慶大霉素對(duì)耳蝸毛細(xì)胞和傳出神經(jīng)損害定量分析

1、慶大霉素對(duì)耳蝸毛細(xì)胞和傳出神經(jīng)損害定量分析研究生： 馬 靜導(dǎo) 師： 隋 軍 教授 昆明醫(yī)學(xué)院第三附屬醫(yī)院耳鼻咽喉頭頸外科中 文 詳 細(xì) 摘 要目的 慶大霉素(gentamicin，GM) 由于其性質(zhì)穩(wěn)定、抗菌譜廣、抗菌殺菌力強(qiáng)、使用方法簡(jiǎn)便、價(jià)格便宜等優(yōu)點(diǎn)，至今仍是世界范圍內(nèi)，尤其是發(fā)展中國(guó)家成為最常使用的抗生素。致命弱點(diǎn)是具有嚴(yán)重的耳毒和腎毒性，?？稍斐赡I功衰竭、聽(tīng)力障礙、平衡失調(diào)等。其耳毒性主要是由于其在內(nèi)耳的蓄積，導(dǎo)致內(nèi)耳毛細(xì)胞損傷。本文主要研究慶大霉素慢性耳中毒豚鼠內(nèi)側(cè)橄欖耳蝸（media olivochlear, MOC）傳出神經(jīng)形態(tài)和功能的改變，印證慶大霉素耳中毒時(shí)傳出神經(jīng)和毛細(xì)胞

2、損害之間的相關(guān)性，為臨床治療藥物性神經(jīng)性耳聾提供新的實(shí)驗(yàn)依據(jù)。材料和方法 72只耳廓反射靈敏的健康花色豚鼠(體重300400g)隨機(jī)分成實(shí)驗(yàn)組和對(duì)照組，實(shí)驗(yàn)組采取肌肉注射慶大霉素150mg/kg，對(duì)照組肌肉注射生理鹽水2ml，連續(xù)10天。兩組豚鼠分別于停藥后2周、4周、10周采用乙酰膽堿酯酶組織化學(xué)染色，硝酸銀染色，全耳蝸鋪片觀察健康對(duì)照組和實(shí)驗(yàn)組耳蝸MOC傳出神經(jīng)和OHC的分布特征，并測(cè)量耳蝸MOC傳出神經(jīng)纖維末梢及OHC的數(shù)量，電鏡下觀察毛細(xì)胞和神經(jīng)纖維的形態(tài)，實(shí)驗(yàn)前分別行畸變產(chǎn)物耳聲發(fā)射和聽(tīng)性腦干反應(yīng)測(cè)聽(tīng)檢測(cè)。 SPSS10.0軟件行統(tǒng)計(jì)學(xué)分析。結(jié)果 （1）對(duì)照組MOC纖維及末梢分布均勻

3、，比較密集，在耳蝸各回的分布不一致。鉤回、第一、二回傳出神經(jīng)分布差別不大，第三、第四回傳出神經(jīng)末梢逐漸減少，底回神經(jīng)纖維較粗，而頂回纖維較細(xì)。（2）慶大霉素停藥后2周，鉤回、第一回至第四回神經(jīng)纖維數(shù)量無(wú)明顯變化；4周后，鉤回、第一回神經(jīng)纖維大部分消失，第二至第四回神經(jīng)纖維部分消失；10周后，鉤回和第一回神經(jīng)纖維絕大部分消失，僅殘留少量的神經(jīng)末梢，第二回至第四回的神經(jīng)纖維數(shù)量也明顯減少。損傷在各回都有發(fā)現(xiàn)，但主要發(fā)生在第一回和第二回。（3）對(duì)照組耳蝸毛細(xì)胞呈三排排列，分布均勻。慶大霉素組外毛細(xì)胞的損害程度因觀察時(shí)間和部位而異，停藥后2周后開(kāi)始出現(xiàn)OHC散在或灶性缺乏，隨觀察時(shí),間延長(zhǎng)而逐漸加重，

