中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2018, Vol. 13 ›› Issue (2): 298-303.doi: 10.4103/1673-5374.226400

• 原著:脑损伤修复保护与再生 • 上一篇    下一篇

胶原结合结构域修饰的胰岛素样生长因子1可用于神经再生

  

  • 收稿日期:2018-01-11 出版日期:2018-02-15 发布日期:2018-02-15
  • 基金资助:

    中国国家自然科学基金项目（81350013），吉林省科技项目（20160101027JC，SC201502001），吉林大学创新基金（2017031 ，2017176）

Modified insulin-like growth factor 1 containing collagen-binding domain for nerve regeneration

Jian-an Li1, 3, Chang-fu Zhao2, Shao-jun Li3, Jun Zhang1, Zhen-hua Li3, Qiao Zhang2, Xiao-yu Yang1, Chun-fang Zan1   

  1. 1 Department of Orthopedics, Second Hospital of Jilin University, Changchun, Jilin Province, China
    2 Department of Orthopedics, China-Japan Union Hospital, Jilin University, Changchun, Jilin Province, China
    3 Department of Orthopedics, Affiliated Hospital of Changchun University of Traditional Chinese Medicine, Changchun, Jilin Province, China
  • Received:2018-01-11 Online:2018-02-15 Published:2018-02-15
  • Contact: Xiao-yu Yang, M.D. or Chun-fang Zan,181974183@qq.com or 861916323@qq.com.
  • Supported by:

    This study was supported by the National Natural Science Foundation of China, No. 81350013; a grant from the Jilin Provincial Science and Technology Plan of China, No. 20160101027JC & SC201502001; and the Graduate Innovation Fund of Jilin University in China,No. 2017031 & 2017176.

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摘要：

胰岛素样生长因子1是一种潜在的神经修复营养因子，但由于半衰期短、清除速度快且靶向目标特异性有限，因而不能成为一种有效的治疗方法。作者希望为其绑定一个特异性结构，以实现进行有针对性、持久治疗的可能。为此，实验将胰岛素样生长因子1与胶原结合结构域融合，并使用酶标仪检测其与胶原蛋白的亲和能力，并转染在PC12细胞和许旺细胞中，以MTT测定细胞增殖情况表达靶蛋白，免疫荧光染色检测神经丝和微管相关蛋白2表达，以实时PCR检测胰岛素样生长因子1受体和神经生长因子mRNA表达情况。结果显示，与胰岛素样生长因子1相比，重组蛋白的胶原结合活性明显增加，且该蛋白可促进PC12细胞和许旺细胞的增殖，以及PC12细胞中神经丝和微管相关蛋白2的表达。此外该重组蛋白还能刺激胰岛素样生长因子1受体和神经生长因子mRNA的长期稳定表达。表明构建的胶原结合结构域修饰的胰岛素样生长因子1可到达靶向持续治疗的目的，可能成为一种临床上使用的具有持续治疗作用的促进神经再生因子。

orcid:0000-0001-9388-3794(Xiao-yu Yang)
         0000-0002-4557-293X(Chun-fang Zan)

关键词: 神经再生, 胰岛素样生长因子1, 胶原结合结构域, 融合蛋白, 胶原酶, 靶向治疗

Abstract:

Insulin-like growth factor 1 (IGF-1) is a potential nutrient for nerve repair. However, it is impractical as a therapy because of its limited halflife,rapid clearance, and limited target specificity. To achieve targeted and long-lasting treatment, we investigated the addition of a binding structure by fusing a collagen-binding domain to IGF-1. After confirming its affinity for collagen, the biological activity of this construct was examined by measuring cell proliferation after transfection into PC12 and Schwann cells using a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay. Immunofluorescence staining was conducted to detect neurofilament and microtubule-associated protein 2 expression, while real time-polymerase chain reaction was utilized to determine IGF-1 receptor and nerve growth factor mRNA expression. Our results demonstrate a significant increase in collagen-binding activity of the recombinant protein compared with IGF-1.Moreover, the recombinant protein promoted proliferation of PC12 and Schwann cells, and increased the expression of neurofilament and microtubule-associated protein 2. Importantly, the recombinant protein also stimulated sustained expression of IGF-1 receptor and nerve growth factor mRNA for days. These results show that the recombinant protein achieved the goal of targeting and long-lasting treatment,and thus could become a clinically used factor for promoting nerve regeneration with a prolonged therapeutic effect.

Key words: nerve regeneration, insulin-like growth factor 1, collagen-binding domain, fusion protein, collagenase, targeted therapy, neural regeneration