崔丽丽,余芳芳,左丽
CUI Lili,YU Fangfang,ZUO Li

• 1:贵阳医学院免疫学教研室

摘要(Abstract)：

目的:优化人脐静脉内皮细胞(HUVECs)的分离及培养方法。方法:在传统HUVECs分离的基础上进行改进,通过Ⅰ型胶原酶消化分离HUVECs,利用倒置显微镜观察其生长状况、免疫组织化学方法及流式细胞术检测所培养细胞的纯度。结果:改良实验步骤和精简操作后,HUVECs分离不受影响,分离的HUVECs在体外2~4 d可长成单层,倒置显微镜下可见细胞呈"鹅卵石"状排列,Ⅷ因子免疫组化实验阳性,流式细胞术检测细胞纯度达99.37%。结论:用胶原酶消化法分离HUVECs具有培养周期短,所获得的HUVECs纯度较高的优点,有利于体外血管内皮细胞模型的建立。
Objective: To optimize the isolation and nurturing method of the human umbilical vein endothelial cell( HUVECs),to conclude identification methods of HUVEC. Method: Improving the traditional isolation of HUVECs,isolating HUVECs by type I collagenase. Then the growth of cells was observed under an inverted microscope,the cultured cells were identified by immunocytochemistry,and the purity of cells was identified by flow cytometry. Result: Improved experimental procedures and simplified the operation,the isolation of HUVECs was not affected,the primary HUVECs began to grow into a single layer about 2 ~ 4 days in vitro,arrayed like " cobblestone" shape under the inverted microscope. Ⅷ factor in the HUVECs showed positive reaction by immunocytochemistry,the purity was99. 37% by flow cytometry. Conclusions: Collagenase procedure is convenient and high purified HUVECs can be acquired. It can successfully establish the model for endothelial cells.

关键词(KeyWords)： 内皮,血管;脐静脉;免疫组织化学;细胞培养;细胞分离;流式细胞术
endothelium,vascular;umbilical veins;immunocytochemistry;cell culture;cell separation;flow cytometry

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金(31260224)

作者(Author): 崔丽丽,余芳芳,左丽
CUI Lili,YU Fangfang,ZUO Li

DOI: 10.19367/j.cnki.1000-2707.2015.01.006

参考文献(References)：

• [1]Xiao L,Liu Y,Wang N.New paradigms in inflammatory signaling in vascular endothelial cells[J].Am J Physiol Heart Circ Physiol,2014(306):H317-H325.
• [2]Sena CM,Pereira AM,Seica R.Endothelial dysfunction-a major mediator of diabetic vascular disease[J].Biochim Biophys Acta(BBA)-Mol Basis Dis,2013(1832):2216-2231.
• [3]Sumpio BE,Riley JT,Dardik A.Cells in focus:endothelial cell[J].International Journal of Biochemistry&Cell Biology,2002(34):1508-1512.
• [4]Deanfield JE,Halcox JP,Rabelink TJ.Endothelial function and dysfunction:testing and clinical relevance[J].Circulation,2007(115):1285-1295.
• [5]Cines DB,Pollak ES,Buck CA,et al.Endothelial cells in physiology and in the pathophysiology of vascular disorders[J].Blood,1998(91):3527-3561.
• [6]Trung DT,Wills B.Systemic vascular leakage associated with dengue infections the clinical Perspective[J].Curr Top Microbiol Immunol,2010(338):57-66.
• [7]Jaffe EA.Culture of human endothelial cells derived from umbilical veins[J].Clin Invest,1973(52):2754.
• [8]张宝庚,陈铁镇,张晶范.人脐带静脉及大鼠主动脉内皮细胞的培养[J].中华心血管病杂志,1985(1):52-54.
• [9]Crampton SP,Davis J,Hughes CC.Isolation of human umbilical vein endothelial cells(HUVEC)[J].J Vis Exp,2007(3):183.
• [10]余继海,许戈良,傅斌生.人脐静脉内皮细胞的培养及鉴定[J].安徽医学,2003(24):4-6.
• [11]钱勇,张励.内皮细胞生长因子和肝素对人脐静脉内皮细胞增殖的影响[J].眼科研究,2003(21):26-28.

文章评论(Comment)：