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. 2018 Apr 25;9:877. doi: 10.3389/fimmu.2018.00877

Figure 1.

Figure 1

Native and oxidized alpha-1 antitrypsin (AAT) inhibit the ATP-dependent release of interleukin (IL)-1β. (A,B,D) Human lipopolysaccharide (LPS)-primed monocytic U937 cells were stimulated with 2′(3′)-O-(4-Benzoylbenzoyl)adenosine-5′-triphosphate (BzATP, 100 µM) in the presence or absence of different concentrations of the AAT preparation Prolastin® [(A) AAT-P], of native AAT purified from the blood of healthy donors (B) and of oxidized AAT (oxAAT) (D). IL-1β released to the supernatant was measured after 30 min. Native (C) and oxAAT (E) were separated by SDS-polyacrylamide gel electrophoresis (7.5% acrylamide) and stained with Brilliant Blue. In addition, native AAT from healthy donors (D1–D4) and oAAT were incubated with neutrophil elastase (NE) before electrophoresis. Arrows are pointing to NE (25 kDa), a fragment of AAT (48 kDa), AAT (52 kDa), oxAAT (52 kDa), and to a covalent complex of cleaved AAT and NE (77 kDa). (F,G) BzATP-mediated release of IL-1β from freshly isolated peripheral blood mononuclear cells (F) or enriched monocytes (G) from healthy human donors in the presence or absence of AAT-P (1 mg/ml). Absolute IL-1β values obtained from individual donors are connected by lines. (H) Rat precision cut lung slices were primed with LPS followed by application of BzATP in the presence or absence of AAT-P (1 mg/ml). Data are expressed as the concentration of IL-1β released per milligram lung tissue protein. Data are presented as individual data points, bar represents median, whiskers encompass the 25th to 75th percentile, n-numbers of independent experiments are indicated in the figure. Experimental groups were compared by Wilcoxon signed-rank test (F,G) or Kruskal–Wallis test followed by Mann–Whitney rank sum test (A,B,D,H) (*p ≤ 0.05 compared to supernatants from cells treated with LPS and BzATP alone).