D. Duval

Dexamethasone-induced inhibition of prostaglandin production does not result from a direct action on phospholipase activities but is mediated through a steroid-inducible factor

Investigations were carried out to define the mechanisms of steroid-induced inhibition of prostaglandin secretion by rat renomedullary cells in tissue culture. Although it was strongly proposed that glucocorticoids may inhibit phospholipase A2 activity, we present several pieces of evidence against a direct action of dexamethasone on phospholipase activities. First, dexamethasone, which significantly decreases the release of labeled material from cells prelabeled with [3H]arachidonate, does not significantly alter the pattern of distribution of the radioactivity among the various classes of cell lipids. In addition, direct measurement of phospholipase A3 activity in dexamethasone-treated cells failed to show any significant decrease in the deacylation capacity. On the other hand, several indications suggest that dexamethasone may induce the secretion of a non-dialysable, transferable factor able to inhibit prostaglandin production, the mechanism of which remains to be investigated.

Differential of sex steroids of prostaglandin secretion by male and female cultured piglet endothelial cells

Abstract The effect of sex steroids, 17β-estradiol and testosterone, on the production of 6-keto-prostaglandin F 1α , prostaglandin F 2α and prostaglandin E 2 was studied in cultures of piglet aorta endothelial cells. In cells isolated from female animals both steroids stimulated the secretion of prostaglandins. In contrast, sex steroids did not affect prostaglandin synthesis by endothelial cells taken from male animals. In addition, female endothelial cells convert testosterone into Estriol, estrone and estradiol. estradiol-induced stimulation of prostacyclin production may explain in part the beneficial role generally attributed to naturally occuring estrogens in cardiovascular diseases.

Calcium and A23187-induced cytolysis of mouse thymocytes

Abstract The cytotoxic effects of ionophore A 23187 were studied in parallel with its action on calcium uptake in isolated mouse thymocytes. Under conditions where the cells were preincubated in a calcium-containing medium prior to ionophore treatment a close relationship could be observed between the extent of cell lysis and the stimulation of calcium uptake in the presence of A 23187 . In addition, increasing concentrations of calcium ions in the incubation medium lead to a pronounced decrease of cell viability and to a stimulation of calcium uptake suggesting that calcium is critical for cell survival.

Glucocorticoid sensitive and resistant cell populations in the mouse thymus

Abstract We have studied in parallel the effects of dexamethasone on in vitro incorporation of [ 3 H]-uridine and [ 3 H]-thymidine and on cell viability in thymocyte subpopulations isolated by centrifugation on a discontinuous bovine serum albumin gradient from intact, adrenalectomized and hydrocortisone-treated mice. In cell populations isolated from intact and adrenalectomized animals, the steroid induces both inhibition of precursor incorporation and a decrease in cell viability, whereas cells isolated from hydrocortisone-treated mice show marked inhibition of precursor incorporation but no cell death after 24 h. Our present results suggest that differences in steroid sensitivity are not directly related to variations in cell receptor content but may be associated with cell differentiation.

Glucocorticoid receptors in corticosensitive and corticoresistant thymocyte subpopulations: II. Studies with hydrocortisone-treated mice

In vitro studies of the residual thymocyte population isolated 48 h after in vivo hydrocortisone injection showed: 1. These cells are partly sensitive to steroid as demonstrated by uridine incorporation inhibition. 2. This residual cell fraction appears heterogeneous after centrifugation on a bovine serum albumine gradient. 3. These cells exhibit low steroid binding and DNA synthesis capacities.

Effect of antiglucocorticoids on dexamethasone-induced inhibition of uridine incorporation and cell lysis in isolated mouse thymocytes

We have compared in isolated mouse thymocytes the action of progesterone, cortexolone, DXH (a 17-beta carboxamide derivative of dexamethasone) and RU 38486 (a new antiglucocorticoid molecule), on dexamethasone-induced inhibition of uridine incorporation and cell lysis, with the affinities of these drugs for glucocorticoid receptors. Our results show that progesterone, cortexolone and DXH which possess similar affinities for glucocorticoid receptors may exhibit variable, weak agonist and antagonist activities according to the parameter studied. RU 38486 was a potent competitor of dexamethasone and was able, when present in a 10-fold excess, to counteract almost completely the inhibitory action as well as the lytic action of 5 X 10(-8) M dexamethasone. This compound which exerts almost no agonist activity may therefore represent a useful tool to investigate the mode of action of antiglucocorticoids.

