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Peritubular transport of ochratoxin A by single rabbit renal proximal tubules
J R Welborn1, C E Groves, S H Wright
1Department of Physiology, College of Medicine, University of Arizona, Tucson 85724, USA.
Abstract:
Epifluorescence microscopy was used to study peritubular transport of the fluorescent mycotoxin ochratoxin A (OTA) into single proximal tubule segments of the rabbit. Initial rates of OTA uptake into S2 segments were saturable and adequately described by Michaelis-Menten kinetics, with an apparent Km of 2.2+/-0.3 microM (SEM). Several lines of evidence indicated that peritubular uptake of OTA in S2 segments was effectively limited to the "classical" organic anion transporter. First, 5 mM p-aminohippurate (PAH) cis-inhibited the uptake of 1 microM OTA into tubules by 96%. Kinetic analysis of the inhibition of OTA uptake by PAH (100 microM to 5 mM) yielded an apparent Ki of 164 microM, similar to the 100 to 200 microM range of Km values previously reported for the peritubular uptake of PAH. Second, efflux of OTA from tubules was trans-stimulated 3.2-fold by the presence of 2.5 mM PAH in the uptake medium. Third, 100 microM alpha-ketoglutarate (alphaKG) trans-stimulated the uptake rate of 1 microM OTA by 1.8-fold. Fourth, besides PAH, other organic anions effectively cis-inhibited the uptake of 1 microM OTA into tubules (inhibitor, % inhibition): 1.5 mM alphaKG, 80%; 1 mM probenecid, 100%; 1 mM piroxicam, 100%; 1 mM octanoate, 100%. In contrast, 1.5 mM tetraethylammonium, an organic cation, blocked uptake of 1 microM OTA by only 7%. The inhibition of OTA uptake into S1 and S3 segments of the proximal tubule was qualitatively similar: 5 mM PAH cis-inhibited the uptake of 1 microM OTA by approximately 95% in both S1 and S3 segments. Thus, peritubular OTA uptake into all segments of the proximal tubule appears to be dominated by its interaction with the classical organic anion transporter. The high-affinity and relatively high capacity of this pathway for OTA suggest that peritubular uptake may be a significant avenue for the entry of this toxin into proximal tubule cells.
Insights
The study reveals that the mycotoxin ochratoxin A (OTA) enters kidney tubule cells via the organic anion transporter. This pathway is crucial for OTA
Area of Science:
- Nephrology
- Toxicology
- Molecular Biology
Background:
- Ochratoxin A (OTA) is a mycotoxin with known nephrotoxic effects.
- Understanding the cellular uptake mechanisms of OTA is crucial for assessing its toxicity and developing interventions.
Purpose of the Study:
- To investigate the peritubular transport mechanism of ochratoxin A (OTA) into rabbit proximal tubule segments.
- To determine the role of the organic anion transporter in OTA uptake.
Main Methods:
- Epifluorescence microscopy was employed to visualize and quantify OTA uptake.
- Kinetic analysis using Michaelis-Menten models was performed.
- Inhibition and trans-stimulation studies with known organic anions and cations were conducted.
Main Results:
- Peritubular OTA uptake into S2 proximal tubule segments exhibited saturable kinetics, consistent with Michaelis-Menten transport.
- Uptake was significantly inhibited by p-aminohippurate (PAH) and other organic anions, but not by organic cations.
- Efflux of OTA was trans-stimulated by PAH, and uptake was trans-stimulated by alpha-ketoglutarate (alphaKG).
- Similar transport characteristics were observed in S1 and S3 proximal tubule segments.
Conclusions:
- Peritubular uptake of OTA into rabbit proximal tubule cells is primarily mediated by the classical organic anion transporter.
- The high affinity and capacity of this transporter for OTA suggest it is a significant route for toxin entry into renal cells.
- These findings have implications for understanding OTA nephrotoxicity.