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Biomedical subjects

D Schindler

Publications and source records attributed to D Schindler.

At least 73 records · Page 4Linked to original sources
PubMed1989

The effects of bacterial lipopolysaccharide, anti-receptor antibodies and recombinant interferon on mouse B cell cycle progression using 5-bromo-2'-deoxyuridine/Hoechst 33258 dye flow cytometry.

Polyclonal stimulation of resting B cells with anti-antigen receptor antibodies [anti-IgM mu chain antibody (anti-mu)] or with bacterial lipopolysaccharide (LPS) stimulates a proportion of B cells to proliferate. Exposure of resting B cells to both LPS and anti-mu activates a larger population of resting B cells than either alone, suggesting a synergistic effect of these two stimuli. Although recombinant interferon (rIFN) either alone or in combination with anti-mu has no apparent proliferative activity (as measured by [3H]thymidine incorporation), application of 5-bromo-2'-deoxyuridine/Hoechst 33258 dye flow cytometry reveals a distinct effect of rIFN on B cell growth. In the presence of anti-mu plus LPS, rIFN causes the majority of B cells to enter the cell cycle (CC), but a subset of B cells remains in the resting stage. Another subset of B cells has extremely rapid CC transit times, with a CC duration of less than 10 h. These studies show that both anti-mu and LPS are competence factors (which move cells from the G0 phase to the G1 phase). LPS acts also as a CC progression factor, while rIFN is a CC potentiating factor.

Animals↗
PubMed1989

Centromeric association of a microchromosome. A new category of non-random arrangement of metaphase chromosomes.

A supernumerary microchromosome measuring 0.5-1 microns found in over half of the metaphases of a CREST scleroderma patient and his daughter has been characterized by various cytogenetic techniques. The microchromosome consisted of constitutive heterochromatin and contained nuclear antigens reacting with specific anti-kinetochore antibodies. The most remarkable property of the microchromosome was its non-random position: it was closely associated with the centromere of any of the normal chromosomes in the majority of the metaphases. Furthermore, an inordinately high rate of Y chromosome aneuploidy was found in the CREST scleroderma patient. The origin and structure of the microchromosome, its possible connection with the CREST variant of scleroderma, and the phenomenon of centromeric association are discussed.

Aged↗
PubMed1989

Chromosome banding and DNA replication patterns in bird karyotypes.

The karyotypes of the domestic chicken (Gallus domesticus), Japanese quail (Coturnix coturnix), and griffon vulture (Gyps fulvus) were studied with a variety of banding techniques. The DNA replication patterns of bird chromosomes, analyzed by incorporation of 5-bromodeoxyuridine (BrdU) and deoxythymidine (dT), are presented here for the first time. In particular, the time sequence of replication of the ZZ/ZW sex chromosomes throughout the S-phase was meticulously analyzed. BrdU and dT incorporation are very useful methods to identify homoeologies between karyotypes, as well as rearrangements that occurred in the macroautosomes during speciation. The Z chromosomes of the three birds displayed the same replication patterns, indicating a high degree of evolutionary conservation. In the homogametic male, BrdU and dT incorporation revealed no evidence of asynchronous replication between euchromatic bands in the ZZ pair. The same was true of the three Z chromosomes in a triploid-diploid chimeric chicken embryo. Minor replication asynchronies between the homologous ZZ or ZZZ chromosomes were restricted to heterochromatic C-bands. These results confirm that, in the ZZ male/ZW female sex-determining system of birds, dosage compensation for Z-linked genes does not occur by inactivation of one of the two Z chromosomes in the homogametic male. The heterochromatic W chromosomes of the three species showed bright labeling with distamycin A/mithramycin counterstain-enhanced fluorescence and exhibited significantly delayed DNA replication. The nucleolus organizers of birds, frequently located in microchromosomes, were also distinguished by bright distamycin A/mithramycin fluorescence.

Animals↗
PubMed1989

Isolation and structural characterization of sialic-acid-containing glycopeptides of the O-glycosidic type from the urine of two patients with an hereditary deficiency in alpha-N-acetylgalactosaminidase activity.

Glycopeptides have been isolated from the urine of two patients, aged 5 and 6, with a new lysosomal storage disease characterized by a deficiency in alpha-N-acetylgalactosaminidase activity. Isolation of these glycopeptides was achieved using gel filtration and ion-exchange chromatography. Structural determination was done using one- and two-dimensional 500 MHz 1H-NMR spectroscopy and FAB mass spectrometry of native and derivatized glycopeptides. The following structures were inferred as being present: Glycopeptide A (up to 140 mg/l urine) (1)-(3) Neu5Ac alpha 2-3Gal beta 1-3 (Neu5Ac alpha 2-6)GalNAc alpha 1-R A1: R = Ser A2: R = Thr A3: R = Thr-Pro Glycopeptide B (up to 80 mg/l urine) (4)-(6) Neu5Ac alpha 2-3Gal beta 1-4GlcNAc beta 1-6 (Neu5Ac alpha 2-3-Gal beta 1-3) GalNAc alpha 1-R B1: R = Ser B2:R = Thr B3: R = Thr-Pro

Amino Acids↗
PubMed1989

Long-term survival of patients with chronic obstructive pulmonary disease following mechanical ventilation.

