鞘内注射Reelin基因干扰慢病毒载体对大鼠慢性神经病理性疼痛的研究

发布时间：2018-07-13 21:55

【摘要】：目的探讨鞘内注射Reelin基因干扰慢病毒载体(LV-Reelin-RNAi)对慢性坐骨神经压迫性损伤(chronic constriction injury,CCI)疼痛大鼠脊髓水平Reelin的表达变化及疼痛行为学的影响。方法鞘内置管成功后无运动障碍的SD雄性大鼠144只,体重200-270g,随机分为4组:Sham组(n=36),CCI组(n=36),RNAi组(n=36)和空载体组(n=36)。Sham组大鼠只暴露右侧坐骨神经后不结扎,而其他三组大鼠在暴露右侧坐骨神经后,用4.0含铬羊肠线轻轻的结扎四道以制造慢性坐骨神经压迫性损伤疼痛模型。术后第1天,各组分别鞘内注射生理盐水15μl、生理盐水15μl、LV-Reelin-RNAi5μl+生理盐水10μl、空载体5μl+生理盐水10μl。各组大鼠均于术前1d、术后1d、4d、7d、10d、14d和21d测量双侧后足底机械缩足阈值(paw withdrawal mechanical threshold,PWMT)和热缩足潜伏期(paw withdrawal thermal latency,PWTL)。各组在第7、14、21天取腰膨大,采用QPCR、Western blot和石蜡切片免疫组化(Immunohistochemistry-paraffin sections,IHC-P)检测脊髓背角Reelin m RNA和蛋白的表达。结果术前各组大鼠PWMT和PWTL无显著性差异;CCI模型大鼠手术测术后各组各时间点PWMT和PWTL均明显降低(P0.05),手术侧脊髓背角Reelin m RNA的表达量和Reelin蛋白表达量CCI术后明显减少(P0.05)。鞘内注射LV-Reelin-RNAi可进一步降低PWMT和PWTL。与之相对应,大鼠右侧脊髓背角Reelin m RNA和蛋白的表达也进一步降低(P0.05)。鞘内注射空载体对慢性神经病理性疼痛大鼠的疼痛行为学、Reelin m RNA和Reelin蛋白表达无明显影响(P0.05)。结论脊髓水平Reelin的表达在慢性压迫坐骨神经诱发慢性神经病理性疼痛后明显下降;鞘内注射LV-Reelin-RNAi可进一步下调Reelin在脊髓背角的表达并且加剧神经病理性疼痛症状。Reelin可能参与慢性压迫性神经病理性疼痛的产生与发展。
[Abstract]:Objective to investigate the effect of intrathecal injection of Reelin gene interference lentivirus vector (LV-Reelin-RNAi) on the expression of Reelin in spinal cord and pain behavior in rats with chronic sciatic nerve compression injury (chronic constriction injura). Methods 144 male Sprague-Dawley rats (200-270g), weighing 200-270 g, were randomly divided into 4 groups: the control group (nong36), the CCI group (nong36) and the empty carrier group (nong36). The rats in the empty carrier group (nong36) .Sham group were exposed to the right sciatic nerve and did not ligate the right sciatic nerve. The other three groups of rats were exposed to the right sciatic nerve, and 4. 0 chromium-containing intestinal cord was ligated gently to make the pain model of chronic sciatic nerve compression injury. On the first day after operation, 15 渭 l saline, 15 渭 l LV-Reelin-RNAi 5 渭 l normal saline and 10 渭 l empty carrier 5 渭 l normal saline were injected intrathecally in each group. The threshold of mechanical foot contraction (paw withdrawal mechanical threshold( paw withdrawal mechanical) and the latent period of thermal contraction (paw withdrawal thermal latencies) were measured 1 day before operation, 1 day after operation, 1 day, 4 days, 7 days, 10 days, 14 days and 21 days, respectively. The expression of Reelin mRNA and protein in the dorsal horn of spinal cord was detected by immunohistochemistry (Immunohistochemistry-paraffin sectionsof IHC-P). Results there was no significant difference in PWMT and PWTL between the groups before and after operation. PWMT and PWTL were significantly decreased at each time point after operation in CCI model rats (P0.05), and the expression of Reelin mRNA and Reelin protein in the dorsal horn of spinal cord were significantly decreased after CCI (P0.05). Intrathecal injection of LV-Reelin-RNAi further reduced PWMT and PWTLI. In contrast, the expression of Reelin mRNA and protein in the right dorsal horn of rat spinal cord was further decreased (P0.05). There was no significant effect on the expression of Reelin mRNA and Reelin protein in pain behavior of chronic neuropathic pain rats by intrathecal injection of empty vector (P0.05). Conclusion the expression of Reelin in spinal cord decreased significantly after chronic sciatic nerve compression induced chronic neuropathic pain. Intrathecal injection of LV-Reelin-RNAi can further down-regulate the expression of Reelin in the dorsal horn of spinal cord and aggravate the symptoms of neuropathic pain. Reelin may be involved in the generation and development of chronic compressive neuropathic pain.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R402

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