Induction of Autophagy and Autophagic Cell Death in Damaged Neural Tissue After Acute Spinal Cord Injury in Mice
Induction of Autophagy and Autophagic Cell Death in Damaged Neural Tissue After Acute Spinal Cord Injury in Mice
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DOI：
10.1097/brs.0b013e3182028c3a
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发表时间：
2011-10-15
期刊：
影响因子：
3
通讯作者：
Itoi, Eiji
中科院分区：
文献类型：
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作者：
Kanno, Haruo;Ozawa, Hiroshi;Itoi, Eiji
研究设计.采用免疫组化和Western印迹法检测了小鼠脊髓损伤（SCI）后轻链3（LC 3）的表达，LC 3是自噬的特征性标志物。电子显微镜分析也进行了检查的解剖形成的自噬和自噬细胞死亡的损伤脊髓。从生物化学和解剖学两方面研究SCI后受损神经组织中自噬的活性。自噬是细胞内蛋白质和细胞器的大量降解，与许多疾病有关。自噬可导致非凋亡的程序性细胞死亡，称为自噬性细胞死亡。最近的研究表明，在各种疾病模型的受损组织中，LC 3的表达增加，自噬和自噬性细胞死亡的解剖形成。然而，以前的研究集中在细胞凋亡过程中，而不是自噬活性作为SCI后神经组织损伤的机制。到目前为止，还没有研究检查LC 3的表达和自噬的解剖形成后SCI。在成年雌性C57 BL/6 J小鼠中，在T10时半横断脊髓。免疫组化和Western blot检测LC 3的表达。用电镜观察自噬活性的解剖结构。免疫组化结果显示，LC 3阳性细胞数量在半切后明显增加。LC 3阳性细胞从4 h开始增加，3d达高峰，并持续至少21 d。在神经元、星形胶质细胞和少突胶质细胞中观察到LC 3阳性细胞。Western blot分析显示，损伤脊髓中LC 3-II蛋白表达水平显著升高。电镜下可见损伤细胞自噬泡形成增多。此外，转移酶介导的dUTP缺口末端标记阳性细胞表达LC 3的细胞核呈圆形，这与自噬性细胞死亡一致，并且它们既不像凋亡细胞核那样皱缩也不碎裂。本研究从生物化学和解剖学两个方面表明，SCI后损伤的神经组织中自噬被明显激活，并诱导自噬性细胞死亡。
Study Design. Expression of light chain 3 (LC3), a characteristic marker of autophagy, was examined by immunohistochemistry and Western blot using a spinal cord injury (SCI) model in mice. Electron microscopic analysis was also performed to examine the anatomic formation of autophagy and autophagic cell death in the injured spinal cord.Objective. To examine both biochemically and anatomically the activity of autophagy in the damaged neural tissue after SCI.Summary of Background Data. Autophagy is the bulk degradation of intracellular proteins and organelles, and it is involved in a number of diseases. Autophagy can lead to nonapoptotic programmed cell death, which is called autophagic cell death. Recent researches have revealed the increased expression of LC3 and the anatomic formation of autophagy and autophagic cell death in damaged tissues of various disease models. However, previous studies have focused on apoptotic process but not autophagic activity as mechanism of neural tissue damage after SCI. To date, there has been no study to examine the expression of LC3 and the anatomic formation of autophagy after SCI.Methods. The spinal cord was hemitransected at T10 in adult female C57BL/6J mice. The LC3 expression was examined by immunohistochemistry and Western blot. The anatomic formation of autophagic activity was investigated using electron microscopy.Results. Immunohistochemistry showed that the number of the LC3-positive cells significantly increased at the lesion site after hemisection. The increase of LC3-positive cells was observed from 4 hours and peaked at 3 days, and it lasted for at least 21 days after hemisection. The LC3-positive cells were observed in neurons, astrocytes, and oligodendrocytes. Western blot analysis demonstrated that the level of LC3-II protein expression significantly increased in the injured spinal cord. Electron microscopy showed the formation of autophagic vacuoles to increase in the damaged cells. Furthermore, the nuclei in the transferase-mediated dUTP nick end labeling-positive cells expressed LC3 were round, which is consistent with autophagic cell death, and they were neither shrunken nor fragmented as is observed in apoptotic nuclei.Conclusion. This study suggested both biochemically and anatomically that autophagy was clearly activated and autophagic cell death was induced in the damaged neural tissue after SCI.