1、慶大霉素對耳蝸毛細胞和傳出神經損害定量分析研究生： 馬 靜導 師： 隋 軍 教授 昆明醫學院第三附屬醫院耳鼻咽喉頭頸外科中 文 詳 細 摘 要目的 慶大霉素(gentamicin，GM) 由于其性質穩定、抗菌譜廣、抗菌殺菌力強、使用方法簡便、價格便宜等優點，至今仍是世界范圍內，尤其是發展中國家成為最常使用的抗生素。致命弱點是具有嚴重的耳毒和腎毒性，常可造成腎功衰竭、聽力障礙、平衡失調等。其耳毒性主要是由于其在內耳的蓄積，導致內耳毛細胞損傷。本文主要研究慶大霉素慢性耳中毒豚鼠內側橄欖耳蝸（media olivochlear, MOC）傳出神經形態和功能的改變，印證慶大霉素耳中毒時傳出神經和毛細胞

2、損害之間的相關性，為臨床治療藥物性神經性耳聾提供新的實驗依據。材料和方法 72只耳廓反射靈敏的健康花色豚鼠(體重300400g)隨機分成實驗組和對照組，實驗組采取肌肉注射慶大霉素150mg/kg，對照組肌肉注射生理鹽水2ml，連續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染色，硝酸銀染色，全耳蝸鋪片觀察健康對照組和實驗組耳蝸MOC傳出神經和OHC的分布特征，并測量耳蝸MOC傳出神經纖維末梢及OHC的數量，電鏡下觀察毛細胞和神經纖維的形態，實驗前分別行畸變產物耳聲發射和聽性腦干反應測聽檢測。 SPSS10.0軟件行統計學分析。結果 （1）對照組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、二回傳出神經分布差別不大，第三、第四回傳出神經末梢逐漸減少，底回神經纖維較粗，而頂回纖維較細。（2）慶大霉素停藥后2周，鉤回、第一回至第四回神經纖維數量無明顯變化；4周后，鉤回、第一回神經纖維大部分消失，第二至第四回神經纖維部分消失；10周后，鉤回和第一回神經纖維絕大部分消失，僅殘留少量的神經末梢，第二回至第四回的神經纖維數量也明顯減少。損傷在各回都有發現，但主要發生在第一回和第二回。（3）對照組耳蝸毛細胞呈三排排列，分布均勻。慶大霉素組外毛細胞的損害程度因觀察時間和部位而異，停藥后2周后開始出現OHC散在或灶性缺乏，隨觀察時,間延長而逐漸加重，

4、10周時最明顯,首先損傷基底回外毛細胞，從基底向頂部發展，病變程度由重到輕，三排外毛細胞中以第一排細胞受損最早，逐漸向外排發展，在本實驗中各回內毛細胞未見明顯損傷。（4）實驗組畸變產物耳聲發射幅值隨觀察的時間逐漸下降，10周后耳聲發射未引出；聽性腦干反應測聽潛伏期延長，閾值降低。(5)實驗組停藥2周后電鏡下外毛細胞形態發生改變，細胞質水腫，線粒體腫脹并伴有線粒體嵴消失，隨著時間的延長病變程度呈遞增趨勢,內毛細胞未見明顯異常；實驗組停藥4周后，神經纖維發生改變，表現為軸突內可見髓樣結構和較多空泡；10周后出現髓鞘分離、傳導功能嚴重受損。結論 慶大霉素耳中毒時毛細胞的缺失早于內側橄欖耳蝸傳出神經系

5、統神經損害，兩者損害的特征具有一致性，MOC神經損害在慶大霉素耳中毒中發揮著重要作用。畸變產物耳聲發射可作為一個較早反映外毛細胞功能的生理監測指標，并具有長期、穩定、安全的效果。 關鍵詞 慶大霉素 耳毒性 傳出神經 毛細胞 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 preyers reflex weigh

9、ted 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. The MO

10、C 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 the

11、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 gyrus

12、 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 afer

13、 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 weeks of

14、 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 prol

15、ong, 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 thresho

16、lds 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 l