霉菌毒素赭曲霉毒素A诱导原代人尿道上皮细胞中DNA的非预定合成。

Induction of unscheduled DNA synthesis in primary human urothelial cells by the mycotoxin ochratoxin A.

作者信息

Dörrenhaus A, Flieger A, Golka K, Schulze H, Albrecht M, Degen G H, Föllmann W

机构信息

Institut für Arbeitsphysiologie an der Universität Dortmund, Germany.

出版信息

Toxicol Sci. 2000 Feb;53(2):271-7. doi: 10.1093/toxsci/53.2.271.

DOI:10.1093/toxsci/53.2.271

PMID:10696775

Abstract

Ochratoxin A (OTA) is a widespread contaminant in human staple food. Exposure of humans to this mycotoxin is a matter of concern because OTA is a known rodent carcinogen. As the urothelium is one target tissue of this mycotoxin, primary cultured human urothelial cells (HUC) from adults and children were used to analyze the induction of unscheduled DNA synthesis (UDS) by OTA. HUC were isolated from the ureters or renal pelves of two nephrectomized adults and of two children with ureteropelvic junction stenosis and cultured under serum-free conditions. After a confluency of 70-80% was reached, cell proliferation was suppressed by arginine-deficient medium (ADM), and UDS was assessed autoradiographically by 3H-thymidine incorporation upon exposure to OTA (10-2000 nM), ethyl methanesulfonate (EMS, 5 mM, positive control), or dimethyl sulfoxide (DMSO, 0.2%, solvent control). In control cultures the level of UDS was low. Exposure to EMS resulted in an induction of UDS (2-to 5-fold compared to control), thus allowing the sensitive detection of repair resulting from induction of DNA lesions in all four specimens, and demonstrating that repair of EMS-induced DNA lesions can take place under the chosen culture conditions. In two HUC cultures derived from adults, a significant induction of UDS was observed in the concentration range of 50-500 nM OTA. The highest fraction of cells in repair (CIR) was found at 50 nM OTA for the HUC from the older male (50% CIR). The maximum response in the other specimens from the adult female and the 7-year-old boy were seen at OTA concentrations of 500 and 250 nM, respectively. In contrast to all other specimens, no significant induction of UDS by OTA was found in the HUC cultures derived from an infant's urothelium. Signs of cytotoxicity were observed above 500 nM OTA in all cultures. The varying susceptibility toward OTA observed in vitro may hint at varying predispositions of individuals in vivo.

摘要

赭曲霉毒素A（OTA）是人类主食中广泛存在的污染物。人类接触这种霉菌毒素令人担忧，因为OTA是一种已知的啮齿动物致癌物。由于尿路上皮是这种霉菌毒素的一个靶组织，因此使用来自成人和儿童的原代培养人尿路上皮细胞（HUC）来分析OTA对非预定DNA合成（UDS）的诱导作用。从两名接受肾切除术的成人以及两名患有输尿管肾盂连接部狭窄的儿童的输尿管或肾盂中分离出HUC，并在无血清条件下培养。当细胞汇合度达到70 - 80%后，用精氨酸缺陷培养基（ADM）抑制细胞增殖，通过暴露于OTA（10 - 2000 nM）、甲基磺酸乙酯（EMS，5 mM，阳性对照）或二甲基亚砜（DMSO，0.2%，溶剂对照）后3H - 胸腺嘧啶核苷掺入进行放射自显影评估UDS。在对照培养物中，UDS水平较低。暴露于EMS导致UDS诱导（与对照相比增加2至5倍），从而能够灵敏地检测到所有四个样本中由DNA损伤诱导引起的修复，并证明在所选培养条件下可以发生EMS诱导的DNA损伤修复。在源自成人的两种HUC培养物中，在50 - 500 nM OTA浓度范围内观察到UDS的显著诱导。对于来自老年男性的HUC，在50 nM OTA时发现修复细胞的最高比例（CIR）（50% CIR）。在成年女性和7岁男孩的其他样本中，分别在OTA浓度为500和250 nM时观察到最大反应。与所有其他样本不同，在源自婴儿尿路上皮的HUC培养物中未发现OTA对UDS的显著诱导。在所有培养物中，OTA浓度高于500 nM时观察到细胞毒性迹象。体外观察到的对OTA的不同敏感性可能暗示个体在体内的不同易感性。