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Regulation of C5a and formyl-methionine-leucine-phenylalanine-lysine (fMLPL) receptors on human monocytes has been studied using fluorescein-conjugated derivatives and flow cytometry. Monocytes have receptors for each of these ligands, as evidenced by their ability to bind specifically biologically active fluorescein derivatives of these ligands. Quenching experiments showed that bound fluoresceinated C5a and fMLPL are rapidly internalized at 37 degrees C. Once internalized, monocytes are able to reexpress these receptors, returning to control levels within approximately 90 min. This contrasts with rate differences seen in polymorphonuclear neutrophils (PMNs), where fMLPL receptors return more rapidly (approximately 30 min) than do C5a receptors (approximately 100 min). Monensin inhibited the reexpression of C5a but not fMLPL receptors, suggesting that a receptor recycling process is necessary to replenish C5a receptors on the monocyte surface. Similar although less efficient inhibition of C5a receptor reexpression was observed with NH4Cl treatment. Reexpression of both C5a and fMLPL receptors was independent of extracellular Ca2+. Treatment with various agents known to stimulate monocytes and PMNs increased the expression of fMLPL receptors in both cell types but either had no effect on or reduced the level of C5a receptor expression. This would indicate that monocytes, like PMNs, have intracellular pools of preformed fMLPL receptors, available for reexpression. These studies show that, like PMNs, monocytes modulate C5a and fMLPL receptors through different mechanisms. Furthermore, monocytes are capable of reexpressing these receptors following exposure to ligand, a theoretical requirement for chemotaxis.

The neuropeptides beta-endorphin and met-enkephalin are potent analgesics and have a broad spectrum of biologic activities including the recently described alterations of lymphocyte proliferation and antibody production. The current study demonstrates that beta-endorphin and met-enkephalin stimulate human mononuclear cell chemotaxis, as measured by the in vitro leading front assay for migration. The response to both beta-endorphin and met-enkephalin was bimodal, with peak activities occurring at 10(-12) M and 10(-8) M. The distance migrated in response to optimal concentrations of beta-endorphin or met-enkephalin was approximately 80% of that obtained with 10(-8) M formyl-methionyl-leucyl-phenylalanine (f-MLP) and was blocked by prior incubation with 10(-8) M naloxone. Removal of glass adherent cells resulted in a loss of the response to beta-endorphin. Quantitation of the number of cells responding to beta-endorphin showed that only about 50% as many cells responded to beta-endorphin as compared with f-MLP. Human neutrophils showed some migration in response to beta-endorphin and met-enkephalin, although the average optimal migration was less than 30% of that observed with 10(-8) M f-MLP. Studies of the in vivo infusion of beta-endorphin into the cerebral ventricle of the rat resulted in the immigration of macrophage-like cells and are consistent with the in vitro evidence for a chemotactic effect of beta-endorphin.

Peripheral blood monocytes undergo an oxidative burst similar to that seen in neutrophils. The basis for this response appears to be an NAD(P)H oxidase that utilizes reduced NAD(P)H to form superoxide anion. We utilized the unique UV-stimulated fluorescence property of reduced pyridine nucleotides to analyze NAD(P)H utilization in monocytes. UV-stimulated fluorescence in mononuclear cell preparations indicated two populations of cells with the highly fluorescent cells having a Coulter volume consistent with that of monocytes. Dual laser analysis with monoclonal antibodies confirmed that these highly fluorescent cells are monocytes by showing them to be OKM1+, Leu DR+, and anti-monocyte 0.2+. Natural killer (NK) cells, as defined by Leu 7, were not found in this highly fluorescent population. Stimulation of mononuclear cells with phorbol myristate acetate caused a fluorescence loss indicative of NAD(P)H oxidation in monocytes but not in lymphocytes. Stimulation with suboptimal concentrations of PMA (1-5 ng/ml) resulted in a dose-dependent fluorescence loss in monocytes that occurred in an all-or-none fashion identical to the pattern observed in neutrophils. Simultaneous measurement of H2O2 production using dichlorofluorescein formation with NAD(P)H fluorescence indicates that oxidant production occurs in a graded manner. This method, then, provides a convenient way to study in single cells the metabolic events involved in depletion and replenishment of NAD(P)H during the oxidative burst and demonstrates an additional means by which to distinguish monocytes from lymphocytes using flow cytometry.

