Interleukin-17 expression in the injured site of a rat model of complete spinal cord transection

doi:10.3969/j.issn.2095-4344.2014.18.007

Abstract

Abstract:

BACKGROUND: Intervention using known inflammatory transmitters has limitations on relieving secondary spinal cord injury. Interleukin-17 is an important proinflammatory cytokine, and is gradually paid attention in the pathogenesis of central nervous system diseases.
OBJECTIVE: To investigate the altered rule of interleukin-17 mRNA and protein in a rat model of acute spinal cord injury.
METHODS: Healthy male Sprague-Dawley rats were randomly assigned to two groups. In the model group, rats were made into complete spinal cord transaction models. In the sham surgery group, only spinal dura mater was opened, but parenchyma was not injured. Basso, Beattie, Bresnahan locomotor rating scale was used to observe the effects of acute spinal cord injury on limb motor function of rats. Hematoxylin-eosin staining was used to observe histopathological changes at various time points after spinal cord injury. Real-time fluorescence quantitative PCR and western blotting were used to detect interleukin-17 mRNA and protein levels in each group at various time points after spinal cord injury.
RESULTS AND CONCLUSION: Basso, Beattie, Bresnahan locomotor rating scale: Basso, Beattie, Bresnahan scores were 20 to 21 in the sham surgery group. Basso, Beattie, Bresnahan scores were 0 at 1 and 2 days after spinal cord injury. At 7 days, Basso, Beattie, Bresnahan scores were 0 to 3 (P < 0.05). Hematoxylin-eosin staining results revealed that compared with the sham surgery group, inflammatory cell infiltration, neuronal and glial cell swelling, and a reduced number of neuronal processes were observed at 6 hours after spinal cord injury. Gray matter and white matter were loose and vacuolated at 12 hours. Gliocyte proliferation and tissue fibrosis were apparent at 7 days. Real-time PCR results demonstrated that interleukin-17 mRNA appeared at 3 hours, and peaked at 6 hours (P < 0.01), and then decreased. Interleukin-17 mRNA levels were similar to the sham surgery group at 7 days. Western blotting results revealed that interleukin-17 expression began to increase at 6 hours and peaked at 12 hours (P < 0.05), and then reduced, and reached the levels in the sham surgery group at 7 days. Results indicated that tissue injury was most severe at 12 hours, and showed a time consistency with interleukin-17 expression. It is inferred that interleukin-17 is possibly involved in the process of secondary inflammatory reaction of spinal cord.

中国组织工程研究杂志出版内容重点：肾移植；肝移植；移植；心脏移植；组织移植；皮肤移植；皮瓣移植；血管移植；器官移植；组织工程

全文链接：

Key words: spinal cord injuries, interleukin-17, inflammation, rats, Sprague-Dawley

CLC Number:

R318

Ma Chao, Xu Zhen, Wang Zhuo-qiang, Deng Shi-yuan. Interleukin-17 expression in the injured site of a rat model of complete spinal cord transection[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(18): 2824-2829.

Figures/Tables 2

2.1 实验动物数量分析由于实验取损伤区为中心1 cm的脊髓组织，组织质量不足以同时进行实时荧光定量PCR与Western blot，因此实时荧光定量PCR进入结果分析数量每组8只，Western blot进入结果分析数量每组6只。 2.2 BBB评分评价大鼠脊髓损伤后后肢运动功能变化所有大鼠术前BBB评分均为21分，脊髓损伤后7 d组大鼠均出现了典型的截瘫综合征，表现为双后肢完全瘫痪，腹部贴地，尾部无明显活动，伴有尿潴留、血尿等症状。损伤后1，2 d大鼠BBB评分均为0分，大鼠双后肢呈拖行，足背着地，5 d后部分大鼠后肢运动功能开始恢复，膝或髋关节有轻微活动，BBB评分为0-2分；损伤后7 d，大鼠后肢1或2个关节可有广泛活动，但仍拖行，足背着地，BBB评分为0-3分(P < 0.05，图1)。 2.3 脊髓损伤大鼠局部组织病理改变损伤区为中心 1 cm的脊髓组织苏木精-伊红染色结果：假手术组神经元胞体呈球形、锥形、梭形或星形，细胞形态规则，核大而圆，核异染质少，着色浅(图2A)。脊髓损伤组神经细胞肿大、空泡化、细胞核浓缩、崩解，主要结构消失。脊髓损伤后1 h，损伤区以灰质出血为主，神经元和胶质细胞未见明显改变。脊髓损伤6 h后，出现一些炎性细胞，以中性粒细胞为主，神经元和胶质细胞肿胀，神经元突起减少(图2B)。脊髓损伤12 h后，灰质白质组织结构疏松、空泡化，神经元水肿进一步加剧(图2C)。脊髓损伤后7 d，继发损伤区组织损伤程度较破坏前明显加重，灰、白质结构破坏，细胞凋亡现象逐渐减少，胶质细胞增生，组织纤维化明显(图2D)。 2.4 脊髓损伤大鼠局部组织白细胞介素17 mRNA表达水平采用实时荧光定量PCR技术，检测脊髓损伤大鼠不同时间点白细胞介素17表达变化：假手术组为0.26±0.25，脊髓损伤后1 h组为0.60±0.70，脊髓损伤后3 h组为0.28±0.17，脊髓损伤后6 h组为1.78±1.26，脊髓损伤后12 h组为0.61±0.48，脊髓损伤后24 h组为0.60±0.32，脊髓损伤后72 h组为0.64±0.35，脊髓损伤后7 d组为0.29±0.17。结果表明，与假手术组相比，脊髓损伤后6 h组白细胞介素17 mRNA水平表达显著升高(P < 0.01)，12 h后降低，7 d后表达接近假手术组水平。 2.5 脊髓损伤大鼠局部组织白细胞介素17蛋白表达水平图3结果显示，脊髓损伤6 h后，白细胞介素17表达开始升高，并于损伤后12 h表达出现高峰(P < 0 .05)，24 h后表达逐渐减少，至伤后7 d表达接近假手术组水平。在其他时间点，白细胞介素17蛋白表达与假手术组无明显差异。"

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