Differentiation of Rabbit Bone Mesenchymal Stem Cells into Endothelial Cells In Vitro and Promotion of Defective Bone Regeneration In Vivo
Differentiation of Rabbit Bone Mesenchymal Stem Cells into Endothelial Cells In Vitro and Promotion of Defective Bone Regeneration In Vivo
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兔骨间充质干细胞体外分化为内皮细胞及促进体内缺陷骨再生
DOI：
10.1007/s12013-013-9726-1
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发表时间：
2014-04-01
影响因子：
2.6
通讯作者：
Sun, Hongchen
中科院分区：
文献类型：
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作者：
Liu, Jinzhong;Liu, Chao;Sun, Hongchen
由于血管化不足，组织工程策略在骨再生方面常常失败，尤其是在大段骨缺损的重建中。在成骨过程中与成骨细胞有功能相互作用的大量血管内皮细胞（ECs）很难获取。在本研究中，我们通过在培养板上或聚乳酸 - 羟基乙酸（PLGA）支架上体外共培养分化的内皮细胞和间充质干细胞（MSCs）来模拟骨愈合。我们还在体内评估了成骨作用在修复兔下颌骨缺损中的效果。在本研究中，从兔体内分离间充质干细胞作为种子细胞。传代后，将间充质干细胞在血管内皮细胞条件培养基中培养以分化为内皮细胞。CD34免疫组化染色分析表明，诱导后的细胞具有内皮细胞和间充质干细胞的特征。将诱导后的内皮细胞在体外进行共培养，并以间充质干细胞诱导为成骨细胞作为对照。进行了碱性磷酸酶（ALP）和茜素红（AZR）染色实验，并测量了考马斯亮蓝总蛋白和碱性磷酸酶活性。间充质干细胞通过衍生的内皮细胞和间充质干细胞之间的直接接触增殖并分化为成骨样细胞。将共培养的细胞接种在PLGA支架上以修复兔1厘米的下颌骨缺损。通过软X射线和组织学分析评估修复效果。主要研究结果表明，间充质干细胞在支架上存活良好，支架具有生物相容性且无细胞毒性。结果证明，共培养的间充质干细胞衍生的内皮细胞改善了间充质干细胞的成骨作用并促进了新骨形成。本研究可为将体外共培养技术作为骨组织工程的一种改进方法用于大骨缺损修复提供依据。
Tissue engineering strategies often fail to regenerate bones because of inadequate vascularization, especially in the reconstruction of large segmental bone defects. Large volumes of vascular endothelial cells (ECs) that functionally interact with osteoblasts during osteogenesis are difficult to obtain. In this study, we simulated bone healing by co-culturing differentiated ECs and mesenchymal stem cells (MSCs) either on a culture plate or on a polylactide glycolic acid (PLGA) scaffold in vitro. We also evaluated the effect of osteogenesis in repairing rabbit mandible defects in vivo. In this study, MSCs were separated from rabbit as the seed cells. After passage, the MSCs were cultured in an EC-conditioned medium to differentiate into ECs. Immunohistochemical staining analysis with CD34 showed that the induced cells had the characteristics of ECs and MSC. The induced ECs were co-cultured in vitro, and the induction of MSCs to osteoblast served as the control. Alkaline phosphatase (ALP) and alizarin red (AZR) staining experiments were performed, and the Coomassie brilliant blue total protein and ALP activity were measured. The MSCs proliferated and differentiated into osteoblast-like cells through direct contact between the derived ECs and MSCs. The co-cultured cells were seeded on PLGA scaffold to repair 1 cm mandible defects in the rabbit. The effectiveness of the repairs was assessed through soft X-ray and histological analyses. The main findings indicated that MSCs survived well on the scaffold and that the scaffold is biocompatible and noncytotoxic. The results demonstrated that the co-cultured MSC-derived ECs improved MSC osteogenesis and promoted new bone formation. This study may serve as a basis for the use of in vitro co-culturing techniques as an improvisation to bone tissue engineering for the repair of large bone defects.