骨髓间充质干细胞构建组织工程化小口径血管  

Construction of small diameter tissue engineering blood vessel by bone marrow mesenchymal stem cells

作　　者：杜振宗[1] 任华[2] 刘洪生[2] 宋剑非[1] 郑民[1] 王玮[1] 

机构地区：[1]桂林医学院附属医院胸心外科,541001 [2]中国医学科学院中国协和医科大学北京协和医院胸心外科

出　　处：《中华生物医学工程杂志》2008年第3期166-170,共5页Chinese Journal of Biomedical Engineering

摘　　要：目的采用在动物体外构建初级组织工程化血管、体内强化的方法，探讨构建小口径组织工程化血管的可能性。方法体外培养骨髓间充质干细胞（BMSC），用含全反式维甲酸（AT-RA）、双丁酰环磷酸腺苷（db-cAMP）的DMEM-LG培养液和含血管内皮细胞生长因子（VEGF）的培养液诱导BMSC分别向血管平滑肌样细胞和血管内皮样细胞分化。免疫荧光观察平滑肌样细胞B肌动蛋白的表达和内皮样细胞vWF的表达。电镜观察超微结构的改变。诱导的血管平滑肌样细胞和血管内皮样细胞，分层种植于胶原包埋聚乙醇酸（PGA）的复合支架表面，将细胞和支架复合体种植于动物皮下，于植入后第4、8周再次麻醉动物，取出植入皮下的组织工程化血管，行组织学检查、压力实验及免疫荧光检查。结果诱导14d后，BMSC能够分化为血管平滑肌样细胞和血管内皮样细胞：B肌动蛋白和vWF呈阳性表达，电镜证实细胞出现了相应的形态学改变。人工血管组织学观察见管壁结构清晰。单纯支架组可承受100～150mmHg（1mmHg=0．133kPa）的血管腔内压力，实验组则均可承受200mmHg的血管腔内压力不破裂。实验组皮下培养8周Brdu标记细胞的免疫荧光结果显示部分细胞核呈现明亮的黄绿色荧光。结论以动物皮下为生物反应器可构建出组织工程化血管，其大体结构和天然血管相似。Objective To investigate the possibility of constructing small diameter vessel by means of tissue engineering in vivo. Methods Bone marrow stem cells （ BMSCs ） were cultured in vitro, and differentiated into vascular smooth muscle-like cells and vascular endothelium-like cells by DMEM-LG culture fluid including all trans-retinoic acid （AT-RA） , dibutyryl cyclic adenosine mono-phosphate （rib-cAMP） and culture fluid including vascular endothelial growth factor （VEGF） respectively. 13 actin and vWF expression were observed using immunofluorescence. Ultrastructural change was observed by electron microscope. The mixture of seed cells and scaflbld were produced by separately seeding vascular smooth muscle-like cells and vascular endothelium-like cells derived from BMSCs on polyglycolic acid （PGA）/eollagen scaffold. The two layers were separated by extracellular matrix gel （ECMgel）. The tissue engineering blood vessels were examined by pressure experiment and both gross and histology with HE staining and immunofluorescence. Results BMSCs could be differentiated into vascular smooth muscle-like cells and vascular endothelium-like cells by inducing for 14 days. [3 actin and vWF expression were positive. Uhrastructural related changes were observed by electron microscope respectively. Histological analysis of the tissue engineering blood vessel wall revealed a typical artet＇y structure, including endothelium, media and adventitia. Blood vessel intracavitary pressure was 100-150 mm Hg（ 1 mm Hg =0. 133 kPa） in scaffold group, 200 mm Hg in experimental group and tissue engineering blood vessel did not break. Eight weeks after implantation, the labeled seed cells by 5- bromo-2＇-deoxyuridine （ Brdu ） were found in the three layers of the vessel wall. Conclusions The subcutaneous tissue is a good bioreactor to construct tissue engineering blood vessels. Histology of tissue engineering blood vessels is similar to natural vessel.

关 键 词：骨髓 间质干细胞 生物反应器 组织工程 血管 种子细胞 

分 类 号：R318[医药卫生—生物医学工程]