Alterations in Membrane Transport Function and Cell Viability Induced by ATP Depletion in Primary Cultured Rabbit Renal Proximal Tubular Cells.
10.4196/kjpp.2009.13.1.15
- Author:
Sung Ju LEE
1
;
Chae Hwa KWON
;
Yong Keun KIM
Author Information
1. Department of Physiology, MRC for Ischemic Tissue Regeneration, College of Medicine, Pusan National University, Busan 602-739, Korea. kim430@pusan.ac.kr
- Publication Type:Original Article
- Keywords:
ATP depletion;
Membrane transport;
cPLA2;
Serine/cysteine proteases
- MeSH:
Adenosine Triphosphate;
Amino Acids;
Antipain;
Arachidonic Acid;
Arachidonic Acids;
Catalase;
Cell Death;
Cell Survival;
Cytosol;
Diminazene;
Hydrogen Peroxide;
Hydroxyl Radical;
Iodoacetic Acid;
Membranes;
Ouabain;
Peptide Hydrolases;
Phospholipases A2;
Potassium Cyanide;
Protease Inhibitors;
Reactive Oxygen Species;
RNA, Messenger;
Trypan Blue
- From:The Korean Journal of Physiology and Pharmacology
2009;13(1):15-22
- CountryRepublic of Korea
- Language:English
-
Abstract:
This study was undertaken to elucidate the underlying mechanisms of ATP depletion-induced membrane transport dysfunction and cell death in renal proximal tubular cells. ATP depletion was induced by incubating cells with 2.5 mM potassium cyanide (KCN)/0.1 mM iodoacetic acid (IAA), and membrane transport function and cell viability were evaluated by measuring Na+-dependent phosphate uptake and trypan blue exclusion, respectively. ATP depletion resulted in a decrease in Na+-dependent phosphate uptake and cell viability in a time-dependent manner. ATP depletion inhibited Na+-dependent phosphate uptake in cells, when treated with 2 mM ouabain, a Na+ pump-specific inhibitor, suggesting that ATP depletion impairs membrane transport functional integrity. Alterations in Na+-dependent phosphate uptake and cell viability induced by ATP depletion were prevented by the hydrogen peroxide scavenger such as catalase and the hydroxyl radical scavengers (dimethylthiourea and thiourea), and amino acids (glycine and alanine). ATP depletion caused arachidonic acid release and increased mRNA levels of cytosolic phospholipase A2 (cPLA2). The ATP depletion-dependent arachidonic acid release was inhibited by cPLA2 specific inhibitor AACOCF3. ATP depletion-induced alterations in Na+-dependent phosphate uptake and cell viability were prevented by AACOCF3. Inhibition of Na+-dependent phosphate uptake by ATP depletion was prevented by antipain and leupetin, serine/cysteine protease inhibitors, whereas ATP depletion-induced cell death was not altered by these agents. These results indicate that ATP depletion-induced alterations in membrane transport function and cell viability are due to reactive oxygen species generation and cPLA2 activation in renal proximal tubular cells. In addition, the present data suggest that serine/cysteine proteases play an important role in membrane transport dysfunction, but not cell death, induced by ATP depletion.
