Polymorphonuclear cell function in rheumatoid arthritis and in Felty''s syndrome.

Tests for polymorphonuclear cell (PMN) chemotaxis, adherence, and electrophoretic mobility (EPM) were carried out on blood PMN isolated from 27 normal subjects, 16 patients with uncomplicated rheumatoid arthritis (RA), and 9 patients with Felty's syndrome. Chemotaxis was measured by a modification of the Boyden chamber technique, adherence by retention of cells on nylon fibre columns, and EPM in a cylindrical electrophoretic assembly. There was no significant difference between the chemotactic migration of normal and rheumatoid PMN as assessed by the leading front measurement. However, PMN from patients with Felty's syndrome showed significantly reduced chemotaxis (P less than 0.001). Computerised image analysis showed this impaired migration to be due to an overall reduction in cell motility rather than loss of a subset cells. Activated serum from patients with RA and Felty's syndrome were as good chemoattractants as activated pooled AB serum. There was no significant difference in the adhesiveness of PMN from normal persons and rheumatoid patients, though PMN from patients with Felty's syndrome did show a trend to lower adhesiveness. Both RA and Felty's syndrome patients had an increase in the proportion of PMN of lower surface charge than controls. Direct correlations were observed between cells of high surface charge and nonadhesiveness.     

Phagocytes and C4 in Paraproteinaemia

Immunological studies were performed on patients with multiple myeloma. A defect in polymorphonuclear leucocyte (PMN) function as evidenced by diminished adherence of these cells to nylon fibre columns was detected in 16, and low levels of the fourth component of complement (C4) were observed in 14, of the 26 patients studied. Twelve of the patients with low C4 exhibited the defect in PMN adhesiveness whereas only four of the 12 patients with normal C4 showed the defect. The PMN defect was not caused solely by the low C4, since PMNs from seven patients with hereditary angioedema, which is associated with low levels of C4, did not show the defect. The low C4 and defect in PMN adhesiveness occurred primarily in patients with IgG myeloma; all but one of the patients with IgA myeloma, macroglobulinaemia, or light chain disease were normal in both parameters. Results of skin window studies indicated that patients with the PMN defect also had a defect in the early PMN inflammatory response. The defect in PMN adhesiveness could be completely corrected by incubating the cells in normal plasma. Binding of the C4 to paraprotein could not be demonstrated, and C1 activation was found to be caused only by one of 10 isolated paraproteins studied. These studies indicate that patients with paraproteinaemia have immunological abnormalities in addition to low immunoglobulin levels and suggest that these abnormalities may be involved in the pathogenesis of the recurrent infections commonly associated with this disease.     

Interleukin-15 enhances neutrophil functional activity in patients with human immunodeficiency virus infection

Polymorphonuclear leukocyte (PMN) dysfunction has been reported in human immunodeficiency virus (HIV)-infected patients. Interleukin (IL)-15 is a recently discovered cytokine that potentiates antimicrobial functions of normal PMNs. We evaluated the in vitro effect of IL-15 on chemotaxis and fungicidal activity of PMNs from 9 patients with untreated advanced HIV infection, 8 patients with viral suppression after 52 to 130 weeks of highly active antiretroviral therapy (HAART), and 12 patients with treatment failure. We also studied oxidative burst and apoptosis of PMNs in 5 patients with untreated advanced HIV infection. Twelve healthy donors were included as controls. Chemotaxis and fungicidal activity of unprimed PMNs was significantly lower in patients with untreated HIV infection compared with controls. After incubation with IL-15, a significant increase in PMN chemotaxis and fungicidal activity was found; moreover, IL-15 induced a significant reduction in the number of apoptotic HIV(+) PMNs. IL-15 did not modulate oxidative burst of HIV(+) PMNs as measured by chemiluminescence production. The in vitro priming of PMNs with IL-15 determined a complete reversal of defective chemotaxis and killing in all HAART-treated patients with long-term HIV suppression. IL-15 significantly enhanced chemotaxis and fungicidal activity also in patients with HAART failure. In conclusion, IL-15 is an important cytokine in the activation of the functional properties of HIV(+) PMNs, by delaying apoptosis and enhancing chemotaxis and fungicidal activity. The potent stimulant effect of IL-15 on PMN function was observed in antiretroviral naive patients as well as in individuals who were receiving HAART, including those with treatment failure. (Blood. 2000;96:1979-1984)     

Polymorphonuclear Leucocyte Chemotaxis in Boyden Chambers Effect of Low Concentrations of Vinblastine

It has been reported by previous authors that vinblastine in high and presumably non-specific concentrations (10^-5-10^-4 M) inhibits polymorphonuclear (PMN) chemotaxis in vitro. In the present study, vinblastine in subantimitotic concentration (10^-8M) induced maximal inhibition of PMN chemotaxis in a modified Boyden chamber. It is suggested that the vinblastine inhibition of PMN chemotaxis is due to an interference with the spatial reorganization of centriole-associated microtubules during PMN direction-finding or PMN directional movement. The effect of subantimitotic concentrations of vinblastine and other antitubulins on PMN chemotaxis may have important applications in the treatment of diseases in which the tissue damage is mediated by the polymorphonuclear leucocytes.     

Hemolytically inactive C5b67 complex: an agonist of polymorphonuclear leukocytes

The activity of hemolytically inactive C5b67, designated iC5b67, was evaluated as an agonist for functional responses of human polymorphonuclear leukocytes (PMN). C5b67 was formed from purified human complement components and decayed in phosphate-buffered saline (PBS) until it had no lytic activity for sheep erythrocytes in a standard assay. iC5b67, at nanomolar concentrations, stimulated PMN chemotaxis and Ca2+ fluxes, but inhibited superoxide production and failed to upregulate CR1 and CR3. There was no significant contamination of the iC5b67 with C5a to explain these results. Neither isolated C5b6 nor C7 alone exhibited the activities of iC5b67, while insolubilized anti-C7 could remove the PMN agonist activity from the iC5b67 preparation. Binding studies to define a specific receptor for iC5b67 on PMN were hampered by the very hydrophobic nature of the ligand. 125I-iC5b67, by contrast to hemolytically active 125I-C5b67, was unable to insert in erythrocytes, suggesting that iC5b67 need not insert in the PMN membrane to induce signaling. Two lines of evidence suggest that iC5b67 and C5a and FMLP share common steps in intracellular signaling (1) pretreatment of PMN with iC5b67 deactivates PMN for C5a- and FMLP-induced chemotaxis; and (2) pretreatment of PMN with pertussis toxin inhibits iC5b67-induced chemotaxis. Thus, iC5b67 has important effects on the activity of PMN and G-proteins and Ca2+ are involved in the signaling.     

Endothelial-secreted arachidonic acid metabolites modulate polymorphonuclear leukocyte chemotaxis and diapedesis in vitro

The influence of endothelial cells (ECs) on polymorphonuclear leukocyte (PMN) motility was examined using in vitro assays of PMN diapedesis and chemotaxis. ECs are seen to release arachidonic acid (20:4) metabolites that directly increase or decrease PMN movement, with their general effect being enhanced motility. This effect can be increased or decreased by treating ECs with stimulators or inhibitors of 20:4 metabolism, respectively. The metabolites include thromboxane B2, which increases PMN random motility, chemotaxis, and diapedesis in a dose- responsive manner and which acts as a chemoattractant; 6-keto-PGF1 alpha, which increases chemotaxis and diapedesis at high doses but decreases these responses at low doses; and a lipoxygenase-derived metabolite, suggested to be 5-hydroxyeicosatetraenoic acid, which increases chemotaxis and diapedesis. Thromboxane A2 and prostacyclin mimetics also affect chemotaxis in qualitatively similar manners as TxB2 and 6-keto-PGF1 alpha, respectively, but display greater potency. EC release of these metabolites is also seen to be substratum modulated, with an increased production by cells cultured on extracellular matrices. These results suggest that ECs are capable of modulating PMN motility and suggest a role for ECs in the control of PMN diapedesis.     

Alteration of the Cell Periphery During Granulocyte Maturation: Relationship to Cell Function

Physiologic characteristics of the surface of human granulocytes may beimportant determinants of functionalcapacity. Studies of immature granulocytes and polymorphonuclear neutrophils (PMN’s) from marrow andblood indicated that immature granulocytes are characterized by a high,neuraminidase-susceptible, negativesurface charge density; a high degreeof cellular rigidity, as measured by anelastimeter with a micropipette orificesize that approximated estimates ofaverage bone marrow basement membrane pore size (3.5µ); low adhesiveness to glass and plastic; low propensity to aggregate; a slow rate of cellspreading, pseudopod extension, andmotility; and a very low rate of phagocytosis. At the myelocyte stage, adhesiveness and phagocytosis, althoughnot prominent, became evident. Incontrast PMN’s had a lower negativesurface charge density, a higher degree of surface deformability (theentire PMN readily deforming to entera 3.5µ micropipette whereas the immature granulocyte would not so deform),higher adhesiveness (plastic > glass),a higher propensity to aggregate, ahigher rate of cell spreading, pseudopod formation and motility, and a higher rate of phagocytosis. The increaseddeformability and adhesiveness of thePMN as compared to the immaturegranulocyte may be a reflection of analtered relationship between relaxingand contracting systems at the cellperiphery during maturation and/orchanges in the sol-gel state of macromolecules at the cell periphery. Thisconcept is supported by the change insurface properties of the PMN towardthose of the immature granulocyte atcold temperature (4°C). The increaseddeformability and reduced surfacenegative charge of the PMN couldfacilitate adhesiveness and pseudopodformation and, thereby, increase cell-surface and cell-particle contact withenhancement of motility and phagocytosis. Hence the exit of PMN frombone marrow and circulation and thefunctional capacity of the PMN may bedetermined by a process of cytoplasmic maturation during granulopoiesisthat adapts the PMN for its essentialcell-extracell interactions.

Submitted on July 16, 1971 Revised on September 21, 1971 Accepted on September 22, 1971     

Direct Chemotaxis and Leucocyte-Induced Chemotaxis of Polymorphonuclear Leucocytes

The present study was designed to discriminate and analyze the presence of direct PMN chemotaxis and leucocyte-induced PMN chemotaxis in filter assay systems of PMN chemotaxis, namely the Wilkinson chamber and the Boyden chamber, which yield a more quantified information on leucocyte chemotaxis than filming of vital cells. The PMN chemotaxis was reduced by approximately 30–35 μm after incubation with vinblastine, 0.01, 0.10 and 1.00 μg/ml respectively, as measured by the leading front method in the Wilkinson chamber. This figure was thought to represent the contribution of the leucocyte-induced antitubulin-sensitive PMN chemotaxis to the casein-induced PMN chemotaxis under the experimental conditions prevailing. The remaining 60 μm antitubulin-insensitive PMN migration into the filter probably represented a combination of direct PMN chemotaxis and stimulated PMN random motility. Since the above-mentioned vinblastine inhibition of PMN chemotaxis was recorded in the absence of serum, complement factors included, the vinblastine-inhibited PMN chemotaxis was thought to be due to the release of a leucocyte-derived cytotaxin. The significance of incubation time and chemotactic parameters was further analyzed in the presence of serum in Boyden chambers, in the intercompartmental filters by means of PMN distribution curves and on the bottom filter by cell numbers. The antitubulin inhibition of PMN chemotaxis was evident in the intercompartmental filter during the initial period of incubation and later by cell counts on the bottom filter. These observations suggested that the antitubulin inhibition of PMN chemotaxis was due to antitubulin inhibition of the direction-finding in a minor proportion of fast-moving PMNs.     

