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Lanthanum chloride suppresses oxysterol-induced ECV-304 cell apoptosis via inhibition of intracellular Ca2+ concentration elevation, oxidative stress, and activation of ERK and NF-κB signaling pathways

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Abstract

Experimental studies have demonstrated that oral administration of lanthanum chloride (LaCl3) inhibits the development of atherosclerosis, but the related mechanism has not been fully elucidated. Oxysterols are toxic to the vascular endothelial cells which are important in preventing the formation and progression of atheromatous plaque. In this study, we examined the effect of LaCl3 on oxysterol cholestane-3β,5α,6β-triol (Triol)-induced apoptosis and the related mechanisms in ECV-304 cells, a presumptive endothelial cell line. Incubation with Triol resulted in apoptosis of ECV-304 cells, as determined by Hoechst 33342 staining, fluorescein isothiocyanate labeled annexin V/propidium iodide double staining, and the loss of mitochondrial membrane potential. Triol activated extracellular-signal-regulated kinase (ERK) and nuclear factor κB (NF-κB), and inhibition of Triol-activated ERK and NF-κB signaling by specific inhibitors attenuated apoptosis induction by Triol in ECV-304 cells. Pretreatment with LaCl3 (1 μM) for 12 h before exposure to Triol decreased Triol-mediated apoptosis as well as activation of ERK and NF-κB. In addition, Triol induced oxidative stress in ECV-304 cells, manifested by the increase of intracellular reactive oxygen species generation and malondialdehyde level, and the reduction of the content of total protein thiols and the activity of antioxidant glutathione peroxidases; LaCl3 pretreatment significantly reversed these effects. Finally, LaCl3 pretreatment significantly inhibited the increases of intracellular Ca2+ concentration induced by Triol. Our study suggests that Triol induced ECV-304 cell apoptosis, and LaCl3 could suppress this effect probably by inhibiting intracellular Ca2+ concentration elevation, oxidative stress, as well as activation of ERK and NF-κB signaling pathways.

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Abbreviations

BSA:

Bovine serum albumin

CAPE:

Caffeic acid phenethyl ester

2,7-DCFH-DA:

2,7-Dichlorofluorescein diacetate

DMEM:

Dulbecco’s modified Eagle’s medium

DTNB:

Dithiobis(2-nitrobenzoic acid)

DTT:

Dithiothreitol

ECL:

Enhanced chemiluminescence

ERK:

Extracellular-signal-regulated kinase

FITC:

Fluorescein isothiocyanate

GPx:

Glutathione peroxidase

GSH:

Reduced glutathione

HEPES:

4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid

MDA:

Malondialdehyde

MEK:

Mitogen-activated protein kinase kinase

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NCS:

Newborn calf serum

NF-κB:

Nuclear factor κB

oxLDL:

Oxidized low-density lipoprotein

PBS:

Phosphate-buffered saline

pERK:

Phosphorylated extracellular-signal-regulated kinase

PI:

Propidium iodide

pIκB:

Phosphorylated IκB

PMSF:

Phenylmethylsulfonyl fluoride

Rh-123:

Rhodamine 123

ROS:

Reactive oxygen species

SD:

Standard deviation

TBA:

Thiobarbituric acid

Triol:

Cholestane-3β,5α,6β-triol

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (grant nos. 20637010 and 30700136).

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Authors and Affiliations

  1. Hubei Key Laboratory of Bioinorganic Chemistry and Materia Medica, School of Chemistry and Chemical Engineering, Huazhong University of Science and Technology, Wuhan, 430074, People’s Republic of China

    Hongmei Liu, Congcong Zhang & Kaixun Huang

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  1. Hongmei Liu
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  2. Congcong Zhang
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  3. Kaixun Huang
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Correspondence to Hongmei Liu.

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Liu, H., Zhang, C. & Huang, K. Lanthanum chloride suppresses oxysterol-induced ECV-304 cell apoptosis via inhibition of intracellular Ca2+ concentration elevation, oxidative stress, and activation of ERK and NF-κB signaling pathways. J Biol Inorg Chem 16, 671–681 (2011). https://doi.org/10.1007/s00775-011-0766-y

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  • DOI: https://doi.org/10.1007/s00775-011-0766-y

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