Defective Neutrophil Research Articles

Effects of Influenza A Nucleoprotein on Polymorphonuclear Neutrophil Function

Infection with influenza virus is commonly associated with polymorphonuclear neutrophil (PMNL) dysfunction and consequent secondary bacterial pneumonia. A recently isolated human-derived protein that inhibits PMNL chemotaxis and oxidant production shows a striking homology to the influenza A nucleoprotein. In the present study, the effects of purified influenza A nucleoprotein on PMNL chemotaxis, oxidant production, degranulation, and calcium homeostasis were studied. Results of the study demonstrate that purified nucleoprotein inhibits PMNL chemotaxis as well as superoxide production. In addition, purified nucleoprotein induces a rise in PMNL cytosolic calcium concentration in a manner similar to that demonstrated for crude influenza A lysates. In contrast, no difference in FMLP-stimulated PMNL elastase or beta glucuronidase release was noted after exposure to nucleoprotein. These studies suggest that the influenza A nucleoprotein may account for some of the neutrophil defect associated with cellular infection by this virus.

FcγRII, FcγRIII, and CD18 receptors mediate in part neutrophil activation on a plasma coated expanded polytetrafluoroethylene surface*

A biomaterial-induced polymorphonuclear neutrophil (PMN) defect may predispose the implanted vascular graft to infection. PMNs bind, activate, and undergo morphologic changes when exposed to uncoated or plasma coated expanded polytetrafluoroethylene (ePTFE) surfaces. The purpose of this study was to investigate whether the CD18 integrin receptor or the immunoglobulin receptors Fc gamma RII and Fc gamma RIII mediate either PMN binding or activation on ePTFE. PMN binding and activation were determined after incubation of these cells on human immunoglobulin (IgG) or fibrinogen coated surfaces and uncoated or plasma coated ePTFE. PMN activation was measured by using the ferricytochrome reduction assay. Binding was determined with chromium 51-labeled PMNs. To block the Fc gamma RII, Fc gamma RIII, and CD18 receptors, PMNs were preincubated with the monoclonal antibodies (mAbs) IV.3, 3G8, and IB4, respectively. Irrelevant isotype matched mAbs were used as control. Monoclonal antibody IB4 inhibited binding of activated PMNs to fibrinogen coated surfaces. Binding to IgG was affected by either mAb IB4 or IV.3, but the greatest degree of inhibition was achieved when mAbs IB4 and IV.3 were used in combination. IgG-induced activation was partially inhibited by mAb IV.3 but was fully inhibited by a combination of mAbs IB4 and IV.3 The mAbs did not affect PMN binding to uncoated or plasma coated ePTFE, nor was PMN activation on the uncoated ePTFE surface inhibited. PMN activation on the plasma coated ePTFE surface was, however, partially inhibited by the combination of mAb IB4 with either mAb IV.3 or 3G8. A synergistic interaction between the PMN Fc gamma RII receptor and the CD18 integrin receptor accounts for surface bound IgG-induced cell activation. Both receptors also play a role in mediating PMN activation on the plasma-coated ePTFE surface.

Shwachman-Diamond syndrome associated with hypogammaglobulinemia and growth hormone deficiency

Shwachman-Diamond syndrome is a rare congenital disorder of unknown etiology. Characteristic abnormalities of this disease include pancreatic insufficiency, skeletal anomalies, growth retardation, recurrent infections, and hematologic abnormalities. Significant morbidity and mortality in these patients result from respiratory infections, which are not well explained on the basis of neutrophil defects. We have had the opportunity to perform an in-depth clinical immunologic and endocrinologic evaluation of a patient with this syndrome with recurrent respiratory tract infections. She was found to have profound humoral immunologic defects, and serum thymulin was absent. In addition, endocrinologic evaluation for growth retardation revealed growth hormone deficiency. The patient responded to treatment with supplemental growth hormone and intravenous gammaglobulin with accelerated growth and cessation of infections. This case is unique in that it links growth hormone deficiency and hypogammaglobulinemia in a non-X-linked manner and may provide the basis for treatment of other patients with this rare syndrome. (J A LLERGY C LIN I MMUNOL 1995;96:247-50.)

