Regulation of the processing and release of tumor necrosis factor α in a human macrophage cell line

The proteolytic cleavage of a variety of membrane proteins, including pro-tumor necrosis factor alpha (pro-TNF-alpha), is induced by various reagents such as phorbol 12-myristate 13-acetate (PMA). In this study we generated a human macrophage cell line that constitutively produces TNF-alpha, and examined how the processing and release of TNF-alpha are regulated. The processing and release of TNF-alpha were enhanced by PMA through a protein kinase C-dependent pathway. Although only hydroxamate matrix metalloproteinase (MMP) inhibitors inhibited the basal processing and release of TNF-alpha, the PMA-induced processing and release of TNF-alpha were inhibited not only by MMP inhibitors but also by 1,10-phenanthroline, 3,4-dichloroisocoumarin (3,4-DCI), iodoacetamide, and Nalpha-p-tosyl-L-phenylalanine chloromethyl ketone (TPCK). Hydroxamate MMP inhibitors and 1,10-phenanthroline inhibited the processing of TNF-alpha on the cell surface, whereas 3,4-DCI, iodoacetamide, and TPCK inhibited the transport of TNF-alpha to the cell surface. These results suggest that serine and/or cysteine protease(s) may be involved in PMA-induced processing and/or transport to the cell surface.