EMF radiation at 2450 MHz triggers changes in the morphology and expression of heat shock proteins and glucocorticoid receptors in rat thymus.

AIMS: Electromagnetic fields (EMFs) can act as inducers or mediators of stress response through the production of heat shock proteins (HSPs) that modulate immune response and thymus functions. In this study, we analyzed cellular stress levels in rat thymus after exposure of the rats to a 2.45 GHz radio frequency (RF) using an experimental diathermic model in a Gigahertz Transverse Electromagnetic (GTEM) chamber. MAIN METHODS: In this experiment, we used H&E staining, the ELISA test and immunohistochemistry to examine Hsp70 and Hsp90 expression in the thymus and glucocorticoid receptors (GR) of 64 female Sprague­Dawley rats exposed individually to 2.45 GHz (at 0, 1.5, 3.0 or 12.0 W power). The 1 g averaged peak and mean SAR values in the thymus and whole body of each rat to ensure that sub-thermal levels of radiation were being reached. KEY FINDINGS: The thymus tissue presented several morphological changes, including increased distribution of blood vessels along with the appearance of red blood cells and hemorrhagic reticuloepithelial cells. Levels of Hsp90 decreased in the thymus when animals were exposed to the highest power level (12 W), but only one group did not show recovery after 24 h. Hsp70 presented no significant modifications in any of the groups. The glucocorticoid receptors presented greater immunomarking on the thymic cortex in exposed animals. SIGNIFICANCE: Our results indicate that non-ionizing sub-thermal radiation causes changes in the endothelial permeability and vascularization of the thymus, and is a tissue-modulating agent for Hsp90 and GR.

Ultraviolet radiation regulates cortisol activity in a waveband-dependent manner in human skin ex vivo.

BACKGROUND: 11ß-Hydroxysteroid dehydrogenase type 1 (11ß-HSD1), 11ß-hydroxysteroid dehydrogenase type 2 (11ß-HSD2), and glucocorticoids (GC) and their receptor (GR) play a key role in tissue-specific regulation of GC action. OBJECTIVES: To determine the expression of genes encoding 11ß-HSD1 (HSD11B1), 11ß-HSD2 (HSD11B2) and GR (GRα; also known as NC3R1) and their protein products, and levels of cortisol in human skin explants and/or cocultured keratinocytes/melanocytes after treatment with ultraviolet (UV) A, B or C wavebands. METHODS: Skin from foreskins and/or cocultured human keratinocytes/melanocytes were irradiated with UVA, UVB or UVC (skin) and incubated for 12 and 24 h. Methods of reverse transcription-polymerase chain reaction, Western blotting, enzyme-linked immunosorbent assay and immunohistochemistry (IHC) were used to determine expression and localization of corresponding genes or antigens. RESULTS: UVB enhanced the HSD11B1 gene and protein expression in a dose-dependent manner, while UVA had no effect. Similarly, UVC increased 11ß-HSD1 protein product as measured by IHC. UVB and UVC enhanced cortisol production and decreased epidermal GR expression, while UVA had no detectable effects. Although both UVA and UVB stimulated HSD11B2 gene expression, only UVA increased 11ß-HSD2 protein product levels with UVB and UVC having no effect. CONCLUSIONS: We suggest that these differential, waveband-dependent effects of UV radiation on the expression of cutaneous HSD11B1, HSD11B2 and GRα genes and their corresponding protein products, and cortisol production are to protect and/or restore the epidermal barrier homeostasis against disruption caused by the elevated cortisol level induced by UVB and UVC.

Effect of high-dose total body irradiation on ACTH, corticosterone, and catecholamines in the rat.

Total body irradiation (TBI) or partial body irradiation is a distinct risk of accidental, wartime, or terrorist events. Total body irradiation is also used as conditioning therapy before hematopoietic stem cell transplantation. This therapy can result in injury to multiple tissues and might result in death as a result of multiorgan failure. The hypothalamic-pituitary-adrenal (HPA) axis could play a causative role in those injuries, in addition to being activated under conditions of stress. In a rat model of TBI, we have established that radiation nephropathy is a significant lethal complication, which is caused by hypertension and uremia. The current study assessed HPA axis function in rats undergoing TBI. Using a head-shielded model of TBI, we found an enhanced response to corticotropin-releasing hormone (CRH) in vitro in pituitaries from irradiated compared with nonirradiated rats at both 8 and 70 days after 10-Gy single fraction TBI. At 70, but not 8 days, plasma adrenocorticotrophic hormone (ACTH) and corticosterone levels were increased significantly in irradiated compared with nonirradiated rats. Plasma aldosterone was not affected by TBI at either time point, whereas plasma renin activity was decreased in irradiated rats at 8 days. Basal and stimulated adrenal steroid synthesis in vitro was not affected by TBI. In addition, plasma epinephrine was decreased at 70 days after TBI. The hypothalamic expression of CRH messenger RNA (mRNA) and hippocampal expression of glucocorticoid receptor mRNA were unchanged by irradiation. We conclude that the hypertension of radiation nephropathy is not aldosterone or catecholamine-dependent but that there is an abscopal activation of the HPA axis after 10 Gy TBI. This activation was attributable at least partially to enhanced pituitary ACTH production.

Nuclear export of glucocorticoid receptor is enhanced by c-Jun N-terminal kinase-mediated phosphorylation.

The c-Jun N-terminal kinase (JNK) phosphorylates the glucocorticoid receptor (GR) and inhibits GR-mediated transcription. However, the biological effect of the GR phosphorylation remains unknown. Here we demonstrate that activated JNK phosphorylates human GR at Ser226 and enhances its nuclear export after withdrawal of a ligand for GR, dexamethasone. At 1 h after dexamethasone withdrawal, green fluorescent protein-GR molecules were mostly retained at the nucleus, whereas UV exposure enhanced its nuclear export, and approximately 30-40% of cells revealed distinct nuclear export. JNK overexpression alone mimics UV exposure and enhanced GR export accompanied by inhibition of GR-mediated transcription. However, mutation of the Ser226 JNK phosphorylation site in GR abrogated UV-mediated enhancement of GR nuclear export. Furthermore, overexpression of a dominant negative SEK1 mutant also abrogated the effects of UV exposure on GR export. Taken together, these findings suggest that JNK-mediated phosphorylation of the GR-Ser226 enhances GR nuclear export and may contribute to termination of GR-mediated transcription.

Suppression of mast cell activation by glucocorticoid.

Mast cells play a critical role in allergic diseases. When mast cells are activated by cross-linking of their high affinity IgE receptors by the antigen and IgE antibodies, release of chemical mediators is followed by secretion of multiple cytokines. We report that IL-3-dependent mucosal-type mast cells undergo apoptosis when IL-3 is withdrawn. In addition, cross-linking of high affinity IgE receptors prevents apoptosis of mast cells by paracrine mechanisms, producing IL-3, IL-4 and granulocyte/macrophage colony-stimulating factor (GM-CSF). However, the secretion of endogenous growth factors are not enough for cell survival, whereas IL-4 induces cell aggregation by expressing adhesion molecules such as leukocyte function-associated antigen 1 (LFA-1), and makes it reactive to endogenous growth factors by contact cell to cell interaction. On the other hand, dexamethazone down-regulates the expression of intracelluar adhesion molecule 1 (ICAM-1) and IL-4 in activated mast cells, by which the self-aggregation of mast cells is inhibited and apoptosis is induced. Thus, glucocorticoids suppress mast cell survival by inhibiting IL-4 production and expression of adhesion molecules.

Radiomodification effects of Ukrain, a cytostatic and immunomodulating drug, on intracellular glucocorticoid reception during short-term gamma-irradiation.

The ability of Ukrain, a cytostatic and immunomodulating semisynthetic compound of thiophosphate-modified alkaloids of Chelidonium majus L., to modify the effects of irradiation on intracellular glucocorticoid reception in female rat liver was evaluated after intraperitoneal administration of the drug at 0.4 mg/kg of body weight. Ukrain caused the normalization of the intracellular glucocorticoid-receptor system, influenced by short-term whole-body gamma-irradiation of rats with 1 Gy, beginning on the 10th day after administration of the drug. It was found that Ukrain minimized the consequences of irradiation in the endocrine system of the experimental animals.

