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7-Ketocholesterol Induces Vascular Smooth Muscle Cell Apoptosis via Akt Degradation

7-Ketocholesterol에 의한 Akt 감소와 혈관평활근세포의 세포자멸사

Seo, Kyo Won;Kim, Chi Dae;Lee, Won Suk
서교원;김치대;이원석

  • Received : 2016.01.05
  • Accepted : 2016.02.04
  • Published : 2016.02.25

Abstract

Vascular smooth muscle cell (VSMC) apoptosis has been identified in various vascular diseases, including atherosclerosis and restenosis after angioplasty, and has been known to precipitate atherosclerotic plaque instability and rupture. Oxysterols are known as inducers of apoptosis in VSMC, and 7-ketocholesterol (7KC) is the major nonenzymically formed oxysterol in atherosclerotic lesions. The precise mechanism underlying VSMC apoptosis is still poorly understood. In this study, we investigated whether 7KC causes apoptosis, and characterized its apoptotic mechanisms in primary cultured rat aortic VSMC. Cell viability was assessed by MTT assay and trypan blue assay. Apoptosis was assessed by flow cytometry, immunofluorescence, immunoprecipitation, and Western blot analyses. 7KC markedly decreased the VSMC viability in a time- and concentration-dependent manner, and increased the production of 4-hydroxynonenal (HNE), a major end-product of lipid peroxidation, which also decreased the VSMC viability. Pretreatment with 2,4-dinitrophenylhydrazine, a well-known reagent of lipid peroxidation-derived aldehydes, significantly restored the 7KC-decreased viability of VSMC. Furthermore, HNE, as well as 7KC, reduced the level of total Akt, a major mediator of cell survival. The 7KC-decreased level of total Akt was significantly restored by pretreatments with 2,4-dinitrophenylhydrazine and N-acetylcysteine. Lactacystin, a proteasome inhibitor, protected VSMC against apoptosis and Akt degradation, but did not inhibit HNE production. In the immunoprecipitation assay, 7KC increased HNE-modified Akt. From the results, it seems that, in atherosclerotic lesions, 7KC induces HNE production in VSMC, and this HNE binds to Akt, proceeding to proteasomal degradation of Akt, through which mechanism the atherosclerotic plaque instability may be facilitated.

