Correlation between spontaneous oscillations of cytosolic free Ca2+ and tumor necrosis factor-induced degranulation in adherent human neutrophils

By using a hemolytic plaque assay to detect release of lactoferrin and myeloperoxidase, tumor necrosis factor (TNF) was shown previously to induce secretion of these granule proteins from single adherent neutrophils. Secretion was inhibited by loading neutrophils with calcium chelators, indicating a crucial role of cytosolic free Ca2+] in the signal transduction mechanism of TNF. In the present study, using a microfluorometer technique to follow changes in the cytosolic free Ca2+] in single adherent neutrophils, we were not able to detect any TNF-induced Ca2+] transients. However, these adherent cells exhibited spontaneous oscillations of their cytosolic free Ca2+], as previously reported (Jaconi, M.E.E., Rivest, R.W., Schlegel, W., Wollheim, C.B., Pittet, D., and Lew, P.D. (1988) J. Biol. Chem. 263, 10557-10560). A close correlation was found between a reduced oscillatory activity of cytosolic free Ca2+] and a reduced ability of TNF to induce degranulation, by reducing the extracellular Ca2+] or loading the cells with a calcium chelator (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid). In addition, when the cells were incubated at 37 degrees C for 3 h there was a parallel decline in the spontaneous oscillatory activity of cytosolic free Ca2+] and TNF-induced secretion of lactoferrin. Control experiments showed that phorbol 12-myristate 13-acetate-induced secretion was not affected under the same conditions, indicating that the secretory process per se was not disturbed. We conclude that TNF by itself does not give rise to any changes of the cytosolic free Ca2+] but that the spontaneous oscillatory activity of cytosolic free Ca2+] in adherent neutrophils is necessary for TNF-induced degranulation.