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Biomedical subjects

R Cramer

Publications and source records attributed to R Cramer.

At least 37 records · Page 2Linked to original sources
PubMed1984

A simple reliable assay for myeloperoxidase activity in mixed neutrophil-eosinophil cell suspensions: application to detection of myeloperoxidase deficiency.

Quantitation of myeloperoxidase (MPO) activity by guaiacol peroxidation (GP) assay is profoundly affected by the peroxidase present in eosinophils (EPO) that contaminate the granulocyte suspensions. Inclusion of 3-amino-1,2,4-triazole (AMT) in the GP assay permits quantitation of MPO activity in mixed neutrophil-eosinophil suspension because of the differential inhibition of EPO and MPO by AMT. Results show that: (1) the peroxidase activity of eosinophil-free granulocyte suspensions is not appreciably affected by AMT; (2) in the presence of AMT the peroxidase activities of granulocyte preparations containing different numbers of eosinophils are similar on a neutrophil basis, regardless of the number of eosinophils and correspond with the activity of eosinophil-free granulocyte suspensions; (3) AMT almost completely inhibits the activity of partially purified EPO, only slightly affecting the catalytic activity of partially purified MPO; (4) AMT completely inhibits the residual peroxidase activity of granulocyte suspensions from MPO-deficient subjects contributed by contaminating eosinophils. The GP assay in the presence of AMT was used to study the pattern of hereditary transmission of MPO deficiency. The genealogy derived on the basis of this assay was compatible with an autosomal recessive inheritance, in agreement with previously reported results, while no definite pattern of inheritance could be established by use of the GP assay without AMT. We suggest that the GP assay supplemented with AMT is the method of choice for detection of MPO deficiency, particularly partial deficiency.

Amitrole↗
PubMed1984

Effect of gentamicin and sisomicin on the generation of superoxide by human monocytes.

The effect of gentamicin and sisomicin on the generation of superoxide anions by human monocytes exposed to a phagocytosing stimulus, i.e. serum treated zymosan, or a soluble stimulus, i.e. 4-beta-phorbol-12-myristate-13-acetate, has been studied. Neither sisomicin nor gentamicin affected the superoxide production by stimulated monocytes. The data suggest that the reported variability in the response to aminoglycoside antibiotic therapy in certain clinical situations cannot be attributed to interference with monocyte oxidative burst.

Gentamicins↗
PubMed1982

Simian virus 40 large tumor antigen: a "RNA binding protein"?

Simian virus 40 large tumor antigen was isolated by immunoaffinity chromatography from monkey or mouse cell cultures undergoing lytic or transforming infection. RNase-treated gel-purified large tumor antigen, on hydrolysis with alkali, gave about equimolar amounts of AMP, GMP, CMP, and UMP. Furthermore, RNA fragments of approximately 45 nucleotides could be isolated from large tumor antigen purified by the same procedure. Mapping of the T1 oligonucleotides showed a high complexity, as indicated by the presence of unique sequences of 15-30 nucleotides and of poly(A). This is compatible with the hypothesis that these RNA fragments are derived from cellular pre-mRNAs or mRNAs. Our results suggest that Simian virus 40 large tumor antigen is a RNA-binding protein and might possibly be involved in regulation of synthesis, maturation, or translation of cellular mRNAs.

Antigens, Neoplasm↗
PubMed1982

Incidence of myeloperoxidase deficiency in an area of northern Italy: histochemical, biochemical and functional studies.

Forty-five subjects with a complete deficiency of myeloperoxidase were identified in an area of the region Friuli-Venezia Giulia in north-eastern Italy using the Hemalog D system as the screening technique. Histochemical and biochemical tests performed on the leucocytes of some of these subjects confirmed the defects shown by the Hemalog D system. The defect was of genetic origin in seven subjects. The genetic origin could be suspected in another eight subjects since more than two affected members were present in a given family. Eosinophil peroxidase, which is present in MPO deficient subjects, interfered with the guaiacol assay of MPO, and in several cases masked the genetic transmission. An assay was developed using o-dianisidine as the electron donor which considerably reduced the interference by EPO. With this assay an autosomal recessive pattern of inheritance was found. The MPO deficient leucocytes had a higher respiratory burst than control cells and an impaired bactericidal activity, at early post-phagocytic periods, which became comparable to that of control cells at later stages. Particle ingestion by the MPO-deficient cells was normal.

Blood Bactericidal Activity↗
PubMed1982

New approaches to the detection of myeloperoxidase deficiency.

Family studies on myeloperoxidase (MPO) deficiency have been carried out by quantitating the peroxidase activity of granulocyte preparations with three methods, namely guaiacol peroxidation, alanine decarboxylation, and spectroscopic analysis. The guaiacol assay failed to show a definite pattern of inheritance in two families with MPO-deficient subjects. Surprisingly, the granulocytes of three histochemically MPO-negative subjects had a peroxidase activity either half or even higher than that of control subjects. The peroxidase activity of these granulocyte preparations in these three subjects showed a positive correlation to the number of eosinophils. The possibility then considered was that eosinophils may have obscured the true pattern of inheritance in this assay. Two other methods of MPO assay, which are not influenced by the presence of eosinophil peroxidase (EPO), were therefore devised. One is based on the ability of MPO, but not EPO, to catalyze decarboxylation of L-alanine in the presence of Triton X-100, and the other relies on the different spectral properties of the two peroxidases. The results obtained with these two methods (1) were strictly comparable, (2) allowed detection of both totally and partially MPO-deficient subjects, (3) differed profoundly from those obtained with the guaiacol method when eosinophil-containing granulocyte preparations were used, and (4) revealed a pattern of autosomal recessive inheritance in the two families studied. The results of the three methods were comparable when eosinophil-free granulocyte preparations were assayed. It is concluded that failure to show a pattern of inheritance in some instances of primary MPO deficiency, or deviations from the autosomal recessive mode of transmission of this defect, may be attributed to interference by EPO. It is proposed that peroxidase assay methods not subject to EPO interference, such as the two described in this article, may be used, particularly in the detection of heterozygote subjects for MPO deficiency in the presence of high eosinophil counts.

Alanine↗
PubMed1981

Evidence that eosinophils catalyze the bromide-dependent decarboxylation of amino acids.

Human eosinophils from subjects with or without myeloperoxidase (MPO) deficiency and guinea pig eosinophils are able to decarboxylate L-alanine in the presence of the cationic detergent cetyltrimethylammonium bromide (CTAB) but not in the presence of the nonionic detergent Triton X-100. Instead, both normal human neutrophils and guinea pig neutrophils decarboxylate L-alanine in the presence of either detergent. When the non-bromide-containing cationic detergent cetyltrimethylammonium hydroxide (CTAOH) is used instead of CTAB, the eosinophils from MPO-deficient subjects are unable to decarboxylate L-alanine. Decarboxylation occurs with the combination CTAOH-Br-, but not with the combinations CTAOH-I-, CTAOH-CI-, or CTAOH-F-. Bromide in the absence of CTAOH does not promote decarboxylation. Triton X-100 and deoxycholate are much less effective in promoting decarboxylation in the presence of bromide. L-Lysine and L-aspartic acid are decarboxylated to a considerably lower rate than L-alanine in the presence of CTAOH and Br-. It is concluded that the eosinophils can catalyze the bromide-dependent decarboxylation of the apolar amino acid L-alanine in the presence of a cationic detergent.

Alanine↗
PubMed1980

[A thermosensitive inhibitor of the replication of polyomavirus, an extract from BHK2I cells transformed by the Tsa mutant of this virus].

BHK21 cells transformed by wild type or Ts3 mutant polyoma virus contain an inhibitor of polyoma virus replication when grown at permissive (36 degrees C) as well as non-permissive temperature (39 +/- 0.5 degrees C). Cells transformed by the Tsa mutant contain the inhibitor at the permissive but not at the non-permissive temperature. The inhibitor reappears in the latter cells however, upon shift from the non-permissive to the permissive temperature. If a reversible protein inhibitor (methionyl-adenylate, reversible inhibitor of the aminoacyl-t-RNA synthetase) is applied during the temperature shift experiments, the inhibitor does not reappear indicating that new protein synthesis is required for the recovery of its activity.

Animals↗
PubMed1978

Oxidative metabolism of chicken polymorphonuclear leucocytes during phagocytosis.

The oxidative response to phagocytosis by chicken polymorphonuclear leucocytes was investigated as compared to guinea pig polymorphonuclear leucocytes. The polymorphs from both species respond to phagocytosis with an increased oxygen consumption, an increased generation of O2 and H2O2, and an increased oxidation of glucose through the hexose monophosphate shunt. The rate of oxygen consumption, and generation of O2- and H2O2 by phagocytosing chicken polymorphonuclear leucocytes is considerably lower than with phagocytosing guinea pig polymorphonuclear leucocytes. By contrast, the extent of hexose monophosphate shunt stimulation in chicken polymorphs is comparable to that of guinea pig polymorphs. Evidence is presented suggesting that H2O2 is preferentially degraded in chicken cells through the glutathione cycle, whereas catalase and myeloperoxidase are the two main H2O2 degrading enzymes in guinea pig cells. The 20,000 g fraction of the postnuclear supernatant of chicken polymorphs contains a cyanide-insensitive NADPH oxidizing activity which is stimulated during phagocytosis. Similar properties for the NADPH oxidizing activity of guinea pig polymorphs have been previously reported. It is concluded that the metabolic burst of phagocytosing chicken polymorphonuclear leucocytes is qualitatively similar to that of guinea pig polymorphonuclear leucocytes, but the latter cells are more active in all the biochemical parameters that have been measured. The difference in the H2O2 degradation pathways between the two species is accounted for by the lack of myeloperoxidase and catalase in chicken polymorphs.

Animals↗
PubMed1977

Biochemical studies on the leukocytes in Chediak-Higashi syndrome.

Blood leukocytes from a patient with Chediak-Higashi syndrome (CHS) were compared with normal cells for their capacity of extruding (exocytosis) the lysosomal enzyme myeloperoxidase during phagocytosis or after a treatment with the ionophore A23187 and Ca2+. A decreased rate and extent of exocytosis in phagocytizing CHS cells was observed also with the Ca2+ ionophore. This suggests that a defect in Ca2+ mobilization is not responsible for the impaired secretion of granule content. Isolated granules of CHS cells and of leukocytes were treated with the detergent Triton X-100. Since the solubilization of myeloperoxidase from the CHS granules was much lower than from the normal ones, we suggest that the former organelles have a more resistant membrane.

Calcimycin↗
PubMed1975

Studies on the mechanism of metabolic stimulation in polymorphonuclear leucocytes during phagocytosis. II. Presence of the NADPH2 oxidizing activity in a myeloperoxidase-deficient subject.

1. The biochemical properties of leucocytes from a myeloperoxidase-deficient subject were compared with those of leucocytes from healthy subjects. 2. Myleoperoxidase-deficient leucocytes responded to phagocytosis of heat-killed bacteria with increased respiration, increased oxidation of glucose through the hexose monophosphate shunt and increased production of H2O2 as normal leucocytes do. 3. The ability of granules isolated from myeloperoxidase-deficient leucocytes to oxidize nicotinamide coenzymes was comparable to that of granules isolated from normal leucocytes. 4. The results argue against the hypothesis that oxidation of NADPH2 in leucocytes is performed by myeloperoxidase.

Adult↗