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300～350 g,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1,7,14,21 d 4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80 mg/kg,对照组动物每天腹腔注射生理盐水1 mL,连续10 d.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1,7,14,21 d将实验组动物腹腔注射戊巴比妥50 mg/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1 d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1 d其表达最强,之后逐渐下降,但停药后21 d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7 d表达最强,之后逐渐下降,停药后21 d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.
揹景:在內耳毛細胞髮育中,胰島素樣生長因子1是重要的促有絲分裂因子,也是不可缺少的調節因子.細胞外信號調節激酶1/2在哺乳動物前庭器官也有錶達.目的:觀察慶大黴素損傷後豚鼠前庭上皮胰島素樣生長因子1及其受體、細胞外信號調節激酶1/2的錶達和分佈變化.設計:隨機對照實驗.單位:解放軍成都軍區總醫院耳鼻喉科.材料:選用健康成年豚鼠20隻,體質量300～350 g,由解放軍第四軍醫大學實驗動物中心提供.按隨機數字錶法分為對照組(n=4)和實驗組(n=16),實驗組又按停藥後1,7,14,21 d 4箇時間點進行觀察,每箇時間點4隻.方法:實驗于2002-01/2002-05在解放軍第四軍醫大學西京醫院耳鼻喉科實驗室完成.實驗組豚鼠每天腹腔註射慶大黴素80 mg/kg,對照組動物每天腹腔註射生理鹽水1 mL,連續10 d.豚鼠齣現步態不穩或眼震認為造模成功.每天觀察豚鼠的步態、眼震及進食量.分彆于停藥後1,7,14,21 d將實驗組動物腹腔註射戊巴比妥50 mg/kg過度痳醉後,取齣併剪開雙側聽泡,去除鐙骨底闆,固定、脫鈣、包埋.取與蝸軸平行方嚮連續切片.對照組動物于停止註射生理鹽水後1 d採取相同措施.囌木精-伊紅染色觀察前庭上皮形態改變,免疫組織化學法觀察胰島素樣生長因子1及其受體和細胞外信號調節激酶1/2的錶達及分佈變化.主要觀察指標:①各組動物慶大黴素損傷後行為學和前庭上皮形態學的變化.②胰島素樣生長因子1及其受體、細胞外信號調節激酶1/2的錶達及分佈變化.結果:納入的20隻豚鼠全部進入結果分析.①慶大黴素註射期間,所有豚鼠的前庭功能均受損,停藥後未經任何治療,其前庭功能均部分恢複.②註射慶大黴素後,前庭上皮形態學改變明顯,但停止註射慶大黴素1週後明顯改善.③對照組豚鼠胰島素樣生長因子1及其受體有低水平錶達,慶大黴素損傷後停藥1 d其錶達最彊,之後逐漸下降,但停藥後21 d其錶達仍高于對照組.各組間胰島素樣生長因子1及其受體錶達灰度值比較差異明顯(F=51.8,45.7,P＜0.05).二者變化規律基本一緻.④對照組豚鼠細胞外信號調節激酶1/2有低水平錶達.慶大黴素損傷後其錶達逐漸增彊,停藥後7 d錶達最彊,之後逐漸下降,停藥後21 d其錶達仍高于對照組.各組間細胞外信號調節激酶1/2錶達比較差異明顯(F=103.7,106.4,P＜0.05).二者變化規律相近.結論:慶大黴素損傷後豚鼠前庭上皮胰島素樣生長因子1及其受體,細胞外信號調節激酶1/2錶達增加.胰島素樣生長因子1可能是內源性的促有絲分裂劑,通過徬分泌或自分泌的方式在豚鼠前庭毛細胞脩複的早期髮揮重要作用.細胞外信號調節激酶1/2可能在慶大黴素損傷後豚鼠前庭毛細胞自髮脩複中髮揮重要的信號轉導作用.
배경:재내이모세포발육중,이도소양생장인자1시중요적촉유사분렬인자,야시불가결소적조절인자.세포외신호조절격매1/2재포유동물전정기관야유표체.목적:관찰경대매소손상후돈서전정상피이도소양생장인자1급기수체、세포외신호조절격매1/2적표체화분포변화.설계:수궤대조실험.단위:해방군성도군구총의원이비후과.재료:선용건강성년돈서20지,체질량300～350 g,유해방군제사군의대학실험동물중심제공.안수궤수자표법분위대조조(n=4)화실험조(n=16),실험조우안정약후1,7,14,21 d 4개시간점진행관찰,매개시간점4지.방법:실험우2002-01/2002-05재해방군제사군의대학서경의원이비후과실험실완성.실험조돈서매천복강주사경대매소80 mg/kg,대조조동물매천복강주사생리염수1 mL,련속10 d.돈서출현보태불은혹안진인위조모성공.매천관찰돈서적보태、안진급진식량.분별우정약후1,7,14,21 d장실험조동물복강주사무파비타50 mg/kg과도마취후,취출병전개쌍측은포,거제등골저판,고정、탈개、포매.취여와축평행방향련속절편.대조조동물우정지주사생리염수후1 d채취상동조시.소목정-이홍염색관찰전정상피형태개변,면역조직화학법관찰이도소양생장인자1급기수체화세포외신호조절격매1/2적표체급분포변화.주요관찰지표:①각조동물경대매소손상후행위학화전정상피형태학적변화.②이도소양생장인자1급기수체、세포외신호조절격매1/2적표체급분포변화.결과:납입적20지돈서전부진입결과분석.①경대매소주사기간,소유돈서적전정공능균수손,정약후미경임하치료,기전정공능균부분회복.②주사경대매소후,전정상피형태학개변명현,단정지주사경대매소1주후명현개선.③대조조돈서이도소양생장인자1급기수체유저수평표체,경대매소손상후정약1 d기표체최강,지후축점하강,단정약후21 d기표체잉고우대조조.각조간이도소양생장인자1급기수체표체회도치비교차이명현(F=51.8,45.7,P＜0.05).이자변화규률기본일치.④대조조돈서세포외신호조절격매1/2유저수평표체.경대매소손상후기표체축점증강,정약후7 d표체최강,지후축점하강,정약후21 d기표체잉고우대조조.각조간세포외신호조절격매1/2표체비교차이명현(F=103.7,106.4,P＜0.05).이자변화규률상근.결론:경대매소손상후돈서전정상피이도소양생장인자1급기수체,세포외신호조절격매1/2표체증가.이도소양생장인자1가능시내원성적촉유사분렬제,통과방분비혹자분비적방식재돈서전정모세포수복적조기발휘중요작용.세포외신호조절격매1/2가능재경대매소손상후돈서전정모세포자발수복중발휘중요적신호전도작용.
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.
