Abstract
Consumption of herbal medicines derived from Aristolochia plants is associated with a progressive tubulointerstitial disease known as aristolochic acid (AA) nephropathy. The nephrotoxin produced naturally by these plants is AA-I, a nitrophenanthrene carboxylic acid that selectively targets the proximal tubule. This nephron segment is prone to toxic injury because of its role in secretory elimination of drugs and other xenobiotics. Here, we characterize the handling of AA-I by membrane transporters involved in renal organic anion transport. Uptake assays in heterologous expression systems identified murine organic anion transporters (mOat1, mOat2, and mOat3) as capable of mediating transport of AA-I. Kinetic analyses showed that all three transporters have an affinity for AA-I in the submicromolar range and thus are likely to operate at toxicologically relevant concentrations in vivo. Structure-activity relationships revealed that the carboxyl group is critical for high-affinity interaction of AA-I with mOat1, mOat2, and mOat3, whereas the nitro group is required only by mOat1. Furthermore, the 8-methoxy group, although essential for toxicity, was not requisite for transport. Mouse renal cortical slices avidly accumulated AA-I, achieving slice-to-medium concentration ratios >10. Uptake by slices was sensitive to known mOat1 and mOat3 substrates and the organic anion transport inhibitor probenecid, which also blocked the production of DNA adducts formed with reactive intracellular metabolites of AA-I. Taken together, these findings indicate that OAT family members mediate high-affinity transport of AA-I and may be involved in the site-selective toxicity and renal elimination of this nephrotoxin.
Footnotes
This work was supported by the National Institutes of Health National Institute of Environmental Health Sciences [Grant P01-ES004068] (to K.G.D.) and the National Institutes of Health National Institute of Diabetes and Digestive and Kidney Diseases [Grant R01-DK067216] (to D.H.S.). K.G.D. is the recipient of a Translational Research Scholar award funded through the generous support of the Zickler Family Foundation.
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.111.180984.
↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
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ABBREVIATIONS:
- AA
- aristolochic acid
- AAN
- AA nephropathy
- CYP 1A1/2
- cytochrome P450 1A1/2
- OAT
- organic anion transporter
- mOat
- murine OAT
- hOAT
- human OAT
- CHO
- Chinese hamster ovary
- EV
- empty vector
- HBSS
- Hanks' balanced salt solution
- BSA
- bovine serum albumin
- FCS
- fetal calf serum
- dA
- deoxyadenosine
- dG
- deoxyguanosine
- NR
- nitroreduction.
- Received February 23, 2011.
- Accepted May 3, 2011.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
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