Fifty patients with chronic obstructive pulmonary disease (COPD) who survived their first artificial ventilation were followed for 15 years. Five-year survival was 30%, median survival 23.5 months, and average survival 44.9 months. Sudden death occurred in 47% of the patients (40% at home and 7% in the hospital). Using multivariate analysis, survival correlated positively and significantly with partial pressure of arterial oxygen (PaO2) on discharge (P less than 0.02), presence of wheezes or rhonchi (P less than 0.02), absence of right heart failure on admission (P less than 0.02), and male sex (P less than 0.05), and negatively with age (P less than 0.03). Sixty percent of the patients were artificially ventilated at least twice. Prognosis was better as the number of ventilations increased after the first ventilation. Our finding that discharge PaO2 was the factor that correlated best with survival suggests that treatment for these patients consist of continuous long-term use of domiciliary oxygen.

Cause of Death↗
PubMed1988

Location of sites in human lipocortin I that are phosphorylated by protein tyrosine kinases and protein kinases A and C.

Lipocortin I is a 39-kilodalton membrane-associated protein that in A431 cells is phosphorylated on tyrosine in response to epidermal growth factor (EGF). We have used recombinant human lipocortin I as a substrate for several protein kinases and identified phosphorylated residues by a combination of peptide mapping and sequence analysis. Lipocortin I was phosphorylated near the amino terminus at Tyr-21 by recombinant pp60c-src. The same tyrosine residue was phosphorylated by polyoma middle T/pp60c-src complex, by recombinant pp50v-abl, and with A431 cell membranes by the EGF receptor/kinase. The primary site of phosphorylation by protein kinase C was also near the amino terminus at Ser-27. The major site of phosphorylation by adenosine cyclic 3',5'-phosphate dependent protein kinase was on the carboxy-terminal half of the molecule at Thr-216. These sites are compared to the phosphorylation sites previously located in the structurally related protein lipocortin II.

Amino Acid Sequence↗
PubMed1988

Bromodeoxyuridine amplifies the inhibitory effect of oxygen on cell proliferation.

The BrdUrd-Hoechst method was used to analyze the interaction of various oxygen concentrations with BrdUrd substituted DNA with respect to cellular proliferation. At oxygen concentrations above 5%, human diploid fibroblast-like cells and amniotic fluid fibroblast-like cells showed reduced proliferation rates, which resulted from an increase in noncycling cells and from a permanent arrest of cells in the G2 phase of the cell cycle. At 35% oxygen the increase in noncyling cell fraction and the permanent arrest in G2 was strongly dependent upon the concentration of BrdUrd. Incorporation of BrdUrd into DNA, therefore, amplifies the adverse effects of increasing oxygen concentrations upon cell proliferation. The mechanism of this amplification might involve a free radical attack on DNA similar to the radiation sensitizing effect of BrdUrd.

Bromodeoxyuridine↗
PubMed1988

alpha-N-acetylgalactosaminidase deficiency, a new lysosomal storage disorder.

A new lysosomal storage disease with autosomal recessive inheritance is described in two male siblings of 5 1/2 and 4 years of age. Clinical manifestations started after 9 months of age with neurological symptoms, followed by progressive psychomotor deterioration. Urinary oligosaccharide excretion was abnormal and showed a characteristic pattern on chromatography. Enzyme assays showed a profound deficiency of lysosomal alpha-N-acetylgalactosaminidase in cultured fibroblasts, leukocytes and plasma from the patients and reduced activity in material from the parents. The deficiency was demonstrated both with an artificial substrate and a natural one, the blood group A trisaccharide. Excessive intra-lysosomal storage of alpha-N-acetylgalactosamine-containing material was demonstrated in cultured fibroblasts from the patients, using the lectin from Helix pomatia which is specific for terminal alpha-N-acetylgalactosamine residues.

Acetylgalactosamine↗
PubMed1988

BrdU-Hoechst flow cytometry: a unique tool for quantitative cell cycle analysis.

Unlike other techniques, flow cytometric analysis of BrdU-quenched 33258 Hoechst fluorescence may be used to measure cell activation and the G1, S, and G2/M compartment distributions in each of three successive cell cycles after growth stimulation of human peripheral blood lymphocytes. Cell cycle kinetic curves can be constructed from the BrdU-Hoechst flow data which allow the simultaneous assessment of growth fraction, lag-time, compartment exit rate, compartment duration, and compartment arrest. Applications of this new versatile technique include the evaluation of drug and growth factor effects, cell aging, and diagnosis in medicine and immunology.

Benzimidazoles↗
PubMed1988

The phytohemagglutinin response of human peripheral blood lymphocytes as a function of donor age: a re-examination using BrdU-Hoechst flow cytometry.

