刘雅蓉;邵倩;张慧慧;贾颖;戴敏;
LIU Ya-rong;SHAO Qian;ZHANG Hui-hui;JIA Ying;DAI Min;Department of Pharmacy, Anhui University of Traditional Chinese Medicine;Key Laboratory of Xin′an Medicine;

• 1:安徽中医药大学药学院
• 2:新安医学教育部重点实验室

摘要(Abstract)：

为探讨丹皮酚是否能通过调控主动脉小凹蛋白-1的表达,影响NF-κB通路,从而抑制动脉粥样硬化模型大鼠血管内皮炎症反应来发挥抗动脉粥样硬化的作用。采用高脂饮食+维生素D_2诱导大鼠动脉粥样硬化模型;采用组织块预消化贴壁法原代培养血管内皮细胞(VECs),并建立脂多糖(LPS)刺激的VECs损伤模型,加入小凹蛋白抑制剂filipin进行对照。HE染色法观察主动脉病理形态学变化,ELISA法分别检测血清中及VECs分泌的肿瘤坏死因子α(TNF-α)、白细胞介素-6(IL-6)、血管细胞黏附分子-1(VCAM-1)水平变化,Western blot法分别检测主动脉及VECs中小凹蛋白-1、p65蛋白表达水平变化。整体动物结果显示,丹皮酚显著减轻动脉粥样硬化模型大鼠主动脉斑块面积及病变程度,减少内膜增生和血管壁脂质沉积;明显降低血清中的TNF-α,IL-6,VCAM-1水平;升高主动脉中小凹蛋白-1相对表达量并降低p65蛋白水平(P<0.05或P<0.01)。细胞实验结果显示,丹皮酚可以明显改善损伤的VECs细胞形态,显著降低损伤的VECs分泌的TNF-α,IL-6,VCAM-1水平,明显增加小凹蛋白-1的表达并减少p65蛋白表达(P<0.05或P<0.01);加入小凹蛋白-1抑制剂filipin后,可以显著逆转丹皮酚对炎症因子及蛋白表达的作用(P<0.05或P<0.01)。该研究结果表明,丹皮酚可以通过上调主动脉血管小凹蛋白-1的表达,抑制NF-κB通路的激活,从而减轻动脉粥样硬化模型大鼠血管内皮炎症反应,最终发挥抗动脉粥样硬化的作用。
To explore whether paeonol can play an anti-atherosclerotic role by regulating the expression of aortic caveolin-1 and affecting NF-κB pathway, so as to inhibit the inflammatory response of vascular endothelium in atherosclerotic rats. The atherosclerotic model of rats was induced by high-fat diet and vitamin D_2. The primary culture of vascular endothelial cells(VECs) was carried out by tissue block pre-digestion and adherent method. The injury model of VECs was induced by lipopolysaccharide(LPS), and filipin, a small concave protein inhibitor, was added for control. HE staining was used to observe pathological changes of aorta. TNF-α, IL-6 and VCAM-1 were detected by ELISA. Western blot assay was used to detect the protein expression levels of caveolin-1 and p65 in aorta and VECs. The results showed that as compared with model group, paeonol significantly reduced aortic plaque area and lesion degree in rats, decreased the level of serum TNF-α, IL-6 and VCAM-1 in the rats and enhanced the relative expression level of caveolin-1, decreased p65 expression conversely(P<0.05 or P<0.01). In vitro, as compared to model group, paeonol obviously improved cell morphology, decreased the secretion of TNF-α, IL-6 and VCAM-1 in VECs, increased caveolin-1 expression, and decreased p65 protein expression(P<0.05 or P<0.01). Furthermore, filipin could reverse the effect of paeonol on expression of inflammatory factors and proteins(P<0.05 or P<0.01). According to the results, it was found that paeonol could play the role of anti-atherosclerosis by up-regulating the expression of caveolin-1 and inhibiting the activation of NF-κB pathway to reduce vascular inflammation in atherosclerotic rats.

关键词(KeyWords)： 小凹蛋白-1;丹皮酚;动脉粥样硬化;主动脉;血管内皮;炎症反应
caveolin-1;paeonol;atherosclerosis;aorta;vascular endothelial cell;inflammation

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(81773937);; 安徽中医药大学自然科学研究项目(2018zrd06)

作者(Authors): 刘雅蓉;邵倩;张慧慧;贾颖;戴敏;
LIU Ya-rong;SHAO Qian;ZHANG Hui-hui;JIA Ying;DAI Min;Department of Pharmacy, Anhui University of Traditional Chinese Medicine;Key Laboratory of Xin′an Medicine;

