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Lee, Kang, Choi, Lee, Rhyu, and Yoon: Cytotoxic Effects of Lysophosphatidylcholine on Vascular Smooth Muscle Cells
Original Article
The Journal of Korean Society of Menopause

The Journal of Korean Society of Menopause 2012; 18(3): 139-146.

Published online: 31 December 2012

DOI: https://doi.org/10.6118/jksm.2012.18.3.139

Cytotoxic Effects of Lysophosphatidylcholine on Vascular Smooth Muscle Cells

Dong-Yun Lee, M.D.1, Young-Hee Kang, M.S.2, Doo Seok Choi, M.D.1, 2, Young Joo Lee, Ph.D.3, Mee-Ra Rhyu, Ph.D.4, Byung-Koo Yoon, M.D., Ph.D.1, 2

1Department of Obstetrics and Gynecology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

2Samsung Biomedical Research Institute, Seoul, Korea.

3Department of Bioscience and Biotechnology, Sejong University, Seoul, Korea.

4Divisions of Metabolism and Functionality Research, Korea Food Research Institute, Seongnam, Korea.

Address for Correspondence: Byung-Koo Yoon, Department of Obstetrics and Gynecology, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-ro Gangnam-gu, Seoul 135-710, Korea. Tel: +82-2-3410-3519, Fax: +82-2-3410-0630, bkyoon@skku.edu

Received 29 June 2012       Revised 4 September 2012       Accepted 19 September 2012

Copyright © 2012 by The Korean Society of Menopause

(open-access):

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/).

Abstract

Objectives

To investigate the cytotoxic effects of lysophosphatidylcholine (lysoPC), an active component of oxidized low-density lipoproteins (LDL), on vascular smooth muscle cells (VSMCs).

Methods

VSMCs were derived from rat aorta. Cell death was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay, lactic dehydrogenase (LDH) assay, and DNA fragmentation assay. Apoptosis was quantified by propidium iodide staining and fluorescent activated cell sorting (FACS) analysis, and intracellular free radical production was determined using 2',7'-dichlorofluorescin diacetate (DCF-DA). In addition, the changes in caspases, bcl-2 and bax proteins were evaluated by western blot analysis.

Results

LysoPC over 25 µM induced more than 50% of the cell death at 10 hours on MTT assay with no change in the level of LDH. The DNA ladder pattern showed that cell death induced by lysoPC was caused by apoptosis, which was associated with increased free radical production. Vitamin E, a potent antioxidant and caffeic acid phenylethyl ester (CAPE), an inhibitor of nuclear factor-kappaB (NF-κB), blocked apoptosis. The casepase-3 precursor decreased and the active form of caspase-8 increased. Total bcl-2 and bax proteins did not change with lysoPC treatment, but translocation of bax from cytosole to the mitochondria membrane was observed.

Conclusion

LysoPC induces apoptosis in VSMCs via an oxidant mechanism, dependent on NF-κB.

Keywords: Apoptosis, Atherosclerosis, Lysophosphatidylcholines, Vascular smooth muscle cell

Figures and Tables

Fig. 1
Cytotoxic effects of lysophosphatidylcholine (lysoPC) on vascular smooth muscle cells (VSMCs). (A) Viability of VSMC treated with lysoPC, (B) cell necrosis induced by lysoPC, (C) lysoPC induced DNA fragmentation.
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Fig. 2
Mechanisms for vascular smooth muscle cell (VSMC) apoptosis induced by lysophosphatidylcholine (lysoPC). (A) Involvement of reactive oxygen species and nuclear factor-kappaB (NF-κB), as assessed by propidium iodide (PI) staining, (B) kinetics of reactive oxygen species (ROS) production determined by 2',7'-dichlorofluorescin (DCF) staining. DSF: defined serum-free medium, CAPE: caffeic acid phenylethyl ester. *P < 0.001 vs. control.
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Fig. 3
Mechanisms for vascular smooth muscle cell (VSMC) apoptosis induced by lysophosphatidylcholine (lysoPC), evaluated by western blot analyses. (A) expression of caspases, (B) expression of bcl-2, (C) expression of bax.
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Fig. 4
Pathway of apoptosis.
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