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Biomedical subjects

N A Westerdaal

Publications and source records attributed to N A Westerdaal.

12 recordsLinked to original sources

On the interaction of IgG subclasses with the low affinity Fc gamma RIIa (CD32) on human monocytes, neutrophils, and platelets. Analysis of a functional polymorphism to human IgG2.

An allotypic form of the low affinity IgG Fc receptor Fc gamma RIIa (CD32), termed low responder (LR) because of its weak reactivity with mouse (m) IgG1, interacts efficiently with human (h) IgG2. Fc gamma RIIaLR is the first known human FcR that binds this IgG subclass. In this study, we analyzed the role of Fc gamma RIIa in binding of stable hIgG-subclass dimers, and in induction of T cell mitogenesis using chimeric anti-CD3 mAb. We demonstrate that the functional polymorphism to hIgG2 is expressed on the majority of Fc gamma R-bearing peripheral blood cells: monocytes, neutrophils, and platelets. We were able to assess Fc gamma RII-mediated IgG-binding without interference of other Fc gamma R-classes, by blockade of Fc gamma RI on monocytes, and by using neutrophils of an individual deficient for the Fc gamma RIIIB gene. This study indicates as subclass specificity: hIgG3 >hIgG1,hIgG2 >> hIgG4 for Fc gamma RIIaLR and hIgG3,hIgG1 >> hIgG2 > hIgG4 for Fc gamma RIIaHR. Comparing the serum hIgG levels of individuals homozygous for the two fc gamma RIIa allotypic forms, we observed significantly lower hIgG2 serum levels in individuals expressing the hIgG2-binding LR allotypic form. This observation may implicate that Fc gamma RIIa regulates hIgG subclass production or turnover in man.

Antibodies, Monoclonal

A single amino acid in the second Ig-like domain of the human Fc gamma receptor II is critical for human IgG2 binding.

The low-affinity human Fc gamma RIIa is encoded by a single gene with allelic variation, defined by low-responder and high-responder alleles (LR and HR). The HR Fc gamma RIIa transcript interacts strongly with murine (m) IgG1 complexes, in contrast to the LR Fc gamma RIIa. Furthermore, the transcripts can be discriminated by mAb 41H16, which recognizes an epitope expressed on the HR Fc gamma RIIa molecule. We report that this receptor is also polymorphic in its reactivity with human (h) IgG2. Binding studies using well-defined hIgG dimers revealed that LR Fc gamma RIIa molecules can efficiently bind hIgG2, in contrast to HR Fc gamma RIIa. Previous work of others showed one amino acid difference between the allelic forms of Fc gamma RII. We, however, found a second amino acid difference between both allelic forms. In this study, hybrid Fc gamma RIIa molecules were constructed to determine the epitope for mAb 41H16 and the binding domain for mIgG1 and hIgG2 complexes. Our data point to the importance of the amino acid at position 131, located in the second Ig-like domain of Fc gamma RIIa. When an arginine residue is present at amino acid position 131, the receptor is recognized by mAb 41H16. Furthermore, the receptor can bind mIgG1-sensitized indicator E, but binds hIgG2 dimers only weakly. When a histidine residue is present at this amino acid position, hIgG2 dimers do bind efficiently to Fc gamma RII, whereas mIgG1-sensitized E and mAb 41H16 exhibit a strongly diminished binding.

Alleles

Measurement of the humoral immune response against Streptococcus pneumoniae type 14-derived antigens by an ELISA and ELISPOT assay based on biotin-avidin technology.

A Streptococcus pneumoniae type 14-specific ELISA and ELISPOT assay have been developed based on the use of biotinylated type 14 capsular polysaccharide (S14PS-biotin). A major advantage of this application over other methods is the use of 10-100-fold less antigen than that reported in the literature for other similar assays. Moreover, the prepared biotinylated polysaccharides are very stable and it is possible to use the same procedures for other pneumococcal polysaccharide antigens (e.g., S6BPS) with no major changes necessary in the ELISA and ELISPOT protocols. Furthermore, a simple thin layer chromatography method has been developed as a method for quality control of the biotinylated polysaccharide. Immunization with the thymus-independent antigen S14PS resulted in the induction of IgM spot-forming cells (SFC) and antibodies while S14PS-protein conjugates induced a thymus-dependent response. The immune response to the conjugates was enhanced by the addition of the adjuvant Quil A resulting in high levels of both IgG SFC and antibodies at day 14 after immunization. The developed assays are reliable and reproducible tools for studying the humoral immune response against Streptococcus pneumoniae type 14 capsular polysaccharide derived antigens.

