol inhibits γ-secretase proteolytic activity in vascular smooth muscle cells.
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DOI：
10.1111/acer.12875
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发表时间：
2015-11
期刊：
影响因子：
--
通讯作者：
Redmond EM
中科院分区：
文献类型：
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作者：
Hatch E;Morrow D;Liu W;Cahill PA;Redmond EM
Ethanol (EtOH) inhibits Notch-mediated vascular smooth muscle cell (SMC) proliferation, an event that is key in vessel remodeling and atherogenesis. The object of this study was to determine if EtOH inhibits Notch signaling in SMC at the level of γ-secretase, a protease that in concert with α-secretase, catalyzes the release of the intracellular domain of the Notch receptor necessary for signaling. Human coronary artery SMC (HCASMC) were treated with a recombinant soluble Notch ligand, DLL4 (2 μg/ml), or transfected with a constitutively active Notch 1 intracellular domain (N1ICD), in the absence or presence of EtOH. Ethanol (25 mM) treatment inhibited DLL4-stimulated CBF-1/RBP-Jk dependent promoter activity (determined by luciferase assay) and downstream target gene HRT3 mRNA levels. In contrast, ethanol had no effect on N1ICD-driven CBF1/RBP-Jk dependent promoter activity or HRT-3 expression. These data suggest that EtOH inhibits Notch signaling at, or prior to, NICD generation. Gamma-secretase activity was determined in solubilized membrane preparations from HCASMC treated with/without EtOH (25 mM) or the γ-secretase inhibitor DAPT (20 μM) using (i) a fluorometric assay and (ii) western blot detection of cleavage products using a Flag-tagged Notch based substrate, N100Flag. EtOH inhibited basal and DLL4-stimulated γ-secretase activity, and SMC growth to a similar extent as DAPT, whereas it had no effect on α-secretase (TACE/ADAM 17) activity also determined by fluorometric assay. Moreover, EtOH treatment inhibited the expression of caveolin-1, a lipid raft protein implicated in regulating γ-secretase activity, and altered its cellular distribution in HCASMC. Ethanol inhibits Notch signaling in vascular smooth muscle cells at the level of γ-secretase activity, possibly by affecting lipid raft function. Such a response might be expected to result in attenuation of pathologic vessel remodeling and thus may contribute to moderate alcohols’ cardioprotective effects.
