全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...
-  2015 

慢病毒载体表达外源PON1对小鼠巨噬细胞的影响 Effects of Exogenous PON1 Expressed by Lentivirus Vector on Macrophages

Keywords: 对氧磷酯酶1,慢病毒载体,巨噬细胞,TNF-α

Full-Text   Cite this paper   Add to My Lib

Abstract:

目的:为探讨对氧磷酯酶1(PON1)在机体代谢性疾病中的作用及机制,本实验构建含人源性PON1基因的慢病毒表达载体,研究其表达分泌PON1的能力及对小鼠巨噬细胞的影响。方法:根据人肝cDNA文库的PON1序列设计特定引物,定点诱变PCR获得两端为PmeⅠ内切酶位点的目的基因,经酶切连接到pWPI-GFP载体上,筛选及DNA测序鉴定。磷酸钙法瞬时转染293T细胞,荧光显微镜观察GFP表达反映转染效率,Western Blotting检测细胞培养基中PON1分泌表达量,采用对氧磷为底物检测PON1活性。含PON1的培养基与小鼠腹腔巨噬细胞共孵育,ELISA法检测氧化条件下细胞分泌肿瘤坏死因子α(TNF-α)、白细胞介素-6(IL-6)的水平。结果:构建的pWPI-PON1慢病毒表达载体序列正确,高效转染293T细胞(转染效率可达到80%-90%)可有效分泌表达PON1至培养基,显著提高培养基PON1活性,与对照组相比,培养基中PON1可明显抑制巨噬细胞炎症因子TNF-α、IL-6的分泌(P<0.01)。结论:pWPI-PON1慢病毒载体高效表达分泌的外源PON1能有效降低巨噬细胞的炎症反应

References

[1]  Loued S,Isabelle M,Berrougui H,et al.The anti-inflammatory effect of paraoxonase 1against oxidized lipids depends on its association with high density lipoproteins[J].Life Sci,2012,90(1-2):82-88.
[2]  Mineo C,Shaul PW.PON-dering differences in HDL function in coronary artery disease[J].J Clin Invest,2011,121(7):2 545-2 548.
[3]  Wang M,Lang X,Cui S,et al.Quantitative assessment of the influence of paraoxonase 1activity and coronary heart disease risk[J].DNA Cell Biol,2012,31(6):975-982.
[4]  Vakili L,Navab KD,Shabihkhani M,et al.Systemic inflammation,intestine,and paraoxonase-1[J].Adv Exp Med Biol,2014,824:83-88.
[5]  Shih DM,Gu L,Xia YR,et al.Mice lacking serum paraoxonase are susceptible to organophosphate toxicity and atherosclerosis[J].Nature,1998,394:284-287.
[6]  Aharoni S,Aviram M,Fuhrman B.Paraoxonase 1(PON1)reduces macrophage inflammatory responses[J].Atherosclerosis,2013,228(2):353-361.
[7]  Zhao Y,Ma Y,Fang Y,et al.Association between PON1activity and coronary heart disease risk:A metaanalysis based on 43studies[J].Mol Genet Metab,2012,105:141-148.
[8]  Efrat M,Aviram M.Paraoxonase 1interactions with HDL,antioxidants and macrophages regulate atherogenesis-aprotective role for HDL phospholipids[J].Adv Exp Med Biol,2010,660:153-166.
[9]  Litvinov D,Mahini H,Garelnabi M.Antioxidant and anti-inflammatory role of paraoxonase 1:implication in arteriosclerosis diseases[J].N Am J Med Sci,2012,4(11):523-532.
[10]  Zhou CY,Cao J,Shang L,et al.Reduced paraoxonase1activity as a marker for severe coronary artery disease[J].Disease Markers,2013,35(2):79-103.
[11]  She ZG,Zheng W,Wei YS,et al.Human Paraoxonase Gene Cluster Transgenic Overexpression Represses Atherogenesis and Promotes Atherosclerotic Plaque Stability in ApoE-Null Mice[J].Circulation Research,2009,104(10):1 160-1 168.

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133