中国组织工程研究 ›› 2016, Vol. 20 ›› Issue (47): 7057-7063.doi: 10.3969/j.issn.2095-4344.2016.47.009

• 药物控释材料 drug delivery materials • 上一篇    下一篇

骨形态发生蛋白缓释微球结合富血小板凝胶对骨髓基质干细胞增殖分化的影响

卢晓郎,郑亦静,程  涛,叶  超,洪建军
  

  1. 温州医科大学附属第二医院,浙江省温州市  325000
  • 收稿日期:2016-09-23 出版日期:2016-11-18 发布日期:2016-11-18
  • 通讯作者: 洪建军,硕士,主任医师,温州医科大学附属第二医院,浙江省温州市 325000
  • 作者简介:卢晓郎,男,1987年生,浙江省永嘉县人,汉族,2013年温州医科大学毕业,硕士,主要从事骨科方面的研究。

Bone morphogenetic protein sustained-release microspheres combined with   platelet-rich gel contribute to the proliferation and differentiation of bone marrow stromal stem cells

Lu Xiao-lang, Zheng Yi-jing, Cheng Tao, Ye Chao, Hong Jian-jun
  

  • Received:2016-09-23 Online:2016-11-18 Published:2016-11-18
  • Contact: Hong Jian-jun, Master, Chief physician, the Second Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, Zhejiang Province, China
  • About author:Lu Xiao-lang, Master, the Second Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, Zhejiang Province, China

摘要:

文章快速阅读:

 

文题释义:
血小板凝胶
:是富含血小板的血浆加凝血酶、氯化钙激活形成凝胶状物质。由于它不仅具有加速止血、封闭创面的特点,而且含有丰富的生长因子,能加速创面愈合,加快骨的生长。
复乳法:肽类以及蛋白质等水溶性物质的包封,主要使用W/O/W法,该方法是将肽类以及蛋白质溶解在水溶液中,将得到的水溶液分散在含有可生物降解性高分子的有机溶剂中,形成1次乳剂后,再将其分散在水相中。在此过程中,高分子的溶解度降低,使其固化,形成微球。

背景:在骨组织愈合过程的不同阶段需要不同种类的生长因子协同参与,目前组织工程中对生长因子的递送大多局限于单一种类和单一浓度,其结果往往差强人意。新一代的组织工程材料的研究热点之一就是2种或者多种生长因子的仿生程序式释放,然而目前相关的研究还非常少。
目的:观察包埋骨形态发生蛋白的聚乳酸-羟基乙酸共聚物微球结合富血小板凝胶对人骨髓基质干细胞增殖分化的影响。
方法:①复乳法制作负载骨形态发生蛋白2的聚乳酸-羟基乙酸共聚物微球,测定微球包封率、骨形态发生蛋白2的体外释放率及微球形态和体积,将制作满意的微球与激活的富血小板凝胶结合,在聚乳  酸-羟基乙酸共聚物三维支架上与人骨髓间充质干细胞的共培养;②将骨髓基质干细胞分为6组:乏血小板血浆组、血小板凝胶组(加入富血小板血浆)、骨形态发生蛋白+乏血小板血浆组、骨形态发生蛋白+血小板凝胶组、骨形态发生蛋白微球+乏血小板血浆组和骨形态发生蛋白微球+血小板凝胶组。各组加入经预处理的聚乳酸-羟基乙酸共聚物支架,观察各组细胞增殖量、碱性磷酸酶活性以及成骨细胞成熟分化的标志物的变化。
结果与结论:加入富血小板血浆的骨髓基质干细胞增殖水平明显高于乏血小板血浆组;联合应用骨形态发生蛋白2的细胞中碱性磷酸酶活性明显高于其他4组;骨形态发生蛋白微球+乏血小板血浆组和骨形态发生蛋白微球+血小板凝胶组中骨桥蛋白和骨钙蛋白mRNA的表达水平明显高于其他4组。提示骨形态发生蛋白缓释微球结合富血小板凝胶有促进骨髓基质干细胞增殖分化的作用。

关键词: 生物材料, 缓释材料, 骨形态发生蛋白2, 骨髓基质干细胞, 富血小板血浆, 聚乳酸-羟基乙酸共聚物微球, 程序式释放, 骨缺损修复, 增殖, 分化

Abstract:

BACKGROUND: Various growth factors together participate in the different stages of bone healing, but the current tissue-engineered materials limited on single type and concentration cannot obtain satisfactory outcomes. There is still a lack of study on the novel tissue-engineered material releasing two or more growth factors in order.
OBJECTIVE: To observe the effect of platelet-rich gel (PRG) combined with poly(lactide-glycolide acid) (PLGA) microspheres encapsulating bone morphogenetic protein-2 (BMP-2) on the proliferation and osteoblastic differentiation of bone marrow stromal stem cells.
METHODS: PLGA microspheres loading BMP-2 were prepared by multiple emulsion method, and the encapsulation rate, morphology and volume of microspheres as well as in vitro releasing rate of BMP-2 were measured. The eligible microspheres combined with the activated PRG were co-cultured with human bone marrow stromal stem cells on the PLGA three-dimensional scaffold. Additionally, bone marrow stromal stem cells were divided into six groups including platelet poor gel, PRG, BMP-2/platelet poor gel, BMP-2/PRG, BMP-2 microspheres/platelet poor gel and BMP-2 microspheres/PRG groups, and the preprocessed PLGA scaffold was added into each group. The DNA content, alkaline phosphatase activity and mRNA expression levels of osteopontin and osteocalcin were observed.
RESULTS AND CONCLUSION: The DNA content of bone marrow stromal stem cells was higher in all PRG scaffolds compared with the platelet poor gel scaffolds. The alkaline phosphatase activity and mRNA expression levels of osteopontin and osteocalcin in the BMP-2 microspheres/platelet poor gel and BMP-2 microspheres/PRG groups were markedly higher than those in the other four groups. These results suggest that the BMP-2 sustained-release microspheres combined with PRG are able to enhance the proliferation and differentiation of bone marrow stromal stem cells.

Key words: Platelet-Rich Plasma, Bone Morphogenetic Proteins, Tissue Engineering

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