胎猪BMSC体外构建软骨的实验研究  

Porcine Fetal Bone Marrow Mesenchymal Stem Cells in Cartilage Tissue Engineering

作　　者：刘李娜[1] 何爱娟[1] 周广东[1] 曹卫刚[1] 

机构地区：[1]上海交通大学医学院附属第九人民医院整复外科,上海市200011

出　　处：《组织工程与重建外科杂志》2013年第4期181-185,共5页Journal of Tissue Engineering and Reconstructive Surgery

基　　金：国家自然科学基金(81000677;31271046;50830105)

摘　　要：目的比较胎猪骨髓间充质干细胞(Bone Marrow Mesenchymal Stem Cells,BMSCs)和成年猪BMSCs构建软骨能力的差异,寻找合适的同种异体组织工程软骨种子细胞来源。方法通过剖腹产手术获得胎龄为70 d的胎猪,胎猪骨髓液贴壁培养获得胎猪BMSCs;抽取成年猪骨髓液,经贴壁培养法获得成年猪BMSCs。两种细胞体外扩增培养后,观察第3代细胞形态,并进行成骨、成脂和成软骨诱导。分别取两种细胞以1×108cells/mL的细胞终浓度,接种于聚乳酸包埋的聚羟基乙酸支架,体外诱导培养8周后取材。通过大体观察、糖胺聚糖(GAG)含量测定、总胶原含量测定、组织学,以及免疫组化等方法,对两种细胞构建的组织工程软骨的相关生物学特性进行比较。结果胎猪BMSCs比成年猪BMSCs具有更好的增殖和成骨、成脂和成软骨能力。胎猪BMSCs构建的软骨有良好的软骨外观,而且GAG含量和总胶原含量均高于成年猪BMSCs构建的软骨(P<0.01)。组织学和免疫组化显示,胎猪BMSCs构建的软骨组织结构致密,基质及Ⅱ型胶原显色程度均明显强于成年猪BMSCs构建的软骨。结论胎猪BMSCs是组织工程软骨较好的种子细胞来源。Objective To investigate the optimal seed cell for cartilage engineering by comparing the chondrogenesis capability of porcine fetal bone marrow mesenchymal stem cells （fBMSCs） and adult porcine BMSCs （aBMSCs）. Methods Fetal pigs with gestational age of 70 days were obtained by the uterine-incision delivery, and primary fBMSCs were isolated from the bone marrow. Primary aBMSCs were isolated from the bone marrow which was aspirated from iliac. Cell morphology of the two kinds of cells at passage 3 were observed after in vitro proliferation. The two kinds of cells at passages 3 were characterized by their osteogenic, adipogenic and chondrogenic differentiation potential. Then the fBMSCs and aBMSCs were separately seeded onto a polyglycolic acid/polylactic acid （PGA/PLA） scaffold with the concentration of 1 ×108 cells/mL. All specimens were harvested after 8 weeks＇ culture in vitro. Gross observation, glycosaminoglycan （GAG） quantification, total collagen quantification and histology were used to compare related characteristic differences of engineered cartilage formed by the two kinds of cells. Results fBMSCs had better proliferation and multiple differentiation capacity than aBMSCs. The two kinds of cells both formed mature cartilage after 8 weeks of culture in vitro, and the engineered cartilage of aBMSCs group had better appearance. The GAG content and total collagen content of the cartilage formed by fBMSCs were both higher than the cartilage formed by aBMSCs （P〈0.01）. Histology and immunohistochemistry demonstrated that the cartilage formed by fBMSCs have more compact tissue structure. The cartilage matrix staining of cartilage formed by fBMSCs were stronger than that of cartilage formed by aBMSCs. Conclusion The fBMSCs seems to be the optimal seed cells for cartilage tissue engineering.

关 键 词：骨髓间充质干细胞 胎猪 组织工程 软骨 种子细胞 

分 类 号：Q813.11[生物学—生物工程]