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目的通过检测细胞生长抑制和凋亡相关的DNA损伤蛋白在大鼠原代培养海马细胞中的表达与原代培养海马细胞DNA损伤的相关性,探讨铅的神经毒理作用机制。方法原代培养的海马神经细胞,于培养7 d全量更换培养液后加入不同浓度醋酸铅0.2、1.0、10μmol/L,置于37℃、5%CO2培养箱继续培养24 h;蛋白免疫印迹(west-ern blot)方法检测生长抑制DNA损伤诱导因子(GADD 153)表达;彗星实验检测大鼠海马原代培养神经细胞DNA损伤程度。结果与对照组比较,随着染铅浓度增高,原代培养的海马神经细胞中的DNA的尾距(OTM)和彗尾值增大,0.2、1.0μmol/L染铅组GADD 153蛋白表达较弱,染铅浓度为10μmol/L时,表达明显增加。结论细胞DNA损伤与GADD 153蛋白表达呈一定相关性。
OBJECTIVE: To investigate the neurotoxic effect mechanism of lead by detecting the relationship between cell growth inhibition and apoptosis-related DNA damage proteins in rat primary cultured hippocampal cells and the DNA damage of primary cultured hippocampal cells. Methods Primary cultures of hippocampal neurons were cultured in full medium for 7 days. Different concentrations of lead acetate (0.2, 1.0, 10μmol / L) were added into culture medium and incubated at 37 ℃ in 5% CO 2 incubator for 24 hours. Western blotting -ern blot assay was used to detect the expression of growth-inhibitory DNA damage-inducing factor (GADD 153). The DNA damage of hippocampal primary cultured neurons was detected by comet assay. Results Compared with the control group, the DNA tail length (OTM) and the tail of the DNA in primary cultured hippocampal neurons increased with the lead concentration increasing. The expression of GADD 153 protein in 0.2 and 1.0 μmol / L lead-exposed groups was significantly higher than that of the control group Weak, lead concentration was 10μmol / L, the expression was significantly increased. Conclusion DNA damage and GADD 153 protein expression are related to each other.