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Mature polymorphonuclear leukocytes (PMN) are capable of mediating phorbol myristate acetate (PMA)- and antibody (A)-dependent cellular cytotoxicity (DCC) against ox red blood cells (ORBC) by using oxidative means. The purpose of the present study was to investigate the acquirement of these cytotoxic functions during PMN ontogeny, using the promyelocytic HL-60 cell line as a model for PMN differentiation. HL-60 cells were induced to differentiate along the PMN pathway by exposure to dimethyl sulfoxide (DMSO). Uninduced HL-60 cells were found to be completely devoid of PMA-DCC and ADCC activity. DMSO-induced cells progressively acquired the capacity to kill ORBC and to undergo the activation of oxidative metabolic burst when triggered by PMA. Despite approximately 40% of them also were capable of binding IgG-sensitized ORBC, no ADCC activity and respiratory burst activation was observed: this finding indicates that maturing HL-60 cells require a more complete maturation than that induced by DMSO to actually exert ADCC. Together the results suggest that: a. the acquirement of both PMA-DCC and ADCC potential is a post-promyelocytic event; b. the cytotoxicity activating stimuli, PMA and IgG-coated targets, follow different post-receptor transductional pathways to trigger the effector cell lytic systems: only the PMA receptor-linked pathway develops during DMSO-driven differentiation of HL-60 cells.

A-35-year-old woman with a long-lasting history of neutropenia and recurrent infections was found to have defective neutrophil chemotaxis, random motility, and in vivo migration. Although the bone marrow granulocyte reserve was normal, the patient failed to release an appropriate amount of granulocytes after injection of etiocholanolone. These features are characteristic of the so-called "Lazy leukocyte syndrome". The clinical presentation of the five cases of this syndrome so far reported and its pathophysiological aspects are discussed.

Normal human neutrophils were found to destroy ox red blood-cell targets when incubated on micropore filters coated with aggregated IgG, as determined by the 51Cr release method. An intact neutrophil oxidative metabolism was essential for the cytotoxic event, since cells from patients with chronic granulomatous disease failed to exert any cytolysis. The target-cell destruction was prevented by catalase, azide, and cyanide and was enhanced by superoxide dismutase, suggesting involvement of the myeloperoxidase-hydrogen peroxide system. Neutrophil-mediated cytotoxicity was markedly amplified by the chemotactic peptide N-formyl-methionyl-leucylphenylalanine, as a result of an increased activity of the myeloperoxidase-hydrogen peroxide cytolytic system itself. This system of cytotoxicity provides a direct evidence for the neutrophil capacity of destroying bystander target cells under conditions simulating the in vivo immunologically mediated tissue injury and offers an excellent model to study events occurring during immune complex diseases.

The locomotion of human neutrophils deactivated by pretreatment with appropriate concentration of chemoattractant was investigated in experimental conditions capable of dissociating random from truly directional motility. It was shown that deactivated neutrophils lose true chemotactic responsiveness, being unaffected by the rate of locomotion. Ascorbic acid, when present in appropriate concentrations during the deactivation step, completely prevented the loss of true chemotactic responsiveness by cells--that is, their chemotactic deactivation.

Leukaemic blast cells from 20 patients with acute leukaemia were examined for their capacity to mediate cytotoxicity against ox red blood cells in the presence of phorbol myristate acetate (PMA), a system widely employed as an in vitro model of tissue damage by metabolically activated mature phagocytes. Blasts from certain patients with myelomonocytic and monocytic leukaemia behaved like efficient killer cells. Conversely, leukaemic myeloblasts and promyelocytes as well as leukaemic lymphoblasts were ineffective. Blast cells capable of inducing the target cell lysis were also capable of mounting an oxidative respiratory burst upon challenge with PMA, as detected by the superoxide anion release. N-ethyl-maleimide, superoxide dismutase and catalase completely inhibited the cytotoxicity by monocytoid blast cells, suggesting the involvement of oxygen reactive products in the lethal hit itself. The cytolytic potential of blasts committed to monocytic differentiation might be an additional factor contributing to the tissue damage in a subpopulation of leukaemic patients.

Experiments were performed to investigate the in vitro effects of inosiplex on the locomotion and oxidative metabolism of human neutrophils. The drug, when used at the concentration of 500 micrograms/ml, significantly increased neutrophil random migration and chemotaxis. Enhancement of chemotaxis was due to an increased rate of locomotion as well as toan increased true chemotactic response. Inosiplex appeared to exert its effect on the whole migrating cell population. No significant effect on the oxidative metabolism of both resting and phagocytizing neutrophils was found. The results suggest that the effect of inosiplex may be related chiefly to its effect on neutrophil locomotion.

Human neutrophils, incubated in the presence of zymosan-activated serum ( ZAS ), were able to damage ox red blood cells ( ORBC ) as assessed by a 4-hr 51Cr-release assay. Cells from a patient with chronic granulomatous disease were incapable of mediating ZAS -induced cytolysis. Cytotoxicity by normal neutrophils was prevented by N-ethyl-maleimide as well as by catalase, but heat-inactivated catalase and superoxide dismutase were completely ineffective. Heme enzyme inhibitors, azide and cyanide, only partially reduced the target cell lysis. The results indicate the involvement of products of the oxidative metabolism, mainly hydrogen peroxide, as cytolytic mediators. These mediators released by neutrophils may contribute to the tissue injury occurring during inflammation.

