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C Stournaras

Publications and source records attributed to C Stournaras.

At least 19 recordsLinked to original sources
PubMed2005

Dehydroepiandrosterone sulfate and allopregnanolone directly stimulate catecholamine production via induction of tyrosine hydroxylase and secretion by affecting actin polymerization.

Adrenal cortical cells of zona reticularis produce the neuroactive steroids dehydroepiandrosterone (DHEA), its sulfate ester dehydroepiandrosterone sulfate (DHEAS), and allopregnanolone (ALLO). An interaction between zona reticularis and adrenal medulla has been postulated based on their close proximity and their interwoven borders. The aim of this paper was to examine in vitro the possible paracrine effects of these steroids on catecholamine production from adrenomedullary chromaffin cells, using an established in vitro model of chromaffin cells, the PC12 rat pheochromocytoma cell line. We have found the following: 1) DHEA, DHEAS, and ALLO increased acutely (peak effect between 10-30 min) and dose-dependently (EC50 in the nanomolar range) catecholamine levels (norepinephrine and dopamine). 2) It appears that the acute effect of these steroids involved actin depolymerization/actin filament disassembly, a fast-response cellular system regulating trafficking of catecholamine vesicles. Specifically, 10(-6) m phallacidin, an actin filament stabilizer, completely prevented steroid-induced catecholamine secretion. 3) DHEAS and ALLO, but not DHEA, also affected catecholamine synthesis. Indeed, DHEAS and ALLO increased catecholamine levels at 24 h, an effect blocked by L-2-methyl-3-(-4-hydroxyphenyl)alanine and 3-(hydrazinomethyl)phenol hydrochloride, inhibitors of tyrosine hydroxylase and L-aromatic amino acid decarboxylase, respectively, suggesting that this effect involved catecholamine synthesis. The latter hypothesis was confirmed by finding that DHEAS and ALLO increased both the mRNA and protein levels of tyrosine hydroxylase. In conclusion, our findings suggest that neuroactive steroids exert a direct tonic effect on adrenal catecholamine synthesis and secretion. These data associate the adrenomedullary malfunction observed in old age and neuroactive steroids.

Actins↗
PubMed2001

Opioids suppress basal and nicotine-induced catecholamine secretion via a stabilizing effect on actin filaments.

Catecholamine secretion and actin filament disassembly are closely coupled in chromaffin cells. Opioid suppression of catecholamine secretion is fast and transient, both characteristics of actin filament involvement. The aim of the present work was to test the hypothesis that opioids suppress catecholamine secretion via an inhibitory effect on actin filament disassembly. For this purpose we used the PC12 rat pheochromocytoma cell line. Norepinephrine and dopamine were measured by enzyme-linked immunosorbent assay or RIA. Polymerized actin was measured by rhodamine-phalloidin and visualized by confocal laser scanning microscopy. Opioids suppressed basal catecholamine secretion. The onset of this effect was fast and transient, peaking at 2 min, and was reversible by opioid antagonists. Synchronously, opioids suppressed actin filament disassembly; this was also reversible by opioid antagonists. Cytochalasin B prevented the inhibitory effect of opioids on catecholamine secretion. In addition, opioids suppressed the stimulatory effect of nicotine on catecholamine secretion and actin depolymerization. Changes in actin cytoskeleton in neuron-like PC12 cells make them resistant to both effects of opioids, i.e. on catecholamine secretion and actin disassembly. In conclusion, our data suggest that the suppressive effect of opioids on basal and nicotine-induced catecholamine secretion may result from an opioid-provoked stabilization of cortical actin. It also appears that basal catecholamine secretion is associated with opioid-sensitive machinery regulating the continuous formation of short-lived areas of cortical actin filament disassembly.

Actins↗
PubMed2000

Corticotrophin-releasing hormone (CRH) interacts with inflammatory prostaglandins and interleukins and affects the decidualization of human endometrial stroma.

