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Biomedical subjects

L Harvath

Publications and source records attributed to L Harvath.

12 recordsLinked to original sources
PubMed1991

Selected antibodies to leukocyte common antigen (CD45) inhibit human neutrophil chemotaxis.

The CD45 Ag family is a group of high m.w. glycoproteins that are expressed on the plasma membranes of all leukocytes. CD45 has protein tyrosine phosphatase activity and appears to regulate signal transduction and lymphocyte activation by specific association with receptor molecules on T and B lymphocytes. However, little is known about CD45 function in neutrophils (PMN). In this study, PMN were incubated with CD45 mAb and tested for their chemotactic responses to four unrelated chemo-attractants: FMLP, leukotriene B4 (LTB4), recombinant human C5a (C5a), and recombinant human neutrophil-activating protein-1, recently designated IL-8. A panel of CD45 mAb including an IgM mAb, AHN-12.1, and six IgG1 mAb, AHN-12, AHN-12.2, AHN-12.3, AHN-12.4, HLe-1, and KC56(T200), were tested for their effects on PMN chemotaxis. PMN chemotaxis was evaluated with two different membrane assays; one assay quantified the total number of migrating PMN and the other assayed the leading front of migrating PMN. AHN-12.1 and KC56(T200) significantly inhibited PMN chemotaxis to LTB4 and C5a. AHN-12.1 slightly inhibited PMN chemotaxis to FMLP, but KC56(T200) did not. In contrast, AHN-12 and HLe-1 did not significantly inhibit PMN chemotaxis to any of the chemoattractants. None of the CD45 mAb inhibited PMN chemotaxis to neutrophil-activating protein-1/IL-8. None of the CD45 mAb inhibited PMN superoxide production. These results suggest that PMN CD45 epitopes may interact with LTB4 and C5a receptor-associated molecules and regulate chemotactic responses.

Animals↗
PubMed1991

cAMP and human neutrophil chemotaxis. Elevation of cAMP differentially affects chemotactic responsiveness.

Neutrophils (PMN) treated with cAMP elevating agents were evaluated for their chemotactic responsiveness to FMLP and leukotriene B4 (LTB4). PGE1 and isoproterenol, increased PMN cyclic AMP production and inhibited chemotaxis to both FMLP and LTB4. In contrast, forskolin, which activates adenylate cyclase directly, inhibited chemotaxis to FMLP but not to LTB4. The phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX), was required for inhibition of PMN chemotaxis to FMLP by forskolin, PGE1, and isoproterenol. Isoproterenol and PGE1 inhibited PMN chemotaxis to LTB4 in the absence of IBMX and chemotaxis was further inhibited in the presence of IBMX. PMN cAMP levels were stimulated 2- to 3-fold with isoproterenol, 6- to 10-fold with PGE1, and 5- to 7-fold with forskolin over basal levels in the presence of IBMX. These observations demonstrate that total cellular cAMP concentration is not correlated with inhibition of PMN chemotaxis to all stimuli; forskolin, which increased cyclic AMP 5- to 7-fold over basal levels, did not inhibit chemotaxis to LTB4, whereas isoproterenol, which increased cyclic AMP only 2- to 3-fold over basal levels, inhibited chemotaxis to LTB4. PMN cAMP extrusion was determined under basal conditions and in the presence of PGE1, isoproterenol, or forskolin. PMN extruded cAMP under all conditions examined.

1-Methyl-3-isobutylxanthine↗
PubMed1991

Neutrophil chemotactic factors.

Polymorphonuclear leukocytes (neutrophils) are recruited to inflammatory sites by a variety of soluble mediators (chemoattractants) that stimulate neutrophil directed migration (chemotaxis). Many neutrophil chemoattractants such as neutrophil activating proteins, leukotriene B4 (LTB4), platelet activating factor, and complement-derived C5a, are generated endogenously by host cells or enzymatic cleavage of host proteins. Other chemoattractants such as N-formyl peptides are generated exogenously by bacteria that invade the host. Oxidative modification of methionine residues or changes in the amino acid sequence of peptide chemoattractants dramatically alter their chemoattractive properties. Many of the well-defined neutrophil chemotactic factors and studies of their structure-function relationships will be reviewed.

