Present management of Graves' hyperthyroidism: an effective but symptomatic approach.
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Graves Disease↗
Biomedical subjects
P Thomopoulos
Publications and source records attributed to P Thomopoulos.
[Salivary steroids. Simple and significant approach for the measurement of hormone activity].
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Humans↗
The adhesiveness of monocytic U937 cells is stimulated by pro-inflammatory agents and inhibited by adenosine 3':5'-cyclic monophosphate.
We investigated the regulation of the adhesiveness of the human promonocytic cell line U-937, differentiated along the monocytic pathway either by 1,25-(OH)2-cholecalciferol or a combination of retinoic acid and dibutyryl cAMP. Adhesion to untreated polystyrene plastic was induced by inflammatory agents like PAF, fMLP or LTB4. The response to PAF first appeared after 48hr of differentiation and was inhibited by PAF antagonists and protein kinase C inhibitors indicating involvement of the phosphatidyl-inositol pathway in the stimulating effect. On the other hand, all the c-AMP raising agents tested inhibited PAF-induced cell adhesion, whatever their target membrane receptors, the Gs transducing protein, the catalytic unit of adenylate cyclase or cAMP phosphodiesterase. Direct stimulation of protein kinase A by Br8-cAMP had a similar effect. Moreover, PAF was able to increase cAMP levels. This suggests the existence of a cAMP based negative control mechanism limiting the action of PAF.
2-Chloroadenosine↗
Forskolin inhibition of cyclic AMP generation in J774 macrophages.
Forskolin inhibited cyclic AMP generation in J774 macrophage cells in response to isoproterenol. Forskolin, 10 nM-0.1 mM, also inhibited the adenylate cyclase activity of membrane preparations. The basal activity and the isoproterenol-, cholera toxin-, fluoride- or GppNHp-stimulated activities were maximally depressed by 10 microM forskolin (30-70% inhibition, EC50 = 0.3-0.5 microM). This effect was achieved similarly in membranes from pertussis toxin-treated cells. Forskolin required guanine nucleotides for inhibition. In the absence of GTP the decrease in basal activity was reversed into stimulation (EC50 = 10 microM forskolin). Reversal of inhibition into activation also followed the addition of 1 mM MnCl2 (EC50 = 10 microM forskolin). 1,9-Dideoxyforskolin was ineffective to alter adenylate cyclase activity. In contrast, a water-soluble derivative of forskolin was as active as forskolin to regulate activity. The results suggest that forskolin may interact with adenylate cyclase to cause either activation or inhibition depending on the degree of activation of Ns and on its interaction with the catalyst.
Cell Line↗
Biphasic regulation of macrophage attachment by activators of cyclic adenosine monophosphate-dependent kinase and protein kinase C.
A method is described that enabled us to study the adhesiveness of J-774 murine macrophages. Cell attachment was stimulated by activators of kinase C (i.e., phorbol esters) as well as kinase A (cyclic adenosine monophosphate; cAMP). This novel effect of cAMP was observed when its levels were increased via receptor triggering (prostaglandin E1, beta-adrenergic agonists), activation of Ns (cholera toxin), or inhibition of phosphodiesterase (Ro 20-1724) or when the kinase was directly activated by Br8-cAMP. The simultaneous treatment with kinase A and kinase C activators at the time of attachment resulted in a partially additive response. On the other hand, preincubation of the cells in suspension with one of the activators rendered them refractory to subsequent stimulation at the onset of the adhesion assay, whatever agent was used. Such a refractoriness was also observed in cells preincubated with oleoyl-acetyl-glycerol (OAG). On the other hand, when added at the time of attachment, this near-physiological activator of kinase C evoked a biphasic response: the early stimulation of cell attachment was followed by an accelerated rate of "detachment." In conclusion, kinase C and kinase A play a role in the sequence of events leading to cell adhesion. The cross desensitization observed is distal and takes place at or beyond the kinase step.
8-Bromo Cyclic Adenosine Monophosphate↗
Insulin effect on GABA uptake in astroglial primary cultures.
Astroglial cultures from newborn mouse cerebral cortex contain [125I]Insulin binding sites. Binding was specific, reversible, time dependent and reached equilibrium after 45 min. Insulin analogues compete for this [125I]Insulin binding. Incubation of cerebral cortex astroglial cultures with insulin induced a time- and dose-dependent inhibition of the [3H]GABA high affinity uptake. A decrease in the Vmax rather than an effect on the Km was observed. This effect was dose-dependent and effective at 10(-10) M. Autoradiographic observations on the cell monolayer showed the presence of two groups of cells: one which strongly takes up [3H]GABA and consist in smaller GFAP positive process-bearing cells and another group of much flatter and larger GFAP positive cells which uptake was lower. The smaller stellate cells were apparently the most sensitive to insulin effect. These results: 1) confirm the presence of insulin binding sites on astroglial primary cultures, 2) show an effect of insulin on [3H]GABA high affinity uptake of these cells; this effect being optimal on a stellate-like population of astrocytes, and 3) indicate that insulin may interfere in neuromodulation through astroglial signals.
Animals↗
Evening urinary free corticoids: a screening test in Cushing's syndrome and incidentally discovered adrenal tumours.
In this study, we investigated the usefulness of the determination of evening urinary free corticoids/creatinine in samples collected from 20.00 to 24.00 h as a screening test in Cushing's syndrome. In controls (N = 61) the ratio values ranged from 1.1 to 9.4 mumol/mol, whereas in patients with Cushing's syndrome (N = 20), they ranged from 27.5 to 855.5 mumol/mol. However, in 28% of patients with major obesity (greater than 50% overweight) and no hypercortisolism, the ratio values were between 9.4 to 27.8 mumol/mol. A short (10 days) hypocaloric diet induced a decrease in the values in 75% of these patients; the normal range was reached in 50% of them. In addition, the evening urinary free corticoids/creatinine was slightly abnormal in 8 out of 10 patients with incidentally discovered 'silent' adrenal adenomas, whereas it was normal in all 6 with other adrenal masses. In conclusion, evening urinary free corticoids/creatinine is easy to obtain and it reaches a higher sensitivity (100%) and specificity (97%) than the 24 h urinary free corticoids. In the case of borderline values, the presence of overweight should be taken into account.
Adrenal Cortex Hormones↗
Protein kinase C potentiates isoproterenol-mediated cyclic AMP production without modifying the homologous desensitization process in J774 cells.
The J774 murine macrophage cells possess a beta 2-adrenergic receptor coupled to adenylate cyclase, which can be regulated by homologous desensitization. Stimulation of protein kinase C by phorbol esters or oleoyl acetyl glycerol potentiates two-to-threefold the isoproterenol-induced cyclic AMP accumulation. These promoters act at a post-receptor level, since the number and affinity of the beta-adrenergic receptors, measured by use of the hydrophilic ligand [3H]CGP-12177, are not modified. In addition, the effect of cholera toxin is similarly increased and pretreatment of the cells with pertussis toxin prevents the action of phorbol esters. On the other hand, these promoters are ineffective on isoproterenol-induced desensitization and the rates of receptor segregation and recovery remain unchanged. Therefore, protein kinase C modulates the isoproterenol-stimulated adenylate cyclase, whereas it is inactive on the homologous desensitization process.
1-Methyl-3-isobutylxanthine↗
Specific modulation of surface receptors in J.774 macrophages by anchorage.
The J.774 murine macrophage cells were cultured in suspension in Teflon flasks. When allowed to attach on culture plastic dishes, a 2-3-fold increase in transferrin binding was observed. This occurred in 10 min, reached a steady state at 60 min, remained stable for several hours and was reversible after resuspension of the cells at 37 degrees C. The phenomenon was not dependent on the synthesis of new protein. An opposite change of acetyl LDL receptors was observed, with a threefold decrease of the binding 1 h after the attachment of the cells. The increase of transferrin binding affected almost equally the cell surface and the intracellular sites; therefore it could not be related to a simple shift between these two compartments. It is suggested that the attachment of the cells induced a recruitment of binding sites from a 'silent' pool of receptors. Serum factors, as well as phorbol esters and db-cAMP potentiated the effect of anchorage.
