Effect of acute adrenalectomy on rat liver glucocorticoid receptor

In order to improve current clinical treatment of human hypocortisolism, it is necessary to understand molecular aspects of this pathophysiology. In this study liver tissues from male Wistar rats were used as an experimental model to study structural and functional properties of glucocorticoid receptor (GR) in the absence of glucocorticoid hormones (GC). Results show that acute adrenalectomy (ADX) significantly increases the number of GR binding sites and GR protein content. In addition, acute ADX stimulates increase in stability of the GR, decrease in stability of the glucocorticoid- receptor complex (G-R), and changes in accumulation of the G-R complex in nuclei and its cellular distribution.

Kinetics of thyroxine (T-4) and triiodothyronine (T-3) transport in the isolated rat heart

The dynamics and kinetics of thyroid hormone transport in the isolated rat heart were examined using the modified unidirectional paired tracer dilution method. The uptake of I-125-thyroxine (I-115-T-4) and I-125-triiodothyronine (I-125-T-3) from the extracellular space into heart cells was measured relative to the extracellular space marker H-3-mannitol. The thyroid hormone maximal uptake was 54.4 % for I-125-T-4 and 52.15 % for I-125-T-3. The thyroid hormone net uptake was 25.69 % for I-125-T-4 and 25.49 % for I-125-T-3. Backflux from the intracellular space was 53.17 % for I-125-T-4 and 61.59 % for I-125-T-3. In the presence of unlabelled thyroid hormones, I-125-T-4 and I-125-T-3 maximal uptakes were reduced from 10.1 to 59.74 % and from 34.6 to 65.3 %, respectively, depending on the concentration of the unlabelled hormone, suggesting a saturable mechanism of the thyroid hormone uptake by the heart cells, with K-m(T4) = 105.46 muM and the maximal rate of I-125-thyroid hormone flax from the extracellular space to heart cells (V-max(T4))= 177.84 nM min(-1) for I-125-T-4 uptake, and K-m(T3) = 80.0 muM and V-max(T3) = 118.5 nM min(-1) for I-125-T-3 uptake

Molekulska osnova delovanja glikokortikoida

Glucocorticoid hormones are involved in regulation of cell processes and coordinate physiological response to diverse signals. These hormones, through interaction with specific intracellular receptors, coordinate components of physiological repertoires by activating the expression of gene networks. Thus hormone-receptor complexes may function as key constituent in regulation of specific cell functions as well as in provoking differentiation in already determined cells. Analysis of steroid receptors are important for understanding of molecular details of transcriptional control as well as providing the insight as to how an individual transcriptional factor such as glucocorticoid receptor, contributes to cell identity and function. The purpose of this review is to establish the general molecular mechanism of glucocorticoid action and mechanism associated hormone-receptor complexes with the control of differential patterns (i.e. "positive" and "negative") of gene expression. One of the examples of two signal pathways regulating opposite gene expression are NF-kB and GR-mediated signal pathways. These pathways have important and opposite roles in the immune function. NF-kB is transcription factor which induces the expression of many genes that participate in immune and inflamatory response, while GR is transcription factor that serves as antiinflammatory agent and immune suppressor. Their interactions within the cell, although not yet completely understood, appear to be an important, possibly even the primary mechanism of immune homeostasis. It has not been established that glucocorticoid sensitivity can be caused by mechanisms other than changes of GR number and properties, although recent studies have indicated that receptor isoforms and transcriptional factors may modulate glucocorticoid responsiveness by interacting with receptor protein or directly at the site of DNA binding. The aim of this review is also to describe the role of glucocorticoid receptors in mechanism of glucocorticoid action on cell functions, including immune responses, as well as to present emerging issues on clinical aspects of molecular mechanisms of glucocorticoid action.Glikokortikoidni hormoni su uključeni u regulaciju ćelijskih procesa koji koordiniraju fiziološke odgovore na različite signale. Ovi hormoni u kompleksu sa specifičnim ćelijskim receptorima preko aktivacije ekspresije mreže različitih gena učestvuju u koordinaciji komponenti koje su u osnovi fizioloških odgovora. Na taj način kompleksi hormona i receptora funkcionišu kao ključni faktor u regulaciji specifičnih ćelijskih funkcija, a takođe podstiču i procese diferencijacije u već determinisanim ćelijama. Analize glikokortikoidnih receptora (GR) su značajne, kako za bolje upoznavanje transkripcione kontrole, tako i za objašnjenje kako pojedini transkripcioni faktori, kao što su GR, doprinose identitetu ćelije i njenom funkcionisanju. U ovom radu su prikazani opšti principi molekulskog mehanizma delovanja glikokortikoida, kao i mehanizmi koji povezuju komplekse hormona i receptora sa kontrolom različitog tipa („pozitivnom” ili „negativnom”) genske ekspresije. Jedan od primera za signalne puteve koji suprotno regulišu aktivnost gena su nukleusni faktor kapaB (NFκB) i signalni putevi posredovani sa GR. Ovi putevi imaju suprotne uloge u regulaciji funkcionisanja imunskog sistema. NFκB je transkripcioni faktor koji indukuje ekspresiju gena uključenih u imunske i zapaljenjske procese. GR je takođe transkripcioni faktor, ali deluje kao antiinflamacioni i imunosupresivni agens. Njihove interakcije su značajne u ćelijama imunskog sistema, iako još nisu potpuno objašnjene; one su možda čak i primarne u mehanizmu imunske homeostaze. Do sada nije dokazano da ćelijska senzitivnost na delovanje glikokortikoida zavisi od mehanizama koji ne uključuju promene količine i funkcionalnih osobina GR. Međutim, novija istraživanja pokazuju da izoforme receptora i transkripcioni faktori mogu da menjaju ćelijski odgovor na glikokortikoide preko interakcije sa receptornim proteinom ili direktno sa mestima vezivanja na DNK. U ovom radu su takođe prikazani podaci iz literature o ključnoj ulozi glikokortikoidnih receptora u mehanizmu delovanja glikokortikoida u regulaciji ćelijskih funkcija, uključujući i ćelije imunskog sistema, kao i podaci o kliničkim aspektima molekulskog delovanja glikokortikoida

