Clusterin protects renal tubular epithelial cells from gentamicin-mediated cytotoxicity.
Clusterin is a heterodimeric secreted glycoprotein that is upregulated after acute renal injury. In aminoglycoside nephrotoxicity, clusterin is induced in the tubular epithelium and increased levels are found in the urine. In this study, we developed an in vitro model of gentamicin-induced cytotoxicity in renal proximal tubule cells and tested whether clusterin protected these cells from injury. LLC-PK(1) cells were incubated with varying concentrations of gentamicin in serum-free media, and cytotoxicity was quantified by lactate dehydrogenase release and confirmed by vital dye exclusion. A dose-dependent increase in cytotoxicity occurred with gentamicin concentrations up to 27 mg/ml. Clusterin decreased cytotoxicity in a dose- and time-dependent manner at 6, 12, and 24 h, whereas albumin, used as a control protein, had no effect. In contrast to the aminoglycoside model, when cells were injured by depletion of ATP, clusterin had only a minimally protective effect. LLC-PK(1) cells did not express megalin, a receptor that can mediate the uptake of both clusterin and aminoglycosides into proximal tubule cells. Uptake of gentamicin into LLC-PK(1) cells was observed despite the absence of megalin. In conclusion, clusterin specifically protects against gentamicin-induced renal tubular cell injury by a megalin-independent mechanism.
Girton
Richard A
RA
Dept. of Medicine, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Sundin
David P
DP
Rosenberg
Mark E
ME
D000255:Adenosine Triphosphate / Q000502:physiology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D000968:Antimycin A / Q000037:antagonists & inhibitors / Q000633:toxicity; D015153:Blotting, Western; D002470:Cell Survival / Q000187:drug effects; D051152:Clusterin; D003169:Complement Inactivator Proteins / Q000494:pharmacology*; D004336:Drug Antagonism; D004847:Epithelial Cells / Q000187:drug effects*; D005839:Gentamicins / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D006023:Glycoproteins / Q000494:pharmacology*; D007684:Kidney Tubules / Q000187:drug effects* / Q000473:pathology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D018374:LLC-PK1 Cells; D026561:Low Density Lipoprotein Receptor-Related Protein-2 / Q000378:metabolism; D018832:Molecular Chaperones / Q000494:pharmacology*; D013552:Swine
D000900:Anti-Bacterial Agents; D051152:Clusterin; D003169:Complement Inactivator Proteins; D005839:Gentamicins; D006023:Glycoproteins; D026561:Low Density Lipoprotein Receptor-Related Protein-2; D018832:Molecular Chaperones; D000968:Antimycin A; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase