Neutrophil apoptosis and dysfunction in uremia

Abstract

The high prevalence of bacterial infections among patients with end- stage renal disease suggests that 'professional' phagocytes such as neutrophils are functionally impaired. This dysfunction has been ascribed to uremic toxins, malnutrition, and dialysis. The aim of this study was to investigate the contribution of apoptosis to neutrophil dysfunction in uremia. Neutrophils harvested from uremic patients (n = 6) and age-/gender- matched healthy control subjects (n = 6) were incubated with either 50% autologous plasma or 10% fetal calf serum. After 24-h incubation, apoptosis was quantified by flow cytometry by using propidium iodide nuclear staining. Neutrophils from healthy volunteers were also incubated with either 50% heterologous normal or uremic plasma. After 24-h incubation, apoptosis was quantified by flow cytometry and transmission electron microscopy. In addition, superoxide production was determined by measuring the capacity to reduce ferri- to ferro-cytochrome C by using 4-β-phorbol 12-β-myristate 13- α-acetate or N-formyl methiony-leucyl-phenylalanine (fMLP) for stimulus. Phagocytosis was determined by the uptake of 14C-labeled heat-killed Staphylococcus aureus. Compared with normal neutrophils, uremic neutrophils demonstrated greater apoptosis in the presence of autologous plasma (9 ± 4 versus 19 ± 6%, P = 0.01) as well as 10% fetal calf serum (19 ± 7 versus 31 ± 6%, P = 0.03). Furthermore, compared with normal neutrophils exposed to heterologous normal plasma, those exposed to heterologous uremic plasma exhibited higher apoptosis rates (19 ± 3 versus 40 ± 5%, P = 0.002), lower fMLP-stimulated superoxide production (22.6 ± 2.5 versus 15.5 ± 1.1 nmol O2·-/3.12 x 105 cells/30 min, P = 0.01), and a lower phagocytosis index (38 ± 3% versus 27 ± 5%, P = 0.04). Apoptosis correlated inversely with fMLP-stimulated superoxide production (r = -0.60, P = 0.04) and phagocytosis (r = -0.57, P = 0.05). These results suggest that uremic neutrophils undergo accelerated in vitro apoptosis. Furthermore, uremic plasma accelerates apoptosis of normal neutrophils, resulting in a dysfunctional pattern that is similar to that observed in uremia.