Intracellular Secretory Leukoprotease Inhibitor Modulates Inositol 1,4,5-Triphosphate Generation and Exerts an Anti-Inflammatory Effect on Neutrophils of Individuals with Cystic Fibrosis and Chronic Obstructive Pulmonary Disease
Figure 3
rhSLPI inhibits neutrophil degranulation. Neutrophils (5 × 106/mL) isolated from healthy individuals were either untreated (Con) or exposed to SLPI (480 nM), followed by stimulation with fMLP (1 M) and IL-8 (1.2 nM) at 37°C. The use of equal cell numbers in each reaction is demonstrated by the identical electrophoretic profile of whole cell lysates in the Coomassie blue stained gel (a). Cell free supernatants were collected at 10 (□) or 20 min (■) and Western blotted for markers of tertiary ((b), MMP-9), secondary ((c), hCAP-18), or primary granule release ((d), MPO). (e) and (f), Healthy control neutrophils were fixed in 4% (w/v) paraformaldehyde following 10 min IL-8/fMLP stimulation in the presence or absence of SLPI (480 nM) and analysed by flow cytometry employing a FITC-labeled CD66b (e) or CD63 antibody (f). Data are represented as mean fluorescent intensity (MFI). All results (expressed as relative densitometry units) were performed in triplicate, and each bar is the mean ± S.E. A representative Western blot is illustrated. * between SLPI treated and untreated at the respective time points calculated by Student’s -test.