Fucose binding lectin (FBL) from Lotus tetragonolobus seeds has previously been shown by fluorescence microscopy to bind to human neutrophils. This study shows using highly sensitive flow cytometry that this lectin binds both to human peripheral blood neutrophils and monocytes but not to lymphocytes. This binding is blocked by the presence of free L-fucose and is reversible when neutrophils or monocytes stained with fluorescent FBL are subsequently incubated in 0.05 M L-fucose. Quantitative comparison of neutrophils and monocytes from the same individual show that neutrophils bind approximately 2.6 times more FBL than monocytes and that FBL binding is more efficiently reversed with neutrophils, as compared with monocytes, by L-fucose. Additional double-labeling studies of cells with FBL and the OKM1 monoclonal antibody, which identifies monocytes and granulocytes, show that all cells binding FBL also stain with the OKM1 monoclonal antibody. This study shows that qualitatively, FBL may be utilized as a human myeloid cell marker to differentiate peripheral blood monocytes and neutrophils from lymphocytes.

Chronic renal failure patients maintained on dialysis have an increased risk for infection. This article summarizes research that has been done on the function of neutrophils (PMNs) and monocytes from chronic hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) patients. The studies involving the HD patients showed that there is a decreased PMN in vitro chemotactic response, decreased C5a receptors on both PMNs and monocytes, and decreased oxidative metabolic responses of PMNs and monocytes to the chemotactic stimuli C5a and formyl-met-leu-phe (fMLP), but not to nonchemotactic factors. The results of studies involving phagocytosis have been conflicting and are discussed in this paper. Due to the basic principles of peritoneal dialysis, this treatment approach depletes the peritoneum of phagocytic cells, adversely affects the function of peritoneal WBCs, dilutes the existing opsonins, and alters the physiologic environment of the peritoneal cavity. Studies of peripheral PMN and monocyte function in CAPD patients have shown that, similar to HD patients, they also have decreased C5a receptors and decreased oxidative metabolic responses to the chemotactic factors C5a and fMLP. Although the factors contributing to the risk of infection in chronic dialysis patients are multifaceted, there are definitely alterations in PMN and monocyte function.

Peripheral neutrophil and monocyte function was studied in 24 pediatric patients with recurrent lower respiratory tract infections referred to the University of New Mexico Pediatric Pulmonary Center. The age range of the patients was 4 months to 12 yr with a mean of 32 months. Depressed neutrophil chemiluminescence activity was observed in 46% of the 24 patients studied, depressed neutrophil chemotactic responsiveness was observed in 27% of 22 patients, depressed monocyte chemiluminescence was observed in 31% of 16 patients, and depressed monocyte chemotaxis was observed in 45% of 20 patients. Of the 11 patients with abnormal neutrophil chemiluminescence, 56% also had abnormal neutrophil chemotaxis. Eight of the 11 patients with depressed neutrophil chemiluminescence were retested 9 months to 1 yr after initial testing, and in 5 of the 8, chemiluminescence had returned to normal, whereas in the other 3, it had remained depressed. The 3 whose neutrophil chemiluminescence remained depressed continued to have repeated pneumonias as did one patient whose values had returned to normal. The estimated probability of return to normal for the neutrophil chemiluminescence after 1 yr was 0.625. These findings suggest that because the ability to move functionally intact neutrophils and monocytes into the lung is important for effective lung defense, that the defective phagocytic cell function defined here may be directly related to the recurrent pneumonias found in these pediatric patients.

The opioid neuropeptides have previously been shown to bind to and affect leukocyte function including lymphocyte proliferation, NK-cell activity, mononuclear cell chemotaxis, immunoglobulin synthesis, and lymphokine production. The effect of the opioid peptides beta-endorphin and Met-enkephalin on interferon gamma (IFN) production by concanavalin A-stimulated human mononuclear cells was examined. Both beta-endorphin and Met-enkephalin enhanced IFN production by the majority of donor mononuclear cells tested and did so at concentrations between 10(-14) and 10(-10) M. When 10(-12) M beta-endorphin or Met-enkephalin were included in concanavalin A-stimulated mononuclear cell cultures, IFN concentrations were significantly enhanced to 205 +/- 45 and 252 +/- 67% of control, respectively. Although the majority of cell preparations tested exhibited an enhanced production of IFN in response to these opioid peptides, some did not. When beta-endorphin or Met-enkephalin were utilized at 10(-11) M, 10 of 15 and 7 of 11 responded with IFN production greater than 20% above the control (untreated) level. There was not an absolute correlation between an enhanced response to beta-endorphin and Met-enkephalin, suggesting the presence of multiple receptor types on these cells for opioids. The opioid receptor antagonist, naloxone, did not significantly prevent the opiate effect. When 10(-8) M naloxone was included in cultures containing 10(-12) M beta-endorphin or Met-enkephalin no significant inhibition of the effect of either opioid on IFN production was observed.

