Relationship of cell cycle to recovery of IL 2 activity from human mononuclear cells, human and mouse T cell lines.

Abstract

Abstract Interleukin 2 (IL 2) supernatant activity was assessed in cultures of human mononuclear cells stimulated with either phorbol myristic acetate (PMA) and/or with a lectin such as concanavalin A (Con A) or phytohemagglutinin (PHA). The degree of stimulation was measured by using flow cytometry of acridine orange stained cells, which differentiates the cellular RNA content of early and late G1 cells from each other and from resting cells in the G0 phase. Supernatants of unstimulated human mononuclear cells in either the Go phase of the cell cycle or of cells stimulated with PMA, which were shifted into the early G1 phase, did not contain IL 2 activity. In contrast, supernatants of mononuclear cells stimulated with Con A or PHA contained IL 2 activity, and these lectin-treated cells were in the late G1 phase. The addition of PMA to the fectin-stimulated cultures increased supernatant IL 2 activity in association with a concomitant increase in cells in the late G1 phase. However, the total supernatant IL 2 activity was also influenced by absorption by lymphocytes in late G1, because lectin-stimulated lymphocytes even when arrested at the G1/S interphase by hydroxyurea adsorbed IL 2. Unstimulated or PMA stimulated cells in G0 or early G1 did not absorb IL 2. Furthermore, maximal utilization of IL 2 occurred during the S phase because the elimination of proliferating cells in stimulated cultures by hydroxyurea was accompanied by a further increase in IL 2 supernatant activity rather than the progressive decrease in IL 2 activity that is associated with proliferation. By using several human T lymphocyte cell lines and murine EL 4 thymoma cells, it was found that lectin and PMA enhanced IL 2 production and was associated with decreased DNA synthesis. Thus, cell lines also produced more IL 2 when lectins or PMA interfered with the cell cycle of these cells. These observations suggested that IL 2 supernatant activities are influenced by: i) the greater rate of production in the late G1 phase that is prolonged by PMA and high doses of mitogens, ii) the absorption of IL 2 before the S phase; and iii) the “utilization” of IL 2 in the S phase.