收稿日期: 2013-10-20
网络出版日期: 2014-08-07
基金资助
云南省教育厅科学研究基金资助项目（2013J122C）; 云南省自然科学基金资助项目（2007C0034R, 2012FB096）
Study on biological characterization of Wharton's jelly mesenchymal stem cells from human umbilical cord
Received date: 2013-10-20
Online published: 2014-08-07
目的 探讨人脐带Wharton’s jelly间充质干细胞的培养方法及生物学特性。 方法 将脐带Wharton’s jelly剪成约0.5 mm3种植到培养瓶，3 h后添加培养液。细胞达90%融合时用胰酶消化传代。用免疫组化检测波形蛋白的表达；用流式检测细胞膜表型；用诱导液检测其分化为脂肪样细胞和神经样细胞的潜能。 结果 培养6 d 左右，细胞从组织块游出，为长梭形的成纤维样细胞，传代后细胞增殖加快，呈放射状分布。间充质干细胞标记物CD90等为阳性，而CD34等为阴性；波形蛋白呈强阳性。成脂诱导后，细胞质出现脂滴，油红O染色为阳性。经黄芪诱导24 h，细胞表达Nestin和NF，而NSE和GFAP为阴性。 结论 人脐带Wharton’s jelly中有间充质干细胞，且有多向分化潜能。
关键词： 间充质干细胞; 脐带; Wharton’s jelly; 分离; 培养
李士欣 , 华映坤 , 王林坪 , 杨慧 , 鲁俊 , 吕超绍 , 董旭东 , 撒亚莲 . 人脐带Wharton’s jelly间充质干细胞生物学特性的研究[J]. 中国临床解剖学杂志, 2014 , 32(4) : 432 -436 . DOI: 10.13418/j.issn.1001-165x.2014.04.015
Objective To establish the culture methods of Wharton’s jelly mesenchymal stem cells from human umbilical cord（hWJ-MSCs）and observe the biological characteristics. Methods The umbilical cord organization of healthy full-term caesarean section was collected under sterile condition. The WJ was diced into cubes of approximately 0.5 mm3, and then plant them in culture flasks which kept still in 3 h before adding L-DMEM with 100 ml/L fetal bovine serum. The WJ-MSCs were passaged using 1.25 g/L Trypsin when they reached 90% confluence. Vimentin was tested by immunocytochemistry. The cell phenotype of WJ-MSCs was detected by flow cytometry. The hWJ-MSCs can differentiate into adipocytes like cell identified by oil-red stain. WJ-MSCs were cultured in medium containing 0.05 g//L Astragalus, then tested the expression of GFAP, NF, Nestin and NSE by immunocytochemical staining. Results Some cells swam out from the tissue block for 6 days or so, which belong to fibroblast-like cell appearing long spindle shaped fibroblast-like cells. WJ-MSCs after subculture presented in a uniform spindle morphology shape, showing parallel growth or vortex growth. The marks of MSCs such as CD90 were positive. The expression of CD34 was negative. Immunocytochemical staining showed WJ-MSCs expressed Vimentin with strong positive. After adipocyte like cells committed induction, these cells with lipid vacuoles accumulated in the cytoplasm gradually could be stained by oil-red O. After committed induction in 24 h by Astragalus, the differentiated cells were stained by Nestin and NF, but GFAP and NSE were negative. Conclusions WJ-MSCs can be effectively obtained from Wharton's jelly with the planting culture, which possess multi-differentiated capability.
Key words： Mesenchymal stem cells; Umbilical cord; Wharton's jelly; Isolation; Culture
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