N-乙酰半胱氨酸对庆大霉素耳毒性损伤的保护作用研究
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摘要
目的：观察庆大霉素耳毒性及N-乙酰半胱氨酸(N-acetylcysteine，NAC)在耳蜗药物性损伤中的作用。
方法：60只健康Wistar大鼠随机分成正常组、对照组、庆大霉素组、NAC干预-1组和NAC干预-2组。庆大霉素组大鼠腹腔注射庆大霉素120 mg／kg／d，连续14d；对照组大鼠腹腔注射等量的生理盐水；NAC干预-1组在给予庆大霉素之前1小时腹腔注射NAC350mg／kg／d；NAC干预-2组在停用庆大霉素后次日开始给予NAC 350mg／kg／d，连续14天。每组大鼠在给药前及最后一次给药后24h检测ABR，并进行结果分析。用HE染色法观察耳蜗各部位的组织学改变；用免疫组织化学法检测耳蜗中核因子-κB(nuclear factor-kappa B，NF-κB)及肿瘤坏死因子-α(tumor necrosis factor-α，TNF-α)的表达，并进行比较和图像分析。
结果：ABR示庆大霉素组阈值为34.6986±11.5396，NAC干预-2组阈值为21.7373±9.9248，与正常组(5.8426±0.7775)之间存在显著性差异(P＜0.05)，NAC干预-1组阈值为11.5102±5.4074，对照组阈值为6.1552±0.9489，与庆大霉素组和NAC干预-2组均存在显著性差异(P＜0.05)，与正常组无显著性差异(P＞0.05)。
HE染色示：庆大霉素组内外毛细胞变形，Corti器内外毛细胞缺损，血管纹变薄；NAC干预组较相对应的庆大霉素组细胞变形、缺损均减轻，血管纹厚度略增加；NAC干预-1组毛细胞结构损害程度较NAC干预-2组轻。
免疫组织化学染色示：NF-κB及TNF-α在正常耳蜗血管纹、螺旋神经节细胞、Corti器的内、外毛细胞、螺旋韧带中有弱阳性表达；在庆大霉素组，NF-κB和TNF-α在血管纹、螺旋神经节和Corti器都存在上调表达。NAC干预能明显抑制此上调表达。
结论：腹腔注射庆大霉素可引起大鼠内耳损伤，NF-κB和TNF-α可能共同参与了耳蜗药物性损伤过程，NAC可能通过抑制氧自由基的产生减轻耳蜗药物性损伤，且在庆大霉素之前用药效果要好于之后用药。
Objective: To investigate the Gentamicin ototoxicity and the effect ofN-acetylcysteine (NAC) in the damage of cochlea.
Methods: Sixty healthy Wistar rats were divided into the normal group, the controlgroup, medicine damage group and group after the intervention of NAC-1/-2.
Group of Gentamicin ototoxicity: peritoneal injection of Gentamicin120mg/kg/d for 14 days; the control group: peritoneal injection of tales dosesSodium Chloride; group of NAC intervention-1: give NAC 350mg/kg 1 hour beforeperitoneal injection of Gentamicin; group of NAC intervention-2: give NAC for 14days the day after the finish of Gentamicin. Detect the ABR before the administrationand 24 hours after the last administration and make an analysis. The histomorphologyof cochlea was observed with hemotoxylineosin (HE) staining. The NF-κB and TNF-αimmunoactivity in cochlea was studied by immunohistochemisty and labelingratio of nuclear factor-kappa B (NF-κB) and tumor necrosis factor-α(TNF-α)expression was studied by image analysis.
Results: The results of ABR showed that the threshold of group of Gentamicinototoxicity was 34.6986±11.5396, the NAC intervention-2 was 21.7373±9.9248, therewas significant difference with the normal group (5.8426±0.7775) (P＜0.05). TheNAC intervention-1 was 11.5102±5.4074 and the control group was 6.1552±0.9489,both had significant difference with the NAC intervention-2 and the group ofGentamicin ototoxicity (P＜0.05), no significant difference with the normal group(P＞0.05).
HE staining showed there were lesions of hair cells in the Gentamicin ototoxicitygroup;Compare with the group of Gentamicin ototoxicity,groups after theintervention of NAC have a less cytomorphosis、defect and thinner stria vascularis.
There was slight positive expression of NF-κB and TNF-αin the hair cell, thestria vascularis, the spiral ligament, the membrane spiralis, the endothelial lining ofcochlear glomeruli and the spiral ganglion. In the Gentamicin ototoxicity groups, atthe same time, the expression of NF-κB and TNF-αin the stria vascularis, thespiral ligament and cochlear glomeruli increased significantly. NAC could inhibitsuch expression obviously.
Conclusion: We can establish the model of Gentamicin ototoxicity in cochlea ofWistar rats by peritoneal injection of Gentamicin. NF-κB and TNF-αmay lead to the cochlea lesions in the Gentamicin ototoxicity together; NAC may inhibit suchlesions by restraining the production of oxygen free radicals, and the earlier the better.
