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Biomedical subjects

A Munck

Publications and source records attributed to A Munck.

At least 127 records · Page 7Linked to original sources
PubMed1980

In vitro actions of glucocorticoids on murine macrophages: effects on glucose transport and metabolism, growth in culture, and protein synthesis.

In vitro exposure of homogeneous monolayer cultures of thioglycollate-elicited mouse peritoneal macrophages to glucocorticoid hormones for up to 96 hr produced a progressive, dose-related inhibition of cell growth and protein synthesis. The reductions in cell protein, incorporation of radiolabeled leucine, and intracellular water space were specific to steroids with glucocorticoid action, with half-maximal inhibition occurring at about 10(-9) M for dexamethasone. Similar dose-related, glucocorticoid-specific decreases in net glucose uptake, lactate production, and 14CO2 production were observed that were only partially explained by the decrease in cell mass. A decrease in the monosaccharide transport system Vmax was also noted but could be completely accounted for by the reduction in cell surface area and the resulting proportional loss in transport sites. No effect on transport system affinity could be demonstrated. The suggestion was made that such long-term culture of macrophages in the presence of glucocorticoids reverses the biochemical and functional changes that comprise the "activated" state of mononuclear phagocytes.

Animals↗
PubMed1979

Heterogeneity of nuclear glucocorticoid receptor interactions.

When thymocytes are incubated with glucocorticoids at 37 degrees, 60--70% of the receptor bound steroid is associated with the nucleus. Under conditions where the rate of steroid-receptor formation is not limiting the transfer of steroid-receptors from the cytoplasm to the nucleus occurs rapidly with a T 1/2 of 30 seconds. These observations have led us to investigate whether or not all glucocorticoid receptor complexes are associated with the nucleus in the same manner. To this end, nuclear glucocorticoid-receptor complexes have been extracted by differential salt extraction and DNase I and DNase II digeston. Of the nuclear dexamethasone receptor complex initially bound, 70--75% is resistant to 0.2 M KCl extraction (designated N2) and 25--30% is resistant to 0.4 extraction (designated N4). N2 can be further extracted with 0.4 M KCl whereas N4 is resistant to reextraction with either 0.2 M KCl, suggesting that N2-N4 (N2-4) and N4 represent distinct physical forms of nuclear dexamethasone receptor. In intact cells, N2 and N4 differ under the following physiological condition. (1) N4 binding occurs prior to N2-4; (2) a cold chase of unlabeled dexamethasone decreases N2-4 by 70% but N4 binding by only 10%; (3) N4 binding decreases more rapidly than N2-4 following a decrease in hormone concentration by dilution; (4) a cold chase of either cortexolone or progesterone preferentially decreases N2-4 and has little effect on N4. In addition, the nuclear N2-4 and N4 distribution differ for cortisol, dexamethasone and triamcinolone acetonide, three steroids having different in vitro biological potencies. DNase I treatment of nuclei solubilizes approximately 60% of nuclear DNA yet releases only 20--30% of nuclear receptor, whereas DNase II solubilizes only 10% of nuclear DNA and releases 76--80% of nuclear receptor. As seen with salt extraction, the resistance of nuclear glucocorticoid-receptor complexes to a DNase I and II is dependent on the steroid molecule which is associated with the receptor. Of the steroids we have tested, nuclear triamcinolone acetonide and dexamethasone receptor complexes are most resistant to nuclease attack. Nuclear cortisol receptor complexes are readily solubilized by either DNase I or II under conditions where little dissociation of steroid from receptor occurs. These data represent evidence for physiologically distinct forms of nuclear glucocorticoid receptor interaction. In addition, they demonstrate the importance of the steroid portion of the steroid receptor in directing the nature and/or location of steroid receptors within or on the nucleus.

Animals↗
PubMed1979

Glucocorticoid receptors and actions in rat thymocytes and immunologically stimulated human peripheral lymphocytes.

After reviewing briefly our earlier studies on glucocorticoid receptors and mechanisms in thymus cells, we have outlined results from the following two areas of current interest in our laboratories: the "life-cycle" of glucocorticoid receptors and complexes in thymus cells, and the levels of glucocorticoid receptors and sensitivity in immunologically stimulated human peripheral lymphocytes. Several of our results on energetics and kinetics of hormone binding to glucocorticoid receptors in rat thymus cells seem to require extension of the simplest model of hormone-receptor transformations in intact cells. ATP-depletion experiments suggest the existence of a nonbinding form of the receptor; "chase" experiments suggest reaction of hormone directly with nuclear-bound receptor; experiments on depletion and replenishment of cytoplasmic receptor using cortisol and dexamethasone suggest the existence of at least two subpopulations of nuclear-bound hormone-receptor complex. We have found that mitogen or immunologic stimulation of human peripheral lymphocytes in culture leads within 24 h or so to a striking increase in the number of glucocorticoid receptor sites per cell. We believe this increase may be due to partial synchronization of the cell population in a phase of the cell cycle in which receptor content is high. Contrary to the widely held view that mitogen-stimulated cells become insensitive to glucocorticoids, our experiments show that with respect to inhibition of thymidine and uridine incorporation and glucose uptake, the cells are highly sensitive to dexamethasone at 24, 48, and 72 h after stimulation with concanavalin A.

Animals↗
PubMed1978

Comparison of glucocorticoid-receptor complex binding to nuclei and DNA cellulose. Evidence for different forms of interaction.

Binding of dexamethasone . receptors with isolated nuclei, DNA-cellulose and cellulose has been compared with respect to dependence on salt concentration and resistance to KCl extraction and DNAse I digestion. A solution of cytoplasmic dexamethasone . receptor complexes was prepared by the incubation of rat thymus cells with steroid at 3 degrees C and breaking the cells by hypotonic lysis. Activation of the complexes was accomplished by warming the solution at 25 degrees C for 15 min. Activation significantly increased the ability of dexamethasone . receptors to bind to nuclei and DNA-cellulose but not to cellulose. Dexamethasone-receptor complexes bound to nuclei at 3 degrees C are completely resistant to extraction with 0.1 M KCl, 76% resistant to 0.2 M KCl and 20% resistant to 0.4 M KCl. Dexamethasone . receptors bound to DNA-cellulose are 45% resistant to extraction with 0.1 M and 0.2 M KCl and 29% resistant to 0.4 M KCl extraction. Cellulose-bound dexamethasone . receptors are not resistant to any of these extractions. DNAase I treatment releases 60% of the dexamethasone . receptors bound to DNA-cellulose but only 13% of those bound to nuclei, though at least 60% of the nuclear DNA is solubilized. The presence of 0.15 M KCl decreases binding of activated dexamethasone . receptors to nuclei by 73% but to DNA-cellulose by only 17%. Pretreatment of nuclei with 0.1--0.4 M KCl reduces their capacity to bind activated dexamethasone . receptors by 90% whereas similar treatment reduces the capacity of DNA-cellulose to bind dexamethasone . receptors by only 29%. Nuclei extracted with 0.1 M KCl appear to have a limited capacity to accept dexamethasone . receptors. These studies demonstrate that binding of dexamethasone . receptors to nuclei and DNA-cellulose differs by (a) the higher resistance of nuclear complexes to KCl and DNAase I treatment; (b) the much greater sensitivity of nuclei to KCl treatment.

Cell Nucleus↗
PubMed1978

Interaction of glucocorticoids with macrophages. Identification of glucocorticoid receptors in monocytes and macrophages.

Glucocorticoid binding was measured in resident and thioglycollate-elicited mouse peritoneal macrophages, rabbit alveolar macrophages, and human monocytes. Two assays of binding were used--an assay with intact cells in suspension or monolayers, and an assay of cytosol and nuclear forms of glucocorticoid receptors. The mononuclear phagocytes contained approximately equal to 4--10 X 10(3) high affinity receptor sites per cell, with dissociation constants of approximately equal to 2--8 nM dexamethasone. The binding to the saturable sites was specific for steroids with glucocorticoid or antiglucocorticoid activity. Cortisol, corticosterone, and progesterone competed with dexamethasone for binding, whereas estradiol, dihydrotestosterone, and 11-epicortisol competed very little. Binding of dexamethasone to cytosol and nuclear forms of the receptor complex and temperature-sensitive translocation of cytosol forms to nuclear forms were shown. At 37 degrees C the predominant form of the hormone-receptor complex was nuclear. These results demonstrate that corticosteroids interact with macrophages at physiological concentrations.

Animals↗
PubMed1978

Glucocorticoid receptors and glucocorticoid-sensitive secretion of neutral proteinases in a macrophage line.

A continuous line of mouse macrophages (P388D1) has been shown to secrete elastase, collagenase, and plasminogen activator at activities comparable to those of macrophages elicited by an inflammatory stimulus in vivo. At physiologic concentrations anti-inflammatory glucocorticoids selectively and reversibly inhibited secretion of the three proteinases but did not inhibit secretion of lysozyme, a constitutive enzyme produced by the P388D1 cells. The secretion of the neutral proteinases was inhibited 50% by 2 to 10 nM dexamethasone. Proliferation of the macrophages was also glucocorticoid sensitive. The P388D1 macrophages contained about 4000 saturable glucocorticoid-binding sites per cell. Concentrations of hormone saturating the high affinity receptor site (for dexamethasone the dissociation constant for steroid-receptor binding, Kd, was 4 nM) correlated well with concentrations inhibiting secretion of the proteinases. Only glucocorticoids and progesterone competed for binding to the specific receptors. Temperature-sensitive translocation of hormone-receptor complexes from "cytoplasm" to nucleus similar to that found with rat thymocytes was demonstrated. Thus, the interaction between glucocorticoids and the P388D1 cell line provides a model for the regulation of macrophage secretion of neutral proteinases under normal and stress conditions.

