Interactions between cell death induced by statins and 7-ketocholesterol in rabbit aorta smooth muscle cells
Interactions between cell death induced by statins and 7-ketocholesterol in rabbit aorta smooth muscle cells
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DOI：
10.1038/bjp.2008.181
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发表时间：
2008-07-01
影响因子：
7.3
通讯作者：
Bult, H.
中科院分区：
文献类型：
--
作者：
Martinet, W.;Schrijvers, D. M.;Bult, H.
背景和目的：7-酮胆固醇是一种存在于动脉粥样硬化病变中的氧固醇，可诱导平滑肌细胞（SMC）死亡，从而破坏斑块的稳定。他汀类药物可诱导SMC细胞凋亡，但可保护患者免于心肌梗死。我们研究了他汀类药物和7-酮胆固醇是否具有累加性细胞死亡效应。实验方法：将培养的兔主动脉SMCs（传代2-6）暴露于含氟伐他汀、辛伐他汀或普伐他汀或不含氟伐他汀的7-酮胆固醇中。检测中性红（NR）的摄取、单层蛋白、泛半胱天冬酶底物Asp-Glu-Val-Asp-rhodamine110的裂解、细胞形态（光镜和电镜）和微管相关蛋白1轻链3 （LC3，免疫印迹）的加工。关键结果：在暴露于25 μ M 7-酮胆固醇（-41 +/- 3%,n = 13）、100 μ M氟伐他汀（-59%）或30-100 μ M辛伐他汀（-28至-74%）18小时后，NR摄取下降。氧甾醇和高他汀类药物浓度具有叠加效应，但较低浓度（氟伐他汀10-30 μ M，辛伐他汀1-10 μ M）部分逆转了活力丧失。7-酮胆固醇引起细胞质强烈的空泡化，LC3-I加工成LC3-II，但caspase很少激活（增加29.5%）。氟伐他汀（10-100亩，增加70-545%）和辛伐他汀（3-100亩，增加43-322%）诱导无LC3加工的caspase激活，但在7-酮胆固醇处理的SMCs中未能激活caspase。普伐他汀≤100mm均无活性。结论和意义：7-酮胆固醇引起SMC死亡，主要通过LC3加工的自噬泡形成，而亲脂性他汀类药物引起SMC凋亡。7-酮胆固醇和低他汀类药物浓度后的细胞死亡并不是相加的，可能是因为自噬过程干扰了他汀类药物诱导的半胱天冬酶激活。这进一步说明，药物对正常SMCs的影响并不一定能预测动脉粥样硬化情况下的活动。
Background and purpose: 7-Ketocholesterol, an oxysterol present in atherosclerotic lesions, induces smooth muscle cell (SMC) death, thereby destabilizing plaques. Statins protect patients from myocardial infarction, though they induce SMC apoptosis. We investigated whether statins and 7-ketocholesterol exerted additive cell death effects.Experimental approach: Cultured rabbit aorta SMCs (passage 2-6) were exposed to 7-ketocholesterol with or without fluvastatin, simvastatin or pravastatin. Uptake of neutral red (NR), monolayer protein, cleavage of the pan-caspase substrate Asp-Glu-Val-Asp-rhodamine110, cell morphology (light and electron microscopy) and processing of microtubule-associated protein 1 light chain 3 (LC3, immunoblot) were determined.Key results: NR uptake declined upon 18 h exposure to 25 mu M 7-ketocholesterol (-41 +/- 3%, n = 13), 100 mM fluvastatin (-59%) or 30-100 mu M simvastatin (-28 to -74%). Oxysterol and high statin concentrations exerted additive effects, but lower concentrations (fluvastatin 10-30 mu M, simvastatin 1-10 mu M) partly reversed viability loss. 7-Ketocholesterol caused intense cytoplasmic vacuolization, processing of LC3-I to LC3-II, but little caspase activation (increase 29.5%). Fluvastatin (10-100 mu M, 70-545% increase) and simvastatin (3-100 mu M 43-322% increase) induced caspase activation without LC3 processing, but failed to activate caspases in 7-ketocholesterol-treated SMCs. Pravastatin up to 100 mM was always inactive.Conclusions and implications: 7-Ketocholesterol caused SMC death, mainly via autophagic vesicle formation with LC3 processing, whereas lipophilic statins evoked SMC apoptosis. Cell death following 7-ketocholesterol and low statin concentrations were not additive, presumably because the autophagic process interfered with statin-induced caspase activation. This further illustrates that drug effects in normal SMCs are not necessarily predictive for activities in atherosclerotic settings.