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). 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. 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. 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.
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). 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. 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. 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.
Authors: Asher A Mendelson; Qiunong Guan; Irina Chafeeva; Gerald A da Roza; Jayachandran N Kizhakkedathu; Caigan Du Journal: Perit Dial Int Date: 2013 Jan-Feb Impact factor: 1.756
Authors: Priscilla Kheddo; Alexander P Golovanov; Kieran T Mellody; Shahid Uddin; Christopher F van der Walle; Rebecca J Dearman Journal: Toxicol In Vitro Date: 2016-02-10 Impact factor: 3.500
Authors: Beatriz Santamaría; Alberto Benito-Martin; Alvaro Conrado Ucero; Luiz Stark Aroeira; Ana Reyero; María Jesús Vicent; Mar Orzáez; Angel Celdrán; Jaime Esteban; Rafael Selgas; Marta Ruíz-Ortega; Manuel López Cabrera; Jesús Egido; Enrique Pérez-Payá; Alberto Ortiz Journal: PLoS One Date: 2009-08-13 Impact factor: 3.240