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 (1): 135-144.doi: 10.4103/1673-5374.224381

• 原著:视神经损伤修复保护与再生 • 上一篇    下一篇

突变型rAAV2为载体的RNA干扰沉默PTEN促进视神经轴突再生

  

  • 收稿日期:2017-10-09 出版日期:2018-01-15 发布日期:2018-01-15
  • 基金资助:

    江苏省卫计委研究基金项目（H201653），常熟市科技局研究基金项目（CS201616）

PTEN knockdown with the Y444F mutant AAV2 vector promotes axonal regeneration in the adult optic nerve

Zheng-ru Huang1, 2, Hai-ying Chen2, Zi-zhong Hu1, Ping Xie1, Qing-huai Liu1   

  1. 1 Department of Ophthalmology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province, China
    2 Department of Ophthalmology, the Second People’s Hospital of Changshu, Changshu, Jiangsu Province, China
  • Received:2017-10-09 Online:2018-01-15 Published:2018-01-15
  • Contact: Qing-huai Liu, Ph.D.,liuqh@njmu.edu.cn.
  • Supported by:

    This study was supported by the Research Foundation of Jiangsu Provincial Commission of Health and Family Planning of China,No. H201653, and the Research Foundation of Changshu Science and Technology Bureau of China, No. CS201616.

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摘要：

轴突再生障碍是成年哺乳动物视神经损伤后视力丧失的主要原因。激活PI3K/AKT/mTOR信号通路可促进中枢神经系统损伤后的轴突再生，而mTOR负调节因子PTEN的缺失可以增强成年皮质脊髓神经元和神经节细胞的再生能力。实验建立了视神经轴突横断大鼠模型，以酪氨酸突变的AAV2为载体行RNA干扰沉默PTEN表达。结果发现，在损伤前4周玻璃体内注射Y444F AAV2-shRNA.PTEN可增加神经节细胞的存活，再生轴突更长。损伤后注射Y444F AAV2-shRNA.PTEN，也有约19％的神经节细胞存活，并诱导轴突再生至视交叉附近。结果表明，酪氨酸突变的AAV2介导的PTEN沉默能够促进视神经再生。

orcid:0000-0003-0351-136X(Qing-huai Liu)

关键词: 神经再生, 视神经, 基因治疗, Mü, ller细胞, 视网膜神经节细胞, AAV2, shRNA, PTEN, GLAST, mTOR

Abstract:

The lack of axonal regeneration is the major cause of vision loss after optic nerve injury in adult mammals. Activating the PI3K/AKT/mTOR signaling pathway has been shown to enhance the intrinsic growth capacity of neurons and to facilitate axonal regeneration in the central nervous system after injury. The deletion of the mTOR negative regulator phosphatase and tensin homolog (PTEN) enhances regeneration of adult corticospinal neurons and ganglion cells. In the present study, we used a tyrosine-mutated (Y444F) AAV2 vector to efficiently express a short hairpin RNA (shRNA) for silencing PTEN expression in retinal ganglion cells. We evaluated cell survival and axonal regeneration in a rat model of optic nerve axotomy. The rats received an intravitreal injection of wildtype AAV2 or Y444F mutant AAV2 (both carrying shRNA to PTEN) 4 weeks before optic nerve axotomy. Compared with the wildtype AAV2 vector, the Y444F mutant AAV2 vector enhanced retinal ganglia cell survival and stimulated axonal regeneration to a greater extent 6 weeks after axotomy. Moreover,post-axotomy injection of the Y444F AAV2 vector expressing the shRNA to PTEN rescued ~19% of retinal ganglion cells and induced axons to regenerate near to the optic chiasm. Taken together, our results demonstrate that PTEN knockdown with the Y444F AAV2 vector promotes retinal ganglion cell survival and stimulates long-distance axonal regeneration after optic nerve axotomy. Therefore, the Y444F AAV2 vector might be a promising gene therapy tool for treating optic nerve injury.

Key words: nerve regeneration, optic nerve, axotomy, gene therapy, Müller cell, retinal ganglion cell, AAV2, shRNA, PTEN, GLAST, mTOR, neural regeneration