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Perit Dial Int 28(6): 655-666
2008
© 2008 International Society for Peritoneal Dialysis
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CYTOTOXICITY OF MONONUCLEAR CELLS AS INDUCED BY PERITONEAL DIALYSIS FLUIDS: INSIGHT INTO MECHANISMS THAT REGULATE OSMOTIC STRESS-RELATED APOPTOSIS

Karine Gastaldello1, Cecile Husson1, Jean-Paul Dondeyne1, Jean-Louis Vanherweghem2 and Christian Tielemans1,2

Laboratoire de Recherche en Nephrologie,1 Université Libre de Bruxelles; Department of Nephrology, Dialysis and Transplantation,2 Hôpital Erasme, Brussels, Belgium

Correspondence to: K. Gastaldello, Laboratoire de Recherche en Nephrologie, 806 Route de Lennik, Brussels, 1070 Belgium. kgastald{at}ulb.ac.be

{diamondsuit} Objective: High glucose content of peritoneal dialysis fluids (PDFs) has been shown to contribute to loss of peritoneal function during long-term peritoneal dialysis. However, hyperosmolality and hypertonicity of PDF are usually seen as similar stress events inducing osmotic stress-induced programmed cell death. In this study, we examined the impact of various osmotic agents on apoptosis induced by hyperosmolar PDFs, focusing on the mechanisms underlying the lethal effects of PDFs on peripheral blood mononuclear cells (PBMCs).

{diamondsuit} Methods: We assessed apoptosis and necrosis by annexin V–propidium iodide (PI) labeling, and caspase-3 activity by fluorescence assay. F-actin remodeling was measured using fluorescent phalloidin labeling.

{diamondsuit} Results: Hyperosmolality does not cause the cytotoxicity observed with PDF, but exposure to agents incapable of permeating cell membranes results in a significant increase in the percentage of apoptotic PBMCs by annexin V–PI labeling, which is confirmed by the increase in caspase-3 activity. Interestingly, inhibition of caspase-3 by Z-VAD-FMK did not suppress apoptosis. Extracellular hypertonicity produced polymerization of filamentous actin and cell shrinkage, which displayed similar time courses. Cell shrinkage was blocked by cytochalasin D, indicating an active role for actin cytoskeleton in hypertonicity-induced cell shrinkage. F-actin polymerization was related to an increase in intracellular ionic strength. Finally, we excluded a direct role for actin remodeling in osmotic stress-induced programmed cell death.

{diamondsuit} Conclusions: Exposure to osmolytes that cannot penetrate cell membranes results in a hypertonicity-induced apoptosis that cannot be blocked by the broad-spectrum caspase inhibitor Z-VAD-FMK. In addition, extracellular hypertonicity induced by impermeant solutes produces F-actin polymerization through an increase in intracellular ionic strength. The remodeling of the cytoskeleton does not modulate apoptosis but participates in cell shrinkage.

KEY WORDS: Peritoneal dialysis fluids; hyperosmolality; hypertonicity; cytotoxicity; apoptosis; caspase-3; cytoskeleton; cell volume regulation.

Received 19 February 2007; accepted 18 April 2008.






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