中国组织工程研究 ›› 2016, Vol. 20 ›› Issue (52): 7788-7795.doi: 10.3969/j.issn.2095-4344.2016.52.005

• 组织工程骨及软骨材料 tissue-engineered bone and cartilage materials • 上一篇    下一篇

丝素蛋白溶液诱导成骨分化及其性能评价

易兵成,张会兰,余哲泡,袁卉华,王先流,沈炎冰,包佳煜,娄向新,张彦中
  

  1. 东华大学化学化工与生物工程学院,上海市  201620
  • 收稿日期:2016-09-25 出版日期:2016-12-16 发布日期:2016-12-16
  • 通讯作者: 张彦中,博士,教授,东华大学化学化工与生物工程学院,上海市 201620
  • 作者简介:易兵成,男,1991年生,江西省鹰潭市余江县人,汉族,在读博士,主要从事生物材料与组织再生的研究工作。
  • 基金资助:

    国家自然科学基金面上项目(51073032,31570969);上海市科委基础研究重点项目(14JC1490100)

Osteoinductivity and performance of silk fibroin solution

Yi Bing-cheng, Zhang Hui-lan, Yu Zhe-pao, Yuan Hui-hua, Wang Xian-liu, Shen Yan-bing, Bao Jia-yu,  Lou Xiang-xin, Zhang Yan-zhong
  

  1. College of Chemistry, Chemical Engineering & Biotechnology, Donghua University, Shanghai 201620, China
  • Received:2016-09-25 Online:2016-12-16 Published:2016-12-16
  • Contact: Zhang Yan-zhong, Ph.D., Professor, College of Chemistry, Chemical Engineering & Biotechnology, Donghua University, Shanghai 201620, China
  • About author:Yi Bing-cheng, Ph.D. candidate, College of Chemistry, Chemical Engineering & Biotechnology, Donghua University, Shanghai 201620, China
  • Supported by:

    the National Natural Science Foundation of China, No. 51073032, 31570969; the Key Project of Basic Research of Shanghai Science and Technology Commission, No. 14JC1490100

摘要:

文章快速阅读:

 

文题释义:
丝素:
作为手术缝合线用于临床已有近百年历史。随着组织工程的兴起,丝素蛋白因综合性能优良而被认为在组织再生领域具有广阔的应用前景。目前,研究人员已证实多种形式(纤维膜、网状结构和海绵状等)的丝素蛋白支架在体外可支持干细胞的黏附、增殖和分化,在体内能促进损伤组织(如骨、韧带和血管等)修复。尤其在用于工程化骨组织时,丝素膜、丝素纳米纤维、丝素水凝胶和丝素多孔支架对骨损伤的修复能力已得到证实,是当前骨组织工程支架构建的热点生物材料之一,但其离临床应用还有一定差距,一些受关注的基本科学问题仍有待解决:①如何构建出机械强度符合骨组织的丝素蛋白支架;②如何调控丝素蛋白支架的降解速度与骨组织再生速率相匹配;③丝素蛋白自身是否具有诱导成骨分化的能力;④如何通过对丝素蛋白支架的精准设计克服传统生长因子诱导种子细胞成骨分化不足等。本次研究尝试探究丝素蛋白本身是否具有成骨诱导特性,将为今后丝素蛋白用于骨组织工程领域提供更充分的选择依据。
成骨分化:也称成骨诱导,是通过不同方式诱导干细胞向成骨细胞分化的现象,从而实现分化细胞对损伤骨组织的修复和再生。目前,在骨组织工程研究中,采用成骨诱导剂来诱导干细胞的成骨分化已是普遍采用的诱导方式,研究生物材料自身(如丝素蛋白)的成骨诱导特性相对较少。

背景:丝素蛋白作为一种高性能生物材料常用于构建骨组织工程支架,但丝素蛋白自身是否具有成骨诱导性能尚未见报道。
目的:体外观察不同浓度丝素蛋白溶液对大鼠骨髓间充质干细胞增殖和分化的影响。
方法:分别从蚕茧和大鼠胫骨中提取丝素蛋白和骨髓间充质干细胞,并进行鉴定。然后用0.01%、0.05%和0.1%的丝素蛋白溶液对骨髓间充质干细胞进行培养,在不同时间点测定骨髓间充质干细胞增殖和碱性磷酸酶的表达情况。
结果与结论:①FTIR结果证实了蚕茧中提取样品具有丝素蛋白的酰胺Ⅰ、酰胺Ⅱ和酰胺Ⅲ基团,为丝素蛋白;②细胞形貌观察、流式细胞术和三系分化鉴定提取的细胞具有骨髓间充质干细胞长梭形或星芒状形貌特征,能表达间充质干细胞表面抗原标志物,并具有三系分化潜能,证实了该细胞为骨髓间充质干细胞;③与不含丝素蛋白溶液培养的对照组相比,0.05%的丝素蛋白能够显著促进骨髓间充质干细胞黏附、铺展和增殖以及碱性磷酸酶表达,差异有非常显著性意义(P < 0.01);④随着丝素蛋白浓度的增加,促进效果有所降低甚至产生抑制现象。这些结果说明丝素蛋白自身具有诱导成骨作用,可为发展高性能丝素蛋白基骨组织工程支架提供科学依据。

关键词: 生物材料, 骨生物材料, 丝素蛋白, 骨髓间充质干细胞, 细胞相容性, 增殖, 成骨分化, 国家自然科学基金

Abstract:

BACKGROUND: Silk fibroin, as a kind of high-performance biomaterial, has been widely used to construct scaffolds in bone tissue engineering. However, whether silk fibroin itself holds osteoinductive ability has not been reported yet.
OBJECTIVE: To investigate the impact of different concentrations of silk fibroin solution on the proliferation and differentiation of rat bone marrow mesenchymal stem cells (BMSCs) in vitro.
METHODS: Silk fibroin and BMSCs were respectively isolated from silkworm cocoon and rat tibia, and were identified. Then, BMSCs were cultured in different concentrations of silk fibroin solution (0.01%, 0.05% and 0.1%), and the cell proliferation and the alkaline phosphatase activity were detected at different time points.
RESULTS AND CONCLUSION: FTIR spectra of the sample extracted from silkworm cocoon showed distinct absorption peaks at 1 653 (amide I), 1 530.5 (amide II) and 1 212.3 cm-1 (amide III), which could be confirmed to be silk fibroin. Thus generated BMSCs showed long fusiform or astral morphology, positive for representative markers (CD29, CD44 and CD90) relating to mesenchymal stem cells, and could differentiate into osteocytes, chondrocytes and adipocytes under specific induction conditions, which further confirmed the extracted cells were BMSCs. Compared with the control group (without silk fibroin), 0.05% silk fibroin not only significantly promoted the cell adhesion, migration and proliferation, but also enhanced the alkaline phosphatase activity (P < 0.01). With the increasing of the silk fibroin concentrations, the osteodifferentiation capacity of the BMSCs was progressively improved within the range of 0-0.05% and then declined at 0.01% of silk fibroin solutions. These results suggest that silk fibroin can promote osteogenesis, thus providing scientific evidence for developing silk fibroin-based tissue-engineered scaffolds.

Key words: Bone Marrow, Mesenchymal Stem Cells, Silk, Biocompatible Materials, Tissue Engineering

中图分类号: