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M J Weidemann

Publications and source records attributed to M J Weidemann.

At least 55 records · Page 3Linked to original sources
PubMed1983

Cyclic AMP and the regulation of prostaglandin production by macrophages.

The ionophore A23187 (6.7 microM) increased the rates of formation of prostaglandins and cyclic AMP in suspensions of thioglycollate-elicited rat peritoneal macrophages. Both effects were inhibited by the calmodulin blocker trifluoperazine (50 microM) and the calcium channel blocker verapamil (500 microM). Inhibitors of phospholipase A2 and cyclo-oxygenase also blocked both actions of A23187. The stimulated prostaglandin formation was markedly reduced when the cells were preincubated with 8-bromo-cyclic AMP (1mM), dibutyryl cyclic AMP (1mM) or cholera toxin (500ng/ml). Addition of exogenous arachidonic acid (30 microM) alleviated this inhibition. We propose that the effect of A23187 on macrophages includes a 'self-limiting' mechanism whereby newly-synthesized prostaglandins can inhibit, via cyclic AMP, a step(s) prior to the transformation of arachidonic acid and thus modulate their own production.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1981

Concanavalin A-induced chemiluminescence in rat thymus lymphocytes. Its origin and role in mitogenesis.

1. The luminol-dependent chemiluminescence of rat thymocytes responding to concanavalin A can be resolved into glucose-dependent and glucose-independent portions. 2. The glucose-dependent portion, supported by D-glucose and D-mannose oxidation, is inhibited by catalase (200 microgram/ml), amobarbital (1 mM) and hexose analogues that block D-glucose uptake. Thus concanavalin A may activate, transiently, an NAD(P)H oxidase that utilizes reducing equivalents derived from the oxidation of exogenous glucose to give dismutation products of O2- (including H2O2) as its major products. 3. The glucose-independent portion is inhibited by eicosa-5,8,11,14-tetraynoic acid but not by indomethacin. It may therefore be associated with the conversion of hydroperoxy intermediates of arachidonic acid metabolism to hydroxy products by the lipoxygenase pathway. 4. Preincubation of thymocytes for 18 h in serum-free medium enhances the subsequent chemiluminescent response to concanavalin A severalfold and evokes the response at a lower threshold concentration. The incorporation of [3H]thymidine by preincubated cells is similarly enhanced at low doses of concanavalin A, whereas the response to optimal doses is unaltered. 5. Catalase does not inhibit the enhanced incorporation of [3H]thymidine obtained in response to concanavalin A, but instead amplifies the response to low doses in the same manner as preincubation.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1980

Chemiluminescence and immune cell activation. II. Enhancement of concanavalin A-induced chemiluminescence following in vitro preincubation of rat thymocytes; dependency on macrophage-lymphocyte interaction.

The immediate chemiluminescence (CL) response to concanavalin A (Con A) of rat thymocytes is enhanced 10 to 20-fold when the cells are preincubated in serum-free medium for 5--20 h. During this period, multiple encounters between lymphocytes and macrophages occur which morphologically appear as rosettes or grape-like cell aggregates. Additionof bone marrow-derived macrophages increases the number of cell aggregates and also the CTL response to Con A. Paradoxically, removal of macrophage-containing cell aggregates after cocultivation leaves a pure thymocyte population which strongly responds to Con A with CL. Our results confirm that macrophage-depleted "competent" lymphocytes are capable of CL and, furthermore, that "competence" is gained during cocultivation with macrophages. We are therefore convinced that measurements of macrophage and lymphocyte CL are a powerful tool for further elucidation of lymphocyte differentiation and of interactions between macrophages and lymphocytes.

Animals↗
PubMed1979

Rapid alterations in cellular morphology and plasma membrane structure induced in rat thymocytes by mitogenic stimuli.

Under conditions where a maximum stimulation of 3-O-methyl-glucose transport is observed, three thymocyte mitogens (concanavalin A, ionophore A23187 and hydrogen peroxide) cause cell rounding and a decrease in the density of intra-membrane particles on the plasma membrane. The early effects of mitogens on the thymocyte plasma membrane are similar to those of osmotic shock.

Animals↗
PubMed1979

Preferential inhibition by quercetin of mitogen-stimulated thymocyte glucose transport.

The ATPase inhibitor quercetin, which inhibits tumor glycolysis, was shown to be a glucose transport inhibitor like the chemically related compound phloretin. Rat thymocyte glucose transport stimulated by the mitogens concanavalin A or ionophore A 23187 was more sensitive than unstimulated transport to quercetin inhibition. The partial inhibition of Na+-, K+- ATPase activity by quercetin observed in tumor cells was confirmed in thymocyte plasma membranes. The specific Na+-, K+- ATPase inhibitor ouabain did not mimic the effect of quercetin on mitogen-stimulated glucose transport but did reduce the effectiveness of concanavalin A as a stimulator of mitochondrial pyruvate oxidation. The results support the idea that glycolytic flux and the activity of plasma membrane ATPase are related but suggest that glucose transport, rather than the Na+-, K+-ATPase, is the rate-limiting reaction in lymphocytes.

Animals↗
PubMed1978

Aerobic glycolysis and lymphocyte transformation.

