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Biomedical subjects

G Valet

Publications and source records attributed to G Valet.

At least 73 records · Page 4Linked to original sources
PubMed1982

Characterization of the activation of the human C1r complement molecule.

The proenzyme form of C1r was isolated by sequential chromatography from the euglobulin fraction of human serum on DEAE-Sepharose 6B-CL, CM-Sepharose 6B-CL and Sepharose S-300-CL. This C1r had the tendency to spontaneously activate within 60-90 min of incubation at 37 degrees C in presence of EDTA and more slowly in the presence of Ca2+. The spontaneous activation of C1r was found to be a bimolecular process and could be completely inhibited by DFP in the pH range 6-9 and in the presence of Ca2+ without affecting the hemolytic C1r activity. [14C]DFP bound to trace proteins in the 60-90 kD range, but not to C1r proenzyme. The spontaneous activation of C1r was diminished in the presence of EDTA by DFP, but could not be completely suppressed. EDTA acts by removing Ca2+ from C1r, thereby changing the conformation of the protein and causing an increased digestibility of the C1r H-chain. At temperatures above 0-4 degrees C this influence destroyed the ability of C1r proenzyme and enzyme to form macromolecular C1 and thereby abolished its hemolytic activity. We conclude from these results that the spontaneous C1r activation in the pH range 6-9 and in the presence of Ca2+ is due to contaminant proteases. C1r activated also spontaneously at higher pH values between pH 9 and 13.2, but the spontaneous activation ceased abruptly at pH 13.4. An intramolecular process of activation cannot be excluded at these high pH values. It is, however, not clear, whether this activation is a suitable model for the C1r activation in the C1 molecule, because the hemolytic activity of C1r was substantially diminished under the high pH conditions.

Complement Activating Enzymes↗
PubMed1981

Conformational changes of the subunits C1q, C1r and C1s of human complement component C1 demonstrated by 125I labeling.

C1s and C1r proenzymes and enzymes (C1s, C1r) and C1q were labeled with 125I. The distribution of the 125I label between H- and L-chain of C1s was only slightly dependent on the state of activation of C1s, and approx. 90% of the label was found in the H-chain. In the C1r proenzyme molecules 50% of the label was incorporated into the H-chain. The C1r H-chain label was reduced to 10% on activation of C1r to C1r, while the L-chain label increased to 90% of the total label. The presence of either C1s, C1q or C1qs during labeling reduced the C1r H-chain level, although C1r remained in the proenzyme form. The presence of C1s or C1rs enhanced the 125I uptake of C1q in Ca2+ or EDTA medium. This was unexpected because one would have anticipated a diminution of the C1q label due to the apposition of C1r and C1s, similarly as it occurs during C1rs complex and C1s dimer formation for the H-chain label of C1s. The results show that C1r and C1q alter their conformation during activation and C1 complex formation.

Calcium↗
PubMed1981

Sensitive three-parameter flow-cytometric detection of abnormal cells in human cervical cancers: a pilot study.

The cell volume, the DNA, and the carcino-embryonic (CEA) or epithelial-membrane (EMA) antigen of formaldehyde-fixed human cervical cells from 21 malignant cervix tumors and 11 normal patients were measured simultaneously with a Fluvo-Metricell flow cytometer. The simultaneous cell volume and DNA measurement provided the distinction of morphologically intact cells from cell debris, the determination of the cell cycle phase combined with the detection of aneuploid cells, and the distinct of inflammatory cells from parenchymal and tumor cells. Malignant samples were recognized because they contained more than 0.5% CEA positive cells which were of intermediate size. CEA and EMA expression in the malignant samples was not linked. The false positive rate in a total of 32 samples was 6.3% when the sum of CEA and EMA positive cells of each cell sample was calculated. No false negative malignant sample was observed.

Adult↗
PubMed1981

Flow-cytometric measurements of the transmembrane potential, the surface charge density and the phagocytic activity of Guinea pig macrophages after incubation with lymphokines.

The effects of lymphokines on guinea pig peritoneal macrophages were measured via flow cytometry utilizing the three-parameter FLUVO-METRICELL flow cytometer. On the basis of cell volume three distinct macrophage populations could be distinguished. Three to 5 min after starting the incubation with lymphokines a hyperpolarization of all three macrophage populations took place which was followed by depolarization. After 60 min the transmembrane potential reached again its control values. The negative charge density of the cell membrane decreased shortly after beginning of the incubation to 70-80% of the initial value and then remained unchanged for the following 120 min. The phagocytic activity of the macrophages was diminished during the depolarisation phase but increased over control values after restoration of the transmembrane potential.

