Abstract
Oxysterols, oxygenated derivatives of cholesterol, are found abundantly in the plasma and atherosclerotic plaques, a common risk factor for thoracic aortic aneurysms (TAAs). Among the oxysterols, namely 7-ketocholesterol (7-KC) and 25-hydroxycholesterol (25-OHC), lead both to induction of reactive oxygen species (ROS) in cells and to apoptosis in smooth muscle cells (SMCs) probably due to increased oxidative stress. Since loss of SMCs through apoptosis is a major event in TAA formation, it is important to understand the molecular pathways of apoptosis in response to ROS in TAAs. Very little is known about the effect of oxysterols on TAA SMCs. Therefore, we investigated molecular pathways participating in the oxysterol induced cell death of TAAs. Our results showed that TAA SMCs died mainly as a result of apoptosis as suggested by cellular shrinkage, blebbing, DNA condensation/fragmentation in response to oxysterol treatment. There was no significant difference in oxysterol induced cell death between TAA and control SMCs. Addition of antioxidant molecules prevented cell death, hence ROS appears to be involved in the apoptosis of these cells. While oxysterol treatment increased caspase 3 activity, cell death was not rescued in its absence. Efficient silencing of other targets including apoptotic proteins (p53, Bax), and survival proteins (Akt1, Akt2) showed that apoptosis can occur through p53, and Bax independent pathways. Silencing Akt1 or Akt2 did not lead to further cell death. These results indicate that oxysterols can induce several cell death pathways in TAA SMCs.
Similar content being viewed by others
References
Lesauskaite V et al (2001) Smooth muscle cells of the media in the dilatative pathology of ascending thoracic aorta: morphology, immunoreactivity for osteopontin, matrix metalloproteinases, and their inhibitors. Hum Pathol 32(9):1003–1011
Isselbacher EM (2005) Thoracic and abdominal aortic aneurysms. Circulation 111(6):816–828
Blunder S et al (2012) Characteristics of TAV- and BAV-associated thoracic aortic aneurysms–smooth muscle cell biology, expression profiling, and histological analyses. Atherosclerosis 220(2):355–361
Booher AM, Eagle KA (2011) Diagnosis and management issues in thoracic aortic aneurysm. Am Heart J 162(1):38–46e1
El-Hamamsy I, Yacoub MH (2009) Cellular and molecular mechanisms of thoracic aortic aneurysms. Nat Rev Cardiol 6(12):771–786
Ejiri J et al (2003) Oxidative stress in the pathogenesis of thoracic aortic aneurysm: protective role of statin and angiotensin II type 1 receptor blocker. Cardiovasc Res 59(4):988–996
Thomas M et al (2006) Deletion of p47phox attenuates angiotensin II-induced abdominal aortic aneurysm formation in apolipoprotein E-deficient mice. Circulation 114(5):404–413
Xiong W et al (2009) Inhibition of reactive oxygen species attenuates aneurysm formation in a murine model. Atherosclerosis 202(1):128–134
Madamanchi NR, Runge MS (2007) Mitochondrial dysfunction in atherosclerosis. Circ Res 100(4):460–473
Irani K (2000) Oxidant signaling in vascular cell growth, death, and survival : a review of the roles of reactive oxygen species in smooth muscle and endothelial cell mitogenic and apoptotic signaling. Circ Res 87(3):179–183
Li PF, Dietz R, von Harsdorf R (1997) Reactive oxygen species induce apoptosis of vascular smooth muscle cell. FEBS Lett 404(2–3):249–252
Rajagopalan S et al (1996) Reactive oxygen species produced by macrophage-derived foam cells regulate the activity of vascular matrix metalloproteinases in vitro. Implications for atherosclerotic plaque stability. J Clin Investig 98(11):2572–2579
Acilan C et al (2012) Smooth muscle cells isolated from thoracic aortic aneurysms exhibit increased genomic damage, but similar tendency for apoptosis. DNA Cell Biol 31(10):1523–1534
Baykal AT et al (2013) Proteomic evidence for the plasticity of cultured vascular smooth muscle cells. Turk J Biol 37(4):414–425
Absi TS et al (2003) Altered patterns of gene expression distinguishing ascending aortic aneurysms from abdominal aortic aneurysms: complementary DNA expression profiling in the molecular characterization of aortic disease. J Thorac Cardiovasc Surg 126(2):344–357; discussion 357
Vejux A, Malvitte L, Lizard G (2008) Side effects of oxysterols: cytotoxicity, oxidation, inflammation, and phospholipidosis. Braz J Med Biol Res 41(7):545–556
Appukuttan A et al (2013) Oxysterol-induced apoptosis of smooth muscle cells is under the control of a soluble adenylyl cyclase. Cardiovasc Res 99(4):734–742
Miyashita Y et al (2002) Oxysterol-induced apoptosis of vascular smooth muscle cells is reduced by HMG-CoA reductase inhibitor, pravastatin. J Atheroscler Thromb 9(1):65–71
Lordan S, Mackrill JJ, O’Brien NM (2009) Oxysterols and mechanisms of apoptotic signaling: implications in the pathology of degenerative diseases. J Nutr Biochem 20(5):321–336
Nishio E, Watanabe Y (1996) Oxysterols induced apoptosis in cultured smooth muscle cells through CPP32 protease activation and bcl-2 protein downregulation. Biochem Biophys Res Commun 226(3):928–934
Iuliano L et al (2003) Measurement of oxysterols and alpha-tocopherol in plasma and tissue samples as indices of oxidant stress status. Anal Biochem 312(2):217–223
