scholarly journals Monoclonal antibody 7D5 raised to cytochrome b558 of human neutrophils: immunocytochemical detection of the antigen in peripheral phagocytes of normal subjects, patients with chronic granulomatous disease, and their carrier mothers

Blood ◽  
1987 ◽  
Vol 69 (5) ◽  
pp. 1404-1408 ◽  
Author(s):  
M Nakamura ◽  
M Murakami ◽  
T Koga ◽  
Y Tanaka ◽  
S Minakami
Keyword(s):  
Chronic Granulomatous Disease ◽  
Human Peripheral Blood ◽  
Normal Subjects ◽  
Human Neutrophils ◽  
Granulomatous Disease ◽  
Cytochrome B558 ◽  
Normal Human ◽  
Male Patients ◽  
Normal Human Peripheral Blood ◽  
Cytochrome P 450

We have established a monoclonal hybridoma clone that produces IgG1 against the cytochrome b558 of human neutrophils. The antibody 7D5, secreted by the hybridoma, bound to solubilized cytochrome b of the neutrophils but not to other proteins such as hemoglobin, myeloperoxidase, and pig cytochrome P-450. Immunocytochemical studies of normal human peripheral blood showed that 7D5 bound to neutrophils and monocytes but not to lymphocytes or erythrocytes. The neutrophils of male patients but not of a female patient with chronic granulomatous disease lacked the antigen of 7D5 as well as the absorption spectrum for cytochrome b558. A mosaic of the antigen-positive and -negative neutrophils was observed in mothers of the male patients. These biochemical and immunocytochemical results indicate that 7D5 is a specific antibody against cytochrome b558 of human phagocytes.

scholarly journals Monoclonal antibody 7D5 raised to cytochrome b558 of human neutrophils: immunocytochemical detection of the antigen in peripheral phagocytes of normal subjects, patients with chronic granulomatous disease, and their carrier mothers

Blood ◽  
1987 ◽  
Vol 69 (5) ◽  
pp. 1404-1408 ◽  
Author(s):  
M Nakamura ◽  
M Murakami ◽  
T Koga ◽  
Y Tanaka ◽  
S Minakami
Keyword(s):  
Chronic Granulomatous Disease ◽  
Human Peripheral Blood ◽  
Normal Subjects ◽  
Human Neutrophils ◽  
Granulomatous Disease ◽  
Cytochrome B558 ◽  
Normal Human ◽  
Male Patients ◽  
Normal Human Peripheral Blood ◽  
Cytochrome P 450

Abstract We have established a monoclonal hybridoma clone that produces IgG1 against the cytochrome b558 of human neutrophils. The antibody 7D5, secreted by the hybridoma, bound to solubilized cytochrome b of the neutrophils but not to other proteins such as hemoglobin, myeloperoxidase, and pig cytochrome P-450. Immunocytochemical studies of normal human peripheral blood showed that 7D5 bound to neutrophils and monocytes but not to lymphocytes or erythrocytes. The neutrophils of male patients but not of a female patient with chronic granulomatous disease lacked the antigen of 7D5 as well as the absorption spectrum for cytochrome b558. A mosaic of the antigen-positive and -negative neutrophils was observed in mothers of the male patients. These biochemical and immunocytochemical results indicate that 7D5 is a specific antibody against cytochrome b558 of human phagocytes.

scholarly journals NADPH-binding component of the respiratory burst oxidase system: studies using neutrophil membranes from patients with chronic granulomatous disease lacking the beta-subunit of cytochrome b558.

1994 ◽  
Vol 179 (1) ◽  
pp. 291-297 ◽  
Author(s):  
S Tsunawaki ◽  
H Mizunari ◽  
H Namiki ◽  
T Kuratsuji
Keyword(s):  
Chronic Granulomatous Disease ◽  
Respiratory Burst ◽  
Beta Subunit ◽  
Human Neutrophils ◽  
Granulomatous Disease ◽  
Oxidase System ◽  
Cytochrome B558 ◽  
Respiratory Burst Oxidase ◽  
Stimulation Of ◽  
Binding Component