Keywords

4-Hydroxynonenal;7-ketocholesterol;Akt;apoptosis;vascular smooth muscle cell

References

  1. Allard, D., Figg, N., Bennett, M. R. and Littlewood, T. D. 2008. Akt regulates the survival of vascular smooth muscle cells via inhibition of FoxO3a and GSK3. J. Biol. Chem. 283, 19739-19747. https://doi.org/10.1074/jbc.M710098200
  2. Basso, A. D., Solit, D. B., Chiosis, G., Giri, B., Tsichlis, P. and Rosen, N. 2002. Akt forms an intracellular complex with heat shock protein 90 (Hsp90) and Cdc37 and is destabilized by inhibitors of Hsp90 function. J. Biol. Chem. 277, 39858-39866. https://doi.org/10.1074/jbc.M206322200
  3. Bonomini, F., Tengattini, S., Fabiano, A., Bianchi, R. and Rezzani, R. 2008. Atherosclerosis and oxidative stress. Histol. Histopathol. 23, 381-390.
  4. Brown, A. J. and Jessup, W. 1999. Oxysterols and atherosclerosis. Atherosclerosis 142, 1-28. https://doi.org/10.1016/S0021-9150(98)00196-8
  5. Chapple, S. J., Cheng, X. and Mann, G. E. 2013. Effects of 4-hydroxynonenal on vascular endothelial and smooth muscle cell redox signaling and function in health and disease. Redox Biol. 23, 319-331.
  6. Clarke, M. C., Littlewood, T. D., Figg, N., Maguire, J. J., Davenport, A. P., Goddard, M. and Bennett, M. R. 2008. Chronic apoptosis of vascular smooth muscle cells accelerates atherosclerosis and promotes calcification and medial degeneration. Circ. Res. 20, 1529-1538.
  7. Dickson, B. C. and Gotlieb, A. I. 2003. Towards understanding acute destabilization of vulnerable atherosclerotic plaques. Cardiovasc. Pathol. 12, 237-248. https://doi.org/10.1016/S1054-8807(03)00072-3
  8. Doong, H., Rizzo, K., Fang, S., Kulpa, V., Weissman, A. M. and Kohn, E. C. 2003. CAIR-1/BAG-3 abrogates heat shock protein-70 chaperone complex-mediated protein degradation: accumulation of poly-ubiquitinated Hsp90 client proteins. J. Biol. Chem. 278, 28490-28500. https://doi.org/10.1074/jbc.M209682200
  9. Fan, Y., Xie, P., Zhang, H., Guo, S., Gu, D., She, M. and Li, H. 2008. Proteasome-dependent inactivation of Akt is essential for 12-O-tetradecanoylphorbol 13-acetate-induced apoptosis in vascular smooth muscle cells. Apoptosis 13, 1401-1409. https://doi.org/10.1007/s10495-008-0272-z
  10. Guardiola, F., Codony, R., Addis, P. B., Rafecas, M. and Boatella, J. 1996. Biological effects of oxysterols: current status. Food Chem. Toxicol. 34, 193-211. https://doi.org/10.1016/0278-6915(95)00094-1
  11. Korshunov, V. A. and Berk, B. C. 2008. Smooth muscle apoptosis and vascular remodeling. Curr. Opin. Hematol. 15, 250-254. https://doi.org/10.1097/MOH.0b013e3282f97d71
  12. Lawlor, M. A. and Alessi, D. R. 2001. PKB/Akt: a key mediator of cell proliferation, survival and insulin responses? J. Cell Sci. 114, 2903-2910.
  13. Littlewood, T. D. and Bennett, M. R. 2003. Apoptotic cell death in atherosclerosis. Curr. Opin. Lipidol. 14, 469-475. https://doi.org/10.1097/00041433-200310000-00007
  14. Madamanchi, N. R., Hakim, Z. S. and Runge, M. S. 2005. Oxidative stress in atherogenesis and arterial thrombosis: the disconnect between cellular studies and clinical outcomes. J. Thromb. Haemost. 3, 254-267. https://doi.org/10.1111/j.1538-7836.2004.01085.x
  15. Martinet, W. and Kockx, M. M. 2001. Apoptosis in atherosclerosis: focus on oxidized lipids and inflammation. Curr. Opin. Lipidol. 12, 535-541. https://doi.org/10.1097/00041433-200110000-00009
  16. Martinet, W., De Bie, M., Schrijvers, D. M., De Meyer, G. R., Herman, A. G. and Kockx, M. M. 2004. 7-Ketocholesterol induces protein ubiquitination, myelin figure formation, and light chain 3 processing in vascular smooth muscle cells. Arterioscler. Thromb. Vasc. Biol. 24, 2296-2301. https://doi.org/10.1161/01.ATV.0000146266.65820.a1
  17. Martínez-Hervás, S., Vinué, Á., Núñez, L., Andrés-Blasco, I., Piqueras, L., Real, J. T., Ascaso, J. F., Burks, D. J., Sanz, M. J. and González-Navarro, H. 2014. Insulin resistance aggravates atherosclerosis by reducing vascular smooth muscle cell survival and increasing CX3CL1/CX3CR1 axis. Cardiovasc. Res. 103, 324-336. https://doi.org/10.1093/cvr/cvu115
