Direct Effects Of Steroids Research Articles

8693 Dexamethasone Increases Bone Mass In Mice

Abstract Disclosure: F. Sultana: None. M.L. Temple: None. S. Kim: None. V. Ryu: None. F. Korkmaz: None. A. Gumerova: None. P. Georgii: None. U. Cheliadinova: None. D. Vasilyeva: None. V. Laurencin: None. R. Witztum: None. O. Barak: None. L. Cullen: None. A.R. Pallapati: None. S. Rojekar: None. J. Gimenez Roig: None. D. Lizneva: None. W. Zhou: None. T. Yuen: None. M. Zaidi: None. Severe osteoporosis and bone fractures are strongly associated with long-term or recurring glucocorticoid use in people. Steroids have traditionally been used as an anti-inflammatory and immunosuppressive agent, and while their use has been widely implicated in causing bone loss by suppressing osteoblastic bone formation, recent data have consistently shown mixed results in mice. It is also important to stress that dexamethasone (DEX) in particular has been used for over three decades in cell cultures to stimulate osteoblast formation. We therefore posit that mechanisms other than direct steroid effects on osteoblasts contribute to bone loss, and that, at least physiologically, steroids stimulate (rather than inhibit) bone formation. Here, we have aimed to clarify the effects of DEX on bone metabolism. We first studied the effects of low and high doses of DEX (1 mg/kg and 10 mg/kg), given daily as intraperitoneal injections to 8-week-old C57BL/6 mice. Micro-CT showed that DEX markedly increased bone microarchitecture, including the bone volume fraction, connectivity density, and trabecular number, while decreasing trabecular spacing in femoral epiphyses compared with vehicle-injected controls. Serum levels of adrenocorticotropic hormone (ACTH) and corticosterone did not change following DEX. These findings suggest that both low and high DEX doses are anabolic to the skeleton. Reaffirming these data, we found that adrenalectomy (with salt replacement) caused a significant decline in bone mineral density. Likewise, Tpit-/- mice, in which ACTH and corticosterone are both reduced, display reduced bone microarchitectural parameters, together with low mineralizing surface and bone formation rates on dynamic histomorphometry, impaired osteoblast formation in Cfu-f cultures, and lowered expression of the osteoblastogenesis genes Bmp2, Ibsp, Runx2 and Sp7. Finally, deleting the glucocorticoid receptor (Gr) specifically in osteoclasts in Acp5-Cre;Grfl/fl mice had no effect on microarchitectural parameters at the spine or proximal tibial metaphysis. However, and importantly, absent GR in the osteoclast, DEX increased bone mass significantly, suggesting that its notable anabolic action was mediated by the osteoblast. Our findings challenge the longstanding premise that steroids directly impact the osteoblast to cause bone loss, and that other mechanisms, such as reduced ACTH levels and/or central effects of DEX, may play a key role. Presentation: 6/1/2024

Steroid hormones as interkingdom signaling molecules: Innate immune function and microbial colonization modulation

Steroid hormones e.g., estrogen, progesterone, testosterone and dehydroepiandosterone, act as inter-kingdom quorum chemical signaling compounds. All steroids examined exhibit a steroid concentration specific bi-functionality. At one end of the spectrum, the steroids enhance expression of virulence-associated behaviors, most specifically, increased rate of replication and adherence to surfaces. In contrast, the hormones also function as innate immune system compounds providing first-line protection against essential pathogen behaviors e.g., biofilm formation, which plays a role in initiation of the vast majority of infectious processes, especially chronic infections. Mechanistically, this protection is mediated by both direct effects of steroids on microbes, as well as indirect actions which result in expression of nitric oxide at levels reported to inhibit proper biofilm formation and cause return of sessile cells to a planktonic state.

Impact of Steroids on Hepatitis C Virus Replication in Vivo and in Vitro

Chronic hepatitis C virus (HCV) infection is the leading indication for liver transplantation. Transplantation outcome is often compromised by a rapid re-infection of the graft. Several factors have been implicated in the increased severity of recurrence, including steroid-based immunosuppression. Evidence suggests that steroid boluses used to treat acute rejection are associated with an increase in HCV viral load and the severity of recurrence. Two possible mechanisms for a steroid-mediated effect on HCV viral loads can be postulated, the first being a direct effect of steroids on the virus by enhancing its replication. The second, an indirect effect due to the suppression of the HCV immune response, allows unrestricted HCV replication. To investigate the direct effect of steroids on HCV replication, dexamethasone (Dex) and prednisolone (Pred) were tested in an in vitro replicon model. HCV replication was assessed on the basis of luciferase reporter expression (luminescence) and HCV RNA (RT-PCR). At clinically relevant concentrations (1-10 nM), treatment with both Dex and Pred did not enhance, but resulted in a slight reduction of relative luciferase activity (HCV replication), which was independent of increased cellular protein content and reduced cell proliferation. This minor reduction of HCV replication was confirmed by RT-PCR showing more than 41% reduction in HCV RNA levels. In conclusion, despite clinical evidence that the use of steroids aggravates recurrence of HCV, our in vitro study suggests that there is no direct stimulatory effect of steroids on the replication of HCV. As such, the increased viral loads after high-dose steroid treatment are more likely due to a downregulation of the immune response. In such patients, a dampened immune response allows viruses like HCV to replicate free of immune-mediated killing of their host cells. When a change occurs, such as a tapering or an alteration of immunosuppressant drugs, the immune system reinitiates and vigorously attempts to control the virus, resulting in acceleration of liver damage. Therefore, either steroid avoidance or maintaining low levels, coupled with a slow tapering of corticosteroids, may be beneficial to HCV-infected transplantation recipients.

Is there a Glucocorticoid Receptor in the Bovine Lens?

Prolonged glucocorticoid therapy is a risk factor for cataract development. The mechanism remains unknown. If cataract results from the direct effect of steroids on lens function, a glucocorticoid receptor is required. In order to determine whether such a receptor was present in the bovine lens, metabolic and steroid binding experiments were undertaken. Cultured bovine lens epithelial cells were exposed to 10−4and 10−8M dexamethasone or prednisolone and the uptake and incorporation of14C leucine,14C glucose and3H thymidine, examined. Neither glucocorticoid affected cell protein synthesis or glucose uptake. Both dexamethasone concentrations and the lower concentration of prednisolone had no effect on thymidine uptake or incorporation, however, the 10−4M prednisolone exposure reduced these by 15±5%. This regulation is thought to be due to membrane fluidity changes and not the action of the glucocorticoid receptor. As the glucocorticoid receptor is very heat labile in vitro, the effects of increasing temperature on dexamethasone binding by proteins from lens epithelium, lens nucleus and liver were examined. At 0°C, lens epithelial extract bound nine-fold more dexamethasone than liver extract. After exposure to 37°C, liver binding decreased by 66% whereas that for lens epithelium increased by 18%. For both lens extracts, steroid binding increased with temperature up to 50°C. Scatchard analysis of the steroid binding kinetics showed there to be no high affinity sites in lens epithelial extract, with the binding best described as a non-specific partitioning event. Western blotting with a specific glucocorticoid receptor antibody revealed protein bands of approximately 94 and 79kDa in liver, which is known to contain significant levels of receptor. No immunoreactivity was observed for lens epithelial extract. Therefore, within the limits of detection, these results suggest the bovine lens does not contain a glucocorticoid receptor. This raises questions about the validity of receptor-mediated mechanisms proposed for cataract development.

Regulation of the Oxytocin Receptor in Bovine Reproductive Tissues and the Role of Steroids

ContentsThe oxytocin receptor (OTR) plays a central role in the functioning of the reproductive tract in the female ruminant. During the oestrous cycle endometrial OTR is suppressed through most of the cycle, probably in a progesterone‐dependent manner, to be massively up‐regulated during the few days before and after oestrus. At this time, luteal OT triggers a positive feedback loop, the endometrial OTR mediating the release of luteolytic PGF2α which feeds back to the corpus luteum to cause the secretion of more OT and to induce luteolysis. This feedback is only interrupted if a newly formed blastocyst is present. Ruminant blastocysts secrete the cytokine interferon‐τ, which interacts directly with the endometrial epithelial cells, preventing the up‐regulation of OTR in late luteal phase. During early pregnancy the levels of OTR in endometrium remain very low whereas myometrial OTR are less affected. OTR in endometrium are up‐regulated around day 50 and then progressively increase to reach a maximum at the onset of labour; those in the myometrium are upregulated somewhat later in gestation, around day 100. The OTR in the cervix are differently regulated: their concentrations are very low through most of the cycle and gestation and do not increase until pro‐oestrus and at parturition when they abruptly rise to high values. Ligand binding to the mucosal and endometrial OTR causes specific release of prostaglandins, which in turn activate paracrine pathways in the surrounding tissues to induce softening of the connective tissue of the uterine wall during pregnancy, and of the cervix at term. Additionally, local uterine contractions induced by OT and potentiated by prostaglandins may serve a physiological function during pregnancy by improving uterine blood flow. Due to the appearance of gap junctions in myometrium at term the OT‐induced contractions become synchronized and well propagated and because of concomitant cervical softening the contractions become expulsive. Although OTR‐dependent physiology correlates well with the levels of circulating gonadal steroids, there is no evidence for any direct effect of steroids either on the OTR gene or OTR protein. Instead, progesterone appears to exert an indirect inhibitory effect on the OTR gene, probably mediated via other cell types. This is in contrast to the inhibition by interferon‐τ which appears to activate a pathway which directly suppresses the OTR gene expression. The important role of OTR in female uterine physiology, and its value as an indicator molecule, make it imperative to understand the complex molecular mechanisms underlying its regulation. These are likely to be of fundamental importance for many aspects of uterine function both in ruminants and in other species, including the human.