Role of protein adsorption on haemodialysis-induced complement activation and neutrophil defects

The present clinical study investigated the role of protein adsorption on complement activation and neutrophil functions during in vivo haemodialysis. The parameters were measured simultaneously at the arterial and venous sites of a cuprophan (CU) dialyser with or without pretreatment with human albumin, human immunoglobulins or human total plasma proteins (PLP). Leukocyte count, complement activation (C3a des arg), oxygen radical production and chemotaxis were measured at time zero and 15 min at the arterial and venous sites of the dialyser. Leukopenia observed at both sites was prevented only with PLP treatment. Complement activation was maximal at the venous site, but was not prevented by any of the treatments. Neutrophil oxygen radical production and chemotaxis were significantly decreased only at the venous site and restored to normal with any of the three treatments. Complement activation was maximal at the venous site, but was not prevented by any of the treatments. Protein adsorption on the dialyser membrane seems to modulate the bioincompatibility parameters in a different way. Depending on the functions tested, the protein fractions have different protecting effects, indicating the multifactorial mechanism implicated in the CU haemodialysis-induced leukopenia, complement activation and neutrophil defect.

Neutrophil phagocyte dysfunction in a weimaraner with recurrent infections

A five-and-a-half-month-old male weimaraner with severe recurrent bacterial infections was assessed for immunocompetence. Results revealed a low serum immunoglobulin G concentration and defective neutrophil phagocytosis.

TNFα and IL‐1β in serum of LJP patients with normal and defective neutrophil chemotaxis

TNFα and IL‐1β in serum of LJP patients with normal and defective neutrophil chemotaxis

NB4 cells show bilineage potential and an aberrant pattern of neutrophil secondary granule protein gene expression

NB4 is an acute promyelocytic leukemia cell line that has been shown to be inducible to terminal neutrophil maturation with all-trans retinoic acid (ATRA). HL60 cells are differentially inducible with 12-O- tetradecanoylphorbol-13-acetate (TPA) or dimethyl sulfoxide (DMSO) to monocytes or granulocytes, respectively. HL60 cells induced with DMSO undergo defective neutrophil maturation, manifested by a coordinate failure of secondary granule protein gene expression. We observed a similar defect in granulocytic maturation in ATRA-induced NB4 cells. In addition, because normal promyelocytes are known to have bilineage potential, we have investigated differentiation along monocytoid lines induced with TPA. We observed a striking phenotypic change along monocytoid/macrophage lines with TPA induction. Flow cytometry showed a TPA-induced increase in HLA-DR expression, and Northern blot analysis showed induction of expression of CD18, c-fos, and human neutrophil gelatinase (HNG). HNG is unique among the neutrophil secondary granule protein genes in that it is expressed in both the neutrophil and monocyte lineages. This again parallels our findings in TPA-induced HL60 cells, which retain the ability to express HNG. These findings confirm bilineage potential in NB4 cells. They also support the hypothesis of coordinate neutrophil secondary granule protein gene expression and a defect in this control as part of the leukemic phenotype.

NB4 Cells Show Bilineage Potential and an Aberrant Pattern of Neutrophil Secondary Granule Protein Gene Expression

NB4 is an acute promyelocytic leukemia cell line that has been shown to be inducible to terminal neutrophil maturation with all-trans retinoic acid (ATRA). HL60 cells are differentially inducible with 12-O-tetradecanoylphorbol-13-acetate (TPA) or dimethyl sulfoxide (DMSO) to monocytes or granulocytes, respectively. HL60 cells induced with DMSO undergo defective neutrophil maturation, manifested by a coordinate failure of secondary granule protein gene expression. We observed a similar defect in granulocytic maturation in ATRA-induced NB4 cells. In addition, because normal promyelocytes are known to have bilineage potential, we have investigated differentiation along monocytoid lines induced with TPA. We observed a striking phenotypic change along monocytoid/macrophage lines with TPA induction. Flow cytometry showed a TPA-induced increase in HLA-DR expression, and Northern blot analysis showed induction of expression of CD18, c-fos, and human neutrophil gelatinase (HNG). HNG is unique among the neutrophil secondary granule protein genes in that it is expressed in both the neutrophil and monocyte lineages. This again parallels our findings in TPA-induced HL60 cells, which retain the ability to express HNG. These findings confirm bilineage potential in NB4 cells. They also support the hypothesis of coordinate neutrophil secondary granule protein gene expression and a defect in this control as part of the leukemic phenotype.

Nontropical pyomyositis in adults

Pyomyositis (PMS) is a primary infection of striated muscle. Recent scanty reports suggest that non-tropical PMS may differ from classical tropical PMS. To address this question, 12 cases of nontropical PMS seen at two hospitals between 1976 and 1992 were reviewed and an English-literature search of similar cases was conducted. Both the series and reported cases are pooled together and herein reported. The age distribution of the 97 patients showed 30–50 and 60–70-year peaks, with a 3:1 (male-female) ratio. Fever, high erythrocyte sedimentation rate, and muscle stiffness or inflammation were present in more than 75% of patients. Muscles of the thigh (54%), back (13%), buttock (11%), arm (9%), or chest wall (4%) were involved. Staphylococci (61%), gram-negative bacilli (16%), streptococci (12%), and fungi (2%) were isolated from muscle specimens. Human immunodeficiency virus infection, diabetes mellitus, hemopoietic disorders, and other conditions with defective neutrophil function were present in 64 patients (66%). Drainage of pus and antibiotic therapy were the standard treatments. The mortality rate reached 10%. Analysis of patients classified by the comorbid condition showed differences in age, causative microorganisms, clinical features, and death rate. It is concluded that several clinical presentations of nontropical PMS are at variance with that of tropical PMS.

Neutrophil Defects as Risk Factors for Periodontal Diseases

There are several hypotheses proposed for the etiologic mechanisms causing periodontal diseases. These include a paradigm in which all individuals are equally susceptible to one or several pathogenic bacteria; a second paradigm that holds that all bacteria are equally virulent and that host susceptibility determines onset of disease; or a combination of the above. In this review, we analyze the role of neutrophil dysfunction as a risk factor for the onset of periodontitis. Both intrinsic or genetically inherited abnormalities of neutrophils and acquired neutrophil abnormalities are considered. While a large body of data implicates neutrophil dysfunction, either intrinsic or acquired (bacterially or extrinsically induced), as a significant risk factor for the periodontal diseases, clear, prospective, longitudinal epidemiologic studies to evaluate this association remain to be performed.

Neutrophil lysosomal dysfunctions in mutant C57 Bl/6J mice: interstrain variations in content of lysosomal elastase, cathepsin G and their inhibitors.

In this paper we report the serum antiprotease screening and the biochemical and functional characteristics of neutrophils in a variety of mouse strains with different susceptibilities for developing a protease-mediated injury. C57Bl/6J mice and their mutants tight-skin and pallid have a lower serum elastase inhibitory capacity (-30, -65 and -70% respectively) than other inbred strains (i.e. NMRI and Balb/c, which both have similar values). We demonstrate that these values are a consequence of a decreased concentration of the alpha 1-protease inhibitor for elastase [PI(E)], which is the major serum inhibitor of elastase and cathepsin G. In addition, neutrophil lysosomal dysfunctions characterized by abnormally high contents of elastase and cathepsin G, or defective lysosomal secretion are observed in tight-skin and pallid mice respectively. Another C57Bl/6J mutant with lysosomal abnormalities is the beige mouse. Negligible amounts of elastase and cathepsin G, as well as defective neutrophil degranulation, have been described previously in this strain. We found, however, discrete amounts of a latent form of neutrophil elastase that undergoes a spontaneous activation by a protease-dependent mechanism. We also report that neutrophil cathepsin G in this mouse is tightly bound to lysosomal membranes, but is released in near normal quantities during exocytosis. Cytosolic elastase and cathepsin G inhibitors, which were previously reported as being specific for the beige neutrophils, have also been detected in all the examined strains. Neutrophil functions, lysosomal enzyme content and serum antiprotease screening may represent key elements in the protease-antiprotease balance and may explain the different interstrain susceptibility to developing lesions in which an elastolytic activity has been implicated.

