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FRI0007 Effect of Type 1 and 2 Interferons on Neutrophil Apoptosis in naÏve and TNF Alpha Primed Neutrophils
  1. S. Irwin1,
  2. A. Midgley1,
  3. R. Wright1,
  4. M. Peak2,
  5. M. Beresford1
  1. 1Women's and Children's Health, University of Liverpool
  2. 2Alder Hey Children's NHS Foundation, Liverpool, United Kingdom

Abstract

Background Juvenile-onset Systemic Lupus Erythematosus (JSLE) is a multisystem autoimmune disease characterised by the presence of autoantigens. These may result from increased numbers of apoptotic neutrophils and dysregulated clearance of these neutrophils by phagocytes. Serum from JSLE patients contains high levels of type 1 (α and β) and 2 (γ) interferons (IFN). In the context of JSLE being an inflammatory condition, JSLE neutrophils are likely to be more activated in comparison to healthy, non-inflammatory control neutrophils. Tumour necrosis factor (TNF)-α can prime neutrophils (demonstrated by CD62L shedding), and provides an inflammatory setting for neutrophils as seen in JSLE. Gene expression analysis has shown an IFN and granulocyte signature within SLE; however the interaction between IFN and neutrophils in regards to apoptosis, and the pathogenesis of JSLE, is yet to be elucidated.

Objectives To investigate the effect of type 1 and 2 IFNs on neutrophil apoptosis of naïve and primed neutrophils

Methods Neutrophils isolated from healthy control adult donors (n=5) were either unstimulated or primed with TNF-α (1μg/ml) for 30 mins. The neutrophils were subsequently incubated with IFN-α (10μg/ml), IFN-γ (1μg/ml) or IFN-β (1μg/ml) for 6hrs. The expression of CD62L after 30 min TNF-α incubation was measured using flow cytometry using CD62L-APC (DREG-56) antibody staining with IgG-APC isotype control. The percentage of apoptotic neutrophils were measured after 6hr IFN incubation using flow cytometry via annexin V FITC staining.

Results CD62L expression was reduced in neutrophils exposed to TNF-α compared with unstimulated neutrophils (geometric mean: 1.61 ± 0.30 vs 11.43 ± 1.44). Apoptosis (% ± SEM) was significantly reduced upon IFN-α (6.57 ± 1.18%, p=0.04) and IFN-γ (10.79 ± 1.92%, p=0.04) incubation compared to unstimulated (22.01 ± 3.18%), although IFN-β (20.30 ± 5.10%, p=0.35) and TNF-α alone (26.40 ± 4.30%, p=0.50) had little effect. In neutrophils pre-treated with TNF-α, there was significant increase in apoptosis when subsequently exposed to either IFN-α (48.91 ± 4.84%, p=0.04), IFN-γ (43.09 ± 2.99%, p=0.04) or IFN-β (39.41 ± 3.23%, p=0.04) compared to unstimulated neutrophils.

Conclusions There is evidence that both type 1 and 2 IFNs contribute to the development of JSLE. Here we show a differential effect between IFNs on neutrophil apoptosis indicating a potential pro-apoptotic effect of IFNs on neutrophils in a JSLE environment through the pre-incubation with TNF-α. IFNs may contribute to the increase in neutrophil apoptosis in JSLE and lead to an increase in autoantigens that are fundamental to JSLE pathogenesis.

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  2. Kim, T. et al. (1987) Serum levels of interferons in patients with systemic lupus erythematosus. Clin. Exp. Immunol. 70, 562–69.

  3. Midgley, A. et al. (2009). The role of neutrophil apoptosis in juvenile-onset systemic lupus erythematosus. Arthritis Rheum. 60, 2390–2401.

  4. Wittmann, S. et al. (2004) Cytokine upregulation of surface antigens correlates to the priming of the neutrophil oxidative burst response. Cytometry A. 57, 53–62.

Disclosure of Interest None declared

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