Sexual dimorphism of steroid hormone receptor messenger ribonucleic acid expression and hormonal regulation in rat vascular tissue.

Evidence has accumulated suggesting that steroid hormones have a direct effect on the vascular system. Of special interest is the protective effect of estrogens against cardiovascular diseases. One of the aims of the present study was to investigate the messenger RNA (mRNA) expression of the five classic steroid hormone receptors in the great vessels of the rat (aorta, vena cava, and vena portae) to provide a basis to analyze direct steroid effects on vascular tissue. By applying reverse transcription-PCR we compared the expressions of the steroid hormone receptor mRNAs in the respective vessels of male and female rats. Sex differences in the mRNA levels of the mineralocorticoid (MR), estrogen (ER), and progesterone (PR) receptors were found in venous vessels, but not in the aorta. Focussing on the vena cava in the female rat, we investigated whether the ER is active in the vasculature by analyzing regulation of the PR gene. This gene is known to be regulated by estrogens in classic target organs. PR mRNA expression in venous vessels of female rats decreased after ovariectomy. This effect was reversed by chronic sc treatment with estradiol (E2; 1 microgram/animal day). Progester-one (10 mg/animal day, sc) partly inhibited the effect of E2. Besides E2, the partial agonist tamoxifen stimulated PR mRNA expression in ovariectomized rats, whereas the pure antiestrogen ZM 182780 remained inactive in this experiment. Both E2 and tamoxifen caused an autologous down-regulation of ER mRNA. In conclusion, our study demonstrates for the first time that not only the aorta, but also the venous vessels, represented by the vena cava and the portal vein of the rat, are targets for steroid hormones. The ER in vascular tissue is functionally active and mediates direct modulatory effects of estrogens on gene expression in vascular cells.

The neurosteroid progesterone increases the expression of myelin proteins (MBP and CNPase) in rat oligodendrocytes in primary culture

Steroid hormones are known to act in the central nervous system (CNS), affecting brain development and behavior. They have profound influences on the growth, maturation, differentiation and functioning of brain cells. The biological effects of these steroids are mediated by specific high-affinity intracellular receptors, and their presence in the brain has been described by several groups (for review see McEwen et al., 1982). In earlier studies, we have shown that newborn rat glial cells in primary culture can synthesize pregnenolone and progesterone (P) and that these brain cells contain 4 classes of steroid hormone receptors, progesterone, glucocorticoid, estrogen and androgen receptors (PR, GR, ER and AR). After treating glial cells with estrogen, only the PR was induced, and this induction was most significant in cultures established from female rat pups (Jung-Testas et al., 1991). In agreement with the presence of intracellular steroid hormone receptors, we have shown direct effects of steroids on cell multiplication, morphology and differentiation. For instance, after treatment of cells with P, an increased synthesis of myelin proteins was observed in oligodendrocytes (Jung-Testas et al., 1992). In the CNS, oligodendrocytes are responsible for myelin formation and maintenance. In rodents, central myelin is composed of about 70% lipids, among which galactocerebroside (Gal C) is a specific marker for oligodendrocytes (Raft et al., 1987), and of about 30% proteins, mainly proteolipid protein, myelin

