慢病毒介导的脑红蛋白体内基因转染对兔损伤脊髓的作用

The protection of lentivirus-mediated neuroglobin gene transfer in vivo on injured spinal cord in rabbits, an experimental study

中文关键词:  脊髓损伤  脑红蛋白  转基因  丙二醛  一氧化氮

英文关键词:Spinal cord injury  Neuroglobin  Gene transfer  Malondialdehyde  Nitric oxide

中文摘要:

  【摘要】　目的：观察慢病毒介导脑红蛋白（Ngb）体内基因转染兔损伤的脊髓组织后对后肢运动功能的影响，探讨其作用机制。方法：用球囊压迫法制成兔脊髓损伤（ＳＣＩ）模型96只，随机分为对照组（A组）、生理盐水组（B组）、空载体组（C组）和Ngb慢病毒组（D组），每组动物24只，A组SCI后无治疗；B组SCI后向脊髓内注射生理盐水；C组SCI后向脊髓内注射空病毒；D组SCI后向脊髓内注射Ngb重组慢病毒。各组分别在1、3、7、14、21d采用BBB运动功能评分系统检测兔后肢运动功能情况；观察损伤脊髓组织内标记荧光的表达；Real-time PCR和Western blot检测Ngb mRNA及其相应蛋白的表达情况，生化方法检测丙二醛（MDA）和一氧化氮（NO）水平。结果：损伤后14d和21d，D组BBB评分明显高于其他3组（P<0.05），但A、B及C组之间比较无差异（P＞0.05）；C组和D组兔损伤部位脊髓组织均有GFP表达的绿色荧光信号；损伤后7d、14d和21d，D组的Ngb表达与其他3组比较明显增强（P<0.05）；损伤后7d、14d和21d，D组损伤脊髓组织中MDA、NO含量明显低于其他3组（P<0.05）。结论：慢病毒介导脑红蛋白（Ngb）体内基因转染可使Ngb高表达，可能是通过减轻继发性SCI，从而促进SCI后后肢运动功能的恢复。

英文摘要:

  【Abstract】 Objectiveｓ: To investigate the possible protective mechanism of lentivirus-mediated neuroglobin gene transfer in vivo on injured spinal cord (SCI) in rabbits.　Methodｓ:　Ninety-six New Zealand white rabbits were randomly divided into four groups(n=24)　according to intervention after SCI: no intervention(A); normal saline group(B); LV-GFP group (C); and LV-Ngb group(D).SCI was introduced by epidural balloon compression.24 hours after compression, T9 laminectomy was performed, then normal saline, LV-GFP or LV-Ngb was injected in situ respectively.Motor function of hind limbs after injection was evaluated by Basso-Beattie-Bresnahan(BBB) score, the expressions of green fluorescence in injured spinal cord were observed by fluorescence microscope, the expressions of Ngb mRNA and protein in injured spinal cord were detected by real-time PCR and Western blot, and the levels of malondialdehyde (MDA) and nitric oxide (NO) were also tested. Resultｓ：　Significant functional improvement was observed in group D compared with group A, B and C at day 14 and 21.The expressions of green fluorescence in injured spinal cord were observed in group C and D.The Ngb expression in group D increased significantly compared with ｔｈｅ　other 3 groups at day 7, 14 and 21.The MDA and NO levels in group D were lower than ｔｈｅ　other 3 groups(P<0.05)　at day 7, 14 and 21. Conclsionｓ：　Lentivirus-mediated neuroglobin gene transfer in vivo may induce high expression of Ngb, which may be resulted from decreasing secondary spinal cord injury, thus promotｉｎｇ hind limbs motorfunction.