Martha Deierkauf

Toxicity of organotin compounds for polymorphonuclear leukocytes: The effect on phagocytosis and exocytosis

Phagocytosis and concomitant release of enzymes by rabbit polymorphonuclear leukocytes (PMNs) are inhibited by micromolar concentrations of triphenyltin and tributyltin; inhibition by triethyltin occurs at higher concentrations. Chemotactic peptide-induced exocytosis is inhibited at the same concentrations as phagocytosis. Tributyltin causes cell lysis at slightly higher concentrations as required for inhibition of phagocytosis and exocytosis. The organotin compounds have little effect on ATP level in PMNs, which makes an effect on metabolic energy providing processes unlikely. The increase of Ca2+-permeability of the plasma membrane, induced by chemotactic peptide, is inhibited by the organotin compounds. Inhibition of exocytosis by triphenyltin can be counteracted by a number of sulfhydryl compounds. The results suggest that the organotin compounds interfere with PMN function in an early phase of cell activation, where all functions have a common pathway, and where vulnerable sulfhydryl groups play a pivotal role.

The effect of quin2 on chemotaxis by polymorphonuclear leukocytes.

Exposure of rabbit polymorphonuclear leukocytes to micromolar concentrations of quin2-AM results in high intracellular concentrations of quin2, which lead to inhibition of chemotaxis. The loading efficiency of polymorphonuclear leukocytes, being the percentage of quin2-AM which is taken up by the cells and transformed intracellularly into quin2, is very high, reaches a maximum after 30 min, is independent of the presence of extracellular Ca2+ and is fairly independent of cell concentration. As a consequence, inhibition of chemotaxis is strongly dependent on experimental conditions: with a low cell density (3 X 10(6)/ml) exposure to 20 microM quin2-AM results in complete inhibition of chemotaxis, whereas the same concentration of quin2-AM is nearly without effect when an 8-fold higher cell concentration is used. Inhibition by quin2 is dependent on extracellular Ca2+; inhibition is more pronounced in the absence of extracellular Ca2+ than in its presence. It is suggested that quin2 inhibits chemotaxis by interference with intracellular Ca2+.

The effect of verapamil and other calcium antagonists on chemotaxis of polymorphonuclear leukocytes

The calcium antagonists verapamil, D600, prenylamine, diltiazem and perhexiline inhibit chemotaxis and (stimulated) locomotion by rabbit polymorphonuclear leukocytes (PMNs). Inhibition occurs at relatively high concentrations (5.10(-5)-10(-3) M). Inhibition was most pronounced in the absence of extracellular Ca2+. In the presence of extracellular Ca2+ there was also inhibition of chemotaxis, but to a lesser degree than in the absence of Ca2+, thus extracellular Ca2+ partly reverses the inhibition of chemotaxis by calcium antagonists. The results suggest that the mechanism of action of these drugs in PMN chemotaxis differs from that in smooth muscle cell contraction, where much lower concentrations of these drugs are active. The data obtained suggest that calcium antagonists inhibit PMN chemotaxis and locomotion by interference with a Ca2+-dependent intracellular target.

Inhibition of polymorphonuclear leukocyte functions by chlortetracycline

Chortetracycline (CTC) inhibits chemotaxis, exocytosis and metabolic burst in rabbit polymorphonuclear leukocytes (PMNs), when these cells are activated in the absence of extracellular Ca2+. In the presence of extracellular Ca2+ CTC has little or no inhibiting effect on these functions. The inhibiting effect of CTC in the absence of Ca2+ occurs at concentrations which are not cytolytic. The inhibiting effect of CTC can be reversed by washing the cells or by addition of Ca2+ or Mg2+ to CTC-pretreated cells. Inhibition of the metabolic burst by CTC depends on the activator used. When phorbolmyristate acetate is used to activate the cells, the metabolic burst is inhibited at lower CTC concentrations than with chemotactic peptide as an activator. With due observance of the chemical properties of CTC and literature data about the requirement of intracellular Ca2+ for neutrophil functions, the results obtained are consistent with the view that CTC interferes with neutrophil functions, by complexing intracellular Ca2+, and that this inhibition is reversed when sufficient extracellular Ca2+ moves into the cell.

Exocytotic enzyme release from rabbit polymorphonuclear leukocytes after treatment with fluoride and calcium

Treatment of rabbit polymorphonuclear leukocytes (PMNs) with 20 mM sodium fluoride for 10 min, followed by removal of fluoride and addition of Ca2+ results in extensive exocytosis. This is apparent from a strong lysozyme release, together with a slight LDH release. During fluoride-activated Ca2+-dependent exocytosis an increase of indo fluorescence and a strong association of 45Ca with the cells occurs. Different inhibitors inhibit both 45Ca association and lysozyme release. Pretreatment of PMNs with pertussis toxin, or the presence of AI3+ in the medium has little effect on fluoride-activated Ca2+-dependent exocytosis. During pretreatment with fluoride, the ATP level strongly decreases. Exocytosis nevertheless occurs upon addition of Ca2+, indicating that a normal ATP level is not required for exocytosis. The glycogen content of the cell strongly decreases during exposure to Ca2+ after pretreatment with fluoride, but not during pretreatment with fluoride. Breakdown of glycogen and accumulation of 3-phosphoglycerate suggest that glycolysis is blocked at the enolase step, but proceeds as far as that step.

