Increased levels of neutrophil extracellular trap remnants in the circulation of patients with small vessel vasculitis, but an inverse correlation to anti-neutrophil cytoplasmic antibodies during remission.

OBJECTIVES: Neutrophil extracellular traps (NETs) have been visualized at the site of ANCA-associated vasculitis (AAV) lesions. Increased levels of NET remnants in the circulation have been reported in some AAV patients with active disease. The aim of the present study was to analyse NET remnants in a larger cohort of AAV patients with varying degrees of disease activity and to elucidate possible factors responsible for remnant variation. METHODS: Levels of NET remnants in the circulation of healthy controls (HCs; n = 31) and AAV patients (n = 93) were determined with ELISA. NET remnants were then correlated with ANCA levels, spontaneous and induced cell death (NETosis/necrosis) in vitro, neutrophil count and corticosteroid therapy. RESULTS: Patients with active disease showed higher levels of circulating NET remnants compared with patients in remission (P = 0.026) and HCs (P = 0.006). From patients sampled during both remission and active disease, we found increased levels during active disease (P = 0.0010). In remission, ANCA-negative patients had higher levels of NET remnants than ANCA-positive patients and a negative correlation was observed between NET remnants and PR3-ANCA (rs = -0.287, P = 0.048). NET remnants correlated with neutrophil count in HCs (rs = 0.503, P = 0.014) but not in patients during remission. Neutrophils from patients showed enhanced spontaneous cell death (P = 0.043). CONCLUSION: We found increased levels of circulating NET remnants in patients with active AAV. Furthermore, AAV patients exhibited an increased propensity for spontaneous cell death. NET remnant levels seem to be positively related to disease activity and neutrophil count, but inversely related to ANCA at least during remission.

Replacement of acetate with citrate in dialysis fluid: a randomized clinical trial of short term safety and fluid biocompatibility.

BACKGROUND: The majority of bicarbonate based dialysis fluids are acidified with acetate. Citrate, a well known anticoagulant and antioxidant, has been suggested as a biocompatible alternative. The objective of this study was to evaluate short term safety and biocompatibility of a citrate containing acetate-free dialysis fluid. METHODS: Twenty four (24) patients on maintenance dialysis three times per week, 13 on on-line hemodiafiltration (HDF) and 11 on hemodialysis (HD), were randomly assigned to start with either citrate dialysis fluid (1 mM citrate, 1.5 mM calcium) or control fluid (3 mM acetate, 1.5 mM calcium) in an open-labeled cross-over trial (6 + 6 weeks with 8 treatments wash-out in between). Twenty (20) patients, 11 on HDF and 9 on HD were included in the analyses. Main objective was short term safety assessed by acid-base status, plasma ionized calcium and parathyroid hormone (PTH). In addition, biocompatibility was assessed by markers of inflammation (pentraxin 3 (PTX-3), CRP, IL-6, TNF-α and IL-1ß) and thrombogenicity (activated partial thromboplastin time (APTT) and visual clotting scores). RESULTS: No differences dependent on randomization order or treatment mode (HD vs. HDF) were detected. Citrate in the dialysis fluid reduced the intra-dialytic shift in pH (+0.04 week 6 vs. +0.06 week 0, p = 0.046) and base excess (+3.9 mM week 6 vs. +5.6 mM week 0, p = 0.006) over the study period. Using the same calcium concentration (1.5 mM), citrate dialysis fluid resulted in lower post-dialysis plasma ionized calcium level (1.10 mM vs. 1.27 mM for control, p < 0.0001) and higher post-dialysis PTH level (28.8 pM vs. 14.7 pM for control, p < 0.0001) while pre-dialysis levels were unaffected. Citrate reduced intra-dialytic induction of PTX-3 (+1.1 ng/ml vs. +1.4 ng/ml for control, p = 0.04) but had no effect on other markers of inflammation or oxidative stress. Citrate reduced visual clotting in the arterial air chamber during HDF (1.0 vs. 1.8 for control, p = 0.03) and caused an intra-dialytic increase in APTT (+6.8 s, p = 0.003) without affecting post-dialysis values compared to control. CONCLUSIONS: During this small short term study citrate dialysis fluid was apparently safe to use in HD and on-line HDF treatments. Indications of reduced treatment-induced inflammation and thrombogenicity suggest citrate as a biocompatible alternative to acetate in dialysis fluid. However, the results need to be confirmed in long term studies. TRIAL REGISTRATION ISRCTN: ISRCTN28536511.