Role of cytokines in the modulation of neutrophil chemotaxis in localized juvenile periodontitis.

Decreased neutrophil chemotaxis has been implicated in the pathophysiology of the disease, localized juvenile periodontitis (LJP). The biological basis for the altered neutrophil function in LJP has been suggested to be an intrinsic cellular defect, involving a decrease in the number of N-formyl-methionyl-leucyl-phenylalanine (FMLP) receptors on the cell surface. We have investigated the relative contribution of serum-borne factors in the modulation of neutrophil functions in LJP, in a large population of LJP patients and healthy control subjects (HS). Treatment of HS-neutrophils with LJP-sera, resulted in a decreased neutrophil chemotactic response, and down regulation of FMLP receptors on the cell surface. Pretreatment of LJP-sera with anti-TNF and anti-IL-1 antibodies effectively, although incompletely, neutralized the ability of LJP-sera to modulate chemotaxis and FMLP receptor levels in HS-neutrophils. The changes induced by LJP sera were specific and sustained and could not be reversed by placing LJP-serum treated neutrophils in HS-serum. Sera obtained from HS and patients with adult periodontitis (AP), both of which exhibit normal chemotaxis, and patients with clinically diagnosed LJP with normal neutrophil chemotaxis (LJP-nctx) did not modulate HS neutrophil chemotaxis or FMLP receptors. Furthermore, recombinant human TNF-alpha, rhIL-1 alpha and rhIL-1 beta, at very low concentrations (15 pg/ml to 150 pg/ml), modulated the chemotactic response as well as FMLP receptor numbers on HS-neutrophils, in a manner similar to those observed in LJP. The present findings demonstrate that the biologic basis for the altered neutrophil function may not be an intrinsic cellular defect in neutrophils, but at least in part due to quantitatively small but biologically significant elevations in the levels of TNF-alpha and IL-1 in the serum.

Bactericidal activities of synthetic human leukocyte cathepsin G-derived antibiotic peptides and congeners against Actinobacillus actinomycetemcomitans and Capnocytophaga sputigena

Actinobacillus actinomycetemcomitans and Capnocytophaga spp. are gram-negative bacteria implicated in the etiology of periodontal disease (particularly in individuals with neutrophil defects) and life-threatening systemic infections. They are resistant to many antibiotics of microbial origin but are sensitive to the nonoxidative microbicidal action of neutrophils. These organisms are susceptible to the microbicidal effect of cathepsin G but are killed by two distinct mechanisms. The purpose of this study was to assess their sensitivity to the antibiotic effects of IIGGR and HPQYNQR, antimicrobial peptides derived from human neutrophil cathepsin G. The efficacies of the synthetic peptides IIGGR and HPQYNQR were tested by single-dose screening, dose-response, and kinetic assays against three representative strains (each) of A. actinomycetemcomitans and Capnocytophaga spp. and one strain of Eikenella corrodens. Strains of A. actinomycetemcomitans were sensitive to IIGGR and HPQYNQR at equal concentrations (wt/vol), whereas strains of Capnocytophaga and E. corrodens were more sensitive to IIGGR than to HPQYNQR. These differential antibiotic effects occurred over both time and dose ranges too narrow to be of therapeutic significance but are consistent with the premise that cathepsin G kills these oral bacteria by two distinct mechanisms. Except for IVGGR, congeners of IIGGR, including AIGGR, IAGGR, IIAGR, IIGAR, IIGGA, IQGGR, ILGGR, and I-norleucyl-GGR (InLGGR), were microbicidal at 500 micrograms/ml. IIGGR-amide exhibited no antibiotic activity. The D-enantiomer of IIGGR, DIDIGGDR, was as potent as IIGGR itself. APQYNQR exhibited antibiotic activity but somewhat less than HPQYNQR. We conclude that charge distribution, but not chirality or net charge, is an important determinant in the antibiotic efficacy of IIGGR. Moreover, peptide antibiotics derived from cathepsin G may have therapeutic value against periodontal gram-negative, facultative bacteria.

