天津医药

天津医药 ›› 2016, Vol. 44 ›› Issue (6): 662-664.doi: 10.11958/20160032

• 细胞与分子生物学 • 上一篇    下一篇

人脐静脉内皮细胞 1-磷酸鞘氨醇受体鉴定及其应用探讨

梁云 1， 罗光华 2， 刘宏 1， 喻妙梅 2， 王敏 1， 刘洪尧 1， 黄波 1， 徐宁 3， 张晓膺 1   

  1. 1苏州大学附属第三医院心胸外科 （邮编213000）， 2苏州大学附属第三医院综合实验室； 3瑞典隆德大学检验科临床化学与药理学部
  • 收稿日期:2016-01-25 修回日期:2016-02-17 出版日期:2016-06-15 发布日期:2016-07-04
  • 基金资助:
    国家自然科学基金资助项目（81370372）； 江苏省自然科学基金资助项目（BK20130244）； 常州市国际合作资助项目（CZ20120017）

Identification and application of sphingosine-1-phosphate receptor of human umbilical vein endothelial cells

LIANG Yun1, LUO Guanghua2, LIU Hong1, YU Miaomei2, WANG Min1, LIU Hongyao1,HUANG Bo1, XU Ning3, ZHANG Xiaoying1   

  1. 1 Department of Cardiothoracic Surgery， 2 Comprehensive Laboratory, the Third Affiliated Hospital of Soochow University, Changzhou 213000, China; 3 Division of Clinical Chemistry and Pharmacology,Department of Laboratory Medicine, Lund University
  • Received:2016-01-25 Revised:2016-02-17 Published:2016-06-15 Online:2016-07-04

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摘要： 摘要： 目的 鉴定人脐静脉内皮融合细胞株 EA.hy926 和人脐静脉内皮细胞 （HUVEC） CRL-1730 1-磷酸鞘氨醇受体 （S1PR） 亚型和胞浆中 C-myc 及 His 标签蛋白的表达情况， 为研究载脂蛋白 M （ApoM） -1-磷酸鞘氨醇轴 （ApoMS1P 轴） 的功能提供参考。方法 体外培养 EA.hy926 及 CRL-1730 至细胞密度达到 60%～70%时， 采用细胞免疫荧光技术鉴定内皮细胞标志凝血八因子（FⅧ）、 ApoM、 S1PR1~S1PR5、 C-myc 和 His 标签蛋白。结果 （1）两种细胞均表达 FⅧ和 ApoM， FⅧ在 CRL-1730 中呈散在颗粒状分布， 在 EA.hy926 中呈均匀分布。（2）两种细胞均主要表达 S1PR1， 少量表达 S1PR2 和 S1PR3， 不表达 S1PR4 和 S1PR5。（3）两种细胞胞浆中均有 C-myc 和 His 标签蛋白的表达。结论 两种细胞都具有内皮细胞的特性； 因其自身表达 ApoM、 C-myc 和 His 标签蛋白， 在应用这两种细胞研究 ApoM-S1P 轴的功能时， 不适合选用带有 C-myc 和 （或） His 标签的重组 ApoM 蛋白。

关键词: 内皮细胞, 1-磷酸鞘氨醇受体, 载脂蛋白 M, 凝血八因子, 原癌基因蛋白质 c-myc, His 标签蛋白

Abstract: Abstract: Objective To identify the expression of sphingosine-1-phosphate receptor (S1PR) subtypes, C-myc and His tag proteins of human umbilical vein endothelial fusion cell line, EA.hy926 and human umbilical vein endothelial cells(HUVEC), CRL-1730 for studying the function of apolipoprotein M (ApoM)-S1P axis. Methods Two kinds of cells (EA.hy926 and CRL-1730) were cultured to reach the density of 60%-70% in vitro. Immunofluorescence technique was employed to investigate the expressions of coagulation factor Ⅷ (FⅧ), ApoM, S1PR1-S1PR5, C-myc and His tag proteins. Results (1) Two kinds of cells both expressed FⅧ and ApoM. FⅧ presented scattered particle distribution in CRL-1730,while uniform distribution in EA.hy926. However, ApoM was strongly expressed and widely distributed in cytoplasm of twokinds of cells. (2) S1PR1-3 can be detected on their membrane other than S1PR4 and S1PR5. S1PR1 was highly expressed but S1PR2 and S1PR3 were in a low level expression. (3) Two kinds of cells both expressed C-myc and His tag proteins in cytoplasm. Conclusion Two kinds of cells have the properties of endothelial cells and can express FⅧ, ApoM, C-myc and His tag proteins. It is not suitable for choosing C-myc and/or His tag–conjugated recombinant ApoM to study the fuction of ApoM-S1P axis with these two kinds of cells.

Key words: endothelial cells, sphingosine-1-phosphate receptor, apolipoprotein M, coagulation factor Ⅷ, proto-onco? gene proteins c-myc, His tag protein