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Biomedical subjects

K Wrogemann

Publications and source records attributed to K Wrogemann.

At least 55 records · Page 3Linked to original sources
PubMed1983

Mitochondrial calcium content and oxidative phosphorylation in heart and skeletal muscle of dystrophic mice.

Mitochondrial calcium overloading was investigated in the genetically dystrophic mouse (strains 129/ReJ dy/dy) as a possible contributing factor to the development of muscle fiber necrosis. Mitochondrial calcium concentrations were significantly elevated in both skeletal muscle and heart organelles. Because mitochondria were isolated in the presence of ruthenium red this finding was not the result of an artefact of isolation. State 3 respiration rates and concomitantly the respiratory control ratios were slightly decreased in skeletal muscle, but not in heart mitochondria. This abnormality could result from calcium overloading in a small fraction of the mitochondria. Fractionation of skeletal muscle mitochondria on sucrose gradients gave two distinct populations of dystrophic organelles