The hypothalamic neuropeptide, corticotrophin-releasing hormone (CRH), which is also produced by human endometrium, has been shown to induce its decidualization in vitro. This process, induced mainly by progesterone, has characteristics of an aseptic inflammatory reaction, and is modulated by locally produced pro-inflammatory factors. In humans, prostaglandin E(2) (PGE(2)) enhances while interleukin (IL)-1 inhibits the decidualizing effect of progesterone. The aim of the present work was to test the hypothesis that CRH might affect the decidualization of human endometrium interacting with these factors. Therefore, we studied its effects on the production of pro-inflammatory interleukins IL-1, IL-6 and of PGE(2) from human endometrial stromal cells in primary culture. The results strongly suggest that CRH decidualizes stromal cells, as judged by the appearance of cytokeratins and the production of prolactin, two established markers of decidualization. In parallel to its effect on decidualization, CRH also decreased the production of PGE(2), while it increased the production of IL-1 and IL-6. Exposure of endometrial stromal cells to IL-6 also caused decidualization. The data presented here suggest that endometrial CRH regulates the production of local modulators of decidualization, i.e. PGE(2), IL-1 and IL-6. We postulate that, through the regulation of these factors, CRH acts as a local fine-tuner of decidualization initiated by progesterone.

Cell Differentiation↗
PubMed2000

Actin cytoskeleton: a signaling sensor in cell volume regulation.

The actin microfilaments are well known dynamic structures that support and organize the cell membrane and functions associated with the membrane such as ion channels and transporters. In addition, many aspects of cellular physiology seem to be actively modulated by changes in actin cytoskeleton dynamics, which involve reorganization and restructuring of the filaments. For both of these reasons, the actin cytoskeleton has attracted special attention since the early days of cell volume regulation research. Mechanisms controlling the actin equilibrium in response to external stimuli were studied and the signaling cascades leading to the regulation of actin cytoskeleton dynamics have been partially elucidated. They include: a) activation of specific actin binding proteins that regulate actin polymerization dynamics, b) activation of protein kinases or phosphatases regulating phosphorylation of specific cytoskeletal proteins and c) activation of signal transduction pathways leading from membrane receptor activation to actin reorganization involving small GTPases of the Rho and Rac families. These intracellular signal transducers are activated by extracellular stimuli that include hormones, growth factors, cytokines, or ions, many of them in turn are partially known to participate in cell volume regulation. These findings provide strong evidence that the actin cytoskeleton is involved in cell volume regulation by sensing and mediating extracellular signals.

Actins↗
PubMed2000

PLC-gamma1 signaling pathway and villin activation are involved in actin cytoskeleton reorganization induced by Na+/Pi cotransport up-regulation.

BACKGROUND: The brief incubation of opossum kidney (OK) cells with low P(i) results in Na+/P(i) cotransport up-regulation and in substantial, but transient, cytoskeletal reorganization. In this study, we examined signaling events involved in the depolymerization of microfilaments. RESULTS: Confocal laser scanning microscopy, immunoblot and immunoprecipitation experiments revealed villin co-localization with mainly actin short filaments and monomers, indicating that under the conditions used, villin acted as an actin-severing protein. Further analysis revealed that low concentrations of extracellular phosphate resulted in phospholipase Cgammal (PLC-gammal) translocation to the actin cytoskeleton, without increases in its tyrosine phosphorylation. Additionally, tyrosine phosphorylation of a portion of insoluble villin was increased; whereas, only tyrosine phosphorylated villin associated with PLC-gammal. Although, tyrosine phosphorylation of PLC-gammal was not observed during Na+/P(i) cotransport up-regulation, genistein treatment abolished the enzyme's translocation to the actin cytoskeleton, as well as its association with villin. In addition, villin was found to associate with the 85-KDa subunit (p85) of phosphatidylinositol (PI)-3 kinase, concomitant with PLC-gammal, in the cytoskeletal fraction of Na+/P(i) cotransport up-regulated cells. CONCLUSIONS: Our observations suggest a signaling mechanism linking low ambient P(i) levels to the acute up-regulation of its cotransport with sodium and the depolymerization of the subcortical actin cytoskeleton.