Diminished cytochromeb content and toxic oxygen metabolite production in circulating neutrophils from patients with Crohn's disease

Phagocytic, chemotactic, and oxidative metabolic capacity of circulating neutrophils was studied in 20 patients with Crohn's disease. In vitro tests of chemotaxis and phagocytosis of isolated neutrophils from patients did not differ from that of healthy controls. However, superoxide anion production by phorbol-myristate-acetate and formylmethionyl-leucyl-phenylalanine-stimulated neutrophils from patients with Crohn's disease was significantly diminished compared with controls. Measurement of cytochrome b559 in total membranes of neutrophils from patients showed that it was significantly lower than in controls. Disease activity did not correlate either with the production of superoxide anion or with the cytochrome b559 content. It is concluded that oxidative metabolism is impaired in neutrophils from patients with Crohn's disease and that this defect could be caused by a reduced content in membrane b-type cytochrome. Although this defective neutrophil function may contribute to granuloma formation, other factors have to be implicated in disease inflammatory activity.

Molecular Analysis of Myeloperoxidase Deficiency Shows Heterogeneous Patterns of the Complete Deficiency State Manifested at the Genomic, mRNA, and Protein Levels

Myeloperoxidase (MPO) is a glycosylated hemoprotein contained in the azurophil granules of human polymorphonuclear leukocytes (PMNs). MPO is thought to play a role in the oxidative antimicrobial activity of neutrophils by catalyzing the formation of hypochlorous acid, a potent microbicide, from hydrogen peroxide and chloride anions. Seven unrelated individuals with complete MPO deficiency, a relatively common heritable defect of neutrophils, were identified during routine blood tests. Molecular analyses were conducted to determine the level of the abnormality in these individuals. Western blot analysis showed that 6 of the 7 donors were devoid of immunoreactive MPO protein, while neutrophils from one individual contained only the 55-Kd subunit. Northern analysis of bone marrow RNA from one MPO-deficient donor showed the presence of the normal-sized 3.3-kb transcript indicating that the defect in MPO biosynthesis in this case was posttranscriptional. Southern analysis of four MPO-deficient donors showed a normal Bgl II digestion pattern, whereas an abnormal restriction pattern was observed in a fifth individual. Although the Bgl II pattern was similar to that observed in an unrelated subject described by Nauseef (Blood 73:290, 1989), our study strongly suggests that the point mutation does not reflect a polymorphism. Taken together, these analyses show the existence of diverse abnormalities associated with MPO-deficiency that may be detected at the level of the MPO polypeptide, mRNA, and gene.

Molecular analysis of myeloperoxidase deficiency shows heterogeneous patterns of the complete deficiency state manifested at the genomic, mRNA, and protein levels

Myeloperoxidase (MPO) is a glycosylated hemoprotein contained in the azurophil granules of human polymorphonuclear leukocytes (PMNs). MPO is thought to play a role in the oxidative antimicrobial activity of neutrophils by catalyzing the formation of hypochlorous acid, a potent microbicide, from hydrogen peroxide and chloride anions. Seven unrelated individuals with complete MPO deficiency, a relatively common heritable defect of neutrophils, were identified during routine blood tests. Molecular analyses were conducted to determine the level of the abnormality in these individuals. Western blot analysis showed that 6 of the 7 donors were devoid of immunoreactive MPO protein, while neutrophils from one individual contained only the 55-Kd subunit. Northern analysis of bone marrow RNA from one MPO-deficient donor showed the presence of the normal-sized 3.3-kb transcript indicating that the defect in MPO biosynthesis in this case was posttranscriptional. Southern analysis of four MPO-deficient donors showed a normal Bgl II digestion pattern, whereas an abnormal restriction pattern was observed in a fifth individual. Although the Bgl II pattern was similar to that observed in an unrelated subject described by Nauseef (Blood 73:290, 1989), our study strongly suggests that the point mutation does not reflect a polymorphism. Taken together, these analyses show the existence of diverse abnormalities associated with MPO-deficiency that may be detected at the level of the MPO polypeptide, mRNA, and gene.

