Regulation of sex steroid formation by interleukin-4 and interleukin-6 in breast cancer cells.

Sex steroids play a predominant role in the development and differentiation of normal mammary gland as well as in the regulation of hormone-sensitive breast cancer growth. There is evidence suggesting that local intracrine formation of sex steroids from inactive precursors secreted by the adrenals namely, dehydroepiandrosterone (DHEA) and 4-androstenedione (4-dione) play an important role in the regulation of growth and function of peripheral target tissues, including the breast. Moreover, human breast carcinomas are often infiltrated by stromal/immune cells secreting a wide spectra of cytokines. These might in turn regulate the activity of both immune and neoplastic cells. The present study was designed to examine the action of cytokines on 17beta-hydroxysteroid dehydrogenase (17beta-HSD) and 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD) activities in human breast cancer cells. The various types of human 17beta-HSD (five types) and 3beta-HSD (two types), because of their tissue- and cell-specific expression and substrate specificity, provide each cell with necessary mechanisms to control the level of intracellular active androgens and estrogens. We first investigated the effect of exposure to IL-4 and IL-6 on reductive and oxidative 17beta-HSD activities in both intact ZR-75-1 and T-47D human breast cancer cells. In ZR-75-1 cells, a 6 d exposure to IL-4 and IL-6 decreased E2-induced cell proliferation, the half maximal inhibitory effect being exerted at 88 and 26 pM, respectively. In parallel, incubation with IL-4 and IL-6 increased oxidative 17beta-HSD activity by 4.4- and 1.9-fold, respectively, this potent activity being observed at EC50 values of 22.8 and 11.3 pM, respectively. Simultaneously, reductive 17beta-HSD activity leading to E2 formation was decreased by 70 and 40% by IL-4 and IL-6, respectively. Moreover, IL-4 and IL-6 exerted the same regulatory effects on 17beta-HSD activities when testosterone and 4-dione were used as substrates, thus strongly suggesting the expression of the type 2 17beta-HSD ZR-75-1 cells. In contrast, in T-47D cells, IL-4 increased the formation of E2, whereas IL-6 exerts no effect on this parameter. However, we found that T-47D cells failed to convert testosterone efficiently into 4-DIONE, thus suggesting that there is little or no expression of type 2 17beta-HSD in this cell line. The present findings demonstrate that the potent regulatory effects of IL-4 and IL-6 on 17beta-HSD activities depend on the cell-specific gene expression of various types of 17beta-HSD enzymes. We have also studied the effect of cytokines on the regulation of the 3beta-HSD expression in both ZR-75-1 and T-47D human breast cancer cells. Under basal culture conditions, there is no 3beta-HSD activity detectable in these cells. However, exposure to IL-4 caused a rapid and potent induction of 3beta-HSD activity, whereas IL-6 failed to induce 3beta-HSD expression. Our data thus demonstrate that cytokines may play a crucial role in sex steroid biosynthesis from inactive adrenal precursors in human breast cancer cells.

Growth-independent induction of spermidine transport by IL-4 and IL-13 in ZR-75-1 human breast cancer cells.

Polyamine transport is strongly induced by insulin and estradiol (E2) in ZR-75-1 human breast cancer cells. Because signal transduction mechanisms of insulin and interleukin-4 (IL-4) partly overlap, we have compared the ability of these agents as well as that of interleukin-13 (IL-13), a cytokine that often mimics IL-4, to modulate spermidine transport in these cells. In the presence of E2, insulin increased DNA content and the rate of [3H]spermidine uptake by 2.1- and 3.7-fold, respectively, after an 8-day incubation, whereas the sole addition of IL-4 caused a quantitatively similar induction of [3H]spermidine uptake while leaving cell growth unaffected. No comparable induction of spermidine transport was observed with interleukins-1 alpha and -6, and the effect of IL-4 was not additive to that elicited by insulin plus E2. IL-4 and IL-13 stimulated [3H]spermidine uptake to a comparable extent, with half-maximal effects observed at 80 and 400 pg/ml, respectively. Interferon-gamma inhibited IL-4- and IL-13-dependent spermidine uptake to a much greater extent than basal or insulin-induced transport of the polyamine. IL-4 and IL-13 increased the Vmax and K(m) of [3H]spermidine uptake by about 4- and 2.5-fold, respectively. Na(+)-dependent amino acid uptake was increased by insulin but not by IL-4 or IL-13, indicating that the cytokines do not induce a general increase in membrane transport activity. IL-4 and IL-13 did not interfere with feedback inhibition of polyamine uptake, and only modestly decreased polyamine content after prolonged incubation, suggesting that these cytokines stimulate spermidine uptake by increasing total transport capacity rather than by repressing and endogenous inhibitor.

Interleukin-4 and interleukin-13 inhibit estrogen-induced breast cancer cell proliferation and stimulate GCDFP-15 expression in human breast cancer cells.

