Objective To study the protective effects by NT3 gene transfection on postnatal mouse cochlear explant cultures treated with gentamicin. Methods Cultured cochlear explants prepared from postnatal mice were transfected using a NT3-expressing recombinant adenovirus. One day later, gentamicin was added into the organotypic cultures at the concentration of 3 raM for an additional three days of incubation before the cochlear explants were fixed. Finally, the neurofila-ments of the cultures were labeled by the means of immunofluorescence and their densities examined. Results The cultured cochlear explants grew well and were properly transfected by the recombinant adenovirus. The neurofilament density (spiral ganglion peripheral neurites per 100 |I,m) was 16.67±1.63 for those treated with gentamicin only, 16.17±2.31 for those transfected with Ad-DsRed2 and treated with gentamicin, 40.33 ±1.63 for those transfected with Ad-DsRed2-NT3 and treated with gentamicin, and 43.17±0.75 for the controls with no transfection or gentamicin treatment. When the gentamicin treatment only group was compared to the blank control group and the Ad-DsRed2-NT3 transfected group, and when the Ad-DsRed2-NT3 transfected group was compared to the Ad-DsRed2 transfected group, the differences were statistically significant (P<0.05). Conclusion Gentamicin can induce degeneration of spiral ganglion. Overexpression of NT3 can protect spiral ganglion from this ototoxicity.%目的 将含有目的基因NT3的重组腺病毒Ad-DsRed2-NT3转染新生小鼠的离体耳蜗基底膜培养组织,观察其在预防庆大霉素的耳毒性中的作用.方法 取新生小鼠的耳蜗基底膜离体培养1d后,用含有目的基因NT3的重组腺病毒Ad-DsRed2-NT3转染新生小鼠的离体耳蜗基底膜培养组织,转染成功1d后用终浓度为3mM的庆大霉素对离体耳蜗基底膜造模,造模3d后,行离体耳蜗基底膜的神经微丝免疫荧光染色,观察各组耳蜗基底膜神经微丝的密度,以观测NT3对离体耳蜗基底膜上的螺旋神经的保护作用.结果 新生小鼠的离体耳蜗基底膜生长良好,重组腺病毒Ad-DsRed2-NT3能对其有效转染,庆大霉素造模后3d各组耳蜗基底膜神经微丝的密度(每100m距离内的螺旋神经微丝数)分别为(±s):造模组16.67±1.63、Ad-DsRed2组16.17±2.31、Ad-DsRed2-NT3组40.33±1.63、空白组为43.17±0.75,其中,造模组与空白组、Ad-DsRed2-NT3组与造模组、Ad-DsRed2-NT3组与Ad-DsRed2组之间具有显著性统计学差异(P＜0.05).结论 终浓度为3mM的庆大霉素对小鼠离体耳蜗基底膜上的螺旋神经有明显的损害作用,而NT3基因的过表达对这种损害具有保护作用.
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