Tumor-induced variations in a high molecular weight inhibitory monokine

A soluble inhibitor of T cell proliferation was demonstrated in splenic and peritoneal macrophage (M phi) culture supernatants and was variably glycosylated during tumor development. This factor(s) inhibited mixed lymphocyte reaction- (MLR) and concanavalin A-induced T cell proliferation and was not prostaglandin E2 (PGE2), as demonstrated by size (nondialyzable, found in a 30 kilodalton [kd] ultrafiltration retentate and in fractions corresponding to greater than 67 kd on S-200 Sephacryl chromatography) and time course of activity in the MLR. Concentrated supernatants were PGE2-free and yet inhibited proliferation in the MLR. Isoelectric focusing (IEF) revealed normal and tumor-bearing host (TBH) concentrated M phi supernatants' major peaks of inhibitory activity differed in charge, with a pI of 6.5-7.6 for normal hosts and 4.0-6.0 for TBH. Activity in TBH M phi supernatants was found primarily in fractions eluting from hydroxylapatite at 0.3 M sodium phosphate buffer, pH 7.3, was resistant to proteolytic enzymes, but was sensitive to neuraminidase. In contrast, inhibitory activity in normal host M phi supernatants eluted from hydroxylapatite at 0.6 M sodium phosphate buffer and was not sensitive to neuraminidase. Thus, variable glycosylation (presence and absence of sialic residues) could account for the charge difference seen in IEF and may have a bearing on tumor-induced hyporesponsiveness. Kinetic addition of supernatants to the MLR revealed PGE2 may be required for inhibitory activity to be manifested early (0 and 24 h) but not if the high molecular weight (mw) inhibitor was added late (48 and 72 h post initiation). Both normal and TBH M phi supernatants suppressed the generation of interleukin 2 (IL 2) with a dose- and time-dependent difference. Cell-cycle analysis of mitogen-stimulated cells treated with normal and TBH M phi supernatants revealed that TBH M phi supernatants enhanced cell-cycle progression when measured early, but that both normal and TBH M phi supernatants suppressed the number of cells in S phase when measured late in the assay. The production of a high mw inhibitor by both normal and TBH M phi could suggest a homeostatic mechanism, which was upset by increased PGE2 production and production of an inhibitor containing sialic acid, tipping the balance in favor of immunosuppression.