Neutrophils pretreated with a chemotactic factor become deactivated, that is, unresponsive to a subsequent chemotactic stimulus. A decay of surface esterase activity, an increased adhesiveness, an auto-oxidative damage of the cell structures and microtubule hyperpolymerization have been previously proposed as the cause of the chemotactic deactivation. To further understand the cellular mechanisms involved in this process, we have studied the locomotory behaviour of deactivated neutrophils in an experimental system allowing differentiation among the various types of migratory cellular response to chemotactic factors. Thus, we have found that deactivated neutrophils retain a normal capacity of random locomotion, but entirely lose their true chemotactic responsiveness. Diamide, which disrupts assembled microtubules in concanavalin A-treated cells, was able to completely restore the true chemotactic response of deactivated cells. These observations suggest that neutrophil deactivation is a reversible cellular event dependent on the functional state of the microtubular system.

The purpose of the present study was to investigate the general conditions under which neutrophil polymorphonuclears (PMN) mediate antibody-independent cytolysis in the presence of normal human serum (NHS). Normal PMN were found to kill rabbit red blood cells (RRBC) only when cultured with 1% NHS. NHS was per se incapable of lysing RRBC. PMN from a patient with Chronic Granulomatous Disease did not destroy RRBC targets even in the presence of 1% NHS. In addition, cytotoxicity by normal PMN was significantly reduced by scavengers of oxygen metabolites. The results suggest that the target cell lysis by PMN in the presence of NHS requires a synergistic interaction between at least two mediators: serum factor(s) and oxygen metabolite(s).

Granulocyte function was studied in 9 patients with untreated, Ph1-positive chronic myelocytic leukemia (CML). The nitroblue tetrazolium reduction by stimulated granulocytes was impaired in all patients; 4 patients also had diminished phagocytosis and 2 others defective chemotaxis. In spite of this variety of polymorphonuclear (PMN) functional impairments, there is little evidence of increased susceptibility to infections in CML patients. This suggests that CML-PMN leucocytes (PMNs) may be successfully used for transfusion into neutropenic recipients, as previously reported. To evaluate the effects of irradiation and liquid storage on CML-PMNs, 5 of our patients were subjected to leukapheresis by continuous-flow centrifugation in the Aminco Celltrifuge, and granulocyte functional capacities were also evaluated on the cell-rich plasma immediately after collection and after short-term storage at 4 degrees C with or without irradiation (1500 rads). As evaluated by in vitro studies, granulocytes maintained, even after irradiation, functional activities similar to those found immediately after collection up to 24 h of storage at 4 degrees C and presented a moderate loss of function after 48 h. Chemotaxis appeared to be the most sensitive detector for cellular damage of stored leucocytes, irradiated and non-irradiated, so that it might be used for assessment of leucocyte function before transfusion.

A case of ulcerative colitis with defective neutrophil chemotaxis, improved after short sulfasalazine therapy, is reported. This is the first case of this disease in which the chemotactic defect was characterized as a serum inhibitor, directed toward autologous and homologous neutrophils and associated with circulating IgA. Preincubation of normal neutrophils with increasing concentrations of patient's serum resulted in a dose-related inhibition, suggesting a stoichiometric relationship between humoral inhibitory power and neutrophils.

Neutrophilic polymorphonuclear leukocytes (PMNs), triggered by opsonized zymosan (OPZ), lysed chicken red blood cells as measured by a chromium 51 release method. The lysis was prevented by scavengers of hypochlorous acid. When platelets were added to the cytolytic system, a dose-dependent inhibition of the lysis was observed. Moreover, platelets lowered the HOCl recovery from OPZ-triggered PMNs. A positive linear relationship was found between the extent of the lysis mediated by and the amount of HOCl recovered from PMNs. Finally, both the inhibition of the lysis and the reduction of the HOCl recovery induced by platelets were prevented by pulsing platelets with carmustine (BCNU) to block their glutathione cycle. These results suggest that platelets act by consuming via their glutathione cycle significant amounts of PMN-derived hydrogen peroxide, with a consequent impairment of the PMN HOCl production and, in turn, lytic efficiency. Consistent with such a conclusion, platelets were found to consume PMN-derived H2O2 via a BCNU-inhibitable process. Further, the platelet inhibitory activity could be abolished by the addition of an appropriate extra-flux of enzymatically generated H2O2. No evidence for a platelet-induced inhibition of OPZ-PMN interaction, PMN myeloperoxidase release, and H2O2 production was obtained. The present study provides direct evidence for a platelet-dependent mechanism capable of controlling the PMN production of highly reactive oxidants and, in turn, the PMN cytolytic activity.

Rosettes with ox erythrocytes coated with purified IgG antibody were used to detect Fc receptors on neutrophils from 60 patients with solid neoplasias and from 55 normal controls. The mean average of the rosettes in the patients was 48.10%, and that in normal controls was 79.42%, with a highly significant difference according to the Wilcoxon test [negative probability, P(w)-4.47 . 10(-5)]. The low proportion of patients' rosettes ws related to the presence of a serum factor, which also inhibited normal neutrophil rosette formation. Patient neutrophils (or normal neutrophils treated with patient sera) recovered their rosetting capacity when cultured in vitro. No correlation was found between low percentages of rosette-forming cells and the level of circulating immune complexes of the individual patients. Additional evidence also supported the finding that IC and the serum factor are probably unrelated.