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硫酸软骨素酶ABC治疗减少大鼠脊髓损伤引起的酪氨酸蛋白激酶A4的表达

刘喜民^*, 康红钰, 徐纪伟, 孙丹华

河南漯河医学高等专科学校，漯河 462002

摘要

本研究旨在探讨大鼠脊髓损伤(spinal cord impairment, SCI)后硫酸软骨素酶ABC (chondroitinase ABC, ChABC)对酪氨酸蛋白激酶A4 (ephrin A4, EphA4)表达变化的影响。选取成年雌性SD大鼠，随机分为假手术组、生理盐水(NS)组和ChABC组。NS组和ChABC组大鼠行脊髓半切损伤术，并通过蛛网膜下腔分别注射NS和ChABC；假手术组仅接受背部切开术，没有半切脊髓。SCI后1、3、7、14、21 d取损伤段脊髓样品，采用免疫荧光染色和免疫印迹检测各组EphA4表达；SCI后3 d采用免疫荧光双标法检测损伤段脊髓内生长相关蛋白-43 (growth associated protein 43, GAP-43)和胶质纤维酸性蛋白(glial fibrillary acidic protein, GFAP)的表达情况。结果显示，术后1、3、7 d，NS 组EphA4阳性神经元数明显少于假手术组(P < 0.01)，而术后14、21 d，ChABC组和NS组EphA4阳性神经元多于假手术组(P < 0.05)。值得注意的是，在术后各时间点，ChABC组EphA4阳性神经元数均明显低于NS组 (P < 0.01)。免疫印迹结果与免疫荧光术所得结果一致。术后 3 d，各组GFAP标记胶质细胞数之间相比较，假手术组最多，NS组次之，ChABC组最少；另外，各组均未出现GAP-43阳性神经元。以上结果提示，SCI后，ChABC可以降低神经元EphA4的表达，并减少EphA4阳性神经元周围的胶质细胞数，从而改善脊髓的内环境，有利于神经轴突的生长与延伸。

关键词： 脊髓损伤; 硫酸软骨素酶ABC; 酪氨酸蛋白激酶A4

分类号：R322.81; R642

LIU Xi-Min^*, KANG Hong-Yu, XU Ji-Wei, SUN Dan-Hua

Luohe Medical College, Luohe 462002, China

Abstract

The aim of the study was to investigate the effect of chondroitinase ABC (ChABC) on ephrin A4 (EphA4) expression after spinal cord impairment (SCI) in rats. Adult female SD rats were randomly divided into three groups: ChABC group, normal saline (NS) group and sham group. In the ChABC and NS group, the SCI model was produced by the spinal cord hemisection. The rats in sham group received sham operation without the spinal hemisection. ChABC and NS groups were intrathecally injected with ChABC and normal saline, respectively. At different time points after SCI, injured region of spinal cord was taken out as sample. The levels of EphA4 expression were measured by immunofluorescence technique and Western blot. And the expressions of growth associated protein 43 (GAP-43) and glial fibrillary acidic protein (GFAP) were detected using double immunofluorescent staining. Immunofluorescent results showed that, compared with that in sham group, the EphA4 expression was significantly down-regulated on 1, 3 and 7 d after SCI, then up-regulated on 14 and 21 d after SCI in NS group. In ChABC group, the level of EphA4 expression was significantly less than that in the NS group during the whole time after SCI. Western blot showed an identical result to that of immunofluorescent staining. The double labeling results showed that on 3 d after SCI, the number of GFAP, glial cells marker, positive cells in NS group was lower than that in sham group, but higher than that in ChABC group. Moreover, GAP-43 was not detected in all three groups. These results suggest that ChABC can decrease the expression level of EphA4 and reduce the number of astrocytes after SCI, thus improving microenvironment of the injured region and promoting axonal growth and extension.

Key words: chondroitinase ABC; spinal cord injury; ephrin A4

收稿日期：2011-03-07　　录用日期：2011-07-26

通讯作者：刘喜民　　E-mail:

引用本文：

刘喜民, 康红钰, 徐纪伟, 孙丹华. 硫酸软骨素酶ABC治疗减少大鼠脊髓损伤引起的酪氨酸蛋白激酶A4的表达[J]. 生理学报 2011; 63 (6): 498-504.

LIU Xi-Min, KANG Hong-Yu, XU Ji-Wei, SUN Dan-Hua. . Acta Physiol Sin 2011; 63 (6): 498-504 (in Chinese with English abstract).