论文部分内容阅读
目的:研究探讨马兜铃酸Ⅰ(AAⅠ)对猪肾小管上皮细胞(LLC -PK1细胞)DNA损伤和细胞周期阻滞是否具有可逆性。 方法:采用LLC- PK1细胞为实验对象,选择不同浓度的AAⅠ(80, 320,和1 280ng/ml)体外刺激LLC- PK1细胞24h,然后去除含有AAⅠ的培养基,换不含AAⅠ的培养基继续培养细胞24和48h。同时设不加AAⅠ的细胞为对照组。采用单细胞凝胶电泳(又称彗星实验)检测经不同剂量AAⅠ处理的LLC- PK1细胞不同时间点DNA损伤情况,流式细胞仪技术检测它们对LLC -PK1细胞周期的影响。 结果:AAⅠ(80ng/ml)组细胞彗星实验为阴性,但AAⅠ(320和1 280ng/ml)导致细胞DNA损伤,出现明显的彗星拖尾现象,且细胞在G2 /M期的比例也明显增加,均呈剂量依赖性(P<0. 01 )。用不含AAⅠ的培养基继续培养细胞24和48h后,AAⅠ( 320ng/ml)组彗星拖尾现象明显减轻至消失,G2 /M期细胞的比例与同时间对照组比较无明显差异(P>0 .05 )。而AAⅠ(1280ng/ml)组彗星拖尾现象无明显改善(P<0. 01),G2 /M期细胞的比例仍明显高于同时间正常对照组(P<0 .05)。 结论:AAⅠ可导致LLC- PK1细胞DNA损伤,使细胞周期阻滞在G2 /M期。低剂量AAⅠ造成的DNA损伤可完全恢复,细胞进入正常的细胞周期,但高剂量所致DNA损伤不可逆转。
AIM: To investigate whether aristolochic acid Ⅰ (AAⅠ) is reversible in DNA damage and cell cycle arrest in porcine renal tubular epithelial cells (LLC -PK1). METHODS: LLC-PK1 cells were used as experimental subjects and LLC-PK1 cells were stimulated in vitro with different concentrations of AAⅠ (80, 320, and 1 280 ng / ml) for 24 h, then the medium containing AAⅠ was removed and the medium without AAⅠ Continue to culture cells for 24 and 48 h. At the same time set AAⅠ-added cells as control group. The DNA damage of LLC-PK1 cells treated with different doses of AA was detected by single cell gel electrophoresis (also known as comet assay), and their effects on the cycle of LLC-PK1 cells were examined by flow cytometry. Results: The comet assay of AA Ⅰ (80ng / ml) group was negative, but AA Ⅰ (320 and 1 280ng / ml) resulted in DNA damage and obvious comet tailing, and the proportion of cells in G2 / M phase was also significantly increased , All in a dose-dependent manner (P <0.01). After culturing cells with AAⅠ for 24 and 48h, the tailing phenomenon of comet cells in AAⅠ (320ng / ml) group obviously reduced to disappear, and there was no significant difference in G2 / M phase cells compared with the control group at the same time (P> 0 .05). However, there was no significant improvement in tailing of comet cells in AAⅠ (1280ng / ml) group (P <0.01). The proportion of cells in G2 / M phase was still significantly higher than that in normal control group (P <0.05). CONCLUSION: AAⅠ can cause DNA damage in LLC-PK1 cells and block the cell cycle in G2 / M phase. DNA damage caused by low-dose AAI can be completely restored, cells enter the normal cell cycle, but high-dose DNA damage caused by irreversible.