Actin Cytoskeleton↗
PubMed1999

Expression and characterization of Cys374 mutated human beta-actin in two different mammalian cell lines: impaired microfilament organization and stability.

Previous studies have demonstrated that addition of glutathione at the penultimate Cys374 residue of actin results in filaments with diminished mechanical stability. In the present work substitutions introducing a negatively charged (Asp and Glu) or a neutral (Ala) amino acid at position 374 of the human beta-actin and tagged at the N-terminus with the flag epitope were studied by transient transfections into Ishikawa human endometrial and opossum kidney cells. Immunofluorescence revealed that microfilaments which incorporated negatively charged mutants were partially to severely disorganized when compared to the almost well-formed actin-Ala374 filaments or the wild type actin filaments. Furthermore, microfilaments containing either negatively charged mutant were more sensitive to the destabilizing action of cytochalasin B. In addition, Triton fractionation resealed a considerable reduction of flag-actin content in the Triton insoluble fraction for cells expressing Asp374 or Glu374 mutant compared to wild type actin. These results demonstrate that negatively charged amino acid residues at the exposed C-terminal tail strongly affect actin microfilament organization and dynamics in vivo.

Actin Cytoskeleton↗
PubMed1999

Opioid agonists modify breast cancer cell proliferation by blocking cells to the G2/M phase of the cycle: involvement of cytoskeletal elements.

Opioids decrease cell proliferation in different systems including breast, prostate, lung, kidney, and intestine, through an interaction with opioid as well as other membrane-receptor systems (somatostatin, cholinergic), through an unidentified mechanism. Recently, we have reported an interaction of taxol with opioid membrane sites (BBRC 235, 201-204, 1997), and an involvement of opioids to the modification of actin cytoskeleton in renal OK cells (J Cell Biochem. [19981 70:60-69), indicating a possible action of the opioid effect. In the present work, we have examined the effect of two general opioid agonists (ethylketocyclazocine and etorphine) on the cell cycle, in human breast cancer T47D cells, as well as a possible modification of the cellular cytoskeleton under their action, in order to explain the antiproliferative effect of these agents. These two opioids produce a dose-dependent and reversible decrease of the proliferation of T47D cells, with a maximum attained at 10(-8) M. The addition of 10(-8) M of either opioid produced a significant increase of the number of cells arrested in the G2/M phase. Confocal laser microscopy revealed a modification of the actin and tubulin microfilaments, with a clear redistribution at the periphery of the cell, reversed by the addition of the general opioid antagonist diprenorphine. Furthermore, differences between the two opioids were obvious, attributed to the different receptor affinity of each agent. The observed redistribution of actin and tubulin cytoskeletal elements gives therefore a possible answer of the antiproliferative action of opioids. The modification of the cytoskeleton, directly involved to cell division, might provoke a "mechanical" obstacle, which could be the reason of the antiproliferative effect of these agonists. Furthermore, the observed tubulin-opioid interaction by opioids provides a possible explanation of the arrest at the G2/M phase of T47D cells under opioid treatment. Nevertheless, although the observed interaction of opioids with cytoskeletal elements gives a plausible answer of the antiproliferative effects of the agents, this might not be the only action of these agents in cell proliferation. Other, direct or indirect, genomic actions, which which remains to be elucidated, might be taken into consideration.

Actins↗
PubMed1999

Paracrinology of endometrial neuropeptides: corticotropin-releasing hormone and opioids.

Human endometrium possesses remarkable secretory properties and the characteristics of a neuroendocrine organ. Epithelial cells of human endometrium express the corticotropin-releasing hormone (CRH) and opioid peptide precursors genes (i.e., proopiomelanocortin, proenkephalin, and prodynorphin) and their end products. Endometrial neuropeptides are under the control of ovarian steroid hormones and locally produced prostanoids and cytokines. Additionally, neuropeptides participate in local paracrine regulatory loops, facilitating communication between endometrial epithelial and stromal cells as well as the interaction between endometrial and myometrial cells. In view of the proinflammatory cytokine properties of CRH, we postulate that endometrial CRH may participate in intrauterine inflammatory and vascular processes associated with stromal cell decidualization and blastocyst implantation. Additionally, given the myorelaxant actions of opioids these endometrial neuropeptides may participate in the control of myometrial contractility.

