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豚鼠前庭上皮胰岛素样生长因子1及其受体与细胞外信号调节激酶1/2表达在庆大霉素损伤后的变化
引用本文:盛宏申,何跃,徐光,孙景豫,王锦玲. 豚鼠前庭上皮胰岛素样生长因子1及其受体与细胞外信号调节激酶1/2表达在庆大霉素损伤后的变化[J]. 中国组织工程研究与临床康复, 2007, 11(36): 7302-7305
作者姓名:盛宏申  何跃  徐光  孙景豫  王锦玲
作者单位:1. 解放军成都军区总医院耳鼻喉科,四川省成都市,610083
2. 解放军成都军区昆明总医院耳鼻喉科,云南省昆明市,650032
3. 解放军第四军医大学西京医院耳鼻喉科,陕西省西安市,710032
摘    要:背景:在内耳毛细胞发育中,胰岛素样生长因子1是重要的促有丝分裂因子,也是不可缺少的调节因子。细胞外信号调节激酶1/2在哺乳动物前庭器官也有表达。目的:观察庆大霉素损伤后豚鼠前庭上皮胰岛素样生长因子1及其受体、细胞外信号调节激酶1/2的表达和分布变化。设计:随机对照实验。单位:解放军成都军区总医院耳鼻喉科。材料:选用健康成年豚鼠20只,体质量300~350g,由解放军第四军医大学实验动物中心提供。按随机数字表法分为对照组(n=4)和实验组(n=16),实验组又按停药后1,7,14,21d4个时间点进行观察,每个时间点4只。方法:实验于2002-01/2002-05在解放军第四军医大学西京医院耳鼻喉科实验室完成。实验组豚鼠每天腹腔注射庆大霉素80mg/kg,对照组动物每天腹腔注射生理盐水1mL,连续10d。豚鼠出现步态不稳或眼震认为造模成功。每天观察豚鼠的步态、眼震及进食量。分别于停药后1,7,14,21d将实验组动物腹腔注射戊巴比妥50mg/kg过度麻醉后,取出并剪开双侧听泡,去除镫骨底板,固定、脱钙、包埋。取与蜗轴平行方向连续切片。对照组动物于停止注射生理盐水后1d采取相同措施。苏木精-伊红染色观察前庭上皮形态改变,免疫组织化学法观察胰岛素样生长因子1及其受体和细胞外信号调节激酶1/2的表达及分布变化。主要观察指标:①各组动物庆大霉素损伤后行为学和前庭上皮形态学的变化。②胰岛素样生长因子1及其受体、细胞外信号调节激酶1/2的表达及分布变化。结果:纳入的20只豚鼠全部进入结果分析。①庆大霉素注射期间,所有豚鼠的前庭功能均受损,停药后未经任何治疗,其前庭功能均部分恢复。②注射庆大霉素后,前庭上皮形态学改变明显,但停止注射庆大霉素1周后明显改善。③对照组豚鼠胰岛素样生长因子1及其受体有低水平表达,庆大霉素损伤后停药1d其表达最强,之后逐渐下降,但停药后21d其表达仍高于对照组。各组间胰岛素样生长因子1及其受体表达灰度值比较差异明显(F=51.8,45.7,P<0.05)。二者变化规律基本一致。④对照组豚鼠细胞外信号调节激酶1/2有低水平表达。庆大霉素损伤后其表达逐渐增强,停药后7d表达最强,之后逐渐下降,停药后21d其表达仍高于对照组。各组间细胞外信号调节激酶1/2表达比较差异明显(F=103.7,106.4,P<0.05)。二者变化规律相近。结论:庆大霉素损伤后豚鼠前庭上皮胰岛素样生长因子1及其受体,细胞外信号调节激酶1/2表达增加。胰岛素样生长因子1可能是内源性的促有丝分裂剂,通过旁分泌或自分泌的方式在豚鼠前庭毛细胞修复的早期发挥重要作用。细胞外信号调节激酶1/2可能在庆大霉素损伤后豚鼠前庭毛细胞自发修复中发挥重要的信号转导作用。

关 键 词:胰岛素样生长因子1  胰岛素样生长因子1受体  细胞外信号调节激酶1/2  前庭上皮  庆大霉素
文章编号:1673-8225(2007)36-07302-04
修稿时间:2006-12-202007-05-17

Expression changes of insulin-like growth factor-1 and its receptor as well as extracellular signal regulated kinase 1/2 in the vestibular epithelium of guinea pigs following gentamycin toxicity
Sheng Hong-shen,He Yue,Xu Guang,Sun Jing-yu,Wang Jin-ling. Expression changes of insulin-like growth factor-1 and its receptor as well as extracellular signal regulated kinase 1/2 in the vestibular epithelium of guinea pigs following gentamycin toxicity[J]. Journal of Clinical Rehabilitative Tissue Engineering Research, 2007, 11(36): 7302-7305
Authors:Sheng Hong-shen  He Yue  Xu Guang  Sun Jing-yu  Wang Jin-ling
Abstract:BACKGROUND:Insulin-like growth factor-1 (IGF-1) is an important mitogen and an indispensable regulator during normal hair cell development. Extracellular signal regulated kinase 1/2 (ERK1/2) is also expressed in mammalian vestibular organs.OBJECTIVE: To investigate the changes of the expression and the distribution of IGF-1, IGF-1 receptor (IGF-1R) and ERK1/2 in guinea pig's vestibular epithelium following gentamycin injury.DESIGN: A randomly controlled study.SETTING: Department of Otorhinolaryngology, General Hospital of Chengdu Military Area Command of Chinese PLA.MATERIALS: Twenty healthy and adult guinea pigs, weighing 300 to 350 g, provided by the Experimental Animal Center,Fourth Military Medical University of Chinese PLA, were randomly divided into four experimental groups and a control group with 4 in each group.METHODS: The experiment was performed