Biochemical and molecular biological aspects of glucose intolerance in elderly persons

Changes in carbohydrate metabolism in elderly persons have drawn considerable attention but the findings from different studies are in contrast and are even controversial. The insulin receptors in erythrocytes were studied in elderly euglycaemic patients and in a middle-aged control group. The examined persons were also subjected to measurements of blood glucose, insulin and C-peptide concentrations, before and 3 hours after a dietetic meal. In the present study it was found that in spite of the maintained insulin level and C-peptide secretion, some structural and corresponding changes in membrane insulin receptors and the binding site caused differences in postreceptor responses in elderly persons. The examined groups, consisted of 29 males, average age of 66 years (65-70), with normal serum glucose level and 19 middle-aged males, average age of 42 years (32-48), with normal glycoregulation. In basal condition, elderly persons have both normal morning serum insulin (19.68 ± 4.00 mU/L) and C-peptide (2.04 ± 0.78 nmol/L) level. In elderly persons, the number of high affinity insulin receptors in erythrocytes membrane is 22.80 ± 6.18 but the formed insulin-high affinity receptors were not stable. Dissociation constant (Kd1) indicates its elevated dissociation (0.11 ± 0.04). At the same time the number of insulin low affinity binding sites is increased (13 273 ± 5 572) with a fast dissociation of the hormone (13.99 ± 3.37). Food intake raised the number of high affinity receptors compared to the basal value. Alteration in insulin binding affinity suggests the structural and corresponding changes in membrane receptors that may cause differences in postreceptors responses in elderly persons

Comparative analysis of tryptophan oxygenase activity and glucocorticoid receptor under the influence of insulin

This investigation addresses the interaction of insulin (INS) and glucocorticoid (GC) signaling in the hepatic regulation of tryptophan oxygenase (TO) enzyme activity in the rat. Male Wistar rats (200-250 g b.w) received an injection of the different doses of INS (10, 25, 50, 70 and 100 pg/200 g b.w., i.p.) and were used for experiments 3 h and 18 h after INS administration. This study shows that maximum of TO activity was found at dose of 50 mu g of INS with peak increases observed at 3 h and 18 h after injection of INS, while INS had no effect on TO activity in adrenalectomized rats. The analysis of INS effects on glucocorticoid receptor-complex (GC/GR complex) stability shows that complexes from INS-treated rats are less stable than those from control animals. In addition, INS-stimulated stability of glucocorticoid receptor (GR) protein was significantly increased from the controls. Furthermore, the results show that GC/GR complexes from INS-treated rats could be activated and accumulated at higher rate in cell nuclei of control animals. These data support the involvement of INS in modulation of GC signaling pathway which mediates, in part, the activity of TO