Animals↗
PubMed1991

Production of a novel antigen by conjugation of HIV-1 to Brucella abortus: studies of immunogenicity, isotype analysis, T-cell dependency, and syncytia inhibition.

In the present study inactivated human immunodeficiency virus type 1 (HIV-1) was conjugated to Brucella abortus and tested for immunogenicity in normal and anti-L3T4-treated BALB/c mice. HIV-BA was more immunogenic than uncoupled HIV in normal mice, since 6-fold less virus in HIV-BA preparations elicited higher titer responses than HIV-1 alone. Furthermore, the HIV-BA antibody response reached higher levels before the HIV-1 response. Immunoblot analysis showed that most of the HIV-1 antigens were recognized by antibodies induced by either HIV-1 or HIV-BA. Isotype analysis revealed that HIV-1 induced similar levels of IgG1 and IgG2a antibodies, whereas the IgG2a responses to HIV-BA were more pronounced than the IgG1 response. These different IgG subclass patterns suggest that conjugation of HIV-1 to BA changed the immunogenic nature of HIV-1. The requirement for helper T cells was examined by immunizing mice that were depleted of CD4+ T cells by in vivo anti-L3T4 treatment. Under these conditions the IgG responses to HIV-1 were completely eliminated. Although HIV-BA antibody responses were markedly reduced in anti-L3T4-treated mice, anti-HIV-1 antibodies, mainly of the IgG2a isotype, were produced. The antibodies generated by HIV-1 and HIV-BA immunization were also tested for their ability to inhibit syncytia formed by infecting CD4 + CEM cells with gp160 vaccinia. Sera from normal mice, immunized with either HIV-1 or HIV-BA were capable of inhibiting syncytia. In contrast, following anti-L3T4 treatment, only mice immunized with HIV-BA, but not HIV-1, produced antibodies capable of inhibiting syncytia.

Animals↗
PubMed1979

Defective initiation of oxidative metabolism in polymorphonuclear leukocytes.

The polymorphonuclear leukocytes of a two-year-old boy who had multiple episodes of bacterial infections demonstrated defective oxidative metabolism with phagocytic, but not with soluble (non-phagocytic), metabolic stimuli. We used a chemiluminescence assay to examine the patient's polymorphonuclear leukocyte responses to numerous particulate and soluble stimuli. The patient's polymorphonuclear leukocytes had substantially depressed chemiluminescent responses during phagocytosis of opsonized particles (latex, pneumococci, pseudomonas, streptococci and zymosan); however, we observed normal chemiluminescent responses when these leukocytes were stimulated with soluble agents (sodium fluoride, concanavalin A, cytochalasin E, calcium ionophore A23187 or phorbol myristate acetate). Polymorphonuclear leukocyte oxygen consumption and superoxide production were impaired during phagocytosis, even though phagocytosis was normal. In addition to the metabolic defect, this patient's polymorphonuclear leukocytes had depressed chemotactic and bactericidal activities. This study provides evidence that polymorphonuclear leukocytes have more than one mechanism for initiating oxidative metabolism.

Blood Bactericidal Activity↗
PubMed1979

Light generation with Fenton's reagent. Its relationship to granulocyte chemiluminescence.

A simple chemical system consisting of FeSO4 and H2O2 (Fenton's reagent) was shown to emit light (chemiluminescence). The addition of tryptophan to the reaction markedly enhanced light production. Very little chemiluminescence was observed when H2O2 was omitted from the reaction and when ferric, instead of ferrous, ions were used. Hydroxyl radical (OH.) and singlet oxygen (1 deltagO2) quenchers suppressed chemiluminescence of the FeSO4 + tryptophan + H2O2 system; and, deuterium oxide (2H2O) enhanced chemiluminescence of both FeSO4 reactions. These observations suggest that a radical chain reaction involving both OH. and 1 deltag O2 is responsible for the chemiluminescent reactions. Six iron-containing proteins, some of which are located within granulocytes, all emitted light in the presence of H2O2. Since iron and H2O2 are present in metabolically stimulated granulocytes, it is likely that chemiluminescent reactions similar to the ones demonstrated in this study account for part of the chemiluminescence of activated granulocytes.

Chemical Phenomena↗
PubMed1979

The collection, preservation and function of peripheral blood hematopoietic cells in dogs.

Semicontinuous flow centrifugation (SFC) was employed in canines to obtain adequate numbers of cells for autologous marrow repopulation following supralethal cyclophosphamide administration (100 mg per kg). Four cycles using a 225 ml bowl and 30 ml per minute flow rate were carried out for procurement. Collections averaged 8.4 +/- 0.4 x 10(9) (n = 30) leukocytes. Mononuclear cells (MNC) comprised 75 +/- 2% of the population and granulocyte colony-forming units (CFU-C) totaled 1.9 +/- 0.09 x 10(5) colonies per collection. Eighty-six percent of mononuclear cells were "T" cells in the 30 to 90 second fraction compared to 49% at 150 to 120 seconds. CFU-C fractionation revealed a peak at 30-210 seconds and a second peak at 120-210 seconds. Following programed freezing and rapid thawing 77.7 +/- 11.4% of CFU-C were recovered. Using a dose of 1 x10(9) MNC per kg for reinfusion, marrow repopulation and clinical recovery occurred in four out of four dogs. It was concluded that 1) SFC was effective for obtaining adequate numbers of peripheral blood stem cells for autologous marrow repopulation. 2) A rapid thawing and direct transfusion technique appears satisfactory for administration. 3) Some separation of "T", "B" and CFU-C peripheral blood components is possible by centrifugation.

Animals↗
PubMed1978

Chemiluminescence of human and canine polymorphonuclear leukocytes in the absence of phagocytosis.

Polymorphonuclear leukocytes (PMNs) have increased oxidative metabolism during phagocytosis and emit light (chemiluminescence, CL) as a result of metabolic activation. The present study examined PMN CL in the absence of phagocytosis using sodium fluoride (NaF), a nonparticulate agent and known stimulator of cellular oxidative metabolism. Normal human and canine PMNs were assayed in a CL spectrometer which permitted continuous sample mixing and constant temperature regulation during CL measurement. PMNs treated with 20 mM NaF demonstrated maximum CL responses of 10,000-20,000 cpm above background, 13-17 min after addition of NaF at 37 degrees C. Temperature regulation of reaction mixtures was found to be a critical factor in assaying PMN CL responses to NaF, because a small decrease in temperature (i.e. 1.5 degrees C) substantially depressed and delayed the CL response. Superoxide anion production correlated closely with CL responses in NaF-treated human PMNs. CL responses were completely suppressed in the presence of the oxidative metabolic inhibitors, iodoacetamide, and N-ethylmalemide; and were partially suppressed in the presence of either superoxide dismutase or sodium azide.CL responses of NaF-treated PMNs were significantly lower than responses generated by PMNs phagocytizing opsonized yeast. When NaF was evaluated for its effect on light generation from a singlet oxygen dependent CL reaction, it was found that NaF did not quench singlet oxygen light. This study demonstrates that PMN CL can occur in the absence of phagocytosis, and it proposes that a nonphagocytic PMN CL assay may be useful in evaluating leukocyte metabolic defects.

Animals↗
PubMed1976

Evaluation of type-specific and non-type-specific pseudomonas vaccine for treatment of pseudomonas sepsis during granulocytopenia.

The protective role of serotype-specific and non-type-specific active immunity against Pseudomonas aeruginosa infection was assessed in granulocytopenic dogs. Dogs were preimmunized with either specific serotype 6 vaccine (SI) or nonspecific serotype 3 vaccine (NSI) and challenged intravenously with 10(7) viable serotype 6 P. aeruginosa during granulocytopenia. Control dogs (C) having insignificant anti-pseudomonas antibody levels were also tested. Results showed: (i) significant increase in survival of SI dogs (P less than 0.05) compared to C and NSI dogs, with no significant difference between C and NSI animals; (ii) lower febrile responses in SI dogs; and (iii) markedly reduced bacteremia in SI dogs compared to C and NSI animals. SI dog sera from survivor animals did not kill the infecting pseudomonas strain in vitro. The study demonstrated that type-specific immunity to P. aeruginosa induced by active immunization is effective in protection against pseudomonas during granulocytopenia and that non-type-specific immunity offers no cross-reactive protection. The findings suggest that the reticuloendothelial system in conjunction with specific immunity constitute an important defense against pseudomonas infections.

Agranulocytosis↗
PubMed1976

Passive immunity against pseudomonas sepsis during granulocytopenia.

Specific passive immunity against Pseudomonas aeruginosa sepsis was assessed in granulocytopenic dogs. Dogs were infused with either normal or antipseudomonas immune plasma 24 h before pseudomonas challenge. They were challenged intravenously with 10(7) serotype 6 P. aeruginosa during granulocytopenia. Treatment was evaluated by observation of survival periods, febrile responses, type 6 pseudomonas antibody titers, and quantitative cultures of blood and tissues. The results demonstrated that passively immunized dogs did not survive infection. Both normal-plasma and immune-plasma recipients had bacteremia at death, with median values of 980 and 470 pseudomonas per ml of blood, respectively. All dogs had marked febrile responses 24 h after pseudomonas challenge and had high concentrations of pseudomonas in their lung tissue at death, with median values of 10(8) pseudomonas per g of wet tissue weight. After plasma infusion, immune-plasma recipients had high concentrations of anti-pseudomonas antibody, with total antibody titers ranging from 256 to 1,024 and a median value of 1,024. These titers were comparable to titers attained in a previous study from our laboratory using active immunization with pseudomonas lipopolysaccharide vaccine, where the median total anti-pseudomonas antibody titer was 2,048. Actively immunized animals, however, were significantly protected against pseudomonas sepsis and had prolonged survival periods and prevention of bacteremia. The present study demonstrates that circulating type-specific antibody is not solely responsible for the protection afforded to granulocytopenic dogs actively immunized against pseudomonas.

Agranulocytosis↗
PubMed1976

Combined pre-immunization and granulocyte transfusion therapy for treatment of pseudomonas septicemia in neutropenic dogs.

An experimental model was designed to evaluate a combined protocol of active immunization and granulocyte transfusions for treatment of Pseudomonas aeruginosa sepsis in the neutropenic host. One member of a pair of dogs was immunized with P. aeruginosa vaccine. Both dogs were then rendered transiently neutropenic with a single intravenous dose of cyclophosphamide (40 mg. per kilogram) and challenged with an intravenous inoculum of P. aeruginosa. Twenty-four and 48 hours after pseudomonas challenge each animal received granulocyte transfusions. Effectiveness of therapy was evaluated by observation of survival time, febrile response, and quantitative blood cultures. Results showed a significant increase in the survival period (P is less than 0.05), a lower febrile response (P is less than 0.025), negative blood cultures, and a greater recovery rate in the immune group. Immune dogs that died had negative blood cultures or less than or equal to 10 pseudomonas per milliliter of blood despite the presence of P. aeruginosa in tissues. In contrast, control dogs had septic deaths within 67 hours of pseudomonas challenge, marked febrile responses with 24 hours of infection, and positive blood cultures with 4,000 to 25,800 pseudomonas per milliliter of blood. These data show that combined therapy with immunization and granulocyte transfusions is effective in reducing the severity of P. aeruginosa infection and in preventing bacteremia during periods of leukopenia.

Agranulocytosis↗