Zhang Hongjun, Fan Shunwu
Department of Spine, Luoyang Orthopaedic Hospital, Luoyang Henan, 471002, P.R.China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2012 Dec;26(12):1435-41.
To investigate the expression and significance of growth-associated protein 43 (GAP-43) in the dorsal root ganglion (DRG) and intervertebral disc in the rat model of intervertebral disc inflammation.
A total of 103 adult male Sprague Dawley rats (weighing, 200-250 g) were randomly divided into the experimental group (n = 48), the control group (n = 48), and the blank control group (n = 7). Fluoro-gold (F-G) as tracer was injected into the L6, 6 intervertebral disc of 3 groups; after 7 days of F-G injection, complete Freund's adjuvant (50 microL) and the same volume of saline were injected in the experimental group (to prepare the model of intervertebral disc inflammation) and the control group, respectively, and the blank control group had no further treatment. After 1, 3, 7, and 14 days, T13-L6 DRG and L5, 6 intervertebral disc of experimental group and control group were harvested to detect the GAP-43 by using fluorescent immunohistochemistry, in situ hybridization, and RT-PCR. The DRG and intervertebral disc of blank control group were also harvested after 8 days of F-G injection.
Fluorescent immunohistochemistry results showed that the number of F-G-labeled GAP-43 immunoreaction (GAP-43-IR) cells of the DRGs in the experimental group was significantly higher than that in the control group (P < 0.05) at 3 days, and no significant difference was found at the other time points (P > 0.05). There was no significant difference in the cross-sectional area of F-G-labeled GAP-43-IR cells between the experimental group and the control group at each time point (P > 0.05). The co-expression of GAP-43 with calcitonin gene-related peptide (CGRP) and isolectin B4 (IB4)-binding glycoprotein exhibited that the expression of CGRP was 91.4% +/- 7.4% in the control group and was 87.6% +/- 7.8% in the experimental group, showing no significant difference between 2 groups (P > 0.05). There was no IB4-binding glycoprotein expression in GAP-43-IR cells of the DRGs in 2 groups. The expressions of GAP-43, CGRP, and IB4-positive nerve fibers in the intervertebral disc exhibited that the GAP-43-IR nerve fibers in the experimental group were significantly more than that in the control group (P < 0.05), but no significant difference was found in the expression of CGRP between 2 groups (P > 0.05); and there was no IB4-binding glycoprotein expression in GAP-43-IR nerve fibers of the intervertebral disc in 2 group. In situ hybridization and RT-PCR detection showed that the positive expression cells ratio of GAP-43 mRNA and the level of GAP-43 mRNA were significantly higher in the experimental group than in the control group at 1 day (P < 0.05), and no significant difference was found at the other time points (P > 0.05).
Intradiscal inflammatory environment may induce the expression of GAP-43, and potentially promote the nerve fiber ingrowth of rat.
探讨生长相关蛋白43（GAP-43）在椎间盘炎大鼠模型的背根神经节（DRG）和椎间盘中的表达及意义。
将103只成年雄性Sprague Dawley大鼠（体重200-250 g）随机分为实验组（n = 48）、对照组（n = 48）和空白对照组（n = 7）。向3组大鼠的L6、7椎间盘内注射荧光金（F-G）作为示踪剂；F-G注射7天后，实验组（制备椎间盘炎模型）和对照组分别注射完全弗氏佐剂（50 μL）和等体积的生理盐水，空白对照组不再进行进一步处理。在1、3、7和14天后，采集实验组和对照组的T13-L6 DRG和L5、6椎间盘，采用荧光免疫组织化学、原位杂交和RT-PCR检测GAP-43。F-G注射8天后，也采集空白对照组的DRG和椎间盘。
荧光免疫组织化学结果显示，实验组DRG中F-G标记的GAP-43免疫反应（GAP-43-IR）细胞数量在3天时显著高于对照组（P < 0.05），其他时间点无显著差异（P > 0.05）。实验组和对照组在各时间点F-G标记的GAP-43-IR细胞横截面积无显著差异（P > 0.05）。GAP-43与降钙素基因相关肽（CGRP）和isolectin B4（IB4）结合糖蛋白的共表达显示，对照组CGRP的表达为91.4%±7.4%，实验组为87.6%±7.8%，两组间无显著差异（P > 0.05）。两组DRG的GAP-43-IR细胞中均无IB4结合糖蛋白表达。椎间盘内GAP-43、CGRP和IB4阳性神经纤维的表达显示，实验组的GAP-43-IR神经纤维明显多于对照组（P < 0.05），但两组间CGRP表达无显著差异（P > 0.05）；两组椎间盘的GAP-43-IR神经纤维中均无IB4结合糖蛋白表达。原位杂交和RT-PCR检测显示，实验组GAP-43 mRNA的阳性表达细胞比例和GAP-43 mRNA水平在1天时显著高于对照组（P < 0.05），其他时间点无显著差异（P > 0.05）。
椎间盘内炎症环境可能诱导GAP-43的表达，并可能促进大鼠神经纤维向内生长。
