Human Monocytes and Neutrophils Store Transforming Growth Factor-α in a Subpopulation of Cytoplasmic Granules

Abstract

Transforming growth factor-α (TGF-α) exerts several effects on target cells, such as neovascularization promotion and mitogenic signalling. Using immunoelectron microscopy, we show that monocytes and neutrophils, store TGF-α in cytoplasmic granules. In monocytes, TGF-α did not colocalize with components of peroxidase-positive granules or with albumin of secretory vesicles. Furthermore, no colocalization of TGF-α with components of azurophilic or specific granules or secretory vesicles was observed in neutrophils. Activated monocytes and tissue-macrophages contained much less TGF-α–positive granules, suggesting TGF-α release. Western blot analysis showed a protein of 10 kD in lysates of monocytes. TGF-α mRNA was detected in monocytoid cells from the bone marrow by in situ hybridization. This study shows for the first time that monocytes and neutrophils contain TGF-α in all stages of maturation and that TGF-α in monocytes is stored in a large population of peroxidase-negative granules suggesting a function for these granules. Monocytes and neutrophils are important effector cells in inflammatory reactions. The present finding that these cells contain TGF-α might explain complications such as fibrosis and neoplastic transformation, caused by chronic inflammation.

Functional analysis of monocyte MHC class II compartments

“…The ultrastructure of these compartments closely resembles that of the type II MHC class II-positive compartments previously reported in mast cells, characterized by an electron-dense core surrounded by several vesicles juxtaposed to the limiting membrane (19). EDBs were previously described in monocytes as either myeloperoxidase-positive or -negative compartments (20). The former were reported to be particularly enriched in cathepsin G, elastase, and myeloperoxidase, whereas the latter were TGF-␣ positive but myeloperoxidase negative (20).…”

“…EDBs were previously described in monocytes as either myeloperoxidase-positive or -negative compartments (20). The former were reported to be particularly enriched in cathepsin G, elastase, and myeloperoxidase, whereas the latter were TGF-␣ positive but myeloperoxidase negative (20). We now report that the myeloperoxidase-positive EDBs are Lamp-1-and MHC class II-positive compartments, as determined by immunogold staining as well as analysis of endosomal organelles following subcellular fractionation.…”

“…In monocytes, EDBs have been previously subdivided as myeloperoxidase positive or negative (20). The former contains several proteolytic enzymes (cathepsins, elastase myeloperoxidase), whereas the latter have been characterized as storage granules for TGF-␣ (20). Thus, as a next step, the colocalization of HLA-DR with myeloperoxidase or TGF-␣ was examined.…”

“…On the other hand, in granulocytes both true lysosomes and secretory lysosomes are present (17)(18)(19). Secretory granules have been previously identified in monocytes, some containing transforming growth factor ␣ (TGF-␣) and others positive for myeloperoxidase, cathepsin G, and elastase (20). Whether any of these subtypes are also MHC class II positive is still unknown.…”

PIP2 signaling in lipid domains: a critical re-evaluation

“…Transfected cells were fixed for 24 h in 4% paraformaldehyde in 0.1 M PHEM buffer (240 mM PIPES, 100 mM HEPES, 8 mM MgCl 2 , 40 mM EGTA, pH 6.9) and then processed for ultrathin cryosectioning as previously described (Calafat et al, 1997). Ultrathin frozen sections were incubated at room temperature with mouse monoclonal anti-HA 12CA5 (Boehringer) or rabbit anti-GFP (Clontech), followed by incubation with 10 nm protein A-conjugated colloidal gold (EM Lab., Utrecht University, The Netherlands) as described (Calafat et al, 1997). After immunolabeling, cryosections were embedded in a mixture of methylcellulose and uranyl acetate and examined with a Philips CM 10 electron microscope (Eindhoven, The Netherlands).…”