Bakhiya Nadiya, Arlt Volker M, Bahn Andrew, Burckhardt Gerhard, Phillips David H, Glatt Hansruedi
German Institute of Human Nutrition (DIfE) Potsdam-Rehbrücke, Department of Nutritional Toxicology, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.
Toxicology. 2009 Oct 1;264(1-2):74-9. doi: 10.1016/j.tox.2009.07.014. Epub 2009 Jul 28.
Aristolochic acid (AA), present in Aristolochia species, is the major causative agent in the development of severe renal failure and urothelial cancers in patients with AA nephropathy. It may also be a cause of Balkan endemic nephropathy. Epithelial cells of the proximal tubule are the primary cellular target of AA. To study whether organic anion transporters (OATs) expressed in proximal tubule cells are involved in uptake of AA, we used human epithelial kidney (HEK293) cells stably expressing human (h) OAT1, OAT3 or OAT4. AA potently inhibited the uptake of characteristic substrates, p-aminohippurate for hOAT1 and estrone sulfate for hOAT3 and hOAT4. Aristolochic acid I (AAI), the more cytotoxic and genotoxic AA congener, exhibited high affinity for hOAT1 (K(i)=0.6 microM) as well as hOAT3 (K(i)=0.5 microM), and lower affinity for hOAT4 (K(i)=20.6 microM). Subsequently, AAI-DNA adduct formation (investigated by (32)P-postlabelling) was used as a measure of AAI uptake. Significantly higher levels of adducts occurred in hOAT-expressing cells than in control cells: this effect was abolished in the presence of the OAT inhibitor probenecid. In Xenopus laevis oocytes hOAT-mediated efflux of p-aminohippurate was trans-stimulated by extracellular AA, providing further molecular evidence for AA translocation by hOATs. Our study indicates that OATs can mediate the uptake of AA into proximal tubule cells and thereby participate in kidney cell damage by this toxin.
马兜铃酸（AA）存在于马兜铃属植物中，是导致马兜铃酸肾病患者出现严重肾衰竭和尿路上皮癌的主要致病因素。它也可能是巴尔干地方性肾病的病因之一。近端肾小管上皮细胞是AA的主要细胞靶点。为了研究近端肾小管细胞中表达的有机阴离子转运体（OATs）是否参与AA的摄取，我们使用了稳定表达人（h）OAT1、OAT3或OAT4的人肾上皮（HEK293）细胞。AA强烈抑制特征性底物的摄取，对hOAT1而言是对氨基马尿酸，对hOAT3和hOAT4而言是硫酸雌酮。马兜铃酸I（AAI）是细胞毒性和遗传毒性更强的AA同系物，对hOAT1（K（i）=0.6微摩尔）以及hOAT3（K（i）=0.5微摩尔）表现出高亲和力，而对hOAT4的亲和力较低（K（i）=20.6微摩尔）。随后，AAI - DNA加合物形成（通过32P后标记法研究）被用作AAI摄取的指标。在表达hOAT的细胞中出现的加合物水平显著高于对照细胞：在存在OAT抑制剂丙磺舒的情况下，这种效应被消除。在非洲爪蟾卵母细胞中，细胞外AA反式刺激了hOAT介导的对氨基马尿酸流出，为hOATs转运AA提供了进一步的分子证据。我们的研究表明，OATs可以介导AA摄取进入近端肾小管细胞，从而参与这种毒素对肾细胞的损伤。