Paralysis of phagocyte migration due to an artificial blood substitute

We investigated the effect of a candidate artificial blood substitute, Fluosol-DA (FDA), on human neutrophil function in a serum-free medium. In a 50% (vol/vol) mixture with polymorphonuclear cells (PMN), FDA had no effect on PMN viability, phagocytosis, superoxide anion generation, degranulation, or bactericidal activity. In striking contrast, the random migration and chemotaxis of PMN to both f-Met-Leu-Phe (fMLP) and activated serum were inhibited by 98% +/- 2%, 95% +/- 2%, and 88% +/- 6%, respectively. Inhibition of chemotaxis by FDA required no preincubation, was dose-dependent (50% inhibition [ID50] with a 14% vol/vol mixture with FDA), and was fully reversible by washing PMN free of FDA after one hour but not after 18 hours of incubation (32% +/- 11% inhibition of chemotaxis). FDA itself was not chemotactic and did not impair either the chemotactic activity or binding of fMLP to PMN. FDA also inhibited PMN adhesion (ID50, 9 +/- 1 vol/vol%). The inhibitory component of FDA was found to be its detergent additive, Pluronic F-68, which inhibited random migration, chemotaxis, and adhesion with ID50s of 1.4, 2.4, and 2.9 mg/mL, respectively (equivalent to FDA concentrations of 5, 9, and 11 vol/vol%, respectively). All the other components of FDA were noninhibitory. Plasma samples from humans injected with 8 mL/kg FDA and plasma samples from rabbits injected with 16 mL/kg FDA or an equivalent concentration of Pluronic F-68, when mixed with autologous PMN, also severely inhibited PMN chemotaxis. We conclude that exposure of PMN to clinically relevant concentrations of FDA inhibits PMN migration, presumably due to inhibition of adhesion. The inhibitory effect is entirely due to the detergent, Pluronic F-68. Artificial blood substitutes containing Pluronic F-68 may compromise the ability of PMN to prevent or effectively control microbial infections.     

Impaired polymorphonuclear leucocyte chemotaxis in rheumatoid arthritis.

This study has investigated the chemotactic activity of polymorphonuclear cells (PMNs) isolated from the blood of patients with either articular rheumatoid arthritis (RA) or RA with extra-articular manifestations. A double fluorochrome immunofluorescent staining test has been employed to identify cell-associated immunoglobulins, probably immune complexes. The results suggest an inverse relationship between PMN chemotaxis and staining for cell-associated immunoglobulins, either surface bound or internalised. PMNs from RA patients showed reduced chemotaxis, and this was further reduced when RA PMNs were incubated for 30 minutes in autologous serum. A similar reduction in chemotaxis of normal PMNs occurred after incubation in RA sera. Preincubation of both RA and normal PMNs in RA serum (but not normal serum) resulted in an increase in the number of cells in which cell-associated immunoglobulins were demonstrable. This further reduction in RA PMN chemotaxis after exposure to autologous serum, together with an increase in immunoglobulin staining, may indicate selection of certain PMNs at the time of venepuncture due to cell margination. Such a selection process would call for a re-evaluation of previous studies of RA PMN function in relation to the disease process.     

The effect of a therapeutic doxycycline concentration on polymorphonuclear leukocyte migration in vitro

Tetracyclines, and especially doxycycline, have been reported to inhibit polymorphonuclear leukocyte (PMN) migration. The effect of a therapeutic doxycycline concentration (4.33 micrograms/ml) on PMN migration in vitro was tested with the leading-front technique. No significant effect could be demonstrated on PMN chemokinesis and chemotaxis. It is thus concluded that the effect of therapeutic doxycycline concentrations on PMN migration is negligible. This conclusion has a bearing upon the choice of antimicrobial therapy.     