Tarık Kızıltan H, Baltalı M, Kayaselçuk F, Korkmaz ME, Müderrisoğlu H, Taşdelen A, Aşlamacı S (2004) Çıkan Aort Dilatasyonuna Eşlik Eden Histopatolojik Değişiklikler. Türk Göğüs Kalp Damar Cerrahisi Dergisi 4(4):206–210
Alkazemi D et al (2008) Oxysterol as a marker of atherogenic dyslipidemia in adolescence. J Clin Endocrinol Metab 93(11):4282–4289
Acilan C et al (2014) A preliminary proteomic evaluation of smooth muscle cells in thoracic aortic aneurysms. Turk J Biol 38:1–15
Spyridopoulos I et al (2001) Alcohol enhances oxysterol-induced apoptosis in human endothelial cells by a calcium-dependent mechanism. Arterioscler Thromb Vasc Biol 21(3):439–444
Bass DA et al (1983) Flow cytometric studies of oxidative product formation by neutrophils: a graded response to membrane stimulation. J Immunol 130(4):1910–1917
Kacar O et al (2014) Evaluation of the molecular mechanisms of a palladium(II) saccharinate complex with terpyridine as an anticancer agent. Anticancer Drugs 25(1):17–29
Jang ER, Lee CS (2010) 7-ketocholesterol induces apoptosis in differentiated PC12 cells via reactive oxygen species-dependent activation of NF-kappaB and Akt pathways. Neurochem Int 58(1):52–59
Pedruzzi E et al (2004) NAD(P)H oxidase Nox-4 mediates 7-ketocholesterol-induced endoplasmic reticulum stress and apoptosis in human aortic smooth muscle cells. Mol Cell Biol 24(24):10703–10717
Travert C, Carreau S, Le Goff D (2006) Induction of apoptosis by 25-hydroxycholesterol in adult rat Leydig cells: protective effect of 17beta-estradiol. Reprod Toxicol 22(4):564–570
Larsson DA et al (2006) Oxysterol mixtures, in atheroma-relevant proportions, display synergistic and proapoptotic effects. Free Radic Biol Med 41(6):902–910
Yang L, Sinensky MS (2000) 25-Hydroxycholesterol activates a cytochrome c release-mediated caspase cascade. Biochem Biophys Res Commun 278(3):557–563
Choi YK et al (2008) 25-hydroxycholesterol induces mitochondria-dependent apoptosis via activation of glycogen synthase kinase-3beta in PC12 cells. Free Radic Res 42(6):544–553
Prunet C et al (2005) Activation of caspase-3-dependent and -independent pathways during 7-ketocholesterol- and 7beta-hydroxycholesterol-induced cell death: a morphological and biochemical study. J Biochem Mol Toxicol 19(5):311–326
Rusinol AE et al (2004) AKT/protein kinase B regulation of BCL family members during oxysterol-induced apoptosis. J Biol Chem 279(2):1392–1399
Li W et al (2012) Cell death induced by 7-oxysterols via lysosomal and mitochondrial pathways is p53-dependent. Free Radic Biol Med 53(11):2054–2061
de Weille J, Fabre C, Bakalara N (2013) Oxysterols in cancer cell proliferation and death. Biochem Pharmacol 86(1):154–160
Lizard G et al (1999) Characterization and comparison of the mode of cell death, apoptosis versus necrosis, induced by 7 beta-hydroxycholesterol and 7-ketocholesterol in the cells of the vascular wall. Arterioscler Thromb Vasc Biol 19(5):1190–1200
Bennett MR, Evan GI, Schwartz SM (1995) Apoptosis of human vascular smooth muscle cells derived from normal vessels and coronary atherosclerotic plaques. J Clin Investig 95(5):2266–2274
Nury T et al (2013) Induction of oxiapoptophagy, a mixed mode of cell death associated with oxidative stress, apoptosis and autophagy, on 7-ketocholesterol-treated 158N murine oligodendrocytes: impairment by alpha-tocopherol. Biochem Biophys Res Commun 446(3):714–719
He R et al (2006) Characterization of the inflammatory and apoptotic cells in the aortas of patients with ascending thoracic aortic aneurysms and dissections. J Thorac Cardiovasc Surg 131(3):671–678
Larrayoz IM et al (2010) 7-ketocholesterol-induced inflammation: involvement of multiple kinase signaling pathways via NFkappaB but independently of reactive oxygen species formation. Investig Ophthalmol Vis Sci 51(10):4942–4955
Ong JM et al (2003) Oxysterol-induced toxicity in R28 and ARPE-19 cells. Neurochem Res 28(6):883–891
Lee TS, Chau LY (2001) Fas/Fas ligand-mediated death pathway is involved in oxLDL-induced apoptosis in vascular smooth muscle cells. Am J Physiol-Cell Physiol 280(3):C709–C718
Agrawal S et al (2002) Stat1-dependent, p53-independent expression of p21(waf1) modulates oxysterol-induced apoptosis. Mol Cell Biol 22(7):1981–1992
Sung YH et al (2007) Influence of co-down-regulation of caspase-3 and caspase-7 by siRNAs on sodium butyrate-induced apoptotic cell death of Chinese hamster ovary cells producing thrombopoietin. Metab Eng 9(5–6):452–464
Acknowledgments
This work was supported by the FP7 Project entitled “Fighting Aneurysmal Disease” (FAD), Grant No: 200647 and Istanbul University Department of Scientific Research Projects, entitled “Investigation of Gene Expressions Related to Oxidative Stress in Human Thoracic Aort Aneurysms”, Grant No: 10241.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Adiguzel, Z., Arda, N., Kacar, O. et al. Evaluation of apoptotic molecular pathways for smooth muscle cells isolated from thoracic aortic aneurysms in response to oxidized sterols. Mol Biol Rep 41, 7875–7884 (2014). https://doi.org/10.1007/s11033-014-3681-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11033-014-3681-9