Cell Line↗
PubMed1976

Liver hemodynamics and liver function in cats during graded hypoxic hypoxemia.

In 15 cats, anesthetized with chloralose and curarized, liver hemodynamics and liver function were followed during graded hypoxic hypoxemia. Hepatic arterial and intrahepatic portal venous conductance were not influenced by hypoxia, whereas severe hypoxemia increased gastrointestinal conductance. Total liver blood flow remained constant and hypoxemia was compensated for by an increase in hepatic extraction of oxygen approaching 100%. Only when the hepatic venous pO2 fell below 5-10 mmHg did hypoxemia decrease liver function. The results indicate that the sinusoidal perfusion is homogeneous.

Animals↗
PubMed1976

Secretin-like choleretic effect of prostaglandins E1 and E2 in cats.

1. The effects of intraportal and hepatic arterial infusion of prostaglandins E1 and E2 on liver function and circulation was studied in fasting chloralose anaesthetized cats. 2. Infusion of the prostaglandins at rates of 0-1, 1-0 and 5-0 mug/kg.min caused a 35-40% increase in bile flow. This may be explained by a decrease in the reabsorption or a secretion of sodium ions by the ductular cells. The canalicular bile production and bile flow. This may be explained by a decrease in the reabsorption or a secretion of sodium ions by the ductular cells. The canalicular bile production and bile acid excretion was not affected by the prostaglandins. 3. Infused at rates of 1-0 and 5-0 mug/kg.min the prostaglandins caused a transient decrease in mean arterial blood pressure and mesenteric vascular resistance. The resistance in the intrahepatic arterioles and low-pressure vessels was not affected. 4. The prostaglandins did not influence the splanchnic uptake of oxygen and ethanol, whereas a slight increase in the splanchnic glucose output occurred. 5. The effects of the two prostaglandins were identical and not related to the route of administration.

Animals↗
PubMed1975

Imitation of glucagon effects on splanchnic hemodynamics and liver function by N6,2'-O-dibutyryl 3',5'-cyclic AMP (DBcAMP) in cats.

Fasting cats anesthetized with chloralose were used for the experiments. DBcAMP infused at a rate of 340 nmol/kg/min increased the gastrointestinal and intrahepatic portal conductances whereas the hepatic arterial conductance was decreased. The hemodynamic responses to portal and systemic venous administration of DBcAMP were identical. In half of the experiments DBcAMP increased the splanchnic ethanol elimination rate and oxygen consumption and in all experiments there was a decrease in the plasma clearance and extraction ratio of Indocyanine Green. No change in bile flow was observed. DBcAMP infused at a rate of 85 nmol/kg/min was without significant effects on either splanchnic hemodynamics or liver metabolism. DBcAMP infused at a rate of 850 nmol/kg/min accentuated the decrease in hepatic arterial conductanc- but was found to decrease the splanchnic ethanol elimination rate and oxygen cownsumption. Infusion of cAMP, AMP and adenosine at a rate of 340 nmol/kg/min were without measurable effects. Based on these results it is concluded that like the metabolic effects also the vascular effects of glucagon are caused by stimulation of specific glucagon receptors which results in an intracellular release of cAMP.

Animals↗
PubMed1975

Long-term incubation of rat thymus cells: Cytolytic actions of glucocorticoids in vitro.

A system has been developed for long-term incubation of rat thymus cells in sus pension in a chemically defined medium consisting of medium 199 and methylcellulsoe. With this system it is possible to study quantitatively the full time-course of glucocorticoid actions, from the early inhibitory effects on glucose uptake to the slow cytolytic effects. Cells incubated under these conditions preserve their sensitivity to the inhibitory effects of cortisol on glucose uptake for al least 12 h. Glucocorticoids at physiological concentrations lead to cytolysis (measured by reduction in viable cell counts determined by a modification of the pronase-cetrimide method) by 12 h incubation time. The cytolytic effect exhibits well-defined dose-response relationships and specificity for glucocorticoids. By 24 h,cortisol at 10(-6)M consistently reduces the total cell count by about 15%, and the viable cell count by about 40%. Cortisol is active down to 10(-7) M. Dexamethasone is roughly 10 times as active as cortisol, and cortisone completely inactive. Cortexolone is slightly active, but also shows antiglucocorticoid activity by its ability to block the action of cortisol. Under anerobic conditions in Medium 199, no inhibitory effect of cortisol is observed on glucose uptake.

Animals↗