R. R. Strauss

The role of the phagocyte in host-parasite interactions: XIX. Leukocytic glutathione reductase and its involvement in phagocytosis

Abstract Glutathione reductase activity was found in guinea pig peritoneal polymorphonuclear leuckocytes. The cofactor requirement for the enzyme was found to be NADPH. This was determined by the requirement of GSSG for oxidation of the reduced cofactor and the stimulatory effect of KCN on GSH peroxidase which was also found in these cells. Enzyme activity was inhibited by the sulfhydryl antagonist N -ethyl maleimide. The addition of GSSG to the reaction mixture resulted in an increase in cellular GSH content. The activity of the enzyme was confined to the 20,000 g supernatant fraction. Phagocytosis caused a significant increase in GSSG reductase activity within 15 sec after particle addition. The NADPH oxidase stimulation which has been associated with the phagocytic process did not occur until some minutes after the addition of latex particles. Since NADP + is a limiting factor for the hexose monophosphate shunt and GSSG reductase can increase the NADP + content of the cell, one can now postulate that increased activity of this enzyme is an early event involved in the phagocytosis-associated increase in this metabolic pathway.

Peroxidase Mediated Antimicrobial Activities of Alveolar Macrophage Granules

The 20,000g pellet obtained by centrifugation of a homogenate of rabbit alveolar macrophages has antibacterial activity in the presence of a hydrogen peroxide-generating system and iodide. Peroxidase activity has been demonstrated in this fraction. Addition of 3-amino-1,2,4-triazole diminished the antibacterial activity of the pellet-hydrogen peroxide-iodide system.

Direct involvement of nadph oxidase with the stimulated respiratory and hexose monophosphate shunt activities in phagocytizing leukocytes

Abstract Guinea pig peritoneal exudate polymorphonuclear leukocytes (PMN) have KCN-sensitive and insensitive NADPH oxidation activity. These activities can be localized to different homogenate fractions. NADPH oxidation activity of the 19 000 g pellet is KCN-insensitive. Myeloperoxidase (MPO) activity which is also confined to this fraction is completely inhibited by 1 mM KCN. This observation indicates that KCN-insensitive NADPH oxidation is not due to MPO. KCN-sensitive NADPH oxidation activity is found in the 19 000 g supernatant fraction, as is catalase. Purified beef liver catalase does not oxidize NADPH, thus indicating that the NADPH oxidation by the supernatant is not due to the presence of catalase. NADPH oxidation activity of both the 19 000 g pellet and supernatant fractions collected from phagocytizing cells is significantly greater than that found in the corresponding fractions collected from resting cells. The increased oxidative burst associated with phagocytosis can be accounted for by the stimulated NADPH oxidase activity.

The role of the phagocyte in host-parasite interactions. XXIII. Relation of bactericidal activity to peroxidase-associated decarboxylation and deamination

Abstract Peroxidase-mediated decarboxylation and deamination of appropriate substrates appear to be associated with bactericidal activity of the myeloperoxidase-H2O2-chloride antimicrobial system. Products resulting from these reactions, possibly aldehydes, may be the actual bactericidal agents.

The role of the phagocyte in host-parasite interactions: XXV. Metabolic and bactericidal activities of leukocytes from pregnant women☆☆☆

Abstract The hexose monophosphate shunt (HMS) activity of phagocytizing cells collected from pregnant women is approximately twice that found in nonpregnant women. HMS activity in resting leukocytes from bacteriuric pregnant women was lower than that found in leukocytes from normal pregnant women. Phagocytizing cells showed increased activity. The myeloperoxidase (MPO) activity of leukocytes from pregnant women increased significantly during phagocytosis. This increase was not observed in nonpregnant women. Granules containing 20,000 × g MPO isolated from leukocytes from pregnant women have bactericidal activity in the presence of nonbactericidal concentrations of H 2 O 2 in a chloride medium. These granules also decarboxylate alanine in the presence of H 2 O 2 in a chloride medium. Freshly isolated strains of bacteria are not killed as effectively as stock strains. This appears to be due to decreased phagocytosis and/or resistance of the fresh isolates to the MPO-H 2 O 2 -chloride bactericidal system.

The role of the phagocyte in host-parasite interactions. XXXIX. Stimulation of bactericidal activity of myeloperoxidase-containing leukocytic fractions by estrogens.

Abstract Estriol and estrone have been shown to stimulate bactericidal activity of myeloperoxidase (MPO)-containing 20,000 × g granule pellets obtained from guinea pig peritoneal polymorphonuclear leukocytes. Beta estradiol and progesterone were without effect. An MPO-mediated amino acid decarboxylation reaction, optimal at acidic pH and postulated as a mechanism involved with the bactericidal activity, was also stimulated by estriol and estrone but not by beta estradiol and progesterone. In contrast, no stimulation of MPO-mediated guaiacol oxidation at pH 7.0 was observed with any of these hormones. The stimulation of bactericidal activity by estriol and estrone was not observed if highly purified human MPO was substituted for the guinea pig MPO-containing 20,000 × g granule pellets. These observations suggest that estriol and estrone may be stimulating MPO-mediated bactericidal activity through an indirect mechanism. It is possible that hyperbactericidal leukocyte activity observed in pregnancy may be related, at least in part, to stimulation of its MPO-mediated bactericidal activity and amino acid decarboxylation by estriol and/or estrone.

The Biochemical and Antimicrobial Activities of the Phagocyte

Many factors determine whether a parasite can establish itself successfully in a host and initiate an infectious process. An important mechanism that a host can employ to combat disease and that can be studied, in vitro, at the cellular level is phagocytosis. The importance of this host-defense mechanism has been apparent since the process was first described by Metchnikoff in the latter part of the nineteenth century. However, the mechanism(s) involved in engulfment and destruction of ingested microorganisms have been the subject of much study and controversy. The early literature dealing with phagocytosis has been adequately reviewed (1,2). These scholarly works concentrate mainly on the physical forces involved with engulfment and also on the different factors that can effect the phagocytic act. Inexplicably little or no studies on the biochemical aspects of phagocytosis were carried out or reported in this early period. Some scattered reports, however, did appear (3). Somewhat later some excellent work was carried out in Nungester’s laboratory. Unfortunately, most of this work was published in thesis form only (4, 5). These workers all demonstrated that the phagocytic act was accompanied by biochemical changes; specifically, an increased respiratory activity. Suter and his co-workers in the early 1950’s initiated a series of studies concerned with the interaction of tubercle bacilli and guinea pig exudate cells. They described this interaction at the biochemical level (6–8).