Mitochondrial genes as sites of primary action of steroid hormones

Steroid and thyroid hormones act on nuclear gene transcription by activating protein receptors, which in turn bind to hormone response elements (HREs). Among the cell-specific processes regulated by steroid receptors is energy metabolism through increased synthesis of respiratory enzymes. As some of these enzymes are encoded by both nuclear and mitochondrial genes, coordination of their synthesis is probable, inter alia at the transcriptional level. We have postulated a direct effect of steroid hormones on mitochondrial gene transcription and here present the following evidence in support of this hypothesis. 1) The human and rodent mitochondrial genomes contain nucleotide sequences similar both to type I and type II HREs. 2) Glucocorticoid receptors (GR) rapidly translocate from the cytoplasm into mitochondria after administration of glucocorticoids. This process has been reproduced in vitro and deletion of the N-terminal part of the glucocorticoid receptor stops translocation into mitochondria. 3) Gel shift analysis has demonstrated binding of GR to putative mitochondrial GR elements. 4) In transfection experiments, mitochondrial HREs confer dexamethasone inducibility on hybrid reporter constructs, abolished in the presence of excess RU38486. 5) Similar results were obtained for thyroid hormone receptor (TR α) localization, import, and binding to TR elements. These findings, taken with the demonstrated effects of steroid (and thyroid) hormones on mitochondrial transcription and respiratory enzyme biosynthesis, strongly support the hypothesis of a direct effect of steroid (and thyroid) hormones on mitochondrial gene transcription.

Glucocorticoid effects on axonal regeneration of retinal ganglion cells in vitro

Although glucocorticoids are commonly used for the treatment of optic nerve inflammations, little is known about direct effects of steroids on retinal nerve cells. In this study, the neuritogenic effect of prednisolone on retinal ganglion cells was investigated using retinal organ cultures derived from adult rats and embryonic chickens. The numbers of axons growing from retinal ganglion cells (RGC) in vitro were counted to measure the regenerative propensity at various corticosteroid concentrations. Prednisolone exerted a dose-dependent neuritogenic influence on RGC of either origin. The effect was highest at a concentration of 25 μg/ml culture medium, where axon numbers increased to 237% (rat) and 166% (chick) of the control values. This was highly significant (t-test: P < 0.01 and P < 0.005, respectively). High corticosteroid concentrations (>125 μg/ml) produced toxic effects, as revealed by low axon numbers as well as morphological alterations of the explants.

Steroid-Induced Depletion of Mucosal Mast Cells and Eosinophils in Intestine of Athymic Nude Rats

In conventional rats, we have previously demonstrated that corticosteroid treatment caused macrophage engulfment and destruction of intestinal mucosal mast cells and eosinophils by 24 h without evidence of local tissue destruction, inflammation or secretion of rat mast cell protease II. As the growth and survival of these cells appear to be dependent on factors derived from T lymphocytes, we examined the response in congenitally athymic rnu/rnu rats and euthymic rnu/+ rats 35 days after parasitic infection. Rats were injected intraperitoneally with 1 mg dexamethasone and sections of jejunum were examined at 0, 7, 13 and 24 h. The numbers of mucosal mast cells significantly decreased in both groups and became less than 30% of the original values at 24 h. Tissue mast cell protease decreased similarly. However, protease in serum did not increase and there were no inflammatory changes at any time. The numbers of eosinophils also rapidly decreased and became less than 20% at 24 h in both rnu/rnu and rnu/+ rats. By electron microscopy, we saw granular changes (fusion) in mast cells and nuclear changes (apoptosis) in eosinophils by 7 h after corticosteroid in athymic rats. Macrophage engulfment of these cells was observed at 7 and 13 h. Our results suggest that inflammatory cell depletion by macrophages is not dependent on suppression of typical thymus-derived T lymphocytes, and may be due either to direct effects of steroids on the cells themselves, or indirectly upon cells other than T cells which normally supply maintenance and growth factors for them.

Effects of Gonadotropin on Ovarian Intrafollicular Processes during the Development of Oocyte Maturational Competence in a Teleost, the Atlantic Croaker: Evidence for Two Distinct Stages of Gonadotropic Control of Final Oocyte Maturation1

