AIM: To investigate the effects of the potassium-sparing diuretic amiloride on endothelial cell apoptosis during lipopolysaccharide (LPS)-accelerated atherosclerosis. METHODS: Human umbilical vein endothelial cells (HUVECs) were exposed to LPS (100 ng/mL) in the presence of drugs tested. The activity of Na(+)/H(+) exchanger 1 (NHE1) and calpain, intracellular free Ca(2+)level ([Ca(2+)](i)), as well as the expression of apoptosis-related proteins in the cells were measured. For in vivo study, ApoE-deficient (ApoE(-/-)) mice were fed high-fat diets with 0.5% (w/w) amiloride for 4 weeks and LPS (10 μg/mouse) infusion into caudal veins. Afterwards, atherosclerotic lesions, NHE1 activity and Bcl-2 expression in the aortic tissues were evaluated. RESULTS: LPS treatment increased NHE1 activity and [Ca(2+)](i) in HUVECs in a time-dependent manner, which was associated with increased activity of the Ca(2+)-dependent protease calpain. Amiloride (1-10 μmol/L) significantly suppressed LPS-induced increases in NHE1 activity, [Ca(2+)](i). and calpain activity. In the presence of the Ca(2+) chelator BAPTA (0.5 mmol/L), LPS-induced increase of calpain activity was also abolished. In LPS-treated HUVECs, the expression of Bcl-2 protein was significantly decreased without altering its mRNA level. In the presence of amiloride (10 μmol/L) or the calpain inhibitor ZLLal (50 μmol/L), the down-regulation of Bcl-2 protein by LPS was blocked. LPS treatment did not alter the expression of Bax and Bak proteins in HUVECs. In the presence of amiloride, BAPTA or ZLLal, LPS-induced HUVEC apoptosis was significantly attenuated. In ApoE(-/-) mice, administration of amiloride significantly suppressed LPS-accelerated atherosclerosis and LPS-induced increase of NHE1 activity, and reversed LPS-induced down-regulation of Bcl-2 expression. CONCLUSION: LPS stimulates NHE1 activity, increases [Ca(2+)](i), and activates calpain, which leads to endothelial cell apoptosis related to decreased Bcl-2 expression. Amiloride inhibits NHE1 activity, thus attenuates LPS-accelerated atherosclerosis in mice.
AIM: To investigate the effects of the potassium-sparing diuretic amiloride on endothelial cell apoptosis during lipopolysaccharide (LPS)-accelerated atherosclerosis. METHODS:Human umbilical vein endothelial cells (HUVECs) were exposed to LPS (100 ng/mL) in the presence of drugs tested. The activity of Na(+)/H(+) exchanger 1 (NHE1) and calpain, intracellular free Ca(2+)level ([Ca(2+)](i)), as well as the expression of apoptosis-related proteins in the cells were measured. For in vivo study, ApoE-deficient (ApoE(-/-)) mice were fed high-fat diets with 0.5% (w/w) amiloride for 4 weeks and LPS (10 μg/mouse) infusion into caudal veins. Afterwards, atherosclerotic lesions, NHE1 activity and Bcl-2 expression in the aortic tissues were evaluated. RESULTS:LPS treatment increased NHE1 activity and [Ca(2+)](i) in HUVECs in a time-dependent manner, which was associated with increased activity of the Ca(2+)-dependent protease calpain. Amiloride (1-10 μmol/L) significantly suppressed LPS-induced increases in NHE1 activity, [Ca(2+)](i). and calpain activity. In the presence of the Ca(2+) chelator BAPTA (0.5 mmol/L), LPS-induced increase of calpain activity was also abolished. In LPS-treated HUVECs, the expression of Bcl-2 protein was significantly decreased without altering its mRNA level. In the presence of amiloride (10 μmol/L) or the calpain inhibitor ZLLal (50 μmol/L), the down-regulation of Bcl-2 protein by LPS was blocked. LPS treatment did not alter the expression of Bax and Bak proteins in HUVECs. In the presence of amiloride, BAPTA or ZLLal, LPS-induced HUVEC apoptosis was significantly attenuated. In ApoE(-/-) mice, administration of amiloride significantly suppressed LPS-accelerated atherosclerosis and LPS-induced increase of NHE1 activity, and reversed LPS-induced down-regulation of Bcl-2 expression. CONCLUSION:LPS stimulates NHE1 activity, increases [Ca(2+)](i), and activates calpain, which leads to endothelial cell apoptosis related to decreased Bcl-2 expression. Amiloride inhibits NHE1 activity, thus attenuates LPS-accelerated atherosclerosis in mice.
Authors: D B Cines; E S Pollak; C A Buck; J Loscalzo; G A Zimmerman; R P McEver; J S Pober; T M Wick; B A Konkle; B S Schwartz; E S Barnathan; K R McCrae; B A Hug; A M Schmidt; D M Stern Journal: Blood Date: 1998-05-15 Impact factor: 22.113
Authors: Laween Uthman; Antonius Baartscheer; Cees A Schumacher; Jan W T Fiolet; Marius C Kuschma; Markus W Hollmann; Ruben Coronel; Nina C Weber; Coert J Zuurbier Journal: Front Physiol Date: 2018-11-21 Impact factor: 4.566