Effect of alternative peritoneal dialysis solutions on cell viability, apoptosis/necrosis and cytokine expression in human monocytes

Abstract

Effect of alternative peritoneal dialysis solutions on cell viability, apoptosis/necrosis and cytokine expression in human monocytes.BackgroundCellular function, cell viability and the cytokine network of human monocytes are influenced by the specific composition of peritoneal dialysis (PD) fluids. In an in vitro study using isolated human blood monocytes, we investigated the effect of peritoneal dialysates containing amino acids (Amino) or glucose polymer (Glu-poly) instead of glucose (Glu) as the osmotic agent, and bicarbonate (Bic) or PBS instead of lactate (Lac) as a buffer.MethodsThe following parameters were studied: mitochondrial dehydrogenase activity (using the MTT assay), interleukin (IL)-6 and IL-8 release (ELISA) and cellular IL-6 mRNA expression after lipopolysaccharide (LPS) stimulation (using RT-PCR). FACS flow cytometry with annexin V and propidium iodide as markers and fluorescence microscopic methods were used to study the effects of the test fluids on cell necrosis and apoptosis.ResultsGlu/Lac pH 5.5 and Glu-poly/PBS pH 7.4 both significantly reduced mitochondrial dehydrogenase activity by more than 50% after 60 minutes of incubation (30.5 ± 7.6%, 42.5 ± 6.5%, referred to RPMI 1640 as 100%). Amino/Bic and Glu/Bic were both superior (MTT assay > 63%). The rate of necrotic cells after 15 minutes of incubation measured by FACS was mostly increased with Glu/Lac pH 5.5 (29.9 ± 4.0%). The rate of apoptotic cells, however, was not significantly different between the test solutions. The concentration of IL-6 in the supernatant of stimulated monocytes was highest with Glu/Bic (1023 ± 278 pg/ml) and Amino/Bic (776 ± 296 pg/ml) and lowest with Glu/Lac pH 5.5 (46 ± 22 pg/ml) and Glu-poly/PBS (32 ± 13 pg/ml). IL-8 release from stimulated monocytes showed a similar pattern. Glu-poly/PBS showed a suppressive effect on IL-6 mRNA expression (ratio IL-6/β-Actin, 0.4 ± 0.25 vs. RPMI 1.5 ± 3.6).ConclusionsBicarbonate buffered solutions both with glucose or amino acids as osmotic agents were superior when regarding cell metabolism, viability and cytokine release, while lactate buffered solutions and Glu-poly/PBS showed some reduced biocompatibility pattern for monocytes in vitro. Effect of alternative peritoneal dialysis solutions on cell viability, apoptosis/necrosis and cytokine expression in human monocytes. Cellular function, cell viability and the cytokine network of human monocytes are influenced by the specific composition of peritoneal dialysis (PD) fluids. In an in vitro study using isolated human blood monocytes, we investigated the effect of peritoneal dialysates containing amino acids (Amino) or glucose polymer (Glu-poly) instead of glucose (Glu) as the osmotic agent, and bicarbonate (Bic) or PBS instead of lactate (Lac) as a buffer. The following parameters were studied: mitochondrial dehydrogenase activity (using the MTT assay), interleukin (IL)-6 and IL-8 release (ELISA) and cellular IL-6 mRNA expression after lipopolysaccharide (LPS) stimulation (using RT-PCR). FACS flow cytometry with annexin V and propidium iodide as markers and fluorescence microscopic methods were used to study the effects of the test fluids on cell necrosis and apoptosis. Glu/Lac pH 5.5 and Glu-poly/PBS pH 7.4 both significantly reduced mitochondrial dehydrogenase activity by more than 50% after 60 minutes of incubation (30.5 ± 7.6%, 42.5 ± 6.5%, referred to RPMI 1640 as 100%). Amino/Bic and Glu/Bic were both superior (MTT assay > 63%). The rate of necrotic cells after 15 minutes of incubation measured by FACS was mostly increased with Glu/Lac pH 5.5 (29.9 ± 4.0%). The rate of apoptotic cells, however, was not significantly different between the test solutions. The concentration of IL-6 in the supernatant of stimulated monocytes was highest with Glu/Bic (1023 ± 278 pg/ml) and Amino/Bic (776 ± 296 pg/ml) and lowest with Glu/Lac pH 5.5 (46 ± 22 pg/ml) and Glu-poly/PBS (32 ± 13 pg/ml). IL-8 release from stimulated monocytes showed a similar pattern. Glu-poly/PBS showed a suppressive effect on IL-6 mRNA expression (ratio IL-6/β-Actin, 0.4 ± 0.25 vs. RPMI 1.5 ± 3.6). Bicarbonate buffered solutions both with glucose or amino acids as osmotic agents were superior when regarding cell metabolism, viability and cytokine release, while lactate buffered solutions and Glu-poly/PBS showed some reduced biocompatibility pattern for monocytes in vitro.