Animals↗
PubMed1980

Flow cytometric determination of esterase and phosphatase activities and kinetics in hematopoietic cells with fluorogenic substrates.

Viable and formaldehyde fixed rat bone marrow or spleen cells and rat or human blood leukocytes were incubated with fluorescein (fluorescein-diacetate, -dibutyrate and -laurate) or umbelliferone (4-methyl umbelliferyl acetate and -phosphate) fluorogenic substrates for subsequent flow cytometric determination of cell fluorescence and cell volume. Formaldehyde-fixed cells preserved between 14 and 20% of the enzyme activity of the unfixed cells and the number of cell clusters for fixed and unfixed cells was the same. The esterase substrates revealed one cell cluster for spleen cells, two for bone marrow cells and four for peripheral blood leukocytes. Phosphatase activity was only associated with a cell cluster of large cells. The time course of substrate cleavage was linear during the first 10 min of incubation. Later on a plateau was reached. Specific enzyme activities were calculated on a single cell level from the simultaneous cell volume and cell fluorescence measurement. Two enzyme activities could be measured simultaneously by using a substrate mixture of umbelliferone acetate and fluorescein acetate.

Animals↗
PubMed1980

Cation transport in different volume populations of genetically low K+ lamb red cells.

During the first three months after birth lambs produce sequentially three erythrocyte populations of different mean volume as demonstrated by electric sizing methods (Valet, Franz, and Lauf, J. Cell. Physiol. 94 (1978) 215). We separated by centrifugal elutriation the small volume population (type II) red cells of a genotypically low K+ (LK) lamb from the population containing the larger volume type I and III cells, an admixture of fetal (I) and adult (III) erythrocytes. The cells were separated at various time intervals after birth and analyzed with respect to their volumes, cation contents, and cation flux properties by means of 86Rb uptake. The effect of anti-L on K+ pump and leak fluxes was ascertained in unseparated and separated red cells. It was found that the small red cells of population II, transiently present for several weeks, were fully developed LK cells with K+ pumps responding characteristically to the stimulatory action of anti-L. In constrast, the larger cells of population I and III were of high K+ (HK) nature at early time points, the K+ pump activities approximately ten times higher than adult LK cells. These cells constitute an admixture of type I fetal HK cells, and type III reticulocytes which are precursors for the final type III adult LK cells, since anti-L had a small stimulatory effect. At later times, however, only adult type III LK cells predominated. The data directly support our earlier finding that the HK-LK transition in genotypically LK lambs is primarily governed by cellular replacement.

Aging↗
PubMed1980

[Postnatal development of rat thrombocytes: the volume distribution and electrophoretic mobility or rat thrombocytes during the first month of life (author's transl)].

The volume distribution curves of rat thrombocytes show a significant rise of the modal volume between days 5 and 8 after birth. No clear evidence for two distinct thrombocyte populations was obtained by computer analysis of the volume distribution curves. However, the increase of the modal volume of the platelets correlates with the production of a new erythrocyte population. In addition, the platelet concentration in the blood increases significantly during this time. These changes could indicate the production of a second thrombocyte population. The mean electrophoretic mobility of rat thrombocytes decreases only slightly during the first 3 weeks after birth, while it increases for erythrocytes significantly during the same period.

Animals↗
PubMed1980

Epstein-Barr virus-induced increase in the concanavalin-A receptor density of established EBV-negative lymphoma lines in vitro.

Infection of two human EBV-genome negative lymphoma lines with the EBV-variants P3HR-I and B95-8 converts the lines to permanent carriers of EBV-DNA and the EBV-determined nuclear antigen. This process is accompanied by a series of biologic changes similar to those in oncogenic transformation by known tumor viruses (e.g. polyoma, SV40, Rous SV). They include altered growth properties and membrane changes, one of which is expressed in an increased agglutinability by Con-A. Although most transformed cell systems are more agglutinable than the non-transformed counterparts it is controversial whether they possess similar or changed numbers of lectin receptors. Since little was known about lectin receptor densities of virus transformed lymphoma cells, we set out to determine the densities of Con-A receptors on the EBV-negative lymphoma lines BJAB and Ramos and their EBV-converted sublines by simultaneous recording of the cell membrane fluorescence of FITC-Con-A and the cell volume using the FLUVO-METRICELL flow cytometer. We found that EBV infection gave rise to cell lines with a significantly elevated Con-A receptor density compared to the EBV-negative parental lines. The similar changes found in a number of independently converted sublines suggest that they are due to the direct or indirect action of the viral genome.