1. The role of enhanced aerobic glycolysis in the transformation of rat thymocytes by concanavalin A has been investigated. Concanavalin A addition doubled [U-(14)C]glucose uptake by rat thymocytes over 3h and caused an equivalent increased incorporation into protein, lipids and RNA. A disproportionately large percentage of the extra glucose taken up was converted into lactate, but concanavalin A also caused a specific increase in pyruvate oxidation, leading to an increase in the percentage contribution of glucose to the respiratory fuel. 2. Acetoacetate metabolism, which was not affected by concanavalin A, strongly suppressed pyruvate oxidation in the presence of [U-(14)C]glucose, but did not prevent the concanavalin A-induced stimulation of this process. Glucose uptake was not affected by acetoacetate in the presence or absence of concanavalin A, but in each case acetoacetate increased the percentage of glucose uptake accounted for by lactate production. 3. [(3)H]Thymidine incorporation into DNA in concanavalin A-treated thymocyte cultures was sensitive to the glucose concentration in the medium in a biphasic manner. Very low concentrations of glucose (25mum) stimulated DNA synthesis half-maximally, but maximum [(3)H]thymidine incorporation was observed only when the glucose concentration was raised to 1mm. Lactate addition did not alter the sensitivity of [(3)H]-thymidine uptake to glucose, but inosine blocked the effect of added glucose and strongly inhibited DNA synthesis. 4. It is suggested that the major function of enhanced aerobic glycolysis in transforming lymphocytes is to maintain higher steady-state amounts of glycolytic intermediates to act as precursors for macromolecule synthesis.

Acetoacetates↗
PubMed1978

The role of calcium ions in the regulation of rat thymocyte pyruvate oxidation by mitogens.

1. Calcium concentrations in the nanomolar range cause a specific stimulation of the oxidation of pyruvate by isolated mitochondria from rat thymus that is sufficient to account precisely for the stimulation of pyruvate oxidation observed when rat thymocytes are incubated with the mitogens concanavalin A or ionophore A23187. 2. Higher concentrations of Ca2+ (more than 50 nM) inhibit the oxidation of NAD+-linked substrates by rat thymus mitochondria without affecting the oxidation of succinate or ascorbate+ NNN'N'-tetramethyl-p-phenylendiamine. 3. The addition of Ni2+ or Co2+ (2mM) to rat thymocytes prevents the response to concanavalin A at the level of pyruvate oxidation without affecting the stimulation of glycolysis induced by this mitogen. In contrast, the complete metabolic response to the ionophore A23187 is abolished by these ions. Ni2+ and Co2+ interfere with the ability of the ionophore to transport Ca2+ across the plasma membrane. 4. Concanavalin A, but not ionophore A23187, increases the respiratory inhibition induced by Ni2+ and Co2+. 5. These results support the view that mitogens stimulate lymphocyte pyruvate oxidation through an increase in cellular Ca2+ uptake.

Animals↗
PubMed1978

Chemiluminescence and immune cell activation. I. Early activation of rat thymocytes can be monitored by chemiluminescence measurements.

Immediately after the addition of concanavalin A, rat thymocytes respond, in the presence of luminol, with a burst of chemiluminescence (CL) that can be conveniently monitored in an ordinary liquid scintillation spectrometer. Peak CL is reached after 50 sec. Addition of catalase suppresses 65% of the CL, suggesting that H2O2 generation may be its major source. CL with different kinetic characteristics can also be generated by the calcium ionophore A23187. Our rat thymocytes contain approximately 0.1% endogenous macrophages. Bone marrow-derived rat macrophages also respond to concanavalin A or A23187 stimulation with a burst of CL. However, the kinetic properties of CL as well as the inhibition of CL by catalase in these cells differ markedly from those of thymocyte preparations, suggesting that the major portion of the CL in rat thymocytes actually originates in T lymphocytes. Because CL measurements allow the monitoring of very early events in cell activation and because of the simplicity of the technique, CL measurements may become a useful method for the study of lymphocyte activation, of macrophage-lymphocyte interactions, as well as for the rapid screening for specificity and reactivity of T lymphocyte populations.

Animals↗
PubMed1977

Effect of substrates and effectors on the reversible inactivation of pig spleen phosphofructokinase by adenosine triphosphate.

1. To investigate the mechanism of the reversible inactivation of pig spleen phosphofructokinase by ATP, the effect of order of addition of reactants (substrates, effectors and enzyme solution) was studied by preincubating the enzyme before assay with various combinations of its substrates and effectors. 2. Preincubation of the enzyme with MgATP or ATP at pH7.0 before addition of fructose 6-phosphate caused a rapid and much greater inhibition of activity than that observed when the reaction (carried out at identical substrate concentrations) was initiated with enzyme. 3. The rapid inhibition caused by preincubation with ATP, together with the sigmoidal response to fructose 6-phosphate and activation by AMP, were all blocked by prior photo-oxidation of the enzyme with Methylene Blue, which selectively destroys the inhibitory binding site for ATP [Ahlfors & Mansour (1969) J. Biol. Chem.244, 1247-1251]. 4. Fructose 6-phosphate, but not Mg(2+), protected phosphofructokinase from inhibition during preincubation with ATP in a manner that was sigmoidally dependent on the fructose 6-phosphate concentration. 5. Mg(2+), by protecting the enzyme from the inhibitory effect of preincubation at low pH (7.0) and by preventing its activation during preincubation with fructose 6-phosphate, demonstrated both a weak activating effect in the absence of the other substrates and a stronger inhibitory effect in the presence of fructose 6-phosphate. 6. Positive effectors (K(+), NH(4) (+), AMP and aspartate) protected the enzyme from inhibition during preincubation with MgATP in proportion to their potency as activators, but citrate potentiated the ATP inhibition. P(i) significantly slowed the inactivation process without itself acting as a positive effector. 7. The non-linear dependence of the initial rate of the unmodified enzyme on protein concentration (associated with increased positive homotropic co-operativity to fructose 6-phosphate) was intensified by preincubation with ATP and abolished by photo-oxidation. 8. The results are interpreted in terms of an association-dissociation model which postulates that protonation, at low pH, of a photo-oxidation-sensitive inhibitory site for ATP allows more rapid dissociation of an active tetramer to an inactive dimeric species.

Adenosine Monophosphate↗