Figure 7.
Soluble uric acid impairs neutrophil migration in patients with kidney dysfunction. (A) Schematic of study design. Out of 51 individuals, 8 patients were excluded due to the intake of immunosuppressive drugs or xanthine oxidase inhibitors. Of the remaining, 10 patients represented with CKD (CKD stage G2-4), 18 patients with ESKD that were on hemodialysis (CKD stage G5), and 15 healthy individuals without any renal pathologies (healthy, CKD stage 0). (B) Human neutrophils were isolated and identified as CD15+CD66b+ by flow cytometry (with gating strategy) with a purity of ∼99%. (C) Number of human neutrophils that were isolated from patients with kidney dysfunction (CKD and ESKD) as well as healthy individuals. (D) Expression (MFI) of MAC-1 relative to isotype determined by flow cytometry (healthy, n = 15; CKD, n = 10; ESKD, n = 18; 1-way ANOVA). (E-F) The total number of neutrophils isolated from healthy individuals and ESKD patients that migrated toward the chemoattractants human CXCL8 (E) and fMLP (F) (healthy, n = 6-7; ESKD, n = 6; 2-way ANOVA) was determined by flow cytometry after 3 hours. (G-H) Neutrophils were incubated with medium and serum from healthy individuals or CKD/ESKD patients and AnnexinV (AnV)-PI staining performed to quantify cell death by flow cytometry. Representative images of dot plots (G) and the percentage (%) of live (AnV-PI−), apoptotic (AnV+PI−), late apoptotic/early necrotic (AnV+PI+), and necrotic (AnV-PI+) neutrophils (H). (I) Percentage (%) of phagocytosed IgG FITC-latex beads in untreated or CXCL8-stimulated neutrophils with or without preincubation of sera from healthy individuals or CKD/ESKD patients was determined by flow cytometry (n = 4 per group; 2-way ANOVA). (J) Healthy neutrophils were incubated with serum from healthy individuals or patients with kidney dysfunction in the absence or presence of rasburicase, and the total number of neutrophils that migrated toward fMLP was determined by flow cytometry after 3 hours (n = 5-6 per group; 2-way ANOVA). Data are mean plus or minus SD. *P < .05, **P < .01, ***P < .001. ns, not significant; Ctrl, control.

Soluble uric acid impairs neutrophil migration in patients with kidney dysfunction. (A) Schematic of study design. Out of 51 individuals, 8 patients were excluded due to the intake of immunosuppressive drugs or xanthine oxidase inhibitors. Of the remaining, 10 patients represented with CKD (CKD stage G2-4), 18 patients with ESKD that were on hemodialysis (CKD stage G5), and 15 healthy individuals without any renal pathologies (healthy, CKD stage 0). (B) Human neutrophils were isolated and identified as CD15+CD66b+ by flow cytometry (with gating strategy) with a purity of ∼99%. (C) Number of human neutrophils that were isolated from patients with kidney dysfunction (CKD and ESKD) as well as healthy individuals. (D) Expression (MFI) of MAC-1 relative to isotype determined by flow cytometry (healthy, n = 15; CKD, n = 10; ESKD, n = 18; 1-way ANOVA). (E-F) The total number of neutrophils isolated from healthy individuals and ESKD patients that migrated toward the chemoattractants human CXCL8 (E) and fMLP (F) (healthy, n = 6-7; ESKD, n = 6; 2-way ANOVA) was determined by flow cytometry after 3 hours. (G-H) Neutrophils were incubated with medium and serum from healthy individuals or CKD/ESKD patients and AnnexinV (AnV)-PI staining performed to quantify cell death by flow cytometry. Representative images of dot plots (G) and the percentage (%) of live (AnV-PI−), apoptotic (AnV+PI−), late apoptotic/early necrotic (AnV+PI+), and necrotic (AnV-PI+) neutrophils (H). (I) Percentage (%) of phagocytosed IgG FITC-latex beads in untreated or CXCL8-stimulated neutrophils with or without preincubation of sera from healthy individuals or CKD/ESKD patients was determined by flow cytometry (n = 4 per group; 2-way ANOVA). (J) Healthy neutrophils were incubated with serum from healthy individuals or patients with kidney dysfunction in the absence or presence of rasburicase, and the total number of neutrophils that migrated toward fMLP was determined by flow cytometry after 3 hours (n = 5-6 per group; 2-way ANOVA). Data are mean plus or minus SD. *P < .05, **P < .01, ***P < .001. ns, not significant; Ctrl, control.

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