瑞舒伐他汀减轻脂多糖诱导的人晚期内皮祖细胞损伤
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摘要
目的观察瑞舒伐他汀对脂多糖(LPS)诱导的人外周血晚期内皮祖细胞(LOCs)损伤的影响,探讨其可能的机制。方法密度梯度离心法获取人外周血单个核细胞并培养获得LOCs。实验分为对照组、LPS诱导组以及不同浓度瑞舒伐他汀和LPS共处理组。用CCK-8法检测细胞活力;比色法检测细胞培养上清液中丙二醛(MDA)的含量及乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)的活性;流式细胞术检测细胞内活性氧(ROS)水平;ELISA试剂盒检测细胞培养液中IL-6、IL-8水平;Western blot检测Toll样受体4(TLR4)蛋白表达;转录因子活性检测试剂盒检测NFκB p65转录活性。结果瑞舒伐他汀能有效抑制LPS作用下LOCs活力损失(P<0.05),降低细胞培养液中MDA含量及LDH活性(P<0.05),增加SOD活性(P<0.05),抑制LPS引起的ROS水平增高(P<0.01),降低炎性因子IL-6、IL-8的表达水平(P<0.01),并显著抑制LPS诱导的TLR4蛋白表达(P<0.05)和NFκB p65的转录活性(P<0.01)。结论瑞舒伐他汀能提升LPS作用下LOCs的细胞活力,减少LPS诱导的ROS生成和氧化损伤,并减少炎性因子IL-6、IL-8的表达,其抗损伤机制可能与TLR4/NFκB p65信号通路有关。
Objective To investigate the effect and mechanisms of rosuvastatin in lipopolysaccharide(LPS) induced late outgrowth endothelial progenitor cells(LOCs) injury. Methods Mononuclear cells were isolated from human peripheral blood by density gradient centrifugation and cultured to obtain the LOCs. Attached cells were divided into control group, LPS treated group and rosuvastatin with different concentrations and LPS groups. CCK-8 was used to measure the cell viability. Malondialdehyde(MDA), lactate dehydrogenase(LDH) and superoxide dismutase(SOD) were examined by colorimetric assay kits. Flow cytometry was used to analyze the production of reactive oxygen species(ROS). Enzyme-linked immuno sorbent assay was utilized to detect the protein level of IL-6 and IL-8. Western blot was used to determine the TLR4 expression level and the transcription factor assay kit was used to measure the activity of NFκB p65. Results Rosuvastatin significantly inhibited the decrease of cell viability(P<0.05) induced by LPS and reduce the level of MDA(P<0.05), LDH(P<0.05), ROS(P<0.01), increasethe activity of SOD(P<0.05), decreased the expression of IL-6 and IL-8(P<0.01). Moreover, the overexpression of TLR4 and the activation of NFκB p65 induced by LPS were both inhibited by rosuvastatin(P<0.05,P<0.01). Conclusions Rosuvastatin significantly increases LOCs viability and decreases the production level of ROS, reduces the oxidative damage and inhibits IL-6 and IL-8 expression in LOCs induced by LPS. The mechanisms may be associated with TLR4/NFκB p65 signaling pathway.
Objective To investigate the effect and mechanisms of rosuvastatin in lipopolysaccharide(LPS) induced late outgrowth endothelial progenitor cells(LOCs) injury. Methods Mononuclear cells were isolated from human peripheral blood by density gradient centrifugation and cultured to obtain the LOCs. Attached cells were divided into control group, LPS treated group and rosuvastatin with different concentrations and LPS groups. CCK-8 was used to measure the cell viability. Malondialdehyde(MDA), lactate dehydrogenase(LDH) and superoxide dismutase(SOD) were examined by colorimetric assay kits. Flow cytometry was used to analyze the production of reactive oxygen species(ROS). Enzyme-linked immuno sorbent assay was utilized to detect the protein level of IL-6 and IL-8. Western blot was used to determine the TLR4 expression level and the transcription factor assay kit was used to measure the activity of NFκB p65. Results Rosuvastatin significantly inhibited the decrease of cell viability(P<0.05) induced by LPS and reduce the level of MDA(P<0.05), LDH(P<0.05), ROS(P<0.01), increasethe activity of SOD(P<0.05), decreased the expression of IL-6 and IL-8(P<0.01). Moreover, the overexpression of TLR4 and the activation of NFκB p65 induced by LPS were both inhibited by rosuvastatin(P<0.05,P<0.01). Conclusions Rosuvastatin significantly increases LOCs viability and decreases the production level of ROS, reduces the oxidative damage and inhibits IL-6 and IL-8 expression in LOCs induced by LPS. The mechanisms may be associated with TLR4/NFκB p65 signaling pathway.
引文
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[2] Chantzichristos VG,Gkrozou F,Stellos K,et al.Comparative anti-platelet profiling reveals a potent anti-aggre-gatory effect of CD34+ progenitor cell-derived late-outgrowth endothelial cells in vitro[J].J Vasc Res,2018,55:13-25.
[3] Ridker PM,Danielson E,Fonseca FA,et al.Rosuvas-tatin to prevent vascular events in men and women with elevated C-reactive protein [J].N Engl J Med,2008,359:2195-2207.
[4] Hansson GK,Hermansson A.The immune system in atherosclerosis [J].Nat Immunol,2011,12:204-212.
[5] Verma I,Syngle A,Krishan P,et al.Endothelial progenitor cells as a marker of endothelial dysfunction and atherosclerosis in ankylosing spondylitis:a cross-sectional study[J].Int J Angiol,2017,26:36-42.
[6] Giusti L,Gabriele M,Penno G,et al.A fermented whole grain prevents lipopolysaccharides-induced dysfunction in human endothelial progenitor cells[J].Oxid Med Cell Longev,2017,2017:1-13.
[7] Lin PY,Lee FY,Wallace CG,et al.The therapeutic effect of rosuvastatin and propylthiouracil on ameliorating high-cholesterol diet-induced rabbit aortic atherosclerosis and stiffness [J].Int J Cardiol,2017,227:938-949.
[8] Chen YX,Wang XQ,Fu Y,et al.Pivotal role of inflammation in vascular endothelial dysfunction of hyperli-pidemic rabbit and effects by atorvastatin [J].Int J Cardiol,2011,146:140-144.
[9] Yu L,Feng Z.The role of toll-like receptor signaling in the progression of heart failure[J].Mediators Inflamm,2018,2018:1-13.
[10] Knowlton AA.Paying for the tolls:The high cost of the innate immune system for the cardiac myocyte[J].Adv Exp Med Biol,2017,1003:17-34.
[11] Yang Y,Lv J,Jiang S,et al.The emerging role of Toll-like receptor 4 in myocardial inflammation [J].Cell Death Dis,2016,7:e2234.doi:10.1038/cddis.2016.
[12] Edfeldt K,Swedenborg J,Hansson GK,et al.Expression of toll-like receptors in human atherosclerotic lesions:a possible pathway for plaque activation [J].Circulation,2002,105:1158-1161.
[13] Wang Y,Zhang MX,Meng X,et al.Atorvastatin suppresses LPS-induced rapid upregulation of Toll-like receptor 4 and its signaling pathway in endothelial cells [J].Am J Physiol Heart Circ Physiol,2011,300:1743-1752.