Corticotropin-Releasing Hormone↗
PubMed1999

The transcription of corticotropin-releasing hormone in human endometrial cells is regulated by cytokines.

Corticotropin-releasing hormone (CRH), a hypothalamic neuropeptide, is also produced in the human endometrium where it participates in local inflammatory phenomena associated with the decidualization of endometrial stroma and the implantation of the fertilized egg. The inflammatory cytokines interleukin 1 (IL-1), IL-6 and leukemia inhibitory factor (LIF) appear to be the dominant local regulators of these intrauterine inflammatory processes. In the present study we have examined the direct interactions between cytokines and CRH in the endometrium. For this purpose we have measured the effects of IL-1, IL-6 and LIF on the activity of CRH promoter inserted in human endometrial cells in culture. Homologous transient transfection experiments were conducted employing a 0.9-kb fragment of the 5' flanking region of the human CRH gene coupled to the luciferase reporter gene, using Ishikawa human endometrial cells. We have found that IL-1beta increased the activity of CRH gene promoter, in a time- and dose-dependent manner. This effect was antagonized by the IL-1 receptor antagonist IL-1ra and blocked completely by the cyclo-oxygenase inhibitor indomethacin. Similarly, IL-6 increased the activity of CRH promoter in a dose-dependent fashion, an effect partially reversed by indomethacin. LIF did not have any apparent effect. In conclusion, our data suggest that IL-1 and IL-6 exert a strong stimulatory effect on the expression of endometrial CRH. This effect is most probably mediated via prostaglandins. Based on these data we hypothesize that in the human endometrium interleukins, prostaglandins and CRH form a local network regulating the inflammatory phenomena taking place within the uterine cavity.

Adenocarcinoma↗
PubMed1999

Regulation of actin organisation by TGF-beta in H-ras-transformed fibroblasts.

The actin cytoskeleton undergoes architectural changes during the processes of cell transformation and tumourigenesis. Transforming growth factors beta arrest cell cycle progression, regulate differentiation and modulate the onset of oncogenesis and tumourigenesis. Here, we investigated the direct role of transforming growth factor beta-1 in altering the transformed phenotype and regulating the actin organisation of oncogenic fibroblasts that constitutively or inducibly express the H-ras oncogene. Following transforming growth factor beta-1 treatment, these transformed fibroblasts undergo a dramatic morphological alteration that includes a discrete reorganisation of their actin cytoskeleton and focal adhesions. Quantitative biochemical analysis demonstrated that transforming growth factor beta-1 potently induced polymerisation of globular to filamentous actin, thus corroborating the morphological analysis. The effect of transforming growth factor beta-1 on the cytoskeleton correlates with the ability of this cytokine to suppress anchorage-independent growth of the transformed fibroblasts. Furthermore, transforming growth factor beta-1 upregulates considerably the levels of the RhoB small GTPase and less the RhoA levels. Finally, The beta GTPase inhibitor, C3 exotransferase, blocks the ability of TGF-beta1 to induce cytoskeletal reorganisation. These findings indicate that transforming growth factor beta can regulate cell morphology and growth in a concerted manner possibly via mechanisms that control the actin cytoskeleton.

3T3 Cells↗
PubMed1999

TNF-alpha induces actin cytoskeleton reorganization in glomerular epithelial cells involving tyrosine phosphorylation of paxillin and focal adhesion kinase.

