中国神经再生研究(英文版) ›› 2022, Vol. 17 ›› Issue (1): 217-227.doi: 10.4103/1673-5374.314322

• 原著:脊髓损伤修复保护与再生 • 上一篇    下一篇

抑制lncRNA Vof-16有助于促进脊髓损伤后神经再生和功能恢复

  

  • 出版日期:2022-01-05 发布日期:2021-09-22

Inhibition of LncRNA Vof-16 expression promotes nerve regeneration and functional recovery after spinal cord injury 

Xiao-Min Zhang1, #, Li-Ni Zeng1, 2, #, Wan-Yong Yang3, Lu Ding1, 4, Kang-Zhen Chen1, Wen-Jin Fu5, Si-Quan Zeng3, Yin-Ru Liang1, Gan-Hai Chen6, *, Hong-Fu Wu1, *   

  1. 1Key Laboratory of Stem Cell and Regenerative Tissue Engineering, Guangdong Medical University, Dongguan, Guangdong Province, China; 2Biology Research Group, Guangzheng Experimental School, Huizhou, Guangdong Province, China; 3Geriatric Medicine Center, Dongguan Waterfront Zone Central Hospital, Dongguan, Guangdong Province, China; 4Scientific Research Center, the Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, China; 5Clinical Laboratory, Affiliated Houjie Hospital, Guangdong Medical University, Dongguan, Guangdong Province, China; 6Department of Intensive Care Unit, Affiliated Houjie Hospital, Guangdong Medical University, Dongguan, Guangdong Province, China
  • Online:2022-01-05 Published:2021-09-22
  • Contact: Hong-Fu Wu, PhD, hongfuw@126.com or hongfuw@gdmu.edu.cn; Gan-Hai Chen, cgh636@163.com.
  • Supported by:
    This work was financially supported by the National Natural Science Foundation of China, No. 82071374 (to HFW); Characteristic Innovation Project of Colleges and Universities in Guangdong Province of China, No. 2018KTSCX075 (to HFW); the Key Project of Social Development of Dongguan of China, No. 20185071521640 (to HFW); College Students Science and Technology Innovation Cultivation Project in Guangdong of China, Nos. pdjh2020b0257 (to HFW), pdjh2020b0263 (to HFW); College Students Innovative Experimental Project in Guangdong Medical University, China, Nos. ZZDS006 (to HFW), ZYDS005 (to HFW), ZYDB004 (to HFW), FYDY003 (to HFW); College Students’ Science and Technology Innovation Training Project, Nos. 202010571027 (to HFW), 202010571054 (to HFW), 202010571055 (to HFW), 202010571084 (to HFW), 202010571099 (to HFW), GDMU2019054 (to HFW), GDMU2019055 (to HFW), GDMU2019099, GDMU2019123 (to HFW), GDMU2019027 (to HFW), GDMU2019084 (to HFW); and the Scientific and Technological Projects of Dongguan City, No. 202050715023190 (to WJF).

摘要:

作者既往所做RNA测序研究表明,脊髓损伤后长链非编码RNA缺血相关因子Vof-16(lncRNA Vof-16)表达上调,但其在脊髓损伤中的具体作用尚不清楚。(1)实验首先通过生物信息学分析发现lncRNA Vof-16可能参与炎症和凋亡的病理生理过程;(2)利用含pHBLV-U6-MCS-CMV-ZsGreen-PGK-PURO载体的lncRNA Vof-16敲低慢病毒和含pHBLV-U6-MCS-CMV-ZsGreen-PGK-PURO载体的过表达lncRNA Vof-16慢病毒转染PC12细胞,见lncRNA Vof-16过表达可抑制PC12细胞的存活和增殖,并抑制细胞迁移和神经突起延伸,而敲低lncRNA Vof-16则减轻上述变化。Western blot结果显示,lncRNA Vof-16过表达还能提升PC12细胞中白细胞介素6、肿瘤坏死因子α和Caspase-3的表达,并降低Bcl-2的表达;(3)实验还建立了T10完全横断脊髓损伤大鼠模型,在损伤后即刻于损伤部位注射lncRNA Vof-16敲低或过表达慢病毒。损伤后7d时,敲低lncRNA Vof-16的大鼠脊髓组织中神经元的存活和轴突延伸明显增加,损伤后8周时,大鼠的后肢神经功能明显改善;(4)值得注意的是,敲低lncRNA Vof-16的大鼠脊髓组织中Bcl-2表达增加,肿瘤坏死因子α和Caspase-3下调,而过表达lncRNA Vof-16的大鼠则呈现相反的趋势;(5)结果表明,lncRNA Vof-16可能与炎症和细胞凋亡的调节有关,抑制lncRNA Vof-16可能有助于促进脊髓损伤后的神经再生和功能恢复。实验经广东省实验动物管理委员会批准。

https://orcid.org/0000-0002-1115-3681 (Hong-Fu Wu); https://orcid.org/0000-0002-0270-9705 (Gan-Hai Chen)

关键词: 脊髓损伤, 长非编码RNA缺血相关因子Vof-16, 炎症, 凋亡, 神经元存活, 增殖, 突起延伸, 神经修复, 功能恢复, 神经再生

Abstract: Our previous RNA sequencing study showed that the long non-coding RNA ischemia-related factor Vof-16 (lncRNA Vof-16) was upregulated after spinal cord injury, but its precise role in spinal cord injury remains unclear. Bioinformatics predictions have indicated that lncRNA Vof-16 may participate in the pathophysiological processes of inflammation and apoptosis. PC12 cells were transfected with a pHBLV-U6-MCS-CMV-ZsGreen-PGK-PURO vector to express an lncRNA Vof-16 knockdown lentivirus and a pHLV-CMVIE-ZsGree-Puro vector to express an lncRNA Vof-16 overexpression lentivirus. The overexpression of lncRNA Vof-16 inhibited PC12 cell survival, proliferation, migration, and neurite extension, whereas lncRNA Vof-16 knockdown lentiviral vector resulted in the opposite effects in PC12 cells. Western blot assay results showed that the overexpression of lncRNA Vof-16 increased the protein expression levels of interleukin 6, tumor necrosis factor-α, and Caspase-3 and decreased Bcl-2 expression levels in PC12 cells. Furthermore, we established rat models of spinal cord injury using the complete transection at T10. Spinal cord injury model rats were injected with the lncRNA Vof-16 knockdown or overexpression lentiviral vectors immediately after injury. At 7 days after spinal cord injury, rats treated with lncRNA Vof-16 knockdown displayed increased neuronal survival and enhanced axonal extension. At 8 weeks after spinal cord injury, rats treated with the lncRNA Vof-16 knockdown lentiviral vector displayed improved neurological function in the hind limb. Notably, lncRNA Vof-16 knockdown injection increased Bcl-2 expression and decreased tumor necrosis factor-α and Caspase-3 expression in treated animals. Rats treated with the lncRNA Vof-16 overexpression lentiviral vector displayed opposite trends. These findings suggested that lncRNA Vof-16 is associated with the regulation of inflammation and apoptosis. The inhibition of lncRNA Vof-16 may be useful for promoting nerve regeneration and functional recovery after spinal cord injury. The experiments were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.


Key words: apoptosis, functional recovery, inflammation, long non-coding RNA ischemia related factor Vof-16, nerve regeneration, nerve repair, neurite extension, neuronal survival, proliferation, spinal cord injury