The NADPH-binding site of the respiratory burst oxidase system of neutrophils has been proposed to be either at a cytosolic component or at the beta-subunit of cytochrome b558. In this study, affinity labeling of resting and stimulated membranes, the latter having been assembled by all of the oxidase components from both membrane and cytosol, was carried out using [32P]NADPH dialdehyde (oNADPH). Stimulation of human neutrophils with PMA greatly increased O2(-)-generating activity and caused considerable translocation of the cytosolic components p47phox and p67phox. Nevertheless, PMA stimulation did not produce a labeled band which included positions at 47, 67, and approximately 32 kD. The most intense band reflected a molecular mass of 84 kD regardless of the state of activation, but a labeled band was never found near the beta-subunit (91 kD) of cytochrome b558. This 84-kD protein was further confirmed in neutrophils of 14 patients with gp91phox-deficient X-linked chronic granulomatous disease. These results indicate that the NADPH-binding component is not recruited from the cytosol, and also, that a membranous redox component besides cytochrome b558 must be involved in the NADPH oxidase system.

scholarly journals Altered expression of neutrophil peripheral benzodiazepine receptor in X-linked chronic granulomatous disease

Blood ◽  
1990 ◽  
Vol 76 (1) ◽  
pp. 184-188
Author(s):  
F Zavala ◽  
F Veber ◽  
B Descamps-Latscha
Keyword(s):  
Nadph Oxidase ◽  
Chronic Granulomatous Disease ◽  
Host Defense ◽  
Covalent Binding ◽  
Benzodiazepine Receptor ◽  
Mononuclear Phagocytes ◽  
Human Neutrophils ◽  
Granulomatous Disease ◽  
Peripheral Benzodiazepine Receptor ◽  
Cytochrome B558

This study was aimed at determining whether the peripheral benzodiazepine receptor (PBZDR), which is abundantly expressed on mononuclear phagocytes, is involved in host defense mechanisms depending on phagocyte membrane-associated NADPH-oxidase complex. Analysis by reversible and covalent binding of PBZDR expression on human neutrophils shows that it is modulated during NADPH-oxidase activation with phorbol 12-myristate 13-acetate. Based on a series of 17 patients with chronic granulomatous disease (CGD), results show that PBZDR expression is dramatically impaired in X-linked CGD, an inherited disorder due to a mutation on the gene coding for cytochrome b558 NADPH- oxidase component, whereas it is unaffected in autosomal recessive CGD where cytochrome b558 is normally expressed, suggesting a link between PBZDR and cytochrome b558 expressions. PBZDR can be assigned by covalent binding to an 18-Kd membrane protein. These results suggest that the neutrophil PBZDR, which can accommodate the widely prescribed anxiolytic drug Valium (diazepam), is involved in host defense against pathogens, a function that could be affected by neuroimmune interactions.

scholarly journals Separation and functional characterization of human neutrophil subpopulations

Blood ◽  
1978 ◽  
Vol 51 (4) ◽  
pp. 659-669 ◽  
Author(s):  
MS Klempner ◽  
JI Gallin
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Functional Characterization ◽  
Human Neutrophils ◽  
Experimental Conditions ◽  
Homogeneous Population ◽  
Paired Samples ◽  
Normal Human ◽  
Normal Human Peripheral Blood

Abstract Human neutrophils have been considered to be a functionally homogeneous population of cells. We have developed a density sedimentation technique for separation of neutrophils into two populations based on their ability to form rosettes with IgG-coated human erythrocytes (7SEA). Under the experimental conditions 80% +/- 4.3% of normal human peripheral blood neutrophilis form rosettes. Functionally rosette- forming neutrophils are more adherent to nylon wool, able to phagocytize more 14C-labeled Staphylococcus aureus, more efficient in killing S. aureus, and more responsive to endotoxin-activated human serum in a 51-cr chemotaxis assay that the non-rosette forming neutrophils. However, there is no difference among neutrophil subpopulations' ability to phagocytize latex particles. Paired samples of exudate neutrophils from cutaneous abscess fluid and peripheral neutrophils from three patients were investigated for their ability to form 7SEA rosettes. In each case exudate neutrophils contained greater than 96% rosette-forming neutrophils, whereas peripheral blood contained the normal 80% ( less than 0.01). Thus we show that peripheral blood contains at least two distinct populations of neutrophils. However, an essentially homogeneous neutrophil population is present in cutaneous exudate fluid.

scholarly journals Altered expression of neutrophil peripheral benzodiazepine receptor in X-linked chronic granulomatous disease