Full-grown oocytes of Atlantic croaker are insensitive to maturation-inducing steroid (MIS) unless they are primed with gonadotropin (GtH). The objective of this study was to examine the mechanism of GtH-induced maturational competence in croaker oocytes. Specifically, we determined the in vitro secretion of steroids by intact ovarian follicles of unprimed or hCG-primed fish, the direct effects of steroids on maturational competence, and the effects of steroid (cyanoketone), protein (cycloheximide), and RNA (actinomycin D) synthesis inhibitors on hCG-induced maturational competence and steroidogenesis in vitro. The steroid content of the incubation medium after hCG treatment was measured by RIA. The effects of hCG or exogenous steroid treatment on maturational competence were determined by recording the incidence of germinal vesicle breakdown (GVBD) after MIS-induced GVBD in a standard bioassay. Our major findings were: (1) induction of maturational competence occurred after exposure of ovarian follicles to hCG either in vivo or in vitro; (2) MIS secretion was detected in follicles of hCG-primed fish but not unprimed fish, and no MIS secretion was observed during hCG induction of maturational competence in vitro; (3) treatment with cyanoketone blocked the hCG-dependent secretion of testosterone and estradiol but not the development of maturational competence; (4) treatment with MIS or various other exogenous steroids in the absence of hCG did not induce maturational competence; and (5) hCG-induced maturational competence was inhibited by cycloheximide and actinomycin D. Therefore, the mechanisms of GtH induction of oocyte maturation in Atlantic croaker can be described in two distinct stages: a delta-4 steroid-(including MIS) and estrogen-independent priming stage followed by a MIS-mediated GVBD stage. The priming stage may involve mechanisms requiring RNA as well as protein synthesis.

Single-Fiber Electromyography for Studying the Effect of Betamethasone on Normal Neuromuscular Junctions in Man

The effects of steroids on normal end-plate in vivo were evaluated in man by using single-fiber electromyography (SFEMG). SFEMG was performed on voluntarily activated extensor digitorium communis muscle under basal conditions and 1 week later an intramuscular injection of 4 mg betamethasone per day was administered for 7 days. Ten patients with low back pain were examined. Steroid therapy did not induce any statistically significant variation either of the mean jitter value or of the percentage of recordings with abnormally high jitter. The results of this study do not give evidence of a direct effect of steroids on normal neuromuscular junction.

Pituitary steroids in two teleost species: Immunohistological and biochemical studies

Antisera raised against steroid hormones [estradiol-17β (E 2), testosterone (T), 11β-hydroxyandrostenedione (OHA)] were used to localize immunoreactive material in fixed and paraffin-embedded pituitaries of the African catfish and the rainbow trout. Organic extracts of pituitary homogenates were analyzed for steroid hormones by radioimmunoassay and gas chromatography-mass spectrometry (E 2 in female catfish only). With the exception of an E 2-positive cell type in the catfish neurointermediate lobe, steroid immunoreactivity was found to be restricted to the cytoplasm of adenohypophyseal cells, which were also labeled after incubation with catfish α,β-gonadotropin and salmon gonadotropin antisera, respectively. Steroid levels determined by radioimmunoassay in the catfish ranged between 85 and 628 pg/pituitary, while lower levels (2–8 pg/pituitary) were found in the rainbow trout. E 2 was identified by gas chromatography-mass spectrometry at a level of 84 pg/pituitary. The observation that immunolabeling after steroid antiserum incubation is confined mainly to gonadotrops provides morphological evidence for direct steroid effects on this particular cell type.

Prednisone stimulation of erythropoiesis in leukemic children during remission.

Children with acute leukemia in remission manifest reticulocytosis and a significant increase in hemoglobin concentration (mean increment of 2.3 +/- 1.1 g/dl) following prednisone pulse therapy. This hemoglobin increment is not associated with changes in serum erythropoietin activity. It is speculated that this stimulation of red cell production may be a direct effect of steroids on erythroid progenitor cells.

Stage-specific mechanisms regulate the expression of the dopa decarboxylase gene duringDrosophila development

Transcripts of the dopa decarboxylase (Ddc) gene accumulate within 4 h of the administration of 20-OH-ecdysone to mature larvae of Drosophila. The increase can be explained as the sum of a direct steroid effect, independent of protein synthesis, and an indirect effect, dependent on proteins synthesized after an increase in the hormone titer. By contrast, the peaks of DDC activity in embryos, and perhaps in the first two larval instars and in imaginal discs as well, cannot be ascribed to any direct steroid effects. In fact, a decreasing hormone titer might be required before the Ddc gene can be expressed at these stages. The stage-specific differences in the molecular mechanisms regulating Ddc gene activity may be reflected in the phenotype of an activity variant, Ddc+4. Molecular studies now underway on the variant Ddc gene might help us to understand the complex multifunctional region that we believe lies upstream of the gene and regulates its expression during development.