Basal and glucocorticoid induced changes of hepatic glucocorticoid receptor during aging: relation to activities of tyrosine aminotransferase and tryptophan oxygenase

The characteristics of glucocorticoid receptors, their sensitivity to glucocorticoid as well as the basal and glucocorticoid induced thyrosine aminotranferase (TAT) and tryptophan oxygenase (TO) activities were studied in rat liver during aging. The concentration (N) and dissociation constant (K-d) of glucocorticoid receptor (GR) significantly change during the aging both in untreated and dexamethasone treated animals. The level of receptors was lower in dexamethasone treated rats of all analyzed aged groups compared to untreated animals. In comparison to untreated groups, there was no correlation between the changes of N and K-d during the lifespan. According to immunochemical analysis, the decline of receptor protein content occurs during lifespan. Dexamethasone treatment reduced the level of receptor protein compare to respective age group of untreated rats. The glucocorticoid-receptor (G-R) complexes from both untreated and treated animals underwent thermal activation, although the extent of activation was more pronounced in the case of untreated groups compared to treated animals. The magnitude of heat activation of receptor complexes was more pronounced in the liver of the youngest untreated rats compared to elderly ones, while the receptor activation between treated groups of studied ages has shown less significant differences. Besides, basal as well as induced TAT and TO activities after dexamethasone injection also showed age-related alterations. The observed alterations in GR might play a role in the changes of the cell responses to glucocorticoid during the age. This presumption is supported by detected changes in basal and dexamethasone induced TAT and TO activities during aging

Insulin modulates rat liver glucocorticoid receptor

This investigation used cytosol fraction of rat liver to examine the effects of insulin ( INS) on functional properties of glucocorticoid receptor ( GR). Male Wistar rats ( 220 - 250 g b.wt.) were injected with INS ( 50 mu g/200 g b.wt, i.p.) and 18 h after INS administration used for experiments. INS-stimulated dissociation of G-R complexes was significantly increased by 133% compared to control level. However, INS treatment significantly stimulated stability of GR protein by 138% above control value. Furthermore, results show that INS stimulated activation of formed cytosol [ H-3] TA-R complexes by 143% in respect to control. [ H-3] TA-R complexes from INS treated animals could be activated and accumulated at higher rate in cell nuclei of control animals. The physiological relevance of the data was confirmed by INS-related stimulation of Tryptophan oxigenase ( TO) activity. It was observed that INS stimulated TO activity while INS injected to adrenalectomized rats, exhibited less effects compared to control. The results indicate that a glucocorticoid hormone ( CORT) enhances INS induced stimulation of TO activity, as evidenced by enhanced enzyme activity. Presented data suggest: that INS treatment leads to modifications of the GR protein and the nuclear components and that INS activates the rat liver CORT signaling pathway which mediates, in part, the activity of TO

Adrenalectomy and dexamethasone treatment alter the patterns of basal and acute phase response-induced expression of acute phase protein genes in rat liver

Hormonal requirements for full hepatic expression of alpha(2)-macroglobulin (alpha(2)M), alpha(1)-acid glycoprotein (AGP), haptoglobin (Hp) and gamma-fibrinogen (Fb) were assessed at the level of mRNA. Prior to exposure to turpentine-induced inflammation, rats were either depleted of glucocorticoids by adrenalectomy or supplemented with an excess of dexamethasone. Adrenalectomy alone did not affect the basal level of acute phase protein (APP) expression except for alpha(2)M mRNA, the level of which was enhanced. In contrast, dexamethasone treatment alone promoted full induction of alpha(2)M, significant, but not maximal increase of AGP and Hp mRNAs and suppression of Fb. In adrenalectomized rats, acute phase (AP)-cytokines, released in response to inflammation, promoted full expression of Fb and Hp and increased the level of AGP mRNA whereas alpha(2)M mRNA remained at the basal level. Inflammation in dexamethasone pretreated rats elicited changes which, in comparison to mRNA values for dexamethasone unpretreated inflamed rats, were seen as overexpression of alpha(2)M, full expression of AGP and incomplete expression of Hp, whereas Fb mRNA remained at the basal level. These data suggest that glucocorticoids are the principal inducers of a(2)M and AP-cytokines of Fb. For full induction of AGP, additive actions of glucocorticoids and AP-cytokines are required whereas expression of Hp is predominantly controlled by AP-cytokines. (C) 1998 Elsevier Science Ltd. All rights reserved