The role of the MA-sensitive leukocyte chemotaxis in rheumatoid arthritis. A randomized double-blind clinical trial of griseofulvin treatment

Polymorphonuclear leukocyte (PMN) chemotaxis is thought to play an essential role in the pathogenesis of rheumatoid arthritis. PMN chemotaxis is in part sensitive to microtubule antagonists (MAs), e.g. colchicine. The antimycotic antibiotic griseofulvin inhibits the MA-sensitive PMN chemotaxis in vitro in concentrations far below those obtained in serum during antimycotic therapy. The role of the MA-sensitive chemotaxis in rheumatoid arthritis could thus be elucidated by a clinical trial of griseofulvin treatment. Griseofulvin (n = 20) was tested in a randomized double-blind study versus placebo (m = 19) during one year in patients with rheumatoid arthritis of mild-moderate activity. No beneficial effect of griseofulvin treatment was noted on clinical symptoms or laboratory parameters of rheumatoid arthritis. Moreover, the placebo-treated patients showed more improvement than the griseofulvin-treated patients. It is therefore suggested that the MA-sensitive chemotaxis plays a reparative role in the inflammatory lesions of rheumatoid arthritis.     

Insulin differentially regulates monocyte and polymorphonuclear neutrophil functions in healthy young and elderly humans

CONTEXT: Insulin can regulate immune cell function. Aging is associated with various degrees of insulin resistance together with reduced immune cell activity. OBJECTIVE: We investigated the hypothesis that blood monocytes and polymorphonuclear neutrophils (PMNs) are less responsive to the action of insulin in elderly subjects. DESIGN-INTERVENTION: We evaluated the effect of hyperinsulinemia (0.7 mU/kg(-1) fat-free mass per minute(-1)) on monocyte and PMN activity using a 4-h euglycemic clamp technique. PARTICIPANTS: Eight young (24 +/- 6 yr old) and nine elderly (69 +/- 4 yr old) healthy volunteers participated in the study. MAIN OUTCOME MEASURES: Monocyte and PMN receptor expression and density were measured using flow cytometric detection. PMN chemotaxis toward formyl-Met-Leu-Phe (fMLP) was evaluated using a two-compartment chamber. PMN and monocyte phagocytosis was determined by measuring the engulfment of opsonized particles. Microbicidal functions were determined based on the production of reactive oxygen species (ROS) and bactericidal protein by stimulated cells. RESULTS: The density of PMN and monocyte insulin receptors was not affected by age or insulin clamp treatment regardless of the age. Insulin was able to regulate the expression of receptors involved in PMN action in the young-adult group only. PMN chemotaxis was up-regulated by insulin in both groups. In contrast, although insulin stimulated phagocytosis and bactericidal activity in young-adult subjects, the ability of PMN to adapt to physiological hyperinsulinemia was blunted in the older group. The effect of insulin on monocyte bactericidal properties seemed to be limited, although a suppressive action on fMLP-induced ROS production was detected in young adults. CONCLUSIONS: We confirmed the presence of the insulin receptor on monocyte and PMN membranes. We revealed that insulin has a limited action on monocyte function. Insulin has a priming effect on the main PMN functions. Immune cell function adapted poorly to insulin infusion in the elderly subjects.     

Regulatory mechanisms of a chemoattractant receptor on human polymorphonuclear leukocytes

The oligopeptide chemotactic factor receptor in leukocyte membranes exists in two affinity states that are in part interconvertible. Convertibility is regulated by guanine nucleotides, which suggests that a nucleotide regulatory unit allosterically modifies receptor affinity and participates in its transduction mechanisms. Approximately one-third of the high-affinity receptors in polymorphonuclear leukocyte (PMN) membranes are not subject to guanine nucleotide regulation. This fraction can be increased by agonist preincubation and could represent an intermediate form of the receptor prior to signal transduction and/or internalization. Pharmacologic manipulation of viable PMNs demonstrates that the affinity and functional activity of the chemoattractant receptor can be altered divergently by aliphatic alcohols and polyene antibiotics. The alcohols raise the receptors' affinity, enhance chemotaxis, but markedly depress chemoattractant-induced secretory mechanisms. In contrast, polyene antibiotics lower the receptors' affinity, depress chemotaxis, but enhance specific granule secretion. Thus, a relationship exists between the chemoattractant receptors' affinity and their ability to transduce signals for either chemotaxis or secretion.     

