Huang Hui-min, Ma Liang-long, Ren Hong, Wu Shao-feng, Jiang Zu-ming
Shanghai Children's Medical Center, Medical College, Shanghai Jiaotong University, Shanghai 200127, China.
Zhonghua Yi Xue Za Zhi. 2007 Dec 25;87(48):3440-2.
To construct a tissue-engineered graft by using bone marrow cells as seeding cells and heterogeneous acellularized matrix as scaffold.
Mononuclear cells were isolated from the bone marrow from piglets and cultured in different mediums including either vessel endothelial growth factor (VEGF) or platelet derived growth factor BB (PDGF-BB) to observe the differentiation of cells. Immunoassay was used to detect the expression of specific markers of endothelial cells or specific markers of smooth muscle cells. Adult dogs were killed and their thoracic or abdominal aortas were taken out and processed by multi-step decellularizing technique to remove the original cells while the elastic and collagen fibers were preserved. The undifferentiated bone marrow mononuclear cells were seeded onto the acellularized matrix and incubated in vitro. The cell-seeded grafts were then transplanted to the bone marrow donating piglets to substitute part of the native pulmonary artery. Three weeks later right ventriculography was performed. 100 days later the piglets were sacrificed. The transplanted vessels and the nearby tissues of native pulmonary vessels were excised for inspection.
The mononuclear cells cultured in the medium including VEGF showed the morphological features of endothelial cells and were positive of the specific markers of endothelial cells: platelet-endothelial cell adhesion molecule-l, vascular endothelial growth factor receptor Flk-1, VE-cadherin, and plasma factor VIII. The cells cultured in the medium including PDGF-BB showed morphological feature of smooth muscle cells and were positive of he specific marks of smooth muscle cells: alpha-SMA and calponin. One hundred days after transplantation, the inner surfaces of the grafts were smooth without thrombosis, calcification, and aneurysm. The maximal load was 2.76 N and the maximal elongation was 20.31 mm. Under the microscopy a great number of growing endothelial cells and smooth muscle cells could be seen and elastic and collagen fibers were abundant.
The mononuclear cells from bone marrow and acellularized matrix may be used to construct tissue-engineered graft.
以骨髓细胞为种子细胞，异种脱细胞基质为支架构建组织工程移植物。
从小猪骨髓中分离单个核细胞，在含血管内皮生长因子（VEGF）或血小板衍生生长因子BB（PDGF - BB）的不同培养基中培养，观察细胞分化情况。采用免疫分析法检测内皮细胞特异性标志物或平滑肌细胞特异性标志物的表达。处死成年犬，取出其胸主动脉或腹主动脉，采用多步脱细胞技术处理，去除原有细胞，保留弹性纤维和胶原纤维。将未分化的骨髓单个核细胞接种到脱细胞基质上，进行体外培养。然后将接种细胞的移植物移植到供体骨髓小猪体内，替代部分天然肺动脉。3周后进行右心室造影。100天后处死小猪，切除移植血管及天然肺血管附近组织进行检查。
在含VEGF的培养基中培养的单个核细胞呈现内皮细胞形态特征，内皮细胞特异性标志物血小板 - 内皮细胞黏附分子 - 1、血管内皮生长因子受体Flk - 1、VE - 钙黏蛋白和血浆因子VIII呈阳性。在含PDGF - BB的培养基中培养的细胞呈现平滑肌细胞形态特征，平滑肌细胞特异性标志物α - SMA和钙调蛋白呈阳性。移植100天后，移植物内表面光滑，无血栓形成、钙化和动脉瘤形成。最大负荷为2.76 N，最大伸长为20.31 mm。显微镜下可见大量生长的内皮细胞和平滑肌细胞，弹性纤维和胶原纤维丰富。
骨髓单个核细胞和脱细胞基质可用于构建组织工程移植物。