Glomerular permeability for macromolecules depends partially on proper attachment of the glomerular epithelial cells (GEC) to the glomerular basement membrane (GBM). The latter requires integrity of the actin cytoskeleton, which in turn is regulated by specific actin-associated proteins. Since several glomerulopathies characterized by heavy proteinuria are associated with increased glomerular tumor necrosis factor alpha (TNF-alpha) expression, we studied the interaction of TNF-alpha with the actin cytoskeleton of cultured rat GEC. Incubation of GEC with 10 ng/ml TNF-alpha for variable time periods ranging from 15 min to 24 hr demonstrated a marked accentuation and redistribution of actin microfilaments, as shown by direct fluorescence analysis and confocal laser scanning microscopy. Quantitative biochemical determination of the G/total-actin ratio confirmed the above observations. Indeed, this ratio was significantly reduced, indicating substantial polymerization of G-actin and formation of F-actin. Concurrently, TNF-alpha rapidly induced tyrosine phosphorylation of both paxillin and focal adhesion kinase, without affecting the expression levels of these two proteins. In addition, tyrosine phosphorylation of vinculin became evident, indicating involvement of this focal adhesion marker in the observed actin reorganization. Inhibition of tyrosine phosphorylation by genistein prevented the reorganization of the actin cytoskeleton by TNF-alpha. We conclude that TNF-alpha induces substantial reorganization of actin cytoskeleton and focal adhesions. These effects occur simultaneously, with a prompt TNF-alpha-induced tyrosine phosphorylation of paxillin and focal adhesion kinase, indicating that these proteins, known to regulate actin polymerization and formation of focal adhesions, may be directly involved in the mechanism controlling the observed actin redistribution. These findings suggest that the observed TNF-alpha-actin cytoskeleton interactions may relate to the pathogenesis of glomerulopathies with heavy proteinuria, in which increased glomerular expression of TNF-alpha is associated with disturbances in the attachment of podocytes to the GBM.

Actins↗
PubMed1999

Tyrosine phosphorylation of focal adhesion kinase and paxillin regulates the signaling mechanism of the rapid nongenomic action of dexamethasone on actin cytoskeleton.

We have previously shown that dexamethasone (DEX) stimulates rapid polymerization of actin and stabilization of microfilaments in human endometrial adenocarcinoma cells. As the content of total cellular actin and the concentration of the actin transcript did not change, we concluded that polymerization of actin by glucocorticoids involves nongenomic mechanisms. However, the signaling events by which the latter is achieved remain unknown. In the present study we evaluated whether tyrosine phosphorylation is required for the rapid, nongenomic DEX effect on actin assembly. In cells preincubated with the tyrosine kinase inhibitors, genistein or erbstatin analogue (EA), before adding DEX the G-/total actin ratio remained unchanged, whereas DEX in the absence of both inhibitors reduced the ratio by 25%. In addition, when cells were preincubated with the protein tyrosine phosphatase inhibitor pervanadate and subsequently incubated with DEX, the G-/total actin ratio was dramatically reduced by 65%. Furthermore, DEX increased transiently the levels of tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin within 2 to 15 min, without a change in their expression levels. Pervanadate mimicked this effect of DEX and enhanced tyrosine phosphorylation of both proteins. In addition, when cells were exposed to the anticytoskeletal agent cytochalasin B, the basal levels of tyrosine phosphorylation of both proteins were reduced. This effect was reversed by DEX, indicating that actin cytoskeleton integrity is required for the effect of DEX on tyrosine phosphorylation of FAK and paxillin. Finally, we documented enhanced expression of the Ras-related GTP-binding protein Rho-B after long-term (12- and 24-hr) treatment with DEX, whereas Rho-B levels remained unchanged after short-term (3- and 6-hr) treatment. Our observations demonstrate a novel mechanism through which the rapid nongenomic effect of DEX on actin assembly requires tyrosine phosphorylation of the cytoskeleton-associated proteins FAK and paxillin. We also propose that the DEX-induced actin polymerization may constitute a mechanism for transduction of signals resulting in tyrosine phosphorylation of FAK and paxillin. Moreover, the enhanced Rho-B levels observed after long-term treatment with DEX imply a mechanism for the well-described, long-term effects of glucocorticoids on actin cytoskeleton.

Actins↗
PubMed1998

Early alterations of actin cytoskeleton in OK cells by opioids.