Animals

Surface hydrophobicity and opsonic requirements of coagulase-negative staphylococci in suspension and adhering to a polymer substratum.

The opsonic requirements for phagocytosis in suspension of 38 clinical isolates of coagulase-negative staphylococci recovered from neonates with septicemia were found to be related to the degree of surface hydrophobicity of these strains. Sixteen isolates were adequately opsonized only in the presence of complement; this group was significant more hydrophobic (p less than 0.001) than the 22 strains not requiring complement for efficient uptake in suspension. Evidence showed that hydrophobic groups present on the bacterial surface may interfere with IgG opsonization. In contrast, IgG sufficed as an opsonin without complement being required for the efficient uptake of these hydrophobic strains when adhering to a polymer surface.

Bacterial Adhesion

Modulation of adherence of coagulase-negative staphylococci to Teflon catheters in vitro.

The mechanism of adherence of Staphylococcus epidermidis to commercially available catheters was studied in vitro in a quantitative assay employing 3H-labelled bacteria. It was found that adherence to Teflon catheters was significantly related to the degree of hydrophobicity of the strains. When hydrophobic groups were removed from Staphylococcus epidermidis by pepsin treatment, adhesion was almost completely abolished. Preincubation of catheters in human serum also caused a 80-90% reduction of adherence. Preincubation of Staphylococcus epidermidis in serum similarly decreased adhesion. This effect of serum was mainly due to albumin, while IgG and fibronectin were less effective. Culture of Staphylococcus epidermidis in subinhibitory concentrations (0.5 MIC) of cephalothin, clindamycin and vancomycin resulted in a 30-80% reduction in adhesion.

Anti-Bacterial Agents

Opsonic defense to Staphylococcus epidermidis in the premature neonate.

The determinants of opsonic defense to Staphylococcus epidermidis were studied in 47 premature newborns. Opsonic activity for S. epidermidis in serum from premature newborns proved to be proportional to gestational age (r = .664, P less than .001). The level of IgG antibodies to staphylococcal peptidoglycan in neonatal sera was similarly proportional to gestational age (r = .604, P less than .001). However, all opsonic activity of premature neonatal serum proved to be heat labile, i.e., dependent on activation of complement. Thus, no heat-stable, IgG-dependent opsonic activity to S. epidermidis was detected in any of the preterm sera, despite the presence of IgG antibodies to peptidoglycan. Further studies with purified IgG isolated from paired sera from term neonates and their mothers revealed that at similar concentrations the opsonic activity to S. epidermidis of neonatal, transplacentally derived IgG was only 26% of the activity of maternal IgG, a finding that may explain the absence of heat-stable opsonic activity in preterm newborns.

Antibodies, Bacterial

New, simple medium for selective recovery of Klebsiella pneumoniae and Klebsiella oxytoca from human feces.

A culture medium was developed which selectively favored the growth of Klebsiella pneumoniae and Klebsiella oxytoca in Escherichia coli-rich fecal cultures, without the use of antibiotics. The discriminative capacity of this medium was based on the presence of only two carbon sources, citrate and inositol, which can be utilized by nearly all K. pneumoniae and K. oxytoca strains but not by E. coli. The medium consisted of Simmons citrate agar (SCA) with 1% inositol (SCAI). Klebsiella strains from fecal samples subcultured on SCAI grew unhampered as yellow, dome-shaped, often mucoid colonies, whereas E. coli appeared as tiny, watery colonies. Apart from some Enterobacter strains, no other types of bacteria were found to mimic the typical appearance of klebsiellae. Recovery experiments from stool samples revealed a limiting ratio of Klebsiella to E. coli of 1:10(6) or more when samples were plated on SCAI versus ratios of 1:10(2) to 1:10(3) on blood agar or Macconkey agar. Compared with an existing Klebsiella culture method, the combination of SCA and MacConkey-inositol-carbenicillin (MIC) agar, Klebsiella yields with SCAI were not lower than those with the combination of MIC and SCA. Furthermore, the efficiency of the SCAI method was twice that of the latter combination. The SCAI plate could be a valuable tool in studies on the epidemiology of K. pneumoniae and K. oxytoca, for example in nosocomial infections, especially those concerning immunocompromised patients.

Citrates

Activity and synergy of ureido penicillins and aminoglycosides against Pseudomonas aeruginosa.

The in vitro activities of piperacillin, azlocillin, mezlocillin, sulbenicillin and ticarcillin were compared with those of carbenicillin using 88 clinical isolates of Pseudomonas aeruginosa. The minimum inhibitory concentrations (MIC) and the minimum bactericidal concentrations (MBC) were determined by standard techniques. The MIC for 90% of the strains was 7.5 mg/l for piperacillin, 10.0 mg/l for azlocillin, 26.5 mg/l for mezlocillin, 48.4 mg/l for sulbenicillin, 50.0 mg/l for ticarcillin and more than 100 mg/l for carbenicillin. The MBC/MIC ratio was 1.3 for piperacillin, 1.9 for ticarcillin, 2.1 for sulbenicillin, 3.3 for mezlocillin and 4.5 for azlocillin. The susceptibilities of the same strains to four aminoglycosides were tested. The MIC for 90% of the strains was 0.3 mg/l for sisomicin and tobramycin, 1.5 mg/l for amikacin, and 2.2 mg/l for gentamicin. The effect of combining piperacillin, azlocillin and mezlocillin with gentamicin, tobramycin, sisomicin and amikacin was studied using checkerboard titration. The highest degrees of synergy were found with the combinations of piperacillin and an aminoglycoside. Strong potentiation was observed in 85% of the strains with piperacillin - sisomicin and in 50% with piperacillin - gentamicin. The synergistic effects of azlocillin and mezlocillin in combination with an aminoglycoside (observed in 30-65% of the strains) were for the most part moderate or slight. No antagonism was observed.

Aminoglycosides

In vitro susceptibility of pseudomonas to four beta-lactamantibiotics (ampicillin, cephalothin, carbenicillin, piperacillin), to four aminoglycosides (kanamycin, amikacin, gentamicin, tobramycin) and to colimycin.

Of 97 well-defined strains of Pseudomonas, isolated from sputum of patients with cystic fibrosis (CF), the minimum inhibitory concentration (MIC) of several antibiotics was determined with a broth dilution method. The majority of the strains were resistant to ampicillin and cephalothin, moderately susceptible to carbenicillin (70% to 100 microgram/ml) and highly susceptible to piperacillin (100% to 25 microgram/ml, 88% to 6.25 microgram and 60% to 3.12 microgram/ml). If the pharmacological properties of piperacillin are comparable with those of carbenicillin, it can be expected that the sputum level of this drug will be adequate to treat Pseudomonas pulmonary infections. At the lowest concentration tested (0.78 microgram/ml) 3% of the strains were susceptible to kanamycin, 85,5% to amikacin, 95% to gentamicin, 98% of tobramycin, and 80% to colimycin. With regard to clinically attainable concentrations, 98.9% of the strains were susceptible to gentamicin and tobramycin 97.9% to amikacin, 96.9% to colimycin, 88.6% to piperacillin, 38% to carbenicillin, 25.7% to kanamycin, 12.3% to ampicillin, and 1% to cephalothin.

Amikacin

Piperacillin dosage.

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Bacterial Infections

Effects of various coryneforms on mononuclear phagocyte system function and on production of tumour necrosis factor in mice.

Vaccines of 6 strains of the genera Corynebacterium and Propionibacterium were tested for induction of hepatosplenomegaly and stimulation of clearance of injected colloidal carbon and viable Listeria monocytogenes. These effects were compared with the ability of these strains to support the endotoxin-induced liberation of tumour necrosis factor, interferon and heat-stable cytostatic factors in mouse circulation. Five strains caused good stimulation of macrophage functions and induced high levels of all factors. One strain was nearly lacking in macrophage stimulatory capacity and also failed to support factor production. These data are discussed in relation to the production mechanisms of the factors.

Animals