at Department of Otorhinolaryngology Research Laboratory of Xijing Hospital,Fourth Military Medical University of Chinese PLA from January 2002 to May 2002. In the experimental groups,gentamycin was intraperitoneally administered daily (80 mg/kg) for 10 consecutive days. For the control group, 1 mL of 0.9% sterile saline was administered in the same way. The guinea pigs, in which unsteady gait or nystagmus appeared,could be involved in the further experiments. Before being sacrificed, the animals' gait, nystagmus and the amount of food-intake were observed every day. The animals were anesthetized deeply with 50 mg/kg pentobarbital 1, 7, 14 or 21days after the last gentamycin administration and then decapitated. Bilateral otic vesicles were harvested and immersed in 4% paraformaldehyde. Then the specimens were decalcified in 10% ethylenediamine tetraacetic acid (EDTA),thereafter embedded in OCT gel and serially sectioned at a thickness of 10 μm in the cryostat. The behavioral changes of guinea pigs were observed. The morphological changes of vestibular epithelium were examined by hematoxylin and eosin (HE) staining. The expression and distribution changes of IGF-1/1R and ERK1/2 were investigated immunohistochemically. MAIN OUTCOME MEASURES: The behavioral, morphological, expression and distribution changes of IGF-1/1R and ERK1/2 of guinea pigs in each group.RESULTS: Twenty involved guinea pigs entered the stage of final analysis. ① All the guinea pigs' vestibular functions were impaired after gentamycin treatment, then partially restored without any pharmaceutical treatment. ②Morphological injury was obvious after gentamycin treatment, and improved 1 week later. ③The expression of IGF-1/1R was low in the control group, and in the 1-day group it increased significantly to its maximum. After that, its expression decreased gradually, but was still higher in the 21-day group than that in the control group. There were significant differences in the expression of IGF-1/1R among different groups (F =51.8,45.7,P < 0.05). The expression changes of IGF-1 and its receptor were similar. ④The expression of ERK1/2 was low in the control group, and increased gradually after gentamycin toxicity. In the 7-day group, the immunoreactivity reached its maximum. Then the levels of ERK1/2 decreased gradually, but were still higher in the 21-day group than that in the control group. There were significant differences in the expression of ERK1/2 among different groups (F =103.7,106.4, P < 0.01), but no significant differences existed in the expression between ERK1 and ERK2 among different groups (P > 0.05).CONCLUSION: The expression of IGF-1/1R and ERK1/2 in vestibular epithelium increased after gentamycin treatment,and IGF-1 may play an important role as an endogenous mitogen through a paracrine or autocrine manner in the early stage of hair cell self-repair of guinea pigs. ERK1/2 may play an important role in signal transduction during vestibular hair cell self-repair after gentamycin toxicity in guinea pigs.
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