Mononuclear cells were isolated from peripheral blood by a standard Ficoll-Hypaque technique from 127 healthy donors, ranging in age from newborns to 86 years of age. As a measure of their in vitro growth response, the fraction of non-cycling cells was determined at 48 and 72 h after phytohemagglutinin (PHA) exposure by means of BrdU-Hoechst flow cytometry. This technique provides an optimal assay system for the non-cycling cell fraction, since all cycling cells will have incorporated BrdU thereby quenching the fluorescence of the Hoechst 33258 fluorochrome. Lymphocytes from prepubertal donors showed significantly decreased non-cycling cell fractions, as did lymphocytes from an additional group of 14 adults with hypogonadism due to the 45, XO condition (Turner-Syndrome). Much to our surprise, we found no definitive correlation between donor age and the non-cycling fraction of cells from the adult lymphocyte donors. Nor did we find any age-related increase in the variance of the non-cycling cell fraction. These observations suggest that the previously reported age-related decline in the PHA response of human PBL may reflect an increasing delay, rather than an overall diminution, of the PHA response as a function of donor age.

Adolescent↗
PubMed1988

Generation of a mutant form of protein synthesis initiation factor eIF-2 lacking the site of phosphorylation by eIF-2 kinases.

The phosphorylation of the alpha-subunit of initiation factor eIF-2 leads to an inhibition of protein synthesis in mammalian cells. We have performed site-directed mutagenesis on a cDNA encoding the alpha-subunit of human eIF-2 and have replaced the candidate sites of phosphorylation, Ser-48 and Ser-51, with alanines. The cDNAs were expressed in vitro by SP6 polymerase transcription and rabbit reticulocyte lysate translation, and the radiolabeled protein products were analyzed by high-resolution two-dimensional gel electrophoresis. The wild-type and Ser-48 mutant proteins became extensively phosphorylated by eIF-2 kinases present in the reticulocyte lysate, and when additional heme-controlled repressor or double-stranded RNA-activated kinase was present, phosphorylation of the proteins was enhanced. The Ser-51 mutant showed little covalent modification by the endogenous enzymes and showed no increase in the acidic variant with additional eIF-2 kinases, thereby suggesting that Ser-51 is the site of phosphorylation leading to repression of protein synthesis.

Amino Acid Sequence↗
PubMed1988

Fanconi anemia mutation causes cellular susceptibility to ambient oxygen.

The gene defect causing the Fanconi anemia (FA) phenotype appears to be expressed at the cellular level, since FA fibroblasts show a protracted course of explant outgrowth, a diminished in vitro life span, and very poor cloning. We show that exposure of FA fibroblasts to hypoxic (5% v/v oxygen) culture conditions restores their growth in vitro to near normal. Exposure to elevated oxygen tension (35% v/v) causes accumulations of FA cells in the S and G2/M phases of the cell cycle that are in significant excess of those seen in heterozygote and control strains. In the absence of evidence for defective cytoplasmatic radical scavenging systems, these observations suggest increased nuclear susceptibility to ambient oxygen as cause of the FA cellular phenotype.

Adolescent↗
PubMed1988

The "ruler sign"--a semiquantitative physical sign of chronic obstructive pulmonary disease.

This report describes a simple and reproducible physical sign of chronic obstructive pulmonary disease--the "ruler sign." With the patient standing erect, a ruler is placed on the trapezius muscle and the clavicle at the midclavicular line. The ruler forms an angle with the horizontal line. At the end of a normal expiration this angle was 36 degrees +/- 4 SD in healthy control subjects as compared with 15 degrees +/- 4 SD in patients with severe chronic obstructive pulmonary disease. A simple device is described that enables convenient measurement of this angle at the bedside.

Adult↗
PubMed1987

Confirmation of Fanconi's anemia and detection of a chromosomal aberration (1Q12-32 triplication) via BrdU/Hoechst flow cytometry.

Peripheral blood lymphocyte cultures of a patient with clinical evidence of Fanconi's anemia (FA) were investigated by means of BrdU/Hoechst flow cytometry. This technique can be used to differentiate cycling from noncycling cells after PHA stimulation; it also permits the quantitative assessment of cell cycle progression and cell-cycle arrest. A characteristic cell kinetic pattern of combined G2 phase prolongation and G2 phase arrest was observed in the present patient, confirming the clinical diagnosis of FA. An additional finding was the presence of a small subpopulation of peripheral blood cells, refractory to PHA stimulation, and displaying a DNA content 5% greater than that of the diploid cells. BrdU/Hoechst flow cytometry was crucial to the unequivocal demonstration of this subpopulation, which would not have been detected without enrichment by sequential depletion of PHA-responsive cells from the G0/G1 compartment during a 96-h stimulation experiment. The parallel finding of a presumptive triplication affecting the q12-32 segment of chromosome 1 in rare spontaneous divisions of a 40-h peripheral blood culture is consistent with the flow-cytometric detection of elevated DNA content if, as is common in FA, the abnormal cell lineage represents a preleukemic clone of monocytic origin.

Adolescent↗