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| 自噬激活对带状疱疹后神经痛模型小鼠脊髓背角GABA能神经元凋亡的影响 |
| Effects of autophagy activation on the apoptosis of GABA neurons in the dorsal horn of postherpetic neuralgia model mice |
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| DOI：10.12089/jca.2018.03.017 |
| 中文关键词: 带状疱疹后神经痛 GABA能神经元 凋亡 自噬 小鼠 |
| 英文关键词: Postherpetic neuralgia GABA neurons Apoptosis Autophagy Mice |
| 基金项目:广西自然科学基金项目(2016GXNSFAA380172) |
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| 中文摘要: |
| 目的 探讨不同自噬程度对带状疱疹后神经痛（postherpetic neuralgia,PHN）模型小鼠脊髓背角GABA能神经元凋亡的影响。方法 使用SPSS 19.0的随机数字发生器将48只昆明小鼠, 6～8周龄, 体重18～22 g, 随机分为树脂毒素+自噬诱导剂组(PHN+Rapa组)、树脂毒素组(PHN组)、树脂毒素+自噬抑制剂组(PHN+3-MA组)和空白对照组(C组), 每组12只。C组不做任何处理, 其余各组均腹腔注射0.2 μg/g树脂毒素(resiniferatoxin, RTX)制备PHN模型。在构建模型成功后, PHN+Rapa组给予1 μg·kg-1·d-1的自噬诱导剂雷帕霉素(rapamycin, Rapa), PHN组给予生理盐水, PHN+3-MA组给予2 μg·kg-1·d-1的自噬抑制剂3-MA, 连续14 d腹腔注射。检测机械缩足阈值(mechanical withdrawal threshold, MWT)、热缩足潜伏期(thermal withdrawal latency, TWL)至稳定后处死各组小鼠, 采用荧光Tunel法检测脊髓凋亡细胞数;提取脊髓L4～6节段组织, 采用Western blot法检测抑凋亡蛋白bcl-2、促凋亡Bax和自噬相关蛋白LC3的相对表达量;免疫荧光标记脊髓背角GABA能神经元数。结果 与C组比较, PHN+Rapa组、PHN组和PHN+3-MA组的LC3-II/I和Bax的蛋白相对表达量均明显升高, 凋亡细胞数量明显增多, bcl-2蛋白相对表达量明显降低, 脊髓背角GABA能神经元数明显减少(P＜0.05);与PHN组比较, PHN+Rapa组的LC3-II/I比值和Bax的蛋白相对表达量, 凋亡细胞数量明显增多, bcl-2蛋白相对表达量明显降低, 脊髓背角GABA能神经元数明显减少(P＜0.05)。与PHN组比较，PHN+3-MA组的LC3-II/I比值和Bax的蛋白相对表达量明显降低, 凋亡细胞数量明显减少, bcl-2蛋白相对表达量明显升高, 脊髓背角GABA能神经元数明显增多(P＜0.05)。结论 自噬过度激活可能是导致带状疱疹后神经痛脊髓背角GABA能神经元凋亡的机制之一。 |
| 英文摘要: |
| Objective To investigate the effects of different degrees of autophagy on the apoptosis of GABAergic neurons in spinal dorsal horn of postherpetic neuralgia model mice. Methods Forty-eight Kunming mice, approximately 6-8 weeks of age and weighing 18-22 g, were randomly divided into four groups by a random digital generator of SPSS 190: resinotoxin+autophagy induction group (group PHN+Rapa), resinotoxin group (group PHN), resinotoxin+autophagy inhibitor group (group PHN+3-MA) and blank control group (group C), 12 mice in each group. Group C was given no treatment, and the other groups were given intraperitoneal injection of 0.2 μg/g resiniferatoxin (RTX) to prepare PHN model. After successful model establishment, group PHN+Rapa was given Rapamycin (1 μg·kg-1·d-1), physiological saline was given to group PHN, group PHN+3-MA was given 2 μg·kg-1·d-1 autophagy inhibitor 3-MA. The intraperitoneal injection was continued for 14 day. The mechanical withdrawal threshold (MWT) and the latent period of thermal withdrawal latency (TWL) were detected and the mice were killed after stability. The segments of L4-6 spinal cord were extracted and the relative expressions of bcl-2, Bax and autophagy-associated protein LC3 were detected by western blot. Detection of the number of apoptotic cells in the spinal cord by fluorescence Tunel. The number of GABA intermediate neurons in the dorsal horn of the spinal cord was labeled by immunofluorescence. Results Compared with group C, the relative expression level of Bax protein increased significantly, the LC3-II/I ratio and the number of apoptotic cells increased significantly, the relative expression level of Bcl-2 protein and the number of GABA neurons in spinal dorsal horn were significantly reduced in other groups (P＜0.05). Compared with group PHN, the LC3-II/I ratio and the relative expression level of Bax protein, the number of apoptotic cells increased significantly, the relative expression level of Bcl-2 protein and the number of GABA neurons in the spinal dorsal horn were significantly decreased of group PHN+Rapa (P＜0.05). Compared with group PHN, the relative expression level of LC3-II/I ratio and Bax protein, the number of apoptotic cells decreased significantly, the relative expression level of Bcl-2 protein and the number of GABA neurons in spinal dorsal horn increased significantly of group PHN+3-MA, (P＜0.05). Conclusion Over activation of autophagy may be one of the mechanisms leading to the apoptosis of GABA neurons in the dorsal horn of the spinal cord in postherpetic neuralgia. |
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