Apical-to-basolateral transepithelial transport of Ochratoxin A by two subtypes of Madin-Darby canine kidney cells.
In this study we investigated the transepithelial transport of Ochratoxin A (OTA), a potent nephrotoxin, across monolayers of two collecting duct-derived cells clones (Madin-Darby canine kidney cells (MDCK)-C7 and MDCK-C11 cells, resembling principal and intercalated cells, respectively) either from the apical to the basolateral side or vice versa. We cultured cells on permeable supports and compared the transport rates of OTA, p-aminohippuric acid (PAH) and fluorescein-labelled inulin. Monolayers of both cell clones translocated OTA from the apical to the basolateral side but not in the opposite direction. Transport rate across MDCK-C11 cell monolayers was 2.9-fold the transport rate across MDCK-C7 cell monolayers. OTA transport was temperature-dependent being reduced from 77.5 pmol/cm2 per h to 10.1 pmol/cm2 per h in MDCK-C11 and from 27.0 pmol/cm2 per h to 7.6 pmol/cm2 per h in MDCK-C7 cells when temperature was decreased from 37 degrees C to 4 degrees C. In both cell clones, the dipeptides carnosine and glycylsarcosine but not the amino acids glycine or phenylalanine had an inhibitory effect on OTA transport. In both cell clones, transepithelial transport of OTA was dependent on the apical pH (pK(a) of OTA = 7.1). In an environment mimicking the transepithelial in vivo pH gradient to some extent with more acidic pH on the apical side than on the basolateral side, transport was 4-fold higher in both cell clones as compared to conditions when pH was 7.4 in both bath solutions. In the absence of a pH gradient, transport rates were similar to that at 4 degrees C. Apical uptake of [3H]OTA was inhibited by carnosine and by glycylsarcosine and the uptake of [3H]carnosine was inhibited by OTA. Our results indicate that OTA is transported across the apical membrane of MDCK cells by both non-ionic diffusion and by a H+-dipeptide cotransporter. Thus, reabsorption of OTA in the collecting duct contributes to the observed long half life of OTA in the mammalian body.
Schwerdt
G
G
Department of Physiology, University of Würzburg, Germany. phio005@rzroe.uni-wuerzburg.de
Gekle
M
M
Freudinger
R
R
Mildenberger
S
S
Silbernagl
S
S
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002336:Carnosine / Q000494:pharmacology; D002460:Cell Line; D016764:Cell Polarity; D002470:Cell Survival; D004151:Dipeptides / Q000494:pharmacology; D004285:Dogs; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005998:Glycine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007444:Inulin / Q000378:metabolism; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D009183:Mycotoxins / Q000378:metabolism*; D009793:Ochratoxins / Q000378:metabolism*; D010649:Phenylalanine / Q000494:pharmacology; D013696:Temperature; D010130:p-Aminohippuric Acid / Q000378:metabolism
D004151:Dipeptides; D009183:Mycotoxins; D009793:Ochratoxins; C025589:ochratoxin A; C004194:glycylsarcosine; D010649:Phenylalanine; D002336:Carnosine; D007444:Inulin; D005998:Glycine; D010130:p-Aminohippuric Acid