Glucocorticoid receptors in corticosensitive and corticoresistant thymocyte subpopulations I. Characterization of glucocorticoid receptors and isolation of a corticoresistant subpopulation

1. Separation of mouse thymocytes by centrifugation on a discontinuous bovine serum albumin gradient leads to the isolation of four subpopulations of cells. 2. The study of I13H]uridine incorporation in vitro by these subpopulation in the presence of steroid shows that one of them is corticoresistant. 3. However, the binding capacity of these subpopulations measured by incubation with [3H]dexamethasone is very similar. 4. It is therefore concluded that in mouse thymus, in contrast with lymphoma cells, corticoresistance may not be explained by a defect of cytoplasmic glucocorticoid receptors.

Effect of exogenous prostaglandins and nonsteroidal anti-inflammatory agents on prostaglandin secretion and proliferation of mouse embryo fibroblasts in culture

During wound healing, the positive and negative modulation of fibroblast proliferation may be due, in part, to the high prostaglandin concentration of the inflammatory exudates. In vitro, PGF2 alpha has been shown to stimulate, whereas PGE2 inhibits, the growth of different fibroblast cell lines. Therefore, we have investigated the effect of exogenous prostaglandins (PGs) and of various nonsteroidal anti-inflammatory drugs (NSAIDs) on the proliferation and the prostaglandin (PG) synthesis of normal mouse embryo fibroblasts. PGF2 alpha, 6-keto PGF1 alpha and PGE2 increase fibroblast proliferation. On the other hand, PGF2 alpha increases the synthesis of PGE2 and 6-keto PGF1 alpha while 6-keto PGF1 alpha solely inhibits PGF2 alpha release, PGE2 being inactive. The mouse embryo fibroblasts partially transform the prodrug sulindac sulfoxide in the sulfide form, which completely inhibits PG synthesis, as does indomethacin. In contrast, ibuprofen exerts a differential action, according to the type of PG measured. Among the NSAIDs tested, only sulindac (sulfoxide or sulfide) stimulates fibroblast proliferation and this effect appears independent of an alteration of PG synthesis. Therefore, in this model of normal mouse embryo fibroblasts, while endogenous prostaglandins are not involved in the control of cell proliferation, exogenous PGs have the ability to alter fibroblast growth and PG synthesis.

Glucocorticoid receptors and their functions in lymphocytes.

Abstract Most, if not all. the effects of glucocorticoid hormones on lymphoid tissue are thought to be mediated by an initial step of interaction of the steroid with specific cytoplasmic receptors. We have presented results, obtained using whole cell suspensions of mouse thymocytes. showing however that the number of receptors does not accurately reflect the steroid sensitivity of the target cell: both inhibitory effects of steroid on nucleic acid precursor incorporation and steroid-induced cell lysis are not obligatorily related to receptor levels but more likely to cell differentiation. We have therefore also investigated the mechanism of the immunosuppressive effects of steroids and the role of various factors which could be involved in the proliferation or lysis of mouse thymocytes. Dexamethasone and 25-OH cholesterol are the most potent agents, capable of inhibiting cholesterol biosynthesis in non stimulated thymocytes. Although the effect of dexamethasone. which is blocked by actinomycin D. appears to be mediated through receptor occupancy, the action of 25-OH cholesterol is likely to be due to an inhibition of HMG COA reductase activity. Mitogen induced lymphoblastic transformation of thymocytes is blocked by low concentration of dexamethasone but also by sex steroids at higher concentrations. These compounds, which exert immunosuppressive effects in vivo, inhibit concanavalin A induced transformation at concentrations above 10−5 M, at which concentration they also elicit a rapid and transient decrease of uridine uptake in non stimulated thymocytes. In addition, calcium, which has been considered to play an important role in the mechanism of steroid-induced lysis, appears highly toxic by itself.

Chronic lymphatic leukaemia: cellular effects of glucocorticoids in vitro.

Glucocorticoid receptor levels and steroid induced inhibition of nucleic acid precursors have been examined in lymphocytes from 27 patients at different stages of chronic lymphatic leukaemia. No correlation can be found between the level of glucocorticoid receptors and the stage of the disease. On the other hand, a significant difference (P < 0.02) was found between stage 0 and stage III/IV patients, in terms of the in vitro effect of dexamethasone on [3H]uridine incorporation.