顺铂对小鼠耳蜗钙蛋白酶表达的影响

发布时间：2018-05-22 13:43

  本文选题：顺铂 + 耳蜗　； 参考：《辽宁医学院》2012年硕士论文

【摘要】：目的 建立成年BALB/c小鼠顺铂耳毒性模型，研究顺铂对小鼠耳蜗钙蛋白酶（calpain）表达的影响，探讨calpain在顺铂耳毒性中的可能作用，为临床防治顺铂耳毒性聋提供实验依据。 方法 65只健康成年BALB/c小鼠随机分成对照组、顺铂2.5mg/kg组、顺铂3.5mg/kg组、顺铂4.5mg/kg组和顺铂5.5mg/kg组，每组13只，给予腹腔连续注射5d，每天1次。应用免疫荧光染色方法、显微图像分析以及蛋白质印迹技术检测小鼠耳蜗中calpain1和calpain2的表达；同时结合听脑干反应（auditory brainstem response，ABR）测试，观察用药前后小鼠听力的变化。 结果 1．腹腔连续注射5d后，对照组小鼠体重略有增加，而不同剂量顺铂组小鼠体重从第3d起均出现不同程度减少，与对照组比较均有显著性差异（P＜0.05，P＜0.01）；并且随着顺铂给药剂量的增大，小鼠体重减少愈明显（P＜0.05，P＜0.01）。 2．腹腔连续注射5d后，在各刺激频率下，不同剂量顺铂组小鼠ABR阈移均明显增大，与对照组比较均有显著性差异（P＜0.01）；并且随着顺铂给药剂量的增大，ABR阈移亦显著增大，呈现明显的量效关系（P＜0.01）。 3．免疫荧光染色及显微图像分析结果显示，对照组小鼠耳蜗外毛细胞、螺旋神经节和血管纹中均有calpain1和calpain2的微弱表达；不同剂量顺铂组小鼠耳蜗上述各部位中calpain1和calpain2的表达均较对照组明显增强（P＜0.05，P＜0.01），但顺铂组间calpain1的表达无明显差异，而calpain2的表达则随顺铂给药剂量的增大而明显增强（P＜0.01）。 4．蛋白质印迹检测及半定量分析结果显示，对照组小鼠耳蜗calpain1和calpain2的蛋白表达水平较低；不同剂量顺铂组calpain1和calpain2的蛋白表达水平均较对照组明显增高（P＜0.01），但顺铂组间calpain1的蛋白表达水平无显著性差异，而calpain2的蛋白表达水平则随顺铂给药剂量的增大而明显增强（P＜0.01）。 结论 应用成年BALB/c小鼠能建立可靠的顺铂耳毒性模型。正常小鼠耳蜗中仅有少量calpain表达，顺铂可增强小鼠耳蜗中calpain1和calpain2的表达，并且随着顺铂给药剂量的增加，，calpain2的表达亦逐渐增强，提示calpain1和calpain2介导了顺铂的耳毒性损伤，并且calpain2在顺铂耳毒性损伤中可能起主导作用。
[Abstract]:Purpose The ototoxicity model of adult BALB/c mice was established to study the effect of cisplatin on the expression of calpainin in cochlea of mice, to explore the possible role of calpain in cisplatin ototoxicity, and to provide experimental evidence for clinical prevention and treatment of cisplatin ototoxic deafness. Method 65 healthy adult BALB/c mice were randomly divided into control group, cisplatin 2.5mg/kg group, cisplatin 3.5mg/kg group, cisplatin 4.5mg/kg group and cisplatin 5.5mg/kg group. The expression of calpain1 and calpain2 in cochlea of mice was detected by immunofluorescence staining, microimage analysis and Western blotting, and the hearing changes of mice before and after treatment were observed by combining with auditory brainstem response (ABR) test. Result 1. After 5 days of continuous intraperitoneal injection, the body weight of the control group increased slightly, but the weight of the mice in the different doses of cisplatin group decreased in different degrees from the 3rd day, there was significant difference compared with the control group (P < 0.05 P < 0.01), and with the increase of the dosage of cisplatin, the dosage of cisplatin increased. The weight loss of mice was more obvious (P < 0.05) and P < 0.01 (P < 0.01). 2. After 5 days of continuous intraperitoneal injection, the ABR threshold shift of mice in different doses of cisplatin increased significantly, compared with the control group (P < 0.01), and the threshold shift increased with the increase of the dosage of cisplatin. There was a significant dose-effect relationship (P < 0.01). 3. The results of immunofluorescence staining and microscopic image analysis showed that there were weak expressions of calpain1 and calpain2 in the outer hair cells of cochlea, spiral ganglion and stria vascularis in the control group. The expression of calpain1 and calpain2 in the cochlea of mice with different doses of cisplatin was significantly enhanced than that of the control group (P < 0.05 P < 0.01), but there was no significant difference in the expression of calpain1 among the cisplatin groups, while the expression of calpain2 increased significantly with the increase of the dosage of cisplatin (P < 0.01). 4. The results of Western blotting and semi-quantitative analysis showed that the expression of calpain1 and calpain2 in cochlea of control mice was low. The protein expression levels of calpain1 and calpain2 in different dose groups were significantly higher than those in the control group (P < 0.01), but there was no significant difference in the protein expression level of calpain1 between the cisplatin groups, but the protein expression level of calpain2 increased significantly with the increase of the dosage of cisplatin (P < 0.01). Conclusion A reliable ototoxicity model of cisplatin could be established in adult BALB/c mice. Only a small amount of calpain was expressed in the cochlea of normal mice. Cisplatin could enhance the expression of calpain1 and calpain2 in the cochlea of mice, and the expression of calpain2 increased with the increase of the dosage of cisplatin, suggesting that calpain1 and calpain2 mediated the ototoxicity of cisplatin. And calpain2 may play a leading role in cisplatin ototoxicity.
【学位授予单位】：辽宁医学院
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R764

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