  18. Ozaki, H., Miyashita, Y., Watanabe, H. and Shirai, K. 2005. Enhancement of MMP-9 activity in THP-1 cells by 7-ketocholesterol and its suppression by the HMG-CoA reductase inhibitor fluvastatin. J. Atheroscler. Thromb. 12, 308-314. https://doi.org/10.5551/jat.12.308
  19. Clarke, M. and Bennett, M. 2006. Defining the role of vascular smooth muscle cell apoptosis in atherosclerosis. Cell Cycle 5, 2329-2331. https://doi.org/10.4161/cc.5.20.3383
  20. Clarke, M. C, Littlewood, T. D., Figg, N., Maguire, J. J., Davenport, A. P., Goddard, M. and Bennett, M. R. 2008. Chronic apoptosis of vascular smooth muscle cells accelerates atherosclerosis and promotes calcification and medial degeneration. Circ. Res. 102, 1529-1538. https://doi.org/10.1161/CIRCRESAHA.108.175976
  21. Perales, S., Alejandre, M. J., Palomino-Morales, R., Torres, C., Iglesias, J. and Linares, A. 2009. Effect of oxysterol-induced apoptosis of vascular smooth muscle cells on experimental hypercholesterolemia. J. Biomed. Biotechnol. 2009, 456208.
  22. Rusiñol, A. E., Thewke, D., Liu, J., Freeman, N., Panini, S. R. and Sinensky, M. S. 2004. AKT/protein kinase B regulation of BCL family members during oxysterol-induced apoptosis. J. Biol. Chem. 279, 1392-1399. https://doi.org/10.1074/jbc.M308619200
  23. Sampey, B. P., Carbone, D. L., Doorn, J. A., Drechsel, D. A. and Petersen, D. R. 2007. 4-Hydroxy-2-nonenal adduction of extracellular signal-regulated kinase (Erk) and the inhibition of hepatocyte Erk-Est-like protein-1-activating protein-1 signal transduction. Mol. Pharmacol. 71, 871-883.
  24. Schroepfer, G. J. Jr. 2005. Oxysterols: modulators of cholesterol metabolism and other processes. Physiol. Rev. 80, 361-554.
  25. Spiliopoulos, S., Diamantopoulos, A., Katsanos, K., Ravazoula, P., Karnabatidis, D. and Siablis, D. 2014. PolarCath cryoplasty enhances smooth muscle cell apoptosis in a rabbit iliac artery model: an experimental in vivo controlled study. Cryobiology 63, 267-272.
  26. Suizu, F., Hiramuki, Y., Okumura, F., Matsuda, M., Okumura, A. J., Hirata, N., Narita, M., Kohno, T., Yokota, J., Bohgaki, M., Obuse, C., Hatakeyama, S., Obata, T. and Noguchi, M. 2009. The E3 ligase TTC3 facilitates ubiquitination and degradation of phosphorylated Akt. Dev. Cell 17, 800-810. https://doi.org/10.1016/j.devcel.2009.09.007
  27. Lee, J. Y., Jung, G. Y., Heo, H. J., Yun, M. R., Park, J. Y., Bae, S. S., Hong, K. W., Lee, W. S. and Kim, C. D. 2006. 4-Hydroxynonenal induces vascular smooth muscle cell apoptosis through mitochondrial generation of reactive oxygen species. Toxicol. Lett. 166, 212-221. https://doi.org/10.1016/j.toxlet.2006.07.305
  28. Leonarduzzi, G., Chiarpotto, E., Biasi, F. and Poli, G. 2005. 4-Hydroxynonenal and cholesterol oxidation products in atherosclerosis. Mol. Nutr. Food Res. 49, 1044-1049. https://doi.org/10.1002/mnfr.200500090
  29. Tucka, J., Yu, H., Gray, K., Figg, N., Maguire, J., Lam, B., Bennett, M. and Littlewood, T. 2014. Akt1 regulates vascular smooth muscle cell apoptosis through FoxO3a and Apaf1 and protects against arterial remodeling and atherosclerosis. Arterioscler. Thromb. Vasc. Biol. 34, 2421-2428. https://doi.org/10.1161/ATVBAHA.114.304284
  30. Zhao, Y., Miriyala, S., Miao, L., Mitov, M., Schnell, D., Dhar, S. K., Cai, J., Klein, J. B., Sultana, R., Butterfield, D. A., Vore, M., Batinic-Haberle, I., Bondada, S. and St. Clair, D. K. 2014. Redox proteomic identification of HNE-bound mitochondrial proteins in cardiac tissues reveals a systemic effect on energy metabolism after doxorubicin treatment. Free Radic. Biol. Med. 72, 55-65. https://doi.org/10.1016/j.freeradbiomed.2014.03.001
  31. Perales, S., Alejandre, M. J., Palomino-Morales, R., Torres, C. and Linares, A. 2010. Influence of cholesterol and fish oil dietary intake on nitric oxide-induced apoptosis in vascular smooth muscle cells. Nitric Oxide 22, 205-212. https://doi.org/10.1016/j.niox.2009.12.003
  32. Toné, S., Sugimoto, K., Tanda, K., Suda, T., Uehira, K., Kanouchi, H., Samejima, K., Minatogawa, Y. and Earnshaw, W. C. 2007. Three distinct stages of apoptotic nuclear condensation revealed by time-lapse imaging, biochemical and electron microscopy analysis of cell-free apoptosis. Exp. Cell Res. 313, 3635-3644. https://doi.org/10.1016/j.yexcr.2007.06.018