Human breast carcinomas are frequently infiltrated by inflammatory cells secreting several cytokines which may regulate the activity of both immune cells and neoplastic cells. The present study was designed to examine the potential action of interleukin-4 (IL-4) and interleukin-13 (IL-13) in human breast cancer cells. Exposure of ZR-75-1 breast cancer cells to IL-4 or IL-13 for 10 days decreased the amplitude of the mitogenic action of 17 beta-estradiol by 75% and 55%, respectively, while these cytokines failed to change basal cell proliferation. These cytokines also exerted a similar action in T-47D cells. Exposure to IL-4 or IL-13 markedly increased gross cystic disease fluid protein-15 (GCDFP-15) release in both ZR-75-1 and T-47D cells. The half-maximal stimulatory effects of IL-4 and IL-13 on GCDFP-15 secretion were exerted at respective values of 16 +/- 3 pM and 91 +/- 8 pM in T-47D cells incubated for a period of 10 days. The effect of IL-13 was not additive to that elicited by IL-4, whereas the stimulation of GCDFP-15 release by these interleukins were additive to that exerted by maximally effective concentrations of the androgen dihydrotestosterone and the synthetic glucocorticoid dexamethasone. Furthermore, exposure of ZR-75-1 cells of IL-4 and IL-13 increased GCDFP-15 mRNA levels by 5.5- and 6.0-fold, respectively. The present results demonstrate that IL-4 and IL-13 may decrease estrogen-induced breast cancer cell proliferation and induce the expression of a breast cancer marker, thus strongly suggesting that breast cancer cells are targets of both IL-4 and IL-13 action.

Interleukin-6 inhibits the potent stimulatory action of androgens, glucocorticoids and interleukin-1 alpha on apolipoprotein D and GCDFP-15 expression in human breast cancer cells.

Our study was designed to investigate the potential interaction between steroid hormones and interleukin-6 (IL-6) in the regulation of apolipoprotein D (apo-D) and gross cystic disease fluid protein 15 (GCDFP-15) expression in ZR-75-1 human breast cancer cells. We first observed that exposure to IL-6 for 6-14 days decreased basal apo-D and GCDFP-15 secretion by 50% and 23%, respectively. In the same experiment, such treatment with IL-6 decreased cell proliferation by approximately 40% after 6 and 14 days of incubation. Exposure to IL-6 markedly decreased dihydrotestosterone (DHT)-induced apo-D and GCDFP-15 release, with a half-maximal effect measured at 13 U/ml. A similar inhibitory action of IL-6 was observed on the glucocorticoid dexamethasone (DEX)-induced apo-D and GC-DFP-15 secretion. The sensitivity of the apo-D and GCDFP-15 response to the stimulatory action of DHT or DEX was, however, not changed by concomitant exposure to IL-6. The inhibitory effect of IL-6 on the secretion of these two biochemical markers was additive to that of 17 beta-estradiol. In addition, IL-6 blocked the stimulatory effect of interleukin-1 alpha (IL-1 alpha) on apo-D and GCDFP-15 secretion. Our results show that IL-6 is a potent inhibitory of basal as well as androgen-, glucocorticoid- and IL-1 alpha-induced apo-D and GCDFP-15 secretion in ZR-75-1 human breast cancer cells, while cell proliferation is inhibited by this cytokine.

Potent stimulatory effect of interleukin-1 alpha on apolipoprotein D and gross cystic disease fluid protein-15 expression in human breast-cancer cells.

To better understand the multiple hormonal control of the expression of apolipoprotein D (apo-D) and gross cystic disease fluid protein-15 (GCDFP-15, also designated prolactin-inducible protein), which are 2 major proteins found in benign breast-disease fluid, we investigated their regulation by interleukin-1 alpha (IL-1 alpha) in the presence or absence of steroid hormones in ZR-75-1 human breast cancer cells. Exposure of these cells to IL-1 alpha decreased basal cell proliferation by half and markedly reduced the mitogenic action of 17 beta-estradiol (E2), the half-maximal inhibitory effect being exerted at 1.5 pM. In parallel, IL-1 alpha stimulated apo-D and GCDFP-15 secretion with a similar potency. The antiproliferative effect of IL-1 alpha was additive to the inhibition of cell proliferation caused by dihydrotestosterone (DHT) or the glucocorticoid dexamethasone (DEX). In parallel, IL-1 alpha-induced stimulation of apo-D and GCDFP-15 secretion was additive to that exerted by DHT or DEX. The sensitivity of the apo-D and GCDFP-15 responses to the stimulatory action of DHT or DEX was not changed by the presence of IL-1 alpha. IL-1 alpha also increased apo-D and GCDFP-15 mRNA levels. The present findings demonstrate the potent stimulatory effect of IL-1 alpha on basal as well as androgen- and glucocorticoid-induced apo-D and GCDFP-15 expression. The present data strongly suggest that IL-1 alpha and steroids may modulate the secretion of these 2 proteins through different transduction pathways.

Action of spontaneously produced beta interferon in differentiation of embryonal carcinoma cells through an autoinduction mechanism.

In the current study, we have addressed the role of interferons (IFNs) in controlling the differentiation of pluripotent P19 embryonal carcinoma (EC) cells. Blocking IFN activity in the culture medium of differentiating cells with antibodies leads to a strong decrease in the degree of differentiation. The antibodies are active for a relatively short time. During this time, IFN-beta mRNA can be detected in the differentiating cells, as can increases of IFN stimulation response element-binding activity and NF-KB. The timing of IFN action also coincides with the accumulation of cytoplasmic double-stranded RNA (dsRNA) and with a drop in dsRNA unwindase-modificase activity. A model for the involvement of autoinduction of IFN by intracellular dsRNA in the control of differentiation in this system is presented.