The interaction between the immunosuppressive effects of glucocorticoids and the mitogenic effects of prolactin (PRL) were examined in Nb2 lymphoma cells, a pre-T cell line. The synthetic glucocorticoid, dexamethasone (Dex), caused a concentration-dependent (6.25-200 nM) inhibition of basal and ovine PRL (oPRL)-stimulated Nb2 cell proliferation. Although Dex was antiproliferative, the steroid had no effect on cell viability in the presence of PRL. However, when PRL was omitted from the medium, Dex increased the proportion of dead Nb2 cells by 24 hr in a concentration (25-200 nM)-dependent fashion without affecting total cell number. The antiproliferative and cytolytic effects of Dex were mimicked by other corticosteroids (cortisol, corticosterone, aldosterone, and deoxycorticosterone) in the expected order of glucocorticoid potency, but not by other steroids (17-beta-estradiol, progesterone, testosterone, 5-alpha-dihydrotestosterone, and dehydroepiandrosterone) or triiodothyronine. In addition, the antiproliferative and cytolytic effects of glucocorticoids were antagonized by the glucocorticoid receptor antagonist RU 486. Since corticosteroid-induced cytolysis was apparent only in the absence of mitogen, the anticytolytic effects of oPRL were tested. In the presence of Dex (100 nM), oPRL (25-1600 pg/ml) caused a concentration-dependent inhibition of cytolysis without changing cell number. Other lactogenic hormones (human growth hormone, human placental lactogen, rat PRL), but not trophic nonlactogenic hormones (rat growth hormone, human chorionic gonadotropin, ACTH), also inhibited Dex (100 nM)-induced cytolysis. Agarose gel electrophoresis of DNA extracted from Nb2 cells revealed that within 12 hr, 100 nM Dex induced DNA fragmentation, indicative of programmed cell death or apoptosis. Coincubation of cells with Dex and oPRL (1 ng/ml) inhibited Dex-induced fragmentation of Nb2 cell genomic DNA. These studies reveal a complex interaction between glucocorticoids and PRL in Nb2 cells. Although a glucocorticoid receptor-mediated antiproliferative effect is evident, PRL (at concentrations that usually stimulate cell proliferation) has the capacity to protect the cell against glucocorticoid-receptor-mediated induction of apoptosis.

Cultured Nb2 node rat lymphoma cells require lactogenic hormone for their proliferation. We reported previously that dexamethasone (Dex) inhibits prolactin (PRL)-induced mitogenesis and, in the absence of mitogen, induces apoptosis of Nb2 cells. Both antiproliferative and cytolytic effects of Dex on Nb2 cells appear to involve glucocorticoid (Type II) receptor mediation. In this study, we compared Dex effects in PRL-dependent Nb2 cells (Nb2) with SFJCD1 (SF), a clone of Nb2 cells that proliferates independently of exogenous PRL. Proliferative assays involved a 72-hr incubation in a chemically defined, serum-free medium where ovine PRL (1 ng/ml) was added to Nb2 cells but not to SF cells. Both cell lines were responsive to the antiproliferative effects of Dex in a dose (6.25-200 nM)-dependent fashion of comparable sensitivity and magnitude. Co-incubation with the glucocorticoid receptor antagonist, RU 486, prevented the antiproliferative effect of Dex in both cell lines. In the same medium devoid of PRL, Dex was cytolytic to Nb2 cells and fragmented DNA in a fashion reflective of apoptosis, but was ineffective in SF cells. A dual chamber incubation system revealed no evidence that SF cells produced cytokines that were mitogenic or anticytolytic to Nb2 cells. Both Nb2 and SF cells fragmented DNA in a fashion indicative of apoptosis in the presence of the Ca2+ ionophore, A23187 (1 microM). These studies reveal a basic difference in glucocorticoid responsiveness between the PRL-dependent Nb2 cell line and its PRL-independent subclone, SF. While both cell lines exhibit functional glucocorticoid receptors and the necessary intranuclear machinery for apoptosis, the pathway mediating the latter is inhibited or dysfunctional in SF cells.