Direct effects of androgens on lateral hypothalamic neuronal activity in the male rat: I. A microiontophoretic study

Unit neuronal activity in the lateral hypothalamic-medial forebrain bundle region (LHA-MFB) of the male rat is modified following an increase of plasma testosterone level. In order to determine possible direct action of hormones on LHA-MFB neurons, unit discharge frequencies were recorded during local microiontophoresis of testosterone and estradiol and electroosmotic application of cholesterol. Thirteen cells did not respond, 9 were nonspecifically depressed by all the steroids tested, 13 were excited by both sex-steroids and 11 were specifically activated by testosterone. The short latencies of the responses suggest a direct effect of steroids on the plasma membrane sites.

S6A09005) - Direct effects of steroids on the membranes.

S6A09005) - Direct effects of steroids on the membranes.

Glucocorticoid effects on newly synthesized proteins in muscle and non-muscle cells cultured from neonatal rat hearts

To analyze direct effects of steroids on the rates of synthesis (and/or degradation) of newly synthesized proteins of the rat heart, we have used high resolution two-dimensional gel electrophoresis and autoradiography. A collective steroid domain of nineteen proteins, comprising fifteen with an increased rate of synthesis and four with a decreased rate of synthesis, was consistently seen in cultures of cardiac muscle and non-muscle cells from neonatal rats following 24 h incubation with 10 −7dexamethasone. Similarly, incubation with 10 −7M sex steroids, mineralocorticoids, and other glucocorticoids including the highly selective compound RU26988, established the glucocorticoid-specificity of the response. Different subsets of this glucocorticoid domain were seen for collagenase- or trypsin-dispersed primary cultures of cardiac muscle and non-muscle cells or for passaged cultures of cardiac non-muscle cells. Six polypeptides were consistently induced in all cardiac cultures, regardless of cell morphology. Two polypeptides were consistently induced only in those cultures containing cardiac non-muscle cells, whereas protein l, of identical M r, (~ 52 K) and pI (~ 5.3) to desmin, was induced only in cultures of spontaneously contractile cardiac muscle cells. The glucocorticoid domain proteins described herein represent direct steroid effects on cardiac cells and are therefore candidate mediators of physiological glucocorticoid effects on, for example, differentiation and contractility.

Glucocorticoid-induced proteins in bovine endothelial cells

High circulating levels of corticosteroids are associated with elevated blood pressure and an increased incidence of vascular damage. We have used high resolution two-dimensional gel electrophoresis and autoradiography to analyse direct steroid effects on the newly synthesized proteins in bovine aortic endothelial cells. At medium concentrations of 10 −7 M, corticosteroids but not sex steroids increased the synthesis of two endothelial proteins: el (M.W. $ ̃ 43K, pI $ ̃ 6.3) and e2 (M.W. $ ̃ 28K, pI $ ̃ 5.9). The responses were consistent for different endothelial cell cultures, different gel runs, and for natural and synthetic glucocorticoids, including the “pure” synthetic glucocorticoid, RU 26988. Mineralocorticoids (aldosterone, deoxycorticosterone) at 10 −7 M were less potent in their effect on the synthesis of e2, and had no discernible effect on the synthesis of el. These specific effects on cellular protein synthesis demonstrate that glueocorticoids can act directly on endothelial cells. These direct actions may thus mediate altered endothelial cell function in clinical and experimental cardiovascular conditions characterized by corticosteroid excess.

Steroid effects on protein synthesis in cultured smooth muscle cells from rat aorta.

To examine the direct effects of steroids on vascular smooth muscle, we have incubated rat aortic vascular smooth muscle cells in culture either steroid-free, or with natural and synthetic corticosteroids (RU26988, dexamethasone, corticosterone, 9 alpha-fluorocortisol, aldosterone, deoxycorticosterone) or sex steroids (estradiol, 5 alpha-dihydrotestosterone). At the end of 24 h, cultures were pulsed with [35S]methionine for 2 h, the cells lysed, and patterns of incorporation of isotope into protein determined by two-dimensional gel electrophoresis and autoradiography. Neither estradiol nor 5 alpha-dihydrotestosterone altered protein synthetic profiles compared with control (steroid-free) incubations. In contrast, cultures exposed to the six corticosteroids at 10(-7) M showed an identical pattern of response (6 proteins increased, 6 proteins decreased). This response appears to be glucocorticoid specific, since the mineralocorticoids (9 alpha-fluorocortisol, aldosterone, and deoxycorticosterone) did not have any effects over and above those seen with the pure glucocorticoid RU26988. We interpret these data as evidence for a putative glucocorticoid domain of at least 12 proteins in rat vascular smooth muscle cells. In contrast, there appear to be no comparable estrogen-, androgen-, or mineralocorticoid-specific changes in these cells.