Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or VSports app下载. mil. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure V体育官网. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. .

. 2015 Sep;181(3):518-27.
doi: 10.1111/cei.12654. Epub 2015 Jun 22.

Neutrophil extracellular traps can activate alternative complement pathways

H Wang  1   2   3   4   5 , C Wang  1   2   3   4   5 , M-H Zhao  1   2   3   4   5 , M Chen  1   2   3   4   5
Affiliations

VSports - Neutrophil extracellular traps can activate alternative complement pathways

H Wang et al. Clin Exp Immunol. 2015 Sep.

Abstract

The interaction between neutrophils and activation of alternative complement pathway plays a pivotal role in the pathogenesis of anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). ANCAs activate primed neutrophils to release neutrophil extracellular traps (NETs), which have recently gathered increasing attention in the development of AAV. The relationship between NETs and alternative complement pathway has not been elucidated. The current study aimed to investigate the relationship between NETs and alternative complement pathway. Detection of components of alternative complement pathway on NETs in vitro was assessed by immunostain and confocal microscopy. Complement deposition on NETs were detected after incubation with magnesium salt ethyleneglycol tetraacetic acid (Mg-EGTA)-treated human serum. After incubation of serum with supernatants enriched in ANCA-induced NETs, levels of complement components in supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Complement factor B (Bb) and properdin deposited on NETs in vitro. The deposition of C3b and C5b-9 on NETs incubated with heat-inactivated normal human serum (Hi-NHS) or EGTA-treated Hi-NHS (Mg-EGTA-Hi-NHS) were significantly less than that on NETs incubated with NHS or EGTA-treated NHS (Mg-EGTA-NHS) VSports手机版. NETs induced by ANCA could activate the alternative complement cascade in the serum. In the presence of EGTA, C3a, C5a and SC5b-9 concentration decreased from 800·42 ± 244·81 ng/ml, 7·68 ± 1·50 ng/ml, 382·15 ± 159·75 ng/ml in the supernatants enriched in ANCA induced NETs to 479·07 ± 156·2 ng/ml, 4·86 ± 1·26 ng/ml, 212·65 ± 44·40 ng/ml in the supernatants of DNase I-degraded NETs (P < 0·001, P = 0·008, P < 0·001, respectively). NETs could activate the alternative complement pathway, and might thus participate in the pathogenesis of AAV. .

Keywords: ANCA; alternative complement pathway; complement; neutrophil extracellular traps; vasculitis. V体育安卓版.

PubMed Disclaimer

Figures

Fig. 1
Fig. 1
Detection of deposition of components of alternative complement on neutrophil extracellular traps (NETs) in vitro. Neutrophils isolated from healthy donors were primed with tumour necrosis factor (TNF)-α and incubated with 250 µg/ml anti-neutrophil cytoplasmic antibody (ANCA)-positive immunoglobulin (Ig)G. NETs induced by ANCA were identified by co-localization of DNA (blue), elastase, cit-histone H3 or myeloperoxidase (MPO) (green). Deposition of complement factor B (Bb) and properdin (red) on NETs was assessed by confocal microscopy. (a) Deposition of Bb (red) on NETs stained by DNA (blue) and elastase (green). Magnification ×400. (b) Deposition of properdin (red) on NETs stained by DNA (blue) and elastase (green). Magnification ×400. (c) Deposition of Bb (red) on NETs stained by DNA (blue) and cit-histone (green). (d) Deposition of Bb (red) on NETs stained by DNA (blue) and MPO (green). (e) Deposition of properdin (red) on NETs stained by DNA (blue) and cit-histone (green). (f) Deposition of properdin (red) on NETs stained by DNA (blue) and MPO (green). Representative images of six independent experiments are shown. (c–f) Scale bars = 10 μm.
Fig. 2
Fig. 2
Immunofluorescence detection for alternative complement pathway activation on neutrophil extracellular traps (NETs). (a) C3d deposited on NETs during incubation with normal serum. Little or no deposition of C3d from heat-inactivated normal human serum (Hi-NHS) (lacking active complement) could be observed. (b) C3d deposited on NETs during incubation with magnesium salt (Mg)-ethyleneglycol tetraacetic acid (EGTA)-treated serum. Little or no deposition of C3d from Mg-EGTA-treated Hi-NHS (lacking active complement) could be observed. (c) C5b-9 deposited on NETs during incubation with normal serum. Little or no deposition of C5b-9 from Hi-NHS (lacking active complement) could be observed. (d) C5b-9 deposited on NETs during incubation with Mg-EGTA-treated serum. Little or no deposition of C5b-9 from Mg-EGTA-treated Hi-NHS (lacking active complement) could be observed. (a–d) Representative images of six independent experiments are shown. C3d and C5b-9 were stained in green and NETs were stained in red with propidium iodide (magnification ×200).
Fig. 3
Fig. 3
Neutrophil extracellular traps (NETs) induced by anti-neutrophil cytoplasmic antibody (ANCA) could activate the complement cascade in the serum. Neutrophils were stimulated under different conditions for 180 min. Bars represent mean ± standard deviation (s.d.). Each measured on neutrophils of 11 independent experiments and donors. Supernatants from the cell suspensions were incubated for 40 min with normal serum or magnesium salt (Mg)-ethyleneglycol tetraacetic acid (EGTA)-treated normal serum, and C3a, C5a, SC5b-9 generation was measured by specific enzyme-linked immunosorbent assay (ELISA). *P < 0·05, **P < 0·01, ***P < 0·001.
See this image and copyright information in PMC

References

    1. Jennette JC, Falk RJ, Bacon PA, et al. 2012 revised International Chapel Hill consensus conference nomenclature of vasculitides. Arthritis Rheum. 2013;65:1–11. - PubMed
    1. Chen M, Yu F, Zhang Y, Zhao MH. Antineutrophil cytoplasmic autoantibody-associated vasculitis in older patients. Medicine (Baltimore) 2008;87:203–9. - PubMed
    1. Falk RJ, Terrell RS, Charles LA, Jennette JC. Anti-neutrophil cytoplasmic autoantibodies induce neutrophils to degranulate and produce oxygen radicals in vitro. Proc Natl Acad Sci USA. 1990;87:4115–9. - PMC - PubMed
    1. Kessenbrock K, Krumbholz M, Schonermarck U, et al. Netting neutrophils in autoimmune small-vessel vasculitis. Nat Med. 2009;15:623–5. - PMC - PubMed
    1. Lande R, Gregorio J, Facchinetti V, et al. Plasmacytoid dendritic cells sense self-DNA coupled with antimicrobial peptide. Nature. 2007;449:564–9. - PubMed

V体育平台登录 - Publication types

"V体育官网" MeSH terms