异种骨材料浸提液对骨髓间充质干细胞生物学特性的影响

Effect of bone xenograft extract on biological characteristics of bone marrow mesenchymal stem cells

背景:如何在去抗原等处理基础上保留异种骨材料的生物学性能及成骨诱导活性成为研究热点之一。目的:探讨异种骨材料浸提液对骨髓间充质干细胞增殖迁移及成骨分化能力的影响。方法:SD大鼠骨髓间充质干细胞分别用异种骨材料浸提液(实验组)和含体积分数为10%胎牛血清的DMEM/F12培养基培养(对照组)。MTT法、Transwell小室法、流式细胞仪检测骨髓间充质干细胞增殖能力、迁移能力、细胞周期变化。取第3代SD大鼠骨髓间充质干细胞,实验组加入成骨诱导分化浸提液,对照组加入成骨诱导分化培养液,培养至第7,14天进行碱性磷酸酶染色,培养第21天进行茜素红染色,观察钙结节形成情况。结果与结论:实验组培养1,3,5,7 d的细胞增殖活性与对照组比较差异无显著性意义(P>0.05),两组细胞周期差异无显著性意义(P>0.05)。对照组迁移细胞数显著少于实验组(P<0.05)。培养第7,14天时实验组碱性磷酸酶水平均显著高于对照组(P<0.05),成骨诱导第21天进行茜素红染色可见两组均有钙结节形成,实验组钙结节面积更大。结果表明异种骨材料无细胞毒性,具有良好的细胞相容性,可以对骨髓间充质干细胞增殖迁移及定向分化成骨细胞产生有利的促进作用。

BACKGROUND： How to preserve the biological properties and osteogenic activity of bone xenograft materials after antigen removal has become one of the hot spots. OBJECTIVE： To investigate the effect of bone xenograft extract on proliferation, migration and osteogenic differentiation of bone marrow mesenchymal stem cells. METHODS： Bone marrow mesenchymal stem cells from Sprague-Dawely rats were cultured respectively in bone xenograft extract（experimental group） and DMEM/F12 medium containing 10% fetal bovine serum（control group）. MTT, Transwell chamber and flow cytometry were used to detect cell proliferative ability, migration ability and cell cycle changes. Passage 3 bone marrow mesenchymal stem cells were selected in the two groups and cultured in osteogenic induction medium. At 7 and 14 days of osteogenic induction, alkaline phosphatase staining was done; at 21 days of osteogenic induction, alizarin red staining was performed to observe calcium nodule formation. RESULTS AND CONCLUSION： After 1, 3, 5, 7 days of culture, the cell proliferative activity and cell cycle had no significant difference between the two groups, but the number of migrating cells was significantly lower in the control group than the experimental group（P〈0.05）. At 7 and 14 days of culture, the alkaline phosphatase level was significantly higher in the experimental group than the control group（P〈0.05）; at 21 days of culture, calcium nodes formed in both two groups, but the calcium nodule area was larger in the experimental group than thecontrol group. These findings indicate that bone xenograft materials have no cytotoxicity but possess good cytocompatibility, which can promote the proliferation, migration and osteogenic differentiation of bone marrow mesenchymal stem cells.