Phagocytosis-induced chemotaxis receptor cycling in neutrophils is mediated by thiol oxidation

We previously demonstrated that neutrophil (PMN) phagocytosis of opsonized zymosan (OPZ) caused oxygen-dependent inhibition of chemotactic peptide receptor (CPR) binding using the ligand 3H-formyl- methionyl-leucyl-phenylalanine (3H-FMLP). In the current studies we sought to determine the mechanism of CPR inhibition by OPZ. We found that 3 mM cysteine and 5 mM dithiothreitol (DTT) did not decrease PMN phagocytosis, but abolished CPR inhibition by OPZ. Furthermore, incubation of PMN with DTT after OPZ partially restored the suppressed CPR. PMN CPR saturation studies with 3H-FMLP on PMN incubated with or without DTT after phagocytosis indicated that DTT restored receptor numbers to 92% +/- 6% of prephagocytosis values and also improved CPR affinity for 3H-FMLP. In additional studies we found that the cell penetrating thiol reagents mercuric chloride (HgCl2), N-ethylmaleimide (NEM), and diamide, but not the nonpenetrating agents p- chloromercuriphenylsulfonic acid and p-chloromercuribenzoate, caused rapid, dose-dependent, DTT-inhibitable suppression of up to 85% CPR binding. CPR inhibition by HgCl2 and NEM was irreversible. PMN CPR saturation studies showed that NEM decreased total receptor numbers, not CPR affinity. The effect of NEM was not inhibited by receptor occupancy at 4 degree C or at 37 degree C. These studies indicate that: (1) intact thiol groups are a requirement for CPR ligand-binding; (2) the thiol groups are presumptively located on the interior side of, or within hydrophobic portions of, the plasma membrane and are not part of the 3H-FMLP binding site; (3) thiol oxidation-reduction induces reversible alteration in CPR binding; (4) phagocytosis-induced modulation of the CPR is due, in part, to thiol oxidation. These studies suggest that postphagocytic, thiol-mediated reduction in CPR binding may play a physiologic role in the inhibition of PMN chemotaxis at inflammatory sites. The state of CPR-associated thiols may be an important determinant of CPR binding under physiologic conditions other than phagocytosis.     

Modulation of the migration and chemotaxis of PMN cells by hyaluronic acid

Hyaluronic acid (HA) has a dose-related inhibiting effect on the migration and chemotaxis of polymorphonuclear leucocytes (PMN) in vitro. These effects were measured with a new indirect quantitative assay. On average 1 mg HA/ml causes an inhibition to about 80% of the control (spontaneous migration). This effect increased progressively with an increasing HA concentration, and with 4 mg HA/ml only about 19% of the PMN were able to migrate in the in vitro system. Similar results were obtained in the presence of a potent chemotactic factor (leukotriene B4 [LTB4]). In the mean 1 ng LTB4/ml alone stimulated the chemotaxis of PMN by a factor of 3 compared to the spontaneous migration. The highest HA concentration (4 mg/ml) reduced the number of migrating PMN cells to about 17%. From these experiments it may be concluded that the HA in the synovial fluid of the healthy joint has a protective effect against the invasion of PMN cells. This functions is disturbed in inflamed joints by the decrease in the HA concentration and possibly by its depolymerization. The intraarticular application of high molecular weight HA might be an important therapeutic regimen to restore the natural barrier against PMN migration, also in the presence of chemotactic factors and could therefore be helpful for interrupting the inflammatory cascade.     

Fibrinogen-derived peptide B beta 1-42 is a multidomained neutrophil chemoattractant

