Transforming growth factorâ β1 stimulates degranulation and oxidant release by adherent human neutrophils

Abstract

The signal transduction pathways that are activated by cytokines and growth factors binding to their receptors on human neutrophils (PMN) are poorly understood. When PMN in suspension encounter many of these agonists they are not activated, but rather are primed for subsequent activation. We and others reported that when PMN are plated onto fibrinogen and stimulated with cytokines or with the chemotactic peptide Nâ formylâ methionylâ leucylâ phenylalanine (fMLP) they respond by releasing hydrogen peroxide (H2O2) and the specific granule component lactoferrin. Transforming growth factorâ β1 (TGFâ β1) is released by many cells including PMN. It has been reported that TGFâ β1 stimulates chemotaxis but not exocytosis or superoxide production by cells in suspension. We hypothesized that TGFâ β1 would activate PMN to release H2O2 when they were adherent to fibrinogen, a response mediated by β2 integrin receptors. In this study, we determined whether TGFâ β1 stimulated H2O2 and lactoferrin release by PMN adherent to fibrinogen. TGFâ β1 stimulated H2O2 and lactoferrin release from adherent PMN in a concentrationâ dependent manner, with effects seen in the range of 0.1 to 100 pg/mL. Both H2O2 and lactoferrin release were detected by 60 min and continued for at least 180 min. Adhesion and spreading of PMN paralleled H2O2 and lactoferrin release. Ethanol (200 mM) blocked both H2O2 and lactoferrin release, suggesting the involvement of the phospholipase D pathway. In PMN labeled with lyso[3H]phoephatidylcholine, we observed that TGFâ β1 treatment caused an increase in [3H]phoephatidate. Propranolol (150 μM), an inhibitor of phosphatidate phosphohydrolase, blocked both H2O2 and lactoferrin release, suggesting that the conversion of phosphatidic acid to diradylglycerol is an important step in PMN activation by TGFâ β1. Overall, these results are similar to those reported for fMLP activation of adherent PMN and suggest that a common pathway is involved in both chemoattractant and cytokine activation. J. Leukoc. Biol. 60: 772â 777; 1996.