4、10周時(shí)最明顯,首先損傷基底回外毛細(xì)胞，從基底向頂部發(fā)展，病變程度由重到輕，三排外毛細(xì)胞中以第一排細(xì)胞受損最早，逐漸向外排發(fā)展，在本實(shí)驗(yàn)中各回內(nèi)毛細(xì)胞未見(jiàn)明顯損傷。（4）實(shí)驗(yàn)組畸變產(chǎn)物耳聲發(fā)射幅值隨觀察的時(shí)間逐漸下降，10周后耳聲發(fā)射未引出；聽(tīng)性腦干反應(yīng)測(cè)聽(tīng)潛伏期延長(zhǎng)，閾值降低。(5)實(shí)驗(yàn)組停藥2周后電鏡下外毛細(xì)胞形態(tài)發(fā)生改變，細(xì)胞質(zhì)水腫，線粒體腫脹并伴有線粒體嵴消失，隨著時(shí)間的延長(zhǎng)病變程度呈遞增趨勢(shì),內(nèi)毛細(xì)胞未見(jiàn)明顯異常；實(shí)驗(yàn)組停藥4周后，神經(jīng)纖維發(fā)生改變，表現(xiàn)為軸突內(nèi)可見(jiàn)髓樣結(jié)構(gòu)和較多空泡；10周后出現(xiàn)髓鞘分離、傳導(dǎo)功能?chē)?yán)重受損。結(jié)論 慶大霉素耳中毒時(shí)毛細(xì)胞的缺失早于內(nèi)側(cè)橄欖耳蝸傳出神經(jīng)系

5、統(tǒng)神經(jīng)損害，兩者損害的特征具有一致性，MOC神經(jīng)損害在慶大霉素耳中毒中發(fā)揮著重要作用?；儺a(chǎn)物耳聲發(fā)射可作為一個(gè)較早反映外毛細(xì)胞功能的生理監(jiān)測(cè)指標(biāo)，并具有長(zhǎng)期、穩(wěn)定、安全的效果。 關(guān)鍵詞 慶大霉素 耳毒性 傳出神經(jīng) 毛細(xì)胞 定量The quantitative analysis of hair cells and efferents in guinea pigs with chronic gentamicin ototoxicityPostgraduate: Majing Tutor: Prof. Suijun The ENTHead and Neck Surgery Department o

6、f Third Affiliated Hospital of Kunming Medical CollegeAbstractObjective It is well known that the gentamicin(GM) have the side-effects of nephrotoxicity and Ototoxicity. And the GM often gives rise to nephric function decline, balance maladjustment. Even though, they are still widely used worldwide,

7、 especially in the developing country. GM can cause the hair cell injury because the chacter of toxicity accumulation in internal ear. To detect the importance of the medial olivochlear efferent (MOC) in chronic aminoglycoside ototoxicity, the morphological changes and quantitative analysis of the M

8、OC efferent in the cochlea of guinea pig and their correlations with the impairment of the outer hair cells (OHC) are observed. The article can provide original experimental proof in treatment of nerve deafness. Materials and Methods Seventy two healthy guinea pigs with positive preyer's reflex

9、weighted from the 300 to400 grams are randomly assigned to two groups: the normal control group and experimental group. The experimental guinea pigs are injected gentamicin. The dosage is 150mg-1kg-1d, one time a day and it need 10 days. The control guinea pigs are injected 2ml physiologic saline. T

10、he MOC efferent are examined using the histochemical stain for acetylcholinesterase(AchE) and the numbers of OHC were numerateding with silver nitrate staining on the surface preparation. The amorphous of MOC efferent and hair cell are observed with electron microscope. The guinea pigs are performed

11、 the inspection of distortion production otoacoustic emissions and auditory brainstem response audiometry before experiment. The numerical data were statistically analyzed by SPSS10.0 software. Results The MOC efferent fiber and periphery distribute intensively and nerve distribution differ in each

12、gyrus in control group. The nerve of gyrus uncinatus have no difference with its of the first and second gyrus. The nerve periphery of third 、fourth gyrus decrease gradually. The nerve fibers of bottom gyrus are thickness, while parietal gyrus are fine; The nerve of every gyrus have no marked change

13、 afer two weeks of the gentaminic administration. The most of gyrus uncinatus and first gyrus nerve disappear after four weeks of the gentaminic administration. The uncinatus and first gyrus nerve disappear completely and nerve fiber decrease obviously from second gyrus to fourth gyrus after ten wee

14、ks of the gentaminic administration. The nerve injured in each gyrus, especially in the uncinatus and first gyrus. The outer hair cells align three rows and distribute evenly. The lesion of outer hair cell happen two weeks after the gentaminic administration and aggravate gradually as observing time

15、 prolong, especially ten weeks. The outer hair cell of bottom gyrus are injured firstly and first row cell are injured earlier. The inner hair cell are normal.（4）The numeric value of distortion production otoacoustic emissions decreased and latencies of auditive brain response(ABR) lengthened and th

16、resholds decreased in experimental group.（5）The outer hair cell happen cytoplasm and chondrosome edema using electron microscope observation after two weeks of the gentaminic administration . The lesion aggravate gradually as observing time prolong. The nerve fiber axon emerge myelin figure and more vacuoles after four weeks of the gentaminic administration. Medullary sheath abstraction appear in ten weeks after gentaminic administration and nerve conduction injure seriously. Conclusion The results suggest that the OHC lesion is earlier than MOC efferent impairment and the character of le