路丹,丁文惠,彭芬,李军,赵静,叶小巾

• 1:北京大学第一医院心内科

摘要(Abstract)：

目的探索尾加压素Ⅱ(UⅡ)能否促进小鼠单核巨噬细胞系RAW264.7细胞分泌巨噬细胞集落刺激因子(M-CSF),并探讨相关的信号机制。方法培养小鼠单核巨噬细胞系RAW264.7细胞,以Real-time PCR法和ELISA法分别检测细胞的M-CSF mRNA和蛋白表达水平,用细胞免疫印迹法检测p38MAPK和ERK的磷酸化水平。结果 UⅡ呈浓度依赖性和时间依赖性促进RAW264.7细胞中M-CSF mRNA和蛋白水平的表达,最大效应时间均为刺激12 h,M-CSF mRNA水平是对照组水平的2.73倍,蛋白表达水平显著高于对照组水平[(50.04±4.28)pg/ml比(20.39±2.75)pg/ml,P<0.01],是对照组水平的2.45倍;最大效应浓度均为10-8mol/L和10-7mol/L,以10-8mol/L UⅡ刺激12 h,M-CSF的mRNA和蛋白表达水平较对照组分别增加了97.3%和80.8%。采用UⅡ受体(UT)阻断剂Urantide、ERK阻断剂PD98059和p38MAPK阻断剂SB203580分别预处理细胞后,M-CSF蛋白水平较UⅡ组显著降低,分别为[(45±4.04)pg/ml比(57.47±2.93)pg/ml]、[(42.13±4.28)pg/ml比(57.47±2.93)pg/ml]和[(44±5.34)pg/ml比(57.47±2.93)pg/ml],差异均有统计学意义(均P<0.05)。10-8mol/L UⅡ刺激细胞3 min显著促进ERK和p38MAPK蛋白磷酸化,5 min时ERK磷酸化水平达到峰值,至15 min仍持续高值,而p38MAPK在15 min时磷酸化达到峰值;30 min时,两者的磷酸化水平均减弱。结论UⅡ可通过与受体结合活化ERK和p38MAPK信号通路促进RAW264.7细胞中M-CSF的表达。

关键词(KeyWords)： 巨噬细胞集落刺激因子;炎症;尾加压素Ⅱ

Abstract:

Keywords:

基金项目(Foundation): 高等学校博士学科点专项科研基金(20120001120010)

作者(Author): 路丹,丁文惠,彭芬,李军,赵静,叶小巾

参考文献(References)：

• [1]Ames RS,Sarau HM,Chambers JK,et al.Human urotensin-Ⅱis a potent vasoconstrictor and agonist for the orphan receptor GPR14.Nature,1999,401(6750):282-286.
• [2]张丽芳,陈莉,丁文惠,等.血浆尾加压素Ⅱ浓度与冠状动脉病变程度的关系.中日友好医院学报,2008,22(1):3-6.
• [3]Wang ZJ,Shi LB,Xiong ZW,et al.Alteration of vascular urotensinⅡreceptor in mice with apolipoprotein E gene knockout.Peptides,2006,27(4):858-863.
• [4]Shiraishi Y,Watanabe T,Suguro T,et al.Chronic urotensinⅡinfusion enhances macrophage foam cell formation and atherosclerosis in apolipoprotein E-knockout mice.J Hypertens,2008,26(10):1955-1965.
• [5]邸北冰,丁文惠,赵静,等.尾加压素Ⅱ诱导巨噬细胞表达单核细胞趋化蛋白1上调及机制研究.中华老年心脑血管病杂志,2011,13(10):925-928.
• [6]Shi L,Ding W,Li D,et al.Proliferation and anti-apoptotic effects of human urotensinⅡon human endothelial cells.Atherosclerosis,2006,188(2):260-264.
• [7]Segain JP,Rolli-Derkinderen M,Gervois N,et al.UrotensinⅡis a new chemotactic factor for UT receptor-expressing monocytes.J Immunol,2007,179(2):901-909.
• [8]Lee CY,Tsai YT,Loh SH,et al.UrotensinⅡinduces interleukin 8 expression in human umbilical vein endothelial cells.PLo S One,2014,9(2):e90278.
• [9]Rodríguez-Moyano M,Díaz I,Dionisio N,et al.Urotensin-Ⅱpromotes vascular smooth muscle cell proliferation through storeoperated calcium entry and EGFR transactivation.Cardiovasc Res,2013,100(2):297-306.
• [10]Laguna JC,Alegret M.Regulation of gene expression in atherosclerosis:insights from microarray studies in monocytes/macrophages.Pharmacogenomics,2012,13(4):477-495.
• [11]Moulton KS,Vakili K,Zurakowski D,et al.Inhibition of plaque neovascularization reduces macrophage accumulation and progression of advanced atherosclerosis.Proc Natl Acad Sci U S A,2003,100(8):4736-4741.
• [12]Jaipersad AS,Lip GY,Silverman S,et al.The role of monocytes in angiogenesis and atherosclerosis.J Am Coll Cardiol,2014,63(1):1-11.
• [13]Ikonomidis I,Lekakis J,Revela I,et al.Increased circulating Creactive protein and macrophage-colony stimulating factor are complementary predictors of long-term outcome in patients with chronic coronary artery disease.Eur Heart J,2005,26(16):1618-1624.
• [14]Hojo Y,Ikeda U,Takahashi M,et al.Increased levels of monocyte-related cytokines in patients with unstable angina.Atherosclerosis,2002,161(2):403-408.
• [15]Thorpe R,Wadhwa M,Bird CR,et al.Detection and measurement of cytokines.Blood Rev,1992,6(3):133-48.
• [16]Roth P,Stanley ER.The biology of CSF-1 and its receptor.Curr Top Microbiol Immunol,1992,181:141-167.
• [17]Cirillo P,De Rosa S,Pacileo M,et al.Human urotensinⅡinduces tissue factor and cellular adhesion molecules expression in human coronary endothelial cells:an emerging role for urotensinⅡin cardiovascular disease.J Thromb Haemost,2008,6(5):726-736.
• [18]Song N,Ding W,Chu S,et al.UrotensinⅡstimulates vascular endothelial growth factor secretion from adventitial fibroblasts in synergy with angiotensinⅡ.Circ J,2012,76(5):1267-1273.
• [19]Dong X,Ye X,Song N,et al.UrotensinⅡpromotes the production of LTC4 in rat aortic adventitial fibroblasts through NF-kappa B-5-LO pathway by p38 MAPK and ERK activations.Heart Vessels,2013,28(4):514-523.
• [20]Djordjevic T,Bel Aiba RS,Bonello S,et al.Human urotensinⅡis a novel activator of NADPH oxidase in human pulmonary artery smooth muscle cells.Arterioscler Thromb Vasc Biol,2005,25(3):519-525.
• [21]Guan SM,Zhang M,He JJ,et al.Mitogen-activated protein kinases and phosphatidylinositol 3-kinase are involved in Prevotella intermedia-induced proinflammatory cytokines expression in human periodontal ligament cells.Biochem Biophys Res Commun,2009,386(3):471-476.