特异性损伤内耳螺旋神经元小鼠模型的建立

目的：建立小鼠内耳螺旋神经元损伤的耳聋模型，为研究干细胞移植治疗感音神经性聋奠定基础。方法成年雌性SPF级CBA／J小鼠60只，随机分为实验组和生理盐水组，每组30只，实验组动物经圆窗渗透给予10μl哇巴因（ouabain），生理盐水组同法给予等量生理盐水。每组于给药前及给药后7、14及30天分别检测小鼠听性脑干反应（ABR）和畸变产物耳声发射（DPOAE），并用免疫组织荧光技术和基底膜铺片技术分别观察耳蜗螺旋神经元（spiral ganglion neurons ，SGNs）细胞及内耳毛细胞（hair cells ，HCs）的变化。结果①与生理盐水组比较，实验组给药后各时间点 ABR反应阈均明显升高，波Ⅰ潜伏期延长，振幅明显降低，差异有统计学意义（ P＜0．05）。②实验组及生理盐水组小鼠DPOAE均可正常引出。③与生理盐水组相比，实验组耳蜗各回螺旋神经元的数量及密度显著降低，差异有统计学意义（P＜0．05）。④实验组在给药后不同时间点，耳蜗各回内、外毛细胞均排列整齐、形态完整、未见明显损伤或丢失。结论哇巴因经圆窗渗透给药可特异性损伤CBA／J小鼠内耳螺旋神经元及其功能，而不损伤内、外毛细胞，是一种理想的研究干细胞移植治疗感音神经性聋的动物模型。

An Animal Model of Specific Damage to Spiral Ganglion Neurons in the Inner Ear

Objective To establish an animal model of specific loss of spiral ganglion neurons in the mouse in‐ner ear ,and to lay the foundation for the study of stem cell transplantation in the treatment of sensorineural hearing loss .Methods Sixty adult female SPF grade CBA /J mice were randomly divided into experimental group and nor‐mal saline group ,with 30 mice in each .The animals in the experimental group received 0 .5 mM ouabain via the round window membrane ,while the normal saline group used normal saline .Auditory brainstem responses (ABR) and distortion product otoacoustic emissions (DPOAE) were recorded before the administration ,7 days ,14 days and 30 days after the administration respectively .Meanwhile ,immunofluorescence histochemical staining was used to detect spiral ganglion neurons(SGNs)damages ,and basilar membrane preparation was used to observe the chan‐ges of hair cells .Results ABR:compared with those of in the normal saline group ,in the experimental group ,the thresholds were elevated ,the Wave I latency was prolonged ,and the amplitude reduced .The differences were sta‐tistically significant (P<0 .05) .DPOAE :DPOAE was normally recorded in the experimental group and the control group ,without significant differences between the 2 groups .The immunofluorescence results show that compared with the normal saline group ,the number and density of SGNs in the cochlea of the experimental group were signifi‐cantly decreased ,and the differences between the 2 groups were statistically significant (P<0 .05) .The basilar membrane preparation showed that at different time points after the administration of ouabain ,the inner and outer hair cells in the experiment group cochlear were intact ,no damage or loss were observed .Conclusion The applica‐tion of ouabain to the mouse inner ear via the round window membrane can specifically induce the damages of SGNs , but spare the inner and outer hair cells ,thus ,it is an ideal animal model for the research of stem cell transplantation treatment of sensorineural hearing loss .