Recently we identified and characterized opioid binding sites in OK (opossum kidney) cells and observed decreased proliferation of these cells in response to opioids. In the present study we investigated the effects of opioids on the actin cytoskeleton and explored whether their antiproliferative action may relate to alterations in the distribution or the dynamics of actin microfilaments. Exposure of OK cells to the opioids alphaS1 casomorphin and ethylketocyclazocine resulted in a rapid and substantial actin microfilament reorganization. This was documented by a significant dose-dependent decrease in the amounts of F-actin, determined by measurements of quantitative fluorescence, by immunoblot analysis and by a concomitant increase of the G/total-actin ratio measured by the DNase I inhibition assay. These changes were verified by confocal laser scanning microscopy, which showed marked redistribution of the microfilamentous structures in the presence of the opioids without affecting the organization of microtubules or vimentin intermediate filaments. The effect of opioids on actin polymerization dynamics occurred within 15 min and persisted for at least 2 h, while their restoration to control levels was accomplished 6 h later, indicating a reversible phenomenon. Northern blot analysis showed that the concentration of the actin transcript was unaffected. The addition of diprenorphine, a general opioid antagonist, prevented the effects of opioids on the actin cytoskeleton. The inhibition of OK cell proliferation, induced by ethylketocyclazocine and alphaS1 casomorphin was partially prevented in the presence of phallacidin, which stabilizes microfilaments. Our findings demonstrate that opioids, acting via kappa 1 binding sites, induce rapidly modifications in the dynamics of actin polymerization, and in the organization of microfilaments in OK cells, which may relate to their antiproliferative effect on these cells.

Actins↗
PubMed1997

High sensitivity of leukemic peripheral blood lymphocytes to triethyllead action.

In a previous study we reported that triethyllead (Et3Pb+) inhibits cell proliferation of normal human lymphocytes. To further characterize this interaction, we studied herein the effects of Et3Pb+ on the cell viability of normal and leukemic human lymphocytes and analysed the expression and dynamics of the monomer/polymer equilibrium of tubulin in these cells. Short- and long-term cell culture experiments demonstrated significantly different dose-dependent effects of Et3Pb+ on cell viability of leukemic compared to normal lymphocytes. Indeed, in the presence of increasing concentrations of Et3Pb+ (10(-12)-10(-5) M), primary cultures of chronic lymphocytes (CLL) and acute lymphoblastic (ALL) leukemic human Lymphocytes were much more sensitive to Et3Pb+ treatment when compared to normal peripheral blood lymphocytes (PBL). The IC50 values were approximately 5 x 10(-6) M for PBL and 8 x 10(-10) M for both CLL and ALL respectively, when cells were preincubated for 3 h with this agent. These experiments revealed a 1000-fold higher responsiveness of leukemic cells to Et3Pb+ treatment. Quantitative immunoblot analysis showed that leukemic cells express up to 4-fold higher total tubulin amounts. However, the proportion of polymerized tubulin in leukemic compared to normal lymphocytes increased only slightly (up to 1.4-fold). These findings reveal a significant decrease in the polymeric to total tubulin ratio in leukemic lymphocytes, indicating important modifications in tubulin dynamics and reorganization of the microtubular structures. Our results demonstrate that leukemic cells are much more sensitive than normal lymphocytes to Et3Pb+ action. This effect may be due to the altered monomer/polymer dynamic equilibrium of tubulin shown in leukemic cells. It is, therefore, worthwhile exploring future applied uses of Et3Pb+ as a potential suppressor of leukemic cell growth.

Animals↗
PubMed1997

Endometrial corticotropin-releasing hormone. Its potential autocrine and paracrine actions.