Successful alpha‐2b‐interferon therapy for chronic neutrophilic leukemia

The authors report two patients with characteristic features of chronic neutrophilic leukemia (CNL) treated with alpha-2b-interferon (IFN) (Schering-Plough). At initiation of therapy, both patients had progressive disease and presented with large tumors. A rapid reduction of the tumor mass and a long-term stabilisation of the myeloproliferative disorder was obtained (therapy duration 16 and 26 months, respectively, and presently ongoing). In one patient, the dose of IFN could be significantly reduced during maintenance without relapse. Neither presented infectious or hemorrhagic complications under therapy. Alpha-2b-interferon is active and safe in CNL, even pretreated and progressive. It can also correct the neutrophil and natural killer functional defects frequently observed in CNL.

Defective Neutrophil Function in Workers Occupationally Exposed to Lead

Chemotaxis and nitroblue tetrazolium reduction were measured in peripheral blood neutrophils of workers occupationally exposed to lead. These two neutrophil functions were significantly reduced, as compared to controls, even in those workers with blood lead levels and urinary delta-aminolevulinic acid (ALA-U) concentrations below the currently accepted biological limit values of 60 micrograms/dl and 6 mg/l, respectively. The immunosuppressive effects of relatively low level lead absorption suggests that immune dysfunction may be a sensitive indicator of lead exposure.

Defective neutrophil and monocyte functions in glycogen storage disease type Ib: a literature review.

A summary review of leukocyte function in 42 published cases of glycogen storage disease Ib is presented. Polymorphonuclear and monocyte dysfunctions were evidenced in the majority of cases, whereas lymphocytes appeared to be unaffected. Phagocyte dysfunctions comprised in vivo mobilization and motility, in vitro random and directed migration, and one or several component functions of the "metabolic" ("respiratory") burst. On the basis of the available data it is impossible to know whether a primary functional deficit of the glucose 6-phosphate transport protein of the microsomal glucose 6-phosphatase system, as demonstrated in liver, also exists in these phagocytic cells and is responsible for this dysfunction.

Aspergillosis in the immunocompromised: focus on treatment.

Invasive aspergillosis, mainly due to Aspergillus fumigatus and Aspergillus flavus, is one of the most common fatal infections in the immunocompromised host; 25%–40% of autopsy cases in hematologic malignancies have invasive aspergillosis, with more than 80% of them being unrecognized during life (Young et al. 1970). In the normal host, inhaledAspergillus conidia are first phagocytized by alveolar macrophages. Those which escape phagocytosis sporulate and grow into large branching septated hyphae, which can invade lung tissues, but usually they are destroyed by the tissue neutrophils. Therefore, the patients most susceptible to aspergillosis are those with profound and prolonged granulocytopenia, especially those undergoing induction chemotherapy for acute leukemia, intensive chemotherapy for lymphoma, or bone marrow transplantation (Table 1). Patients with defective neutrophil function, particularly those with chronic granulomatous disease, are also at great risk of aspergillosis, as well as those with impaired macrophage function, such those receiving immunosuppressive therapy for organ transplantation or high-dose corticosteroids for vasculitides or connective tissue diseases. Recently, aspergillosis has been reported increasingly among acquired immunodeficiency syndrome (AIDS) patients; traditional factor risks including neutropenia, hematologic malignancies, and corticosteroid therapy were present in the majority of these patients (Pursell et al. 1992).