Animals↗
Expression of transferrin receptors in phytohemagglutinin-stimulated human T-lymphocytes. Evidence for a three-step model.
Resting human T-lymphocytes show an elevated intracellular concentration of ferritin, whereas transferrin receptors are not detectable. Stimulation by phytohemagglutinin markedly lowers their ferritin content, while inducing the synthesis of transferrin receptors. Addition of iron salts (ferric ammonium citrate) in activated T-lymphocyte cultures causes a marked enhancement of both [3H]uridine and [3H]thymidine incorporation. Nevertheless, it also induces a concentration-dependent decrease in transferrin receptor synthesis, associated with a marked rise of ferritin production. Hemin treatment exerts the same effects. Addition of picolinic acid in phytohemagglutinin-stimulated cultures causes a decrease of [3H]thymidine incorporation, whereas transferrin expression is markedly enhanced. The action of iron salts and chelators is specific for transferrin receptors, since the expression of other membrane markers of activated human T-lymphocytes (interleukin-2 receptor, insulin receptor, and HLA-DR antigen) is not modified by treatment with iron or picolinic acid. These observations suggest that expression of transferrin receptors in activated T-lymphocytes is specifically modulated by their intracellular iron level, rather than their proliferative rate. Addition of picolinic acid to resting T-lymphocytes in the absence of mitogen induces a marked decrease of their ferritin content, but not the appearance of transferrin receptors. On the basis of these results, we suggest a three-step model: (a) in resting T-lymphocytes, the gene for transferrin receptor is apparently "closed," in that it is not expressed under both normal conditions and following iron deprivation. (b) After mitogen stimulus, T-lymphocytes are reprogrammed into cell cycle progression, which necessarily entails synthesis of transferrin receptors (c) Expression of these receptors is modulated by the intracellular iron level, rather than the rate of proliferation per se.
Ferric Compounds↗
Phosphorylation of class I HLA antigens in U-937 monocyte-like cells. Role of protein kinase C.
Phorbol esters and 1-oleoyl-2-acetylglycerol induce an increase in phosphorylation of a series of 44-kD proteins in the U-937 human monocyte-like cell line. The phenomenon is rapid at 37 degrees C, starting 0.5 min after adding the inducer, increasing to a maximum of 5-10 min and decreasing to the basal level at 30 min. It is also observed at 4 degrees C, though much more slowly. Immunoprecipitation and two-dimensional electrophoresis demonstrated that these phosphorylated proteins belong to the class I HLA antigens.
Cell Line↗
[Anti-insulin receptor antibodies. Clinical significance in diabetology].
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Autoantibodies↗
Inhibition of insulin receptor binding by A23187: synergy with phorbol esters.
The ionophore A23187 inhibits the binding of insulin on U-937 monocyte-like cells which had been induced to differentiate by 1 alpha, 25-(OH)2 cholecalciferol, while it remains inactive on the undifferentiated cells. A 50% reduction of the specific binding of 125I-insulin is observed within 20 to 30 min at 37 degrees C. The effect is obtained at 10(-7)M to 10(-6)M A23187. It is reversible in 60 min at 37 degrees C. A suboptimal concentration of the ionophore potentiates the inhibitory action of phorbol esters on insulin binding.
Calcimycin↗
Differentiation of U-937 human monocyte-like cell line by 1 alpha,25-dihydroxyvitamin D3 or by retinoic acid. Opposite effects on insulin receptors.
The monocyte-like human cell line U-937 has been differentiated in vitro by incubation with either 1 alpha,25-dihydroxyvitamin D3 or retinoic acid (RA) plus dibutyryl cyclic AMP (db-cAMP). Both methods were effective in inducing the appearance of maturation markers. Their actions on insulin receptors were the opposite, however; 1 alpha,25-dihydroxyvitamin D3 increased the binding of the hormone, while RA plus db-cAMP decreased the binding. These effects were specific for insulin, since the transferrin receptors were reduced by both methods of differentiation. Thus, the changes in insulin receptors during maturation in vitro depend on the inducing agent and are not causally related to the differentiation process.