Protamine sulfate-induced enzyme secretion from rabbit neutrophils

Protamine sulfate induces enzyme secretion from rabbit neutrophils. Enzyme secretion is mainly due to exocytosis but, depending on the experimental conditions, a small amount of cytolysis may occur. As compared with stimulation of neutrophil functions by other activators, protamine sulfate-induced enzyme release by exocytosis is a relatively slow process and is not accompanied by a marked activation of the metabolic burst. For optimal exocytosis, extracellular Ca2+ is required, but there is still some enzyme release in its absence, and other metal ions (Sr2+, Ba2+, Mg2+) can partly mimic the effect of Ca2+. Positive charges on protamine are of primary importance because the polyanion heparin completely inhibits protamine sulfate-induced enzyme release. Protamine linked to agarose beads is able to induce enzyme release; thus the induction of exocytosis is due to an interaction of the positive charges on protamine with the plasma membrane. Sialic acid residues on the membrane, however, seem not to play an important role in this process.

Permeabilization and calcium-dependent activation of rabbit polymorphonuclear leukocytes by poly-L-arginine

In the absence of extracellular Ca2+, poly-l-arginine induces little lysozyme release from rabbit polymorphonuclear leukocytes (PMNs), The polycation causes plasma membrane damage, which is evident from the release of the cytoplasmic enzyme lactate dehydrogenase (LDH). In the presence of Ca2+ concentrations higher than 0.2 mM, poly-l-arginine induces a strong lysozyme release that is superimposed on the membrane-damaging effect. The results suggest that poly-L-arginine permeabilizes the plasma membrane, enabling Ca2+ to enter the cell, which results in the exocytotic release of granule constituents. The GTP analog GTPγS shifts the Ca2+ requirement of exocytosis to slightly higher concentrations, whereas it completely inhibits poly-l-arginine-induced LDH release. Pertussis toxin gives a moderate inhibition, and La3+ completely inhibits poly-L-arginine-induced enzyme release. Whereas poly-l-arginine alone induces little superoxide generation in rabbit PMNs, there is a synergistic enhancement of superoxide production when GTPγS and poly-l-arginine are present together. Guanine nucleotides apparently have a modulating effect on the actions of poly-l-arginine on the PMN, but the nature of this effect remains to be determined.

Latex phagocytosis by polymorphonuclear leukocytes: Role of sialic acid groups

Polystyrene latex particles are rapidly phagocytized by rabbit polymorphonuclear (PMN) leukocytes. The uptake is influenced by macromolecules which have the effect of altering the surface charge of the latex particle. The influence of polylysines of varying chain length on the surface charge of latex particles and of PMN cells was studied by micro-electrophoresis. Charge reversal at the latex surface was found to occur at concentrations considerably below that at which the surface charge of the PMN cells is reversed. Phagocytosis of latex by PMN cells is enhanced in the presence of low concentrations of long-chain polylysines. The enhancement of phagocytosis is strongly reduced if PMN cells are treated with neuraminidase. This suggests participation of sialic acid groups in a stage of particle-cell interaction which precedes engulfment.

Felodipine-induced inhibition of polymorphonuclear leukocyte functions

Felodipine inhibits fMet-Leu-Phe or ionophore A23187-induced exocytosis in rabbit peritoneal polymorphonuclear leukocytes (PMNs), in the concentration range 1-50 microM. Activation of the metabolic burst, and migration of PMNs towards fMet-Leu-Phe are equally inhibited by felodipine in the same concentration range. The effect is not due to blocking of calcium channels in the plasma membrane, because the degree of inhibition remains the same when Ca2+ is omitted from the medium. Felodipine interferes with ionophore A23187-induced association of 45Ca with the PMN but this interference occurs at lower concentrations than the inhibition of exocytosis. Hypotonic hemolysis of erythrocytes is inhibited by felodipine; maximal protection against hemolysis occurs at a concentration of 50 microM felodipine. It is suggested that at least a part of the inhibiting effect on PMN functions might be due to an anesthetic-like membrane effect of felodipine.

Modulation of rabbit neutrophil chemotaxis by cytochalasin A. A comparison with other neutrophil functions

Chemotaxis and stimulated locomotion by rabbit peritoneal neutrophils are inhibited by cytochalasin A; inhibition is complete with 5 × 10−7 M cytochalasin A. Under the same conditions exocytosis and activation of the metabolic burst are stimulated maximally with 5 × 10−7 M cytochalasin A. Inhibition of these function occurs at concentrations higher than 10−6 M, inhibition being complete with 5 × 10−6 M cytochalasin A. The sulfhydryl compound glutathione can prevent the inhibitory effect of cytochalasin A on chemotaxis. Inactivation of the sulfhydryl groups on the outer surface of the plasma membrane does not affect the inhibitory effect of cytochalasin A. It is concluded that the cellular target of cytochalasin A has a different significance for chemotaxis as compared with exocytosis and metabolic burst. The possibility that cytochalasin A reacts with sulfhydryl groups, located on a structure closely connected with the inner surface of the plasma membrane, is discussed.