Chronic renal failure patients on dialysis have an increased susceptibility to infection. Previous studies have demonstrated that these patients have a decreased in vitro neutrophil (PMN) chemotactic response and a reduction in C5a receptor availability on both PMN and monocytes. This study was designed to determine if other chemotactic factor-mediated responses of PMN and monocytes from hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) patients are abnormal. The responses investigated included in vitro chemotaxis, superoxide generation, H2O2 production, and myeloperoxidase (MPO) release. These studies showed that PMN from HD and CAPD patients are significantly decreased in their chemotactic response to both C5a and fMLP when compared to normal controls. The response of HD patient's PMN to C5a was decreased by an average of 55.1% (p less than 0.005) and for CAPD patients by 49.7% (p less than 0.01). Similarly, chemotactic responses to fMLP were decreased by an average of 44.7% (p less than 0.005) for HD patients and 36.3% (p less than 0.02) for CAPD patients. Superoxide anion production by PMN and monocytes from HD and CAPD patients in response to C5a and fMLP was also significantly decreased compared to controls. PMN superoxide production in response to C5a was decreased by an average of 36.5% (p less than 0.001) for HD patients and 32.0% (p less than 0.001) for CAPD patients. fMLP-stimulated production of superoxide was also decreased but to a lesser degree with a mean decrease of 18.0% (p less than 0.01) for HD patients and 24.1% decrease (p less than 0.01) for CAPD patients. This decreased responsiveness was restricted to C5a- and fMLP-stimulated superoxide production since phorbol myristate acetate (PMA)-stimulated responses were comparable to controls. A similar pattern of decreased superoxide production was found with monocytes from these patients. Comparable decreases in chemotactic factor-stimulated responses were also observed in a flow cytometric assay of H2O2 production in both PMN and monocytes and an in vitro assay of MPO release from PMN. Analysis of the binding of fluorescent C5a to PMN showed a direct correlation between decreased C5a binding and decreased O2- production and MPO release. Since all of these chemotactic factor-stimulated events are involved in the inflammatory process and the killing of microorganisms, alterations in these WBC functions in dialysis patients may contribute to their increased susceptibility to infection.

Chronic renal failure patients have an increased risk for infection which may partially be due to altered chemotactic ability of their white blood cells. This study was designed to evaluate chemotactic factor and Fc receptor expression on neutrophils (PMN) and monocytes from chronic renal failure patients on hemodialysis (HD) or continuous ambulatory peritoneal dialysis (CAPD). Analysis of these receptors was performed using flow cytometry and fluorescent chemotactic factors (C5a, f-Met-Leu-Phe-Lys [fMLPL] and casein) and heat-aggregated human IgG. Peripheral blood PMN and monocytes obtained from 14 HD patients (in the predialysis period) and 14 CAPD patients were analyzed for their ability to bind each of the fluoresceinated ligands. PMN and monocytes from both patient groups had a significant reduction in their ability to bind C5a. The average percentage (+/- s.e.m.) of PMN that bound C5a was 93.9 +/- 1.1 for the controls, 72.9 +/- 3.8 for HD patients, and 79.3 +/- 4.0 for CAPD patients. Similar results were obtained with monocytes with 69.7 +/- 1.9% for controls, 54.6 +/- 4.5% for HD patients, and 31.0 +/- 4.5% for CAPD patients. These differences in C5a binding were also reflected in the average intensity of fluorescence. There was no significant difference in the percentage or fluorescence intensity of PMN or monocytes that bound casein or aggregated IgG when either group of dialysis patients was compared to the control values. Binding of fMLPL by PMN and monocytes from the HD patients and PMN from the CAPD patients were similar to control values but the binding of fMLPL by monocytes from CAPD patients was significantly suppressed (p less than 0.03).(ABSTRACT TRUNCATED AT 250 WORDS)