Corticotropin-releasing hormone (CRH) is expressed at several peripheral tissues including normal epithelial cells of human and rodent uterus. However, its biological role is unknown in both species. To clarify this role we studied the regulation of CRH promoter in endometrial cells. We performed homologous transfection experiments in Ishikawa cells, a human endometrial cell line, using a 0.9-kb fragment of the 5'-flanking region of human CRH gene coupled to luciferase. We found that the activity of the 5'-flanking region of the CRH gene is stimulated by cAMP and EGF and inhibited in a receptor-mediated, dose-dependent fashion by estradiol and dexamethasone. The antiglucocorticoid RU 486 acted as a glucocorticoid agonist suppressing CRH gene activation, whereas progesterone was devoid of any activity. Prostaglandin E2 and interleukins-1 and -6 stimulated CRH activation, and the prostanoid inhibitor indomethacin suppressed it, most probably by inhibiting endogenous prostaglandins. These findings suggest that endometrial CRH gene expression may be under the negative control of estrogens and glucocorticoids and under the positive control of PGE2, IL-1, and IL-6. Considering the involvement of CRH in proinflammatory phenomena, we postulate that endometrial CRH, in association with uterine prostanoids and cytokines, may participate in intrauterine inflammatory processes of early pregnancy, such as decidualization and blastocyst implantation.

Animals↗
PubMed1997

Endometrial corticotropin-releasing hormone: expression, regulation, and potential physiological implications.

Our findings show that human and rat uterus express the CRH gene. Epithelial cells of both species are the main source of endometrial CRH, while stroma does not seem to express it, unless it differentiates to decidua. Immunoreactive CRH, produced by endometrial cells, has the chromatographic characteristics of authentic hypothalamic CRH, while the size of its mRNA in both human and rat uterus is similar to or identical with its counterpart, present in placenta and hypothalamus (1.3 kb). Estrogens and glucocorticoids inhibit and prostaglandin E2 stimulates the promoter of human CRH gene in transfected human endometrial cells, suggesting that endometrial CRH gene expression is under the control of these agents. Moreover, in rats, endometrial CRH expression is significantly higher at implantation sites, compared to that at interimplantation uterine regions. Given the proinflammatory/vasoregulatory properties of CRH, we hypothesize that endometrial CRH may participate in the regulation of intrauterine phenomena, such as blastocyst implantation, endometrial vascularization, and myometrial contractility.

Animals↗
PubMed1997

Dexamethasone induces rapid actin assembly in human endometrial cells without affecting its synthesis.

Dexamethasone exerts a stimulatory effect of rapid-onset on the polymerization of actin. This has been documented in human endometrial adenocarcinoma Ishikawa cells, resulting in an acute, dose-dependent decrease in the G/total-actin ratio. In the present study we completely characterized this fast and apparently nongenomic effect of dexamethasone on actin assembly. We followed the morphological alterations of actin cytoskeleton and measured the time-dependent dynamics of actin polymerization both by ruling out any changes of total actin in the cells and by measuring its transcript. Rapid changes in actin polymerization were accurately measured using a highly sensitive and quantitative rhodamine-phalloidin fluorimetric assay. Ishikawa cells, exposed to 0.1 microM dexamethasone for various time periods up to 24 h, showed a highly significant, rapid, and transient increase in the polymerization of actin starting within 15 min of dexamethasone exposure and lasting 2 h. Treated cells showed a significant (1.79-fold) enhancement of the fluorescent signal compared to untreated cells at 15 min. This value decreased continuously in a time-dependent manner, reaching control levels after 120 min and remained so for the next 24 h. Confocal laser scanning microscopy studies confirmed these findings. Intensive coloration of microfilaments over several scanning sections suggested an enhanced degree of actin polymerization in cells preincubated for 15 min with 0.1 microM dexamethasone. Moreover, actin filaments were more resistant to cytochalasin B. Additionally, quantitative immunoblot analysis showed that the content of total cellular actin remained the same during this period, suggesting that the biosynthesis of actin was unaffected. Northern blot analysis showed that the concentration of the actin transcript was also unaffected. Our data suggest that glucocorticoids induce a fast and self-limited polymerization of actin in human endometrial cells without affecting its synthesis. These findings strengthen the hypothesis that glucocorticoids exert rapid, nongenomic cellular effects and that the actin-based cytoskeleton is an integral part of this pathway, playing an essential role in receiving and mediating steroid signals for the modulation of cellular responses.

Actin Cytoskeleton↗