Bucladesine↗
The iron-chelating agent picolinic acid enhances transferrin receptors expression in human erythroleukaemic cell lines.
Picolinic acid, a metal chelating molecule, was administered to human erythroleukaemic cell lines (K 562 and HEL) that were grown in serum-containing media. Picolinic acid inhibited both iron uptake and cell growth. Furthermore, picolinic acid was shown to markedly decrease the level of ferritin in the cells. In spite of the inhibition of cell growth, picolinic acid induced a marked increase in the transferrin-binding capacity of the cells. This phenomenon was due to a two-five-fold enhancement of the rate of transferrin receptor biosynthesis. Other iron-chelating compounds, capable of reducing the level of intracellular iron, also elicited a marked enhancement of the transferrin-binding capacity of the cells. However, the addition of iron, as ferric ammonium citrate, in the culture medium elicited a marked increase in the level of ferritin and a strong decrease in the transferrin-binding capacity of the cells. On the basis of these data we propose that a feed-back mechanism is involved in the regulation of transferrin receptors: when the cells accumulate iron they decrease the number of transferrin receptors in order to prevent further accumulation of iron; when no or low iron is available to the cells, the number of transferrin receptors markedly increases as a compensatory mechanism.
Cell Line↗
Regulation of transferrin receptors in human hematopoietic cell lines.
Cells grown in the presence of ferric ammonium citrate or hemin exhibited a concentration and time-dependent decrease in 125I-transferrin (Trf) binding. In contrast, cells grown in the presence of protoporphyrin IX or picolinic acid (an iron chelator) exhibited a marked increase in Trf binding. The decrease or increase in binding activity observed under these different conditions of culture reflected, respectively, a reduction or increase in receptor number rather than an alteration in ligand receptor affinity. Growth of the cells in the presence of saturating concentrations of apotransferrin only induced a slight reduction in receptor number. Investigation of the Trf receptors' turnover and biosynthesis clearly showed that iron and hemin decreased the synthesis of Trf receptors without any modification of the receptor turnover; in contrast, protoporphyrin IX and picolinic acid markedly increased the synthesis of Trf receptors. Our results suggest that hemin, iron, and protoporphyrin IX may represent the main molecules involved in the regulation of Trf receptors.
Antibodies, Monoclonal↗
The role of iron in the growth of human leukemic cell lines.
The growth requirements of three human leukemic cell lines (K 562, HEL, U937) have been studied in the absence of serum. For growth in serum-free medium, the cells require insulin, transferrin, and albumin. Two highly water-soluble iron salts, ferric ammonium citrate and ferric ammonium sulfate, may completely replace transferrin for supporting the growth of these cell lines. Similar results were obtained when mitogen-stimulated lymphocytes were grown in serum-free media. Iron containing compounds, such as hemin or hemoglobin, were also able to replace transferrin. Experiments using 42/6 monoclonal antibody strongly suggest that free-iron salts are taken up by the cells by a mechanism that is completely independent from transferrin-receptors.
Cell Division↗
Phorbol esters inhibit the binding of low-density lipoproteins (LDL) to U-937 monocytelike cells.
The present study demonstrates that U-937 monocytelike human cells possess specific LDL receptors. 125I-LDL binds at 4 degrees C on the cell surface. The bound molecules are releasable by heparin. The reaction requires Ca2+ and the binding sites are sensitive to proteolysis. Unlabeled LDL compete with 125I-LDL, whereas HDL are ineffective. At 37 degrees C, LDL are internalized and degraded by a chloroquine-sensitive pathway. Tumor-promoting phorbol esters inhibit the binding of 125I-LDL to its receptor on U-937 cells. This inhibition exhibits temperature, time, and concentration dependence. At 37 degrees C, inhibition is 50% at 5 X 10(-9) M of TPA. After removal of phorbol esters, treated cells recover their 125I-LDL-binding activity in 60 min. The inhibitory activities of various phorbol esters are proportional to their tumor-promoting activities. Inhibition appears to be due to a reduction in the number of available LDL receptors rather than a decrease in receptor affinity.
Cell Line↗