方法：60只健康Wistar大鼠随机分成正常组、对照组、庆大霉素组、NAC干预-1组和NAC干预-2组。庆大霉素组大鼠腹腔注射庆大霉素120 mg／kg／d，连续14d；对照组大鼠腹腔注射等量的生理盐水；NAC干预-1组在给予庆大霉素之前1小时腹腔注射NAC350mg／kg／d；NAC干预-2组在停用庆大霉素后次日开始给予NAC 350mg／kg／d，连续14天。每组大鼠在给药前及最后一次给药后24h检测ABR，并进行结果分析。用HE染色法观察耳蜗各部位的组织学改变；用免疫组织化学法检测耳蜗中核因子-κB(nuclear factor-kappa B，NF-κB)及肿瘤坏死因子-α(tumor necrosis factor-α，TNF-α)的表达，并进行比较和图像分析。
结果：ABR示庆大霉素组阈值为34.6986±11.5396，NAC干预-2组阈值为21.7373±9.9248，与正常组(5.8426±0.7775)之间存在显著性差异(P＜0.05)，NAC干预-1组阈值为11.5102±5.4074，对照组阈值为6.1552±0.9489，与庆大霉素组和NAC干预-2组均存在显著性差异(P＜0.05)，与正常组无显著性差异(P＞0.05)。
HE染色示：庆大霉素组内外毛细胞变形，Corti器内外毛细胞缺损，血管纹变薄；NAC干预组较相对应的庆大霉素组细胞变形、缺损均减轻，血管纹厚度略增加；NAC干预-1组毛细胞结构损害程度较NAC干预-2组轻。
免疫组织化学染色示：NF-κB及TNF-α在正常耳蜗血管纹、螺旋神经节细胞、Corti器的内、外毛细胞、螺旋韧带中有弱阳性表达；在庆大霉素组，NF-κB和TNF-α在血管纹、螺旋神经节和Corti器都存在上调表达。NAC干预能明显抑制此上调表达。
结论：腹腔注射庆大霉素可引起大鼠内耳损伤，NF-κB和TNF-α可能共同参与了耳蜗药物性损伤过程，NAC可能通过抑制氧自由基的产生减轻耳蜗药物性损伤，且在庆大霉素之前用药效果要好于之后用药。
Objective: To investigate the Gentamicin ototoxicity and the effect ofN-acetylcysteine (NAC) in the damage of cochlea.
Methods: Sixty healthy Wistar rats were divided into the normal group, the controlgroup, medicine damage group and group after the intervention of NAC-1/-2.
Group of Gentamicin ototoxicity: peritoneal injection of Gentamicin120mg/kg/d for 14 days; the control group: peritoneal injection of tales dosesSodium Chloride; group of NAC intervention-1: give NAC 350mg/kg 1 hour beforeperitoneal injection of Gentamicin; group of NAC intervention-2: give NAC for 14days the day after the finish of Gentamicin. Detect the ABR before the administrationand 24 hours after the last administration and make an analysis. The histomorphologyof cochlea was observed with hemotoxylineosin (HE) staining. The NF-κB and TNF-αimmunoactivity in cochlea was studied by immunohistochemisty and labelingratio of nuclear factor-kappa B (NF-κB) and tumor necrosis factor-α(TNF-α)expression was studied by image analysis.
Results: The results of ABR showed that the threshold of group of Gentamicinototoxicity was 34.6986±11.5396, the NAC intervention-2 was 21.7373±9.9248, therewas significant difference with the normal group (5.8426±0.7775) (P＜0.05). TheNAC intervention-1 was 11.5102±5.4074 and the control group was 6.1552±0.9489,both had significant difference with the NAC intervention-2 and the group ofGentamicin ototoxicity (P＜0.05), no significant difference with the normal group(P＞0.05).
HE staining showed there were lesions of hair cells in the Gentamicin ototoxicitygroup;Compare with the group of Gentamicin ototoxicity,groups after theintervention of NAC have a less cytomorphosis、defect and thinner stria vascularis.
There was slight positive expression of NF-κB and TNF-αin the hair cell, thestria vascularis, the spiral ligament, the membrane spiralis, the endothelial lining ofcochlear glomeruli and the spiral ganglion. In the Gentamicin ototoxicity groups, atthe same time, the expression of NF-κB and TNF-αin the stria vascularis, thespiral ligament and cochlear glomeruli increased significantly. NAC could inhibitsuch expression obviously.
Conclusion: We can establish the model of Gentamicin ototoxicity in cochlea ofWistar rats by peritoneal injection of Gentamicin. NF-κB and TNF-αmay lead to the cochlea lesions in the Gentamicin ototoxicity together; NAC may inhibit suchlesions by restraining the production of oxygen free radicals, and the earlier the better.
引文
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