Comparison of glucocorticoid receptors liganded with dexamethasone or progesterone.

The initial goal of this work was to examine directly the properties of glucocorticoid receptors bound with antagonists. Cortexolone, progesterone, and R-5020 were the antagonists studied. The tritiated agonists, dexamethasone and triamcinolone acetonide, served as controls. Although the three antiglucocorticoids interfered with agonist binding to the glucocorticoid receptor, direct binding of the tritiated antagonists could not be reproducibility demonstrated using either a charcoal assay or rapid techniques like high performance liquid chromatography or vertical tube rotor ultracentrifugation. Ultraviolet radiation was used to attach covalently tritiated steroid to the receptor. This technique allowed the identification of species that bound agonist or antagonist. That the two classes of steroids bound to the same receptor was shown using a monoclonal antibody directed against the glucocorticoid receptor. These labeled species had the same physical properties upon ultracentrifugation, DEAE cellulose chromatography, and high performance liquid chromatography. It is concluded that although the interaction of antiglucocorticoids like progesterone with the glucocorticoid receptor may be fleeting, antagonists do interact with the glucocorticoid receptor and form complexes with grossly similar properties as those derived from an interaction with agonists.

[Effect of irradiation on the intracellular reception of glucocorticoids in tissues with various radiosensitivities]. / Vliianie oblucheniia na vnutrikletochnuiu retseptsiiu glikokortikoidov v tkaniakh razlichnoi radiochuvstvitel'nosti.

The experiments with albino female rats showed that whole-body gamma-irradiation with a dose of 1 Gy caused various alterations in concentrations of cytoplasmic and nuclear glucocorticoid receptors depending on tissue radiosensitivity (liver, thymus) and postirradiation time-intervals (1, 3, 10, 30, and 60 days). There was also a change in the receptor affinity to glucocorticoids.

[Corticosterone receptor binding by various brain structures in extreme body states]. / Retseptornoe sviazyvanie kortikosterona razlichnymi strukturami golovnogo mozga pri ékstremal'nykh sostoianiiakh organizma.

Studies on receptor binding of 3H-corticosterone in different brain structures of mature Wistar rats after a single x-irradiation at a dose of 12.9 mCoul/kg, short-term vertical vibration, glucocorticoid application, and surgical adrenalectomy showed a high sensitivity of all studied structures to the body level of glucocorticoids. Any of these influences caused redistribution of receptor binding of 3H-corticosterone in all brain structures, with a marked effect in the parietal cortex and mediobasal hypothalamus, which characterized the central mechanisms of regulation of the pituitary-adrenal system under extreme influences.

Characterization of the glucocorticoid receptor. Comparison of wild type and variant receptors.

We have measured the size of the glucocorticoid receptors from two murine lymphoid cell lines, one displaying a wild type cytolytic response to hormone, the other a resistant variant. Using radiation inactivation and target analysis, we first compared the nuclear and cytoplasmic forms of the steroid receptors in a wild type line, WEHI 7.1 (W7). Within the variation expected for this type of measurement (+/- 14%), the nuclear and cytoplasmic forms have the same size, 75,000 and 79,000 daltons, respectively. We have also measured the size of the receptor in a hormone-insensitive "nuclear transfer-increased" (nti) variant (S49 143R). In contrast to reports indicating that the nti phenotype is associated with a much smaller cytoplasmic receptor, we found little or no difference in sizes of translocated receptor in wild type and nti cells. We have found significant differences, however, in the release of wild type and nti receptors from nuclei by nuclease digestion, salt, and spermidine. Approximately 80% of the nti receptor was readily released from nuclei incubated with micrococcal nuclease, while only 40-50% of the wild type receptor was released under similar conditions. The wild type nuclei also contained a fraction of receptor (approximately 30%) which was resistant to extraction by NaCl and spermidine. This fraction was greatly diminished in the nti nuclei. Thus, a portion of the wild type receptors appears to be stabilized within the nuclei, possibly through a type of interaction which cannot be sustained by the nti receptor.