Forskolin induces U937 cell line differentiation as a result of a sustained cAMP elevation.

The present study examines the effects of forskolin on U937 cell differentiation. We recently reported that dibutyryl cAMP (dbcAMP), but not cAMP-elevating agents such as histamine, promotes U937 cell differentiation. cAMP production elicited by stimulation of histamine H2 receptors showed a rapid, homologous desensitization, which might explain the dissimilar responses to histamine and dbcAMP. Forskolin induced an increase in cAMP levels in a concentration-dependent manner (EC50=30 microM) for an extended period of at least 24 h. Forskolin but not histamine (up to 100 microM), also inhibited cell growth in a dose-dependent fashion (EC50=22 microM). After 3 days of incubation, 75 microM forskolin induced U937 cell differentiation as judged by an increased rate of reduction of nitrobluetetrazolium (mean+/-S.E.M.: 21.3+/-6.6% in treated cells vs. 3.2+/-1.9% in the control group, P < 0.001) and an augmented chemotactic response to complement 5a (C5a) (33.2+/-5.9% in forskolin-treated vs. 0.34+/-0.12% in control cells, P < 0.01). Furthermore, c-Myc levels decreased following forskolin treatment, while the histamine H2 receptor agonist dimaprit had no effect. We conclude that forskolin induces U937 cell differentiation through a sustained rise in cAMP levels.

Antagonistic actions of phorbol ester in mammalian G0-->G1-->S cell cycle transition.

We have developed a protocol that reveals two antagonistic effects of phorbol-12-myristate-12-acetate (PMA) on the G0-->G1-->S transition of mammalian cell cycle. Balb-3T3 (Clone A31) cells arrested in G0 by serum starvation can be stimulated to traverse the G1 phase and initiate DNA synthesis 12 h later by a 2-h pulse with PMA. In contrast with this early stimulatory effect, PMA has an inhibitory effect when presented to the cells during the last 6 h of G1. PMA is able to inhibit DNA synthesis initiation irrespective of the triggering agent, i.e., serum, fibroblast growth factor, epidermal growth factor, platelet-derived growth factor, or PMA itself (presented as an early pulse). We have established that the critical period for the PMA inhibitory effect is between 6 and 8 h after cell stimulation. This dual effect of PMA is not a peculiarity of Balb-3T3 (clone A31) cells because it is also observed with other fibroblastic cell lines, namely, SWISS 3T3, NIL 8, and RAT 1, and also with the epithelial Y-1 adrenocortical cell line. Treatment with PMA for 0.5 or 2 h activates protein kinase C (PKC) in Balb-3T3-A31 cells, but is not sufficient to down-regulate the enzyme because a second 30-min PMA pulse applied between 6 and 6.5 h activates PKC again. On the other hand, a continuous 6.5-h PMA treatment causes PKC down-regulation; therefore, the inhibitory effect of PMA could be mediated by PKC. Growth factor early response proto-oncogenes c-myc, c-fos, and c-jun are induced transiently by both early and late PMA pulses, suggesting that these genes are not involved in the PMA inhibitory effect.

Haloperidol and oestrogens induce c-myc and c-fos expression in the anterior pituitary gland of the rat.

Decreased dopaminergic and increased oestrogenic effects induce prolactin release and DNA synthesis in prolactin secreting cells of the adult male rats. Oestrogen treatment for 7 days significantly increased the levels of prolactin, c-myc and c-fos mRNAs. The effect of oestrogens was reversed by the administration of the dopaminergic agonist bromocriptine. There was an early gradual increase of c-myc mRNA levels beginning 30 min after the injection of the steroid. c-fos mRNA levels increased sharply 15 min after oestrogen administration and decreased to basal values 15 min later to remain at this level over the period of time evaluated. Administration of the dopaminergic antagonist haloperidol did not change the levels of protooncogenes mRNA. However, in rats previously treated with oestrogens for 7 days c-myc mRNA levels increased 90 min after the injection of haloperidol and decreased to basal values after 2.5 h. c-fos mRNA levels increased sharply 30 min after haloperidol administration and also decreased to basal values 1 h later. We propose that changes in protooncogenes expression may be involved in the stimulation of cell proliferation induced by prolactin secretion.