Erschienen in:American Journal of Physiology-Renal Physiology
Sprache:
Englisch
DOI:
10.1152/ajprenal.90754.2008
ISSN:
1931-857X;
1522-1466
Entstehung:
Anmerkungen:
Beschreibung:
<jats:p>The positively charged fluorescent dyes ethidium (Et<jats:sup>+</jats:sup>) and propidium (Pr<jats:sup>2+</jats:sup>) are widely used as DNA and necrosis markers. Et<jats:sup>+</jats:sup>is cytotoxic and mutagenic. The polyspecific organic cation transporters OCT1 (SLC22A1), OCT2 (SLC22A2), and OCT3 (SLC22A3) mediate electrogenic facilitated diffusion of small (≤500 Da) organic cations with broad specificities. In humans, OCT2 mediates basolateral uptake by kidney proximal tubules (PT), whereas in rodents OCT1/2 are involved. In mouse kidney, perfused Et<jats:sup>+</jats:sup>accumulated predominantly in the S2/S3 segments of the PT, but not Pr<jats:sup>2+</jats:sup>. In cells stably overexpressing human OCTs (hOCTs), Et<jats:sup>+</jats:sup>uptake was observed with K<jats:sub>m</jats:sub>values of 0.8 ± 0.2 μM (hOCT1), 1.7 ± 0.5 μM (hOCT2), and 2.0 ± 0.5 μM (hOCT3), whereas Pr<jats:sup>2+</jats:sup>was not transported. Accumulation of Et<jats:sup>+</jats:sup>was inhibited by OCT substrates quinine, 3-methyl-4-phenylpyridinium (MPP<jats:sup>+</jats:sup>), cimetidine, and tetraethylammonium (TEA<jats:sup>+</jats:sup>). For hOCT1 and hOCT2, the IC<jats:sub>50</jats:sub>values for MPP<jats:sup>+</jats:sup>, TEA<jats:sup>+</jats:sup>, and cimetidine were higher than for inhibition of previously tested transported substrates. For hOCT2, the inhibition of Et<jats:sup>+</jats:sup>uptake by MPP<jats:sup>+</jats:sup>and cimetidine was shown to be competitive. Et<jats:sup>+</jats:sup>also inhibited transport of 0.1 μM [<jats:sup>3</jats:sup>H]MPP<jats:sup>+</jats:sup>by all hOCT isoforms with IC<jats:sub>50</jats:sub>values between 0.4 and 1.3 μM, and the inhibition of hOCT1-mediated uptake of MPP<jats:sup>+</jats:sup>by Et<jats:sup>+</jats:sup>was competitive. In Oct1/2<jats:sup>−/−</jats:sup>mice, Et<jats:sup>+</jats:sup>uptake in the PT was almost abolished. The data demonstrate that Et<jats:sup>+</jats:sup>is taken up avidly by the PT, which is mediated by OCT1 and/or OCT2. Considering the high affinity of OCTs for Et<jats:sup>+</jats:sup>and their strong expression in various organs, strict safety guidelines for Et<jats:sup>+</jats:sup>handling should be reinforced.</jats:p>