Blood ◽  
1990 ◽  
Vol 76 (1) ◽  
pp. 184-188 ◽  
Author(s):  
F Zavala ◽  
F Veber ◽  
B Descamps-Latscha
Keyword(s):  
Nadph Oxidase ◽  
Chronic Granulomatous Disease ◽  
Host Defense ◽  
Covalent Binding ◽  
Benzodiazepine Receptor ◽  
Mononuclear Phagocytes ◽  
Human Neutrophils ◽  
Granulomatous Disease ◽  
Peripheral Benzodiazepine Receptor ◽  
Cytochrome B558

Abstract This study was aimed at determining whether the peripheral benzodiazepine receptor (PBZDR), which is abundantly expressed on mononuclear phagocytes, is involved in host defense mechanisms depending on phagocyte membrane-associated NADPH-oxidase complex. Analysis by reversible and covalent binding of PBZDR expression on human neutrophils shows that it is modulated during NADPH-oxidase activation with phorbol 12-myristate 13-acetate. Based on a series of 17 patients with chronic granulomatous disease (CGD), results show that PBZDR expression is dramatically impaired in X-linked CGD, an inherited disorder due to a mutation on the gene coding for cytochrome b558 NADPH- oxidase component, whereas it is unaffected in autosomal recessive CGD where cytochrome b558 is normally expressed, suggesting a link between PBZDR and cytochrome b558 expressions. PBZDR can be assigned by covalent binding to an 18-Kd membrane protein. These results suggest that the neutrophil PBZDR, which can accommodate the widely prescribed anxiolytic drug Valium (diazepam), is involved in host defense against pathogens, a function that could be affected by neuroimmune interactions.

scholarly journals Separation and functional characterization of human neutrophil subpopulations

Blood ◽  
1978 ◽  
Vol 51 (4) ◽  
pp. 659-669
Author(s):  
MS Klempner ◽  
JI Gallin
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Functional Characterization ◽  
Human Neutrophils ◽  
Experimental Conditions ◽  
Homogeneous Population ◽  
Paired Samples ◽  
Normal Human ◽  
Normal Human Peripheral Blood

Human neutrophils have been considered to be a functionally homogeneous population of cells. We have developed a density sedimentation technique for separation of neutrophils into two populations based on their ability to form rosettes with IgG-coated human erythrocytes (7SEA). Under the experimental conditions 80% +/- 4.3% of normal human peripheral blood neutrophilis form rosettes. Functionally rosette- forming neutrophils are more adherent to nylon wool, able to phagocytize more 14C-labeled Staphylococcus aureus, more efficient in killing S. aureus, and more responsive to endotoxin-activated human serum in a 51-cr chemotaxis assay that the non-rosette forming neutrophils. However, there is no difference among neutrophil subpopulations' ability to phagocytize latex particles. Paired samples of exudate neutrophils from cutaneous abscess fluid and peripheral neutrophils from three patients were investigated for their ability to form 7SEA rosettes. In each case exudate neutrophils contained greater than 96% rosette-forming neutrophils, whereas peripheral blood contained the normal 80% ( less than 0.01). Thus we show that peripheral blood contains at least two distinct populations of neutrophils. However, an essentially homogeneous neutrophil population is present in cutaneous exudate fluid.

scholarly journals Detection of gp91-phox precursor protein in B-cell lines from patients with X-linked chronic granulomatous disease as an indicator for mutations impairing cytochrome b558 biosynthesis

1996 ◽  
Vol 315 (2) ◽  
pp. 571-575 ◽  
Author(s):  
Colin D. PORTER ◽  
KURIBAYASHI KURIBAYASHI ◽  
Mohamed H. PARKAR ◽  
Dirk ROOS ◽  
Christine KINNON
Keyword(s):  
Nadph Oxidase ◽  
B Cell ◽  
Chronic Granulomatous Disease ◽  
Cell Lines ◽  
Binding Domain ◽  
Granulomatous Disease ◽  
Cytochrome B558 ◽  
Stable Association ◽  
P22 Phox ◽  
Fad Binding

NADPH oxidase cytochrome b558 consists of two subunits, gp91-phox and p22-phox, defects of which result in chronic granulomatous disease (CGD). The nature of the interaction between these subunits has yet to be determined. Absence of p22-phox in autosomal CGD patient-derived B-cell lines results in detectable levels of an incompletely glycosylated gp91-phox precursor. We have detected this same precursor species in four cell lines from patients with the X-linked form of the disease due to mutations in gp91-phox. Such mutations should delineate regions of gp91-phox important for its biosynthesis, including stable association with p22-phox. One mutation mapped to the putative FAD-binding domain, one mapped to a potential haem-binding domain, and two involved the region encoded by exon 3.

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