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Am J Physiol Renal Physiol 275: F962-F971, 1998;
0363-6127/98 $5.00
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Vol. 275, Issue 6, F962-F971, December 1998

Extracellular ATP causes apoptosis and necrosis of cultured mesangial cells via P2Z/P2X7 receptors

Eckhard Schulze-Lohoff, Christian Hugo, Sylvia Rost, Susanne Arnold, Angela Gruber, Bernhard Brüne, and Ralf Bernd Sterzel

Nephrologisches Labor, Medizinische Klinik IV, Universität Erlangen-Nürnberg, D-91054 Erlangen, Germany

Mesangial cells undergo cell death both by apoptosis and necrosis during glomerular disease. Since nucleotides are released from injured and destroyed cells in the glomerulus, we examined whether extracellular ATP and its receptors may regulate cell death of cultured mesangial cells. Addition of extracellular ATP (300 µM to 5 mM) to cultured rat mesangial cells for 90 min caused a 5.8-fold increase in DNA fragmentation (terminal deoxynucleotidyl transferase assay) and a 4.2-fold increase in protein levels of the tumor suppressor p53, which is thought to regulate apoptosis. Apoptotic DNA fragmentation was confirmed by the diphenylamine assay and by staining with the DNA-specific fluorochrome Hoechst 33258. The necrotic markers, release of lactate dehydrogenase and uptake of trypan blue, were not positive before 3 h of ATP addition. The effects of ATP on DNA fragmentation and p53 expression were reproduced by the purinergic P2Z/P2X7 receptor agonist, 3'-O-(4-benzoylbenzoyl)-ATP, and inhibited by the P2Z/P2X7 receptor blocker, oxidized ATP. Transcripts encoding the P2Z/P2X7 receptor were expressed by cultured mesangial cells as determined by Northern blot analysis. P2Z/P2X7 receptor-associated pore formation in the plasma membrane was demonstrated by the Lucifer yellow assay. We conclude that activation of P2Z/P2X7 receptors by extracellular ATP causes apoptosis and necrosis of cultured mesangial cells. Activation of purinergic P2Z/P2X7 receptors may play a role in causing death of mesangial cells during glomerular disease.

purinergic receptors; nucleotides; tumor suppressor p53


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