端粒缩短与P53激活在大鼠耳蜗老化中作用的研究

发布时间：2018-08-28 20:44

【摘要】：目的：研究端粒功能障碍与激活P53途径在大鼠耳蜗老化过程中的可能作用，并初步探讨老年性耳聋可能的发病机理。方法：耳廓反射灵敏的1月龄SD大鼠72只，随机分为两组：生理盐水对照组，D-半乳糖组。D-半乳糖组每日接受颈部皮下注射50g/L的D-半乳糖（500mg/Kg体重），连续注射八周。生理盐水对照组每日接受颈部皮下注射等体积生理盐水，连续注射八周。按注射后3、6、9个月将每组分为三个亚组，每亚组12只。实时定量PCR检测耳蜗组织端粒相对长度与P53基因mRNA表达。Western-blotting检测P53蛋白相对表达水平。结果：给药后3月、6月、9月，各时间点D-半乳糖组分别与生理盐水对照组大鼠相比，端粒相对长度检测差异均无统计学意义，但生理盐水对照组大鼠给药后9月与给药后3月相比，端粒相对长度缩短，差异有统计学意义（P0.05），D-半乳糖组大鼠给药后9月与给药后3月相比，端粒相对长度亦缩短，差异有统计学意义（P0.05）。D-半乳糖组大鼠给药后3月、6月分别与对照组相比，D-半乳糖组P53表达增加，与对照组之间差异有统计学意义（P0.05），给药后9月D-半乳糖组和对照组之间P53表达差异无统计学意义（P0.05），生理盐水组大鼠给药后3月、6月、9月P53表达呈增加趋势，给药后6月、9月与给药后3月相比差异有统计学意义（P0.05）；D-半乳糖组大鼠3月、6月、9月P53表达呈也出现增加趋势，给药后9月与给药后3月相比差异有统计学意义（P0.05）。结论：在内耳组织自然老化过程中有端粒的缩短导致其功能障碍的参与，并有抑癌基因P53的激活，说明端粒-P53途径可能参与了ARHL的病程。D-半乳糖诱导的内耳拟老化模型与对照组相比端粒相对长度无明显改变，但P53有一定范围的激活，而端粒缩短与细胞分裂次数有关，因此，我们认为P53表达水平改变可能出现在端粒改变之前，端粒缩短可能需要更长的时间，D-半乳糖诱导的内耳拟老化的作用可能通过其它途径影响P53及其下游信号分子发挥作用，而P53的持续激活有一定的限度，会出现一个平台期，可能为D-半乳糖诱导的线粒体DNA突变与P53依赖的细胞修复相互作用的结果。
[Abstract]:Aim: to investigate the possible role of telomere dysfunction and activation of p53 pathway in cochlear aging in rats and to explore the possible pathogenesis of presbycusis. Methods: 72 1-month old SD rats with sensitive auricle reflex were randomly divided into two groups: normal saline control group, D- galactose group. D- galactose group received subcutaneous injection of 50g/L (500mg/Kg body weight) into the neck for 8 weeks. Saline control group received subcutaneous injection of the same volume of saline into the neck for 8 weeks. Each group was divided into three subgroups with 12 rats in each subgroup. The relative length of telomere and the expression of p53 gene mRNA were detected by real-time quantitative PCR. Western-blotting was used to detect the relative expression of p53 protein. Results: there was no significant difference in telomere relative length between Dgalactose group and normal saline control group at 3 months, 6 months and 9 months after administration. However, the relative length of telomere in the control group was shorter than that in the control group (P 0.05). The relative length of telomere was also shorter in the Dgalactose group than in the control group in 9 months after administration and 3 months after administration. The difference was statistically significant (P0.05). The expression of p53 in D- galactose group was higher than that in control group in 3 months and 6 months after administration. There was no significant difference in p53 expression between Dgalactose group and control group in 9 months after administration (P0.05), but the expression of p53 in saline group showed an increasing trend in 3 months, 6 months and 9 months after administration of normal saline, and there was no significant difference in p53 expression between Dgalactose group and control group (P0.05). The expression of p53 in Dgalactose group was also increased in 6 months, September and 3 months after administration (P0.05), and there was a significant difference in p53 expression between 9 months after administration and 3 months after administration (P0.05). Conclusion: telomere shortening may lead to dysfunction and activation of p53 gene during natural aging of inner ear tissue. The results suggest that telomere p53 pathway may be involved in the course of ARHL. D- galactose induced pseudo aging model of inner ear has no significant change in telomere relative length, but p53 is activated to a certain extent, and telomere shortening is related to the number of cell division. We believe that the changes in p53 expression may occur before telomere changes, and telomere shortening may take longer time to induce pseudoaging of the inner ear induced by D-galactose, which may affect p53 and its downstream signaling molecules in other ways. However, the continuous activation of p53 is limited, and there may be a plateau phase, which may be the result of the interaction between mitochondrial DNA mutation induced by D-galactose and p53 dependent cell repair.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R764.35

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