摘要
目的研究棒曲霉素(PAT)引起的DNA损伤与细胞内ROS水平以及溶酶体的关系,探讨PAT遗传毒性的机制。方法以HEK-293细胞作为试验系统,单细胞凝胶电泳试验检测DNA损伤情况,2',7'-二氢二氯荧光素检测ROS水平,吖啶橙测定溶酶体膜稳定性;采用NAC和NH4Cl对溶酶体进行保护干预;采用NAC、NH4Cl、抑胃肽干预PAT所致的DNA损伤。结果 PAT作用细胞1 h,不同浓度引起细胞DNA链断裂、溶酶体膜稳定性改变和ROS水平增加;经NH4Cl、NAC预处理,PAT引起的溶酶体膜稳定性均增强了;经NH4Cl、NAC和抑胃肽干预处理,PAT引起的DNA链断裂几乎完全被阻止。结论棒曲霉素可致HEK-293细胞DNA链断裂,其作用机制可能与溶酶体途径有关,溶酶体可能是棒曲霉素细胞毒性的生物靶点之一,并由ROS引起溶酶体膜稳定性的破坏。
Objective To study the relationship between patulin induced DNA damage and intracellular ROS levels,also and the lysosomes,to investigate the PAT genotoxic mechanism.Methods HEK-293 cells were selected as test system.The single cell gel electrophoresis（ SCGE） test for detection of DNA damage and used the 2’,7’-dihydro-dichlorofluorescein（ DCFH） to detect intracellular ROS levels; Acridine orange（ AO） was used to measure the changes of lysosomal membrane stability; the use of N-acetylcysteine（ NAC）and ammonium chloride（ NH4Cl） protection of the lysosomal intervention; NAC,NH4Cl,gastric inhibitory peptide（ pepstatina A） were used to interfere with PAT induced DNA damage.Results With different concentrations after being treated with PAT in HEK-293 cells for 1 h,DNA strand has breaks,the change of lysosomal membrane stability and increase of ROS levels.With of NH4Cl and NAC pretreatment of HEK-293 cells,it was significantly protected cell lysosomal membrane stability.With NH4Cl,NAC and pepstatin A pretreatment of HEK-293 cells,PAT induced DNA strand breaks was almost completely blocked.Conclusion Patulin induced DNA strand breaks in HEK-293cells,and its mechanism may be related to lysosomal signaling pathway.Lysosomes may be one of the biological target of patulin cytotoxicity by ROS caused by the destruction of the lysosomal membrane stability.
出处
《毒理学杂志》
CAS
CSCD
北大核心
2014年第3期203-207,共5页
Journal of Toxicology
