【摘要】 目的研究丹参(salvia miltiorrhiza,SM)对庆大霉素(gentamicin,GM)耳中毒豚鼠耳蜗核因子-κ B(nuclear factor-kappa B,NF-κB)活性的影响,探讨SM对GM耳毒性损伤的防护作用及其分子机制。方法豚鼠随机分成正常对照组、GM组、GM+SM组和SM组,应用免疫组织化学SP法检测耳蜗组织中NF-κBP65和P50的活性;同时结合电生理指标听脑干反应(auditory brainstem response,ABR)测试,观察用药前后豚鼠听阈的变化。结果GM组ABR阈值比用药前明显提高,与正常对照组比较,差异有显著性,并且高于GM+SM组。各组NF-κ B P65阳性免疫反应仅见于耳蜗螺旋韧带,而其他部位均呈阴性。GM组、GM+SM组及SM组耳蜗P65阳性免疫反应的分布和正常对照组相同,反应活性与正常对照组比较差异无显著性。各组NF-κ B P50阳性免疫反应见于耳蜗螺旋神经节、神经纤维、螺旋韧带、血管纹、骨螺旋板缘及螺旋器。GM组耳蜗螺旋韧带和血管纹的NF-κ B P50活性较正常对照组明显增强;而同时给予SM的豚鼠,其耳蜗螺旋韧带和血管纹的NF-κ B P50活性较GM组则明显减弱。正常对照组和SM组耳蜗外毛细胞胞浆可见P50阳性染色,胞核则未见染色。给予GM后,耳蜗外毛细胞胞核可见P50阳性染色,而联合应用SM后,外毛细胞核染色消失,在阴性对照切片上未见NF-κ B P50阳性染色。通过直线相关检验证明豚鼠耳蜗螺旋韧带和血管纹NF-κ B P50免疫反应的平均灰度值和ABR阈值二者呈负相关。结论正常豚鼠耳蜗中有NF-κ B P65和P50的表达。注射GM后,NF-κB被诱导激活,进而造成听力损害。SM通过显著抑制GM所致NF-κB的高活性,从而有效拮抗GM的耳毒性,这可能是丹参发挥防护作用的分子机制之一。更多还原

【Abstract】 【Objectives】 To investigate the effect of salvia miltiorrhiza (SM) on gentamicin(GM)-induced activity of nuclear factor-kappa B (NF-κB) in the cochlea of guinea pig, and to explore the protective role and molecular mechanism of SM on GM-induced totoxicity. 【Methods】 Guinea pigs were randomly assigned to four groups: normal control,GM alone,GM plus SM and SM alone. SP method of immunohistochemistry was applied to detect the expression of NF-κB P65 and NF-κB P50,combined with auditory brainstem response (ABR) measurement to observe the changes of auditory threshold.【Results】 Compared with normal controls, ABR thresholds were elevated significantly in GM alone group.Co-treatment with SM resulted in significant reduction of ABR thresholds as compared with GM alone. Immunoreactivity for NF-κB P65 was only observed in the spiral ligament in the cochlea of normal control. Localization of NF-κB P65 in the cochlea of guinea pigs from other three groups was similar to that of con-trol.There were no obvious difference among them. Immunoreactivity for NF-κB P50 was detected in the spiral ligament, stria vascularis, spiral ganglion, nerve fiber, limbus laminae spiralis osseae and spiral organ in the cochlea of normal control.Localization of NF-κB P50 in the cochlea of guinea pigs from GM and GM+SM groups was similar to that of control.But the activity of NF-κB P50 in spiral ligament and stria vascularis in GM group was greater than that in normal control.Whereas the activity of NF-κB P50 in GM+SM group was lower than that in GM group. NFκB P50 immunostaining was observed in cytoplasm but not in nuclei of the outer hair cells in both normal control and SM alone.After treatment with GM,immunostaining with NF-κB P50 was present in nuclei of outer hair cells of GM.However, concomitant injection of SM was no nuclear immunostaining for P50.There was no positive staining of NF-κB P50 in the sections of negative control.Linear correlation test was applied to analyze the average gray of NFκB P50 and ABR threshold,and negative correlation was gained.【Conclusion】 Expression of NF-κB P65 and NFκB P50 was observed in the cochlea in normal guinea pigs. After treatment with GM,NF-κB was activated and hearing was injured.SM could effectively attenuate GM ototoxicity by inhibiting the activity of NF-κB induced by GM, this might be one of the molecular mechanism by which SM could protect against GM ototoxicity.更多还原