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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 investigate the relationship between DNA damage caused by patulin (PAT) and intracellular ROS levels and lysosomes, and to explore the mechanism of PAT genotoxicity. Methods HEK-293 cells were used as the experimental system. The single cell gel electrophoresis was used to detect the DNA damage. ROS levels were detected by 2 ’, 7’-dichlorodihydrofluorescein, and lysosomal membrane stability was measured by acridine orange. NAC And NH4Cl on lysosomal protection intervention; the use of NAC, NH4Cl, gastric inhibitory peptide PAT DNA damage. RESULTS: After treated with PAT for 1 h, DNA strand breaks, changes in lysosomal membrane stability and ROS levels were induced by different concentrations. The stability of lysosomal membranes induced by PAT was enhanced by NH4Cl and NAC pretreatment. NAC and gastric inhibitory peptide intervention, PAT caused DNA strand breaks almost completely blocked. Conclusions Patulin can cause DNA strand breaks in HEK-293 cells, and its mechanism may be related to the lysosomal pathway. Lysosome may be one of the biological targets of patulin cytotoxicity and may be induced by ROS Destruction of body membrane stability.