CAJ | 학술논문

目的：了解年龄相关性听力损失小鼠耳蜗毛细胞的凋亡方式，探讨半胱氨酸蛋白酶（caspase）抑制剂z－VAD－FMK [z － Val － Ala － Asp （Ome）－ fluoromethylketone]防治老年性聋的效果。方法选用新生NMF308nmf／nmf小鼠14只和成年同窝NMF308nmf／nmf小鼠32只，分为新生小鼠腹腔注射组（14只）、成年小鼠腹腔注射组（14只）和成年小鼠圆窗膜注射组（18只），各组又分为治疗组（注射z－VAD －FM K ）和对照组[注射二甲基亚枫（dimethylsulfoxide ，DMSO）]，采用圆窗膜局部和腹腔注射两种方法给药，给药前后行ABR检测，用免疫荧光染色化学技术TUNEL、caspase－3和PI（碘化丙啶）染色标记耳蜗毛细胞，观察各组耳蜗毛细胞的凋亡和存活状况。结果NMF308nmf／nmf小鼠从1月龄开始发生听力减退和毛细胞功能改变，到2月龄时，caspase－3激活表达的毛细胞凋亡现象是毛细胞凋亡出现的最早表现；TUNEL阳性标记特征稍晚出现；PI标记可见毛细胞细胞核固缩和碎片出现的时间从2月龄开始；到3月龄时该种小鼠听力基本丧失，耳蜗毛细胞严重缺失；而应用z－VAD －FM K治疗后，各治疗组小鼠ABR反应阈明显好于对照组；耳蜗毛细胞凋亡数目较对照组减少，存活数明显较对照组多，尤以圆窗膜注射组明显。结论 NMF308nmf／nmf小鼠耳蜗毛细胞的死亡方式以caspase－3凋亡途径为主，应用caspase－3抑制剂z－VAD －FM K定向内耳给药，可以阻断caspase－3信号途径激活所导致的毛细胞凋亡，从而有效防治老年性聋。
목적：료해년령상관성은력손실소서이와모세포적조망방식，탐토반광안산단백매（caspase）억제제z－VAD－FMK [z － Val － Ala － Asp （Ome）－ fluoromethylketone]방치노년성롱적효과。방법선용신생NMF308nmf／nmf소서14지화성년동와NMF308nmf／nmf소서32지，분위신생소서복강주사조（14지）、성년소서복강주사조（14지）화성년소서원창막주사조（18지），각조우분위치료조（주사z－VAD －FM K ）화대조조[주사이갑기아풍（dimethylsulfoxide ，DMSO）]，채용원창막국부화복강주사량충방법급약，급약전후행ABR검측，용면역형광염색화학기술TUNEL、caspase－3화PI（전화병정）염색표기이와모세포，관찰각조이와모세포적조망화존활상황。결과NMF308nmf／nmf소서종1월령개시발생은력감퇴화모세포공능개변，도2월령시，caspase－3격활표체적모세포조망현상시모세포조망출현적최조표현；TUNEL양성표기특정초만출현；PI표기가견모세포세포핵고축화쇄편출현적시간종2월령개시；도3월령시해충소서은력기본상실，이와모세포엄중결실；이응용z－VAD －FM K치료후，각치료조소서ABR반응역명현호우대조조；이와모세포조망수목교대조조감소，존활수명현교대조조다，우이원창막주사조명현。결론 NMF308nmf／nmf소서이와모세포적사망방식이caspase－3조망도경위주，응용caspase－3억제제z－VAD －FM K정향내이급약，가이조단caspase－3신호도경격활소도치적모세포조망，종이유효방치노년성롱。
Objective In this study ,we investigated the apoptosis of hair cell in the cochlea of age -related hearing loss(AHL) generated by ENU mutagenesis ,and to study a pan caspase inhibitor (z-VAD -FMK) which is to protect the cochlea hair cells from hearing loss induced by age-related hearing loss(AHL) .Methods Through z-VAD-FMK intraperitoneal injection and round window membrane (RWM) drug were delivered into the Cdh23 nmf308 nmf/nmf mice 5(postnatal days 2 -32) inner ear .ResuIts The results showed that the nmf308 mice with progressive hair cell loss along a base to apex gradient with age-related hearing loss .The cochlear OHCs reduced from 5% ~10% at 1 month to 100% at 3 month in the basal region .Substantial amounts of TUNEL -positive OHCs nuclei appeared at 1 month age ,and activated caspase-3 labeling demonstrated that most OHCs appeared at 2 months age .These suggested that DNA single strand break was attributed primarily to apoptosis of cochlear le_sions ,whereas in the later stage of lesion ,the expansion led to activation of caspase-3 activity reduced with further progression of nuclear condensation in age-related hearing loss .ConcIusion The addition of a pan caspase inhibitor (z -VAD -FMK ) significantly protected the cochlea against the hair cell loss induced by apoptosis .Our study showed that aspase inhibitor ,Z-VAD-FMK appeared to play a prominent role in age-related hearing loss media_ted hair cell death loss induced by apoptosis .Our study showed that aspase inhibitor ,Z-VAD -FMK appeared to play a prominent role in age-related hearing loss mediated hair cell death .