DOI: 10.19540/j.cnki.cjcmm.20200210.408

参考文献(References)：

• [1] YE Y,NYLANDER S,BIRNBAUM Y.Unraveling the interaction of aspirin,ticagrelor,and rosuvastatin on the progression of atherosclerosis and inflammation in diabetic mice[J].Cardiovasc Drug Ther,2017,31(5/6):489.
• [2] 王迪,王毅.动脉粥样硬化动物模型及其进展[J].心脏杂志,2018,30(4):490.
• [3] 褚现明,李冰,安毅,等.炎症与动脉粥样硬化关系研究进展[J].中国分子心脏病学杂志,2010,11(3):184.
• [4] 廖端芳,覃丽.小凹及小凹蛋白1:胆固醇逆向转运与炎症应答的共同分子平台[J].中国动脉硬化杂志,2012,20(5):385.
• [5] RODRIGUEZ-FEO J A,HELLINGS W E,MOLL F L,et al.Caveolin-1 influences vascular protease activity and is a potential stabilizing factor in human atherosclerotic disease[J].PLoS ONE,2008,3(7):e2612.
• [6] 杨翠,刘先哲.小凹蛋白-1在动脉粥样硬化形成中的研究进展[J].海南医学,2013,24(3):437.
• [7] 李后开,戴敏,汪电雷,等.兔动脉粥样硬化模型的建立及丹皮酚药物作用的实验研究[J].中国中医药科技,2005,12(3):129.
• [8] LIU Y R,LI C,WU H F,et al.Paeonol attenuated inflammatory response of endothelial cells via stimulating monocytes-derived exosomal microRNA-223[J].Front Pharmacol,2018,9:1105.
• [9] 章维志,李国莺,祁芹,等.地奥心血康对动脉粥样硬化大鼠TLR4/MyD88/NF-κB信号通路的调节作用[J].中国中药杂志,2019,45(3):602.
• [10] 吴晶魁,杨乔,李洋洋,等.水蛭通过p38MAPK信号通路对早期动脉粥样硬化大鼠VSMCs的影响[J].中国中药杂志,2017,42(16):3191.
• [11] FERNANDEZ-HERNANDO C,YU J,DAVALOS A,et al.Endothelial-specific overexpression of caveolin-1 accelerates atherosclerosis in apolipo-protein E-deficient mice[J].Am J Pathol,2011,177(2):998.
• [12] 李志乐,徐戈,欧婷,等.烟酸对高脂血症兔脂肪组织Caveolin-1 mRNA表达的影响[J].实用医学杂志,2012,26(6):893.
• [13] 姚贺之.加减活络效灵丹对糖尿病大鼠动脉硬化形成早期的干预作用[D].北京:北京中医药大学,2016.
• [14] 罗小林,钟华,梁霄,等.小凹蛋白1上调人脐静脉内皮细胞钙敏感受体介导一氧化氮合酶激活的机制[J].中国动脉硬化杂志,2013,21(6):486.
• [15] 岳雯,姚树桐,周晓,等.小凹蛋白-1通过激活p38 MAPK抑制巨噬细胞内质网应激凋亡[J].生理学报,2012,64(2):149.
• [16] JOHN H,CHIDLOW J,WILLIAM C S.Caveolae,caveolins,and cavins:complex control of cellular signalling and inflammation[J].Cardiovasc Res,2010,86(2):219.
• [17] HU G,YE R D,DINAUER M C,et al.Neutrophil caveolin-1 expression contributes to mechanism of lung inflammation and injury [J].Am J Physiol Lung Cell Mol Physiol,2008,294:178.
• [18] WANG X M,KIM H P,NAKAHIRA K,et al.The heme oxygenase-1/carbonmonoxide pathway suppresses TLR4 signaling by regulating the interaction of TLR4 with caveolin-1[J].J Immunol,2009,182(6):809.
• [19] GARREAN S,GAO X P,BROVKOVYCH V,et al.Caveolin-1 regulates NF-kappa B activation and lung inflammatory response to sepsis induced by lipopolysaccharide [J].J Immunol,2006,177(7):853.
• [20] 谢先梅,李超,孙颖,等.丹皮酚通过抑制p38 MAPK/N-SMase2通路减少脂多糖诱导的THP-1细胞外泌体分泌[J].中国动脉硬化杂志,2019,27(1):11.
• [21] SONG A W,WU H F,DAI M.Paeonol attenuates progression of atherosclerotic lesion formation through lipid regulation,anti-inflammatory and antioxidant activities[J].J Pharm Sci-US,2018,27(8):565.
• [22] 刘雅蓉,吴鸿飞,戴敏.丹皮酚抑制脂多糖诱导的血管内皮细胞TNF-α释放对共培养体系中血管平滑肌细胞凋亡及p38MAPK信号通路的影响[J].安徽中医药大学学报,2018,37(4):65.
• [23] 胡文君,张振,戴敏.丹皮酚通过抑制PI3K/AKT-NF-κB通路对LPS诱导的与平滑肌细胞共培养的大鼠血管内皮细胞的保护作用[J].中国中药杂志,2016,41(12):2298.