The formation and degradation of fibrin play a central role in hemostasis, but other activities have been associated with fibrin(ogen)- derived peptides, which suggests that products of fibrin(ogen) turnover may be involved in inflammation and wound healing. The present study was undertaken to determine whether the plasmic fibrinogen-derived peptide B beta 1-42 has effects on inflammatory cells and fibroblasts (FB). B beta 1-42 was found to be a potent chemotaxin for neutrophils (PMN) and FB, maximally stimulating PMN migration at 10(-9) mol/L peptide. Unlike the chemotactic factors f-Met-Leu-Phe and C5a, B beta 1- 42 did not induce the release of lysosomal hydrolases and superoxide anion from PMN, nor did it stimulate directed movement of monocytes (MN). These features of B beta 1-42 resemble the properties of human fibrinopeptide B (hFpB), the 14-reside, thrombin-cleaveable fragment that constitutes the amino terminus of B beta 1-42, and suggested that the chemotactic effects of B beta 1-42 are mediated through its hFpB domain. Against this conclusion, however, were observations that (a) desensitization of PMN with 10(-7) mol/L hFpB ablated chemotaxis to hFpB without affecting chemotaxis to B beta 1-42; (b) antiserum to hFpB, which recognizes the B beta 1-14 sequence both free and bound to larger fragments of the B beta chain, blocked hFpB chemotactic activity but did not affect B beta 1-42-mediated chemotaxis; (c) desensitization of PMN with equimolar amounts of hFpB and beta 15-42 (10(-7) mol/L), the isolated carboxyterminal sequence of B beta 1-42 remaining after the removal of hFpB, completely inhibited B beta 1-42-mediated chemotaxis; and (d) beta 15-42 itself was chemotactic for PMN. These data indicate that PMN recognize several independent domains within the amino terminal region of the human fibrinogen B beta chain and that these biologic effects extend to mesenchymal cells.     

Activation of human neutrophils by monoclonal antibody PMN7C3: cell movement and adhesion can be triggered independently from the respiratory burst

Anti-neutrophil monoclonal antibody PMN7C3 (IgG3) recognizes glycoproteins bearing the oligosaccharide lacto-N-fucopentaose III, including the C3bi receptor, LFA-1, and p150,95 on the plasma membrane and a group of granule-associated glycoproteins. We have previously shown that binding of this antibody to polymorphonuclear leukocytes (PMNs) stimulates a transient rise in cytosolic free calcium concentration but does not trigger the neutrophil respiratory burst. We now demonstrate that binding of PMN7C3 (and five other monoclonal antibodies recognizing the same antigen) to human neutrophils activates several other cellular responses. Addition of PMN7C3 to monolayers of neutrophils induces a rapid change in cell shape followed by pseudopod formation and increased migration. With incubation at 37 degrees C, the neutrophils aggregate in clusters (leukoagglutination). Quantitation of cell movement in a multiwell chemotaxis assembly or by migration of PMNs under agarose revealed that PMN7C3 is both chemotactic and chemokinetic. Pretreatment with the antibody inhibits subsequent chemotactic response to other stimuli. Monoclonal antibodies binding to other neutrophil antigens do not mimic these effects. These data suggest that cell movement and adhesion can be triggered independently from the respiratory burst. PMN7C3 may be a useful probe with which to study the events that link receptor-ligand binding to cellular response.     

Vaso-occlusive crisis-associated neutrophil dysfunction in patients with sickle-cell disease

Polymorphonuclear leukocyte (PMN) aggregation and chemotaxis were studied in 27 patients with sickle cell disease (SCD). Pain-free patients with SCD had a significantly impaired aggregation response to stimulation with n-formylmethionyl-leucyl-phenylalanine (FMLP) with or without cytochalasin B (CB), compared with normal volunteers (p less than 0.001). Patients with SCD in vaso-occlusive crisis had PMN aggregation induced by FMLP with or without CB that was significantly increased compared with the cohort of pain-free SCD patients (p less than 0.001). PMN from pain-free patients had normal chemotaxis, whereas patients in vaso-occlusive crisis had a significant impairment in PMN chemotaxis. PMN chemotaxis was inversely related to the PMN aggregation response to FMLP with CB (r = -0.75). Thus, the PMN from pain-free patients with SCD appears to have normal or decreased "stickiness" and to develop increased stickiness during vaso-occlusive crisis. The mechanisms responsible for these changes need further elucidation. Alterations in PMN function may be responsible, in part, for the increased risk of infection noted in individuals with SCD and may play a role in the development of the acute chest syndrome.