人胎盘与骨髓源性间充质干细胞体外培养及生物学特性对比

背景：组织工程修复大块组织缺损需要高浓度、大量的细胞接种，骨髓间充质干细胞是种子细胞的主要来源，但存在数量上的局限性及长期传代后细胞功能老化的问题。 目的：体外分离培养、扩增人胎盘源性间充质干细胞与人骨髓间充质干细胞，并对其生物学性状进行比较。 设计、时间及地点：细胞学体外对照观察，于2008-09/2009-02在东直门医院实验室完成。 材料：胎盘由解放军空军总医院妇产科提供，骨髓来源于健康成人献髓者。 方法：采用密度梯度离心法从胎盘中分离、纯化和传代培养人胎盘源性间充质干细胞；骨髓肝素抗凝后，采用密度梯度离心法分离、纯化和传代培养人骨髓基质干细胞。 主要观察指标：用倒置相差显微镜观察两种细胞形态及细胞生长情况，流式细胞仪分析检测第5代细胞表面标志的表达，Von Kossa染色及油红O染色检测分化能力。 结果：镜下两种细胞均为贴壁生长，形态为均一成纤维细胞样，传5代后细胞的增殖能力随传代次数的增加而有所下降，体外培养10代后细胞生长速度减慢，但人胎盘源性间充质干细胞扩增倍数较人骨髓基质干细胞平均高出两三个数量级。两种细胞具有均一的细胞表型，均表达CD29，CD44，CD166，不表达CD34，CD45，HLA-DR。两种细胞都具有成骨、成脂分化潜能，但人胎盘源性间充质干细胞分化潜能更强。 结论：人胎盘源性间充质干细胞与人骨髓基质干细胞具有相似的生物学特性，且前者具有更强的增殖能力。

In vitro culture and biological characteristics of human placendta mesenchymal stem cells versus human bone marrow mesenchymal stem cells

BACKGROUND: Tissue engineering repair for blocks of tissue defects needs a high concentration and a large number of cells for incubation. Bone marrow mesenchymal stem cells (BMSCs) are a main source of seed cells, but there are some problems: limitation of number; cell functional degradation. OBJECTIVE: To in vitro isolate, culture and amplify human placenta mesenchymal stem cells (hPMSCs) and human BMSCs, and observe their biological characteristics. DESIGN, TIME AND SETTING: The cytological, in vitro controlled study was performed at the Laboratory of Dongzhimen Hospital from September 2008 to February 2009. MATERIALS: Placenta was supplied by Department of Gynaecology and Obstetrics, General Hospital of Air Force of Chinese PLA. Bone marrow was obtained from healthy adult donors. METHODS: HPMSCs were isolated from placenta of fetation human by density gradient centrifuge. Following bone marrow heparin anticoagulation, human BMSCs were isolated, purified and passaged by density gradient centrifuge. MAIN OUTCOME MEASURES: Cell morphology and growth were observed under an inverted phase contrast microscope. The 5th passage cells were detected by flow cytometry. Differentiation of cells was determined using Von Kossa staining and Oil red O staining. RESULTS: HPMSCs and human BMSCs were both uniformly spindle-shaped in appearance under a microscope. The proliferative ability of hPMSCs were quite strong and declined with passages. After cultured 10 passages in vitro, cell growth became slow, but amplified multiple of hPMSCs was over 2-3 order of magnitude compared with human BMSCs. Both hPMSCs and human BMSCs expressed CD29, CD44 and CD166, but did not express CD34, CD45 and HLA-DR. Both cells had the potential of osteogenic and adipocytic differentiation, but hPMSCs had stronger potential. CONCLUSION: HPMSCs and human BMSCs have similar biological characteristics and the proliferative ability of hPMSCs is stronger than human BMSCs.