Burkitt Lymphoma↗
PubMed1979

Fast imaging in flow: a means of combining flow-cytometry and image analysis.

The morphological identification of cells by flow cytometry is difficult. Usually cell sorting and microscopical analysis have to be used in addition. Morphological analysis is simplified by taking cell pictures from a range of particular interest immediately during flow cytometric analysis. Instruments using the video scanning technique for fluorescence imaging are slow and expensive (8, 10). Morphological information can also be obtained by transmission imaging of cells in flow, which requires shorter exposure times. Therefore a cell volume activated flow imaging device has been developed which operates at flow speeds up to 5 m/sec and which depicts transmission images of selected cells on a 16-mm film by a nsec flashlamp illumination. An electronic unit detects the particles in the optically accessible orifice, performs the pulse height analysis, triggers the flashlamp if particles are in the preselcted range of interest and feeds the film. The instrument is capable of delivering up to 150 pictures per second and works either as a flow microscope in which the cells in the preselected volume range are directly observed, or as a picture system in which the cell pictures are stored on the 16-mm film for documentation or for image analysis.

Cell Count↗
PubMed1979

Simultaneous flow cytometric DNA and volume measurements of bone marrow cells as sensitive indicator of abnormal proliferation patterns in rat leukemias.

Simultaneous flow cytometric DNA and volume analysis of normal rat bone marrow cells shows three populations of nucleated cells with different mean volume. Each of these populations proliferates in a distinct cell cycle (alpha, beta, gamma). Normally the alpha-cell cycle has the highest amplitude, the beta-cell cycle is intermediate, and the gamma-cell cycle is low. The alpha-cell cycle was very significantly depressed and the beta + gamma-cell cycle was increased in three different rat leukemias (L5222, Shay, BNML), growing on three different rat strains (BDIX, Holtzmann, Brown Norway). The two parameter analysis further revealed that cells of the beta + gamma-cell cycle were slightly hyperdiploid and hypertetraploid in leukemic animals. The decrease of the alpha-cell cycle and the hyperploidies were more sensitive indicators for the abnormal proliferation pattern than the analysis of one parameter DNA distributions which remained within normal limits in all three leukemias.

Animals↗
PubMed1979

Flow cytometry as a new method for the measurement of electrophoretic mobility of erythrocytes using membrane charge staining by fluoresceinated polycations.

The binding of FITC-labeled poly-L-ornithine and poly-L-lysine to fresh or neuraminidase treated human, rat or rabbit erythrocytes was investigated by simultaneous cell volume and cell membrane fluorescence measurements in a flow cytometer. The cell volume was converted into cell surface and the distribution curve of the fluorescence/micrometer2 cell surface was calculated from all histogram classes by a computer program. The mean fluorescence/micrometer2 cell surface as a measure of the density of the negative charges on the cell surface was directly proportional to the elctrophoretic mobility of the erythrocytes, showing that polycation binding can effectively be used for the measurement of the electrophoretic mobility of erythrocytes. The computer fitting of the experimental two parameter histograms by two dimensional Gaussian normal distributions was found to be a very efficient way of data reduction, and a good separation of overlapping cell clusters was possible even in the case of low total numbers of cells in the histogram.

Animals↗
PubMed1978

Association of low C2 and C4 serum levels with the HLA-DW2 allele in healthy individuals.

HLA typed unrelated healthy individuals (HLA-DW2 positive n = 64, and HLA-DW2 negative n = 72) were investigated for their C2 functional activity and C4 serum protein levels. For the C2 and C4 levels a bimodal distribution was found in HLA-DW2 positive and HLA-DW2 negative individuals. HLA-DW2 positive persons had a significantly higher incidence of low C2 and C4 serum levels. Our data support the concept that genes governing C2 as well as C4 serum levels are in linkage disequilibrium with the HLA-DW2 allele of the major histocompatibility complex.

Complement C2↗