Department of Neurosurgery, Chang Gung Memorial Hospital, Keelung, Taiwan.
Spine (Phila Pa 1976). 2012 Mar 15;37(6):470-5. doi: 10.1097/BRS.0b013e318221e859.
Experimental, controlled trial, animal study.
To assess autophagy expression after rat spinal cord injury (SCI) and investigate the effect of methylprednisolone treatment on autophagy.
Although it is evident that SCI induces necrosis and apoptosis, its relationship to autophagy is uncertain. Autophagy is implicated in various pathological states in the nervous system, such as neurodegenerative diseases, cerebral ischemia, and traumatic brain injury. Up to now, no autophagy expression was evidenced by transmission electronic microscope (TEM) and the autophagy marker, microtubule-associated protein light chain 3 (LC3) in neural tissue after SCI.
Sixty-six Sprague-Dawley rats were used for the experimental procedure. In the SCI group, laminectomy at T9 were performed, followed by impactor contusion of the spinal cord. In the sham group, only a laminectomy was performed without contusion. We used Western blot to analyze LC3 at 2 hours, 4 hours, 1 day, 3 days, and 7 days after SCI. We also investigated the effect of methylprednisolone on autophagy expression of contused spinal cord. Cellular localization and ultrastructural changes after spinal cord injury were compared with those sham-operated rats using immunofluorescent double labeling and TEM, respectively. Data from the Western blot were analyzed using a nonparametric Kruskal-Wallis test with P < 0.05 being considered significant.
We detected significantly elevated level of LC3 2 hours after SCI, and then the level declined until 1 week after SCI. Methylprednisolone decreased LC3 expression at 2 hours after SCI. LC3 positive cells were colocalized with neuronal nuclei, but not with glial fibrillary acidic protein. The existence of autophagy and progress of autophagic cell death after SCI were confirmed by TEM.
Through observing the enhanced autophagy expression in neurons soon after contusion injury and the inhibitive effect of methylprednisolone treatment, this study demonstrates the characteristics of autophagy expression after SCI and suggests that autophagic cell death may play a role in neuronal death after spinal cord trauma.
实验性、对照试验、动物研究。
评估大鼠脊髓损伤（SCI）后的自噬表达，并研究甲泼尼龙治疗对自噬的影响。
尽管 SCI 引起坏死和凋亡是显而易见的，但它与自噬的关系尚不确定。自噬参与了神经系统的各种病理状态，如神经退行性疾病、脑缺血和创伤性脑损伤。到目前为止，在 SCI 后神经组织中，还没有通过透射电子显微镜（TEM）和自噬标志物微管相关蛋白轻链 3（LC3）来证明自噬表达。
66 只 Sprague-Dawley 大鼠用于实验过程。在 SCI 组中，T9 椎板切除术，随后对脊髓进行撞击挫伤。在假手术组中，仅行椎板切除术而不进行挫伤。我们使用 Western blot 在 SCI 后 2 小时、4 小时、1 天、3 天和 7 天分析 LC3。我们还研究了甲泼尼龙对挫伤脊髓自噬表达的影响。使用免疫荧光双标记和 TEM 分别比较了脊髓损伤后细胞定位和超微结构的变化。Western blot 数据采用非参数 Kruskal-Wallis 检验分析，P < 0.05 为差异有统计学意义。
我们在 SCI 后 2 小时检测到 LC3 水平显著升高，然后在 SCI 后 1 周内下降。甲泼尼龙在 SCI 后 2 小时降低了 LC3 的表达。LC3 阳性细胞与神经元核共定位，但与神经胶质纤维酸性蛋白不共定位。通过 TEM 证实了 SCI 后自噬的存在和自噬性细胞死亡的进展。
通过观察挫伤损伤后神经元中自噬表达的增强以及甲泼尼龙治疗的抑制作用，本研究表明了 SCI 后自噬表达的特征，并提示自噬性细胞死亡可能在脊髓创伤后神经元死亡中发挥作用。
