文章摘要
麻凤玉,吴彬,章旭,管大凡,齐豹,孟纯阳.不同浓度透明质酸水凝胶对骨形态发生蛋白2基因修饰的BMSCs特性的影响[J].济宁医学院学报,2017,40(2):87-92
不同浓度透明质酸水凝胶对骨形态发生蛋白2基因修饰的BMSCs特性的影响
Different concentrations of hyaluronic acid hydrogels for the bone morphogenetic protein-2 gene modified BMSCs characteristics
投稿时间:2016-12-13  
DOI:10.3969/j.issn.1000-9760.2017.02.003
中文关键词: 透明质酸水凝胶;骨髓间充质干细胞;骨形态发生蛋白2;组织工程
英文关键词: Hyaluronic acid hydrogel;Bone marrow mesenchymal stem cells;Bone morphogenetic protein-2;Tissue engineering
基金项目:山东省自然科学基金青年基金(ZR2014CQ042);山东省高等学校科技计划项目(J14LL04)
作者单位E-mail
麻凤玉 济宁医学院, 济宁 272067  
吴彬 济宁医学院附属医院, 济宁 272029  
章旭 济宁医学院附属医院, 济宁 272029  
管大凡 安康市中心医院, 安康 725000  
齐豹 济宁医学院, 济宁 272067  
孟纯阳 济宁医学院附属医院, 济宁 272029 chunyangmeng16@163.com 
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中文摘要:
      目的 探讨不同浓度透明质酸水凝胶(hyaluronic acid hydrogel,HAH)包封携带骨形态发生蛋白2(bone morphogenetic protein 2,BMP-2)基因的腺病毒载体(Ad-BMP-2)转染兔骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)后对细胞形态、增殖活性及蛋白表达的影响。方法 提取、培养BMSCs;通过成骨、成脂诱导分化及细胞表面抗原检测对BMSCs进行鉴定。体外制备不同浓度的HAH并构建携带BMP-2腺病毒载体转染至BMSCs。实验分为5组:0g/L HAH+Ad-BMP-2 BMSCs组、2g/L HAH+Ad-BMP-2 BMSCs组、4g/L HAH+Ad-BMP-2 BMSCs组、8g/L HAH+Ad-BMP-2 BMSCs组和12g/L HAH+Ad-BMP-2 BMSCs组。激光共聚焦显微镜观察不同浓度HAH下Ad-BMP-2 BMSCs的形态特征,并进行三维重建。在第1、2、4、6、8、10、12、14天不同时相点使用CCK-8法检测Ad-BMP-2 BMSCs的增殖活性;ELISA检测各组BMP-2的表达情况。结果 激光共聚焦显微镜下三维重建可见Ad-BMP-2 BMSCs在不同浓度HAH中呈现不同的形态:Ad-BMP-2 BMSCs在2、4、8g/L HAH中可多个方向铺展,具有充足的生长空间,在12g/L HAH中呈圆形。细胞增殖活性检测显示:多组间细胞增殖活性比较差异性具有统计学意义(P<0.05);4g/L HAH+Ad-BMP-2 BMSCs组和8g/L HAH+Ad-BMP-2 BMSCs组增殖活性最佳。ELISA检测BMP-2表达结果显示:各组均可表达BMP-2,8g/L HAH+Ad-BMP-2 BMSCs组表达BMP-2最高,且与其他4组相比差异具有统计学意义(P<0.05)。结论 不同浓度HAH中Ad-BMP-2 BMSC细胞形态、增殖活性、BMP-2的表达均不同,8g/L HAH包封下具有最显著的细胞活性。
英文摘要:
      Objective The different concentrations of hyaluronic acid hydrogel (HAH) encapsulated carrying bone morphogenetic protein-2(BMP-2) adenovirus vector transfection of rabbit bone marrow mesenchymal stem cells (BMSCs) and surveyed the effect on cell morphology,proliferation and protein expression.Methods BMSCs were extracted and cultured and identified by the use of osteogenesis and fat cell differentiation and cell surface antigen detection method.The different concentrations of HAH had been preparation in vitro.Adenovirus vector carrying BMP-2 gene were built and transfected into the BMSCs.The experiment was divided into five groups:0g/L HAH+Ad-BMP-2 BMSCs group;2g/L HAH+Ad-BMP-2 BMSCs group;4g/L HAH+Ad-BMP-2 BMSCs group;8g/L HAH+Ad-BMP-2 BMSCs group;12g/L HAH+Ad-BMP-2 BMSCs group.Laser scanning confocal microscopy at different concentrations HAH encapsulating observed the morphological characteristics Ad-BMP-2 BMSCs and the cells were been three-dimensional reconstruction.Ad-BMP-2 BMSCs proliferation activity was detected by CCK-8 assay method and ELISA method were used to detect the expression of BMP-2 in each group at 1,2,4,6,8,10,12,14d.Results Cell morphology was built by the Three-dimensional reconstruction under the laser confocal microscopy and was different in different concentrations HAH:Ad-BMP-2 BMSCs spread out in multiple directions and had plenty of growth space in the 2,4 and 8 g/L HAH,and cells gathered into a round shape in the 12 g/L HAH.The difference was significant statistically with comparison of the cell proliferation activity (P <0.05);4g/L HAH+Ad-BMP-2 BMSCs and 8g/L HAH+Ad-BMP-2 BMSCs had the best proliferation activity.ELISA method was used to detect the expression of BMP-2.BMP-2 could be expressed in each groups.8g/L HAH+Ad-BMP-2 BMSCs was the highest expression of BMP-2 and compared with the other four groups,and the difference was statistically significant (P <0.05).Conclusion The different concentrations of HAH in the Ad-BMP-2 BMSC cell morphology,proliferation activity and the expression of BMP-2 had differences.Cell proliferation activity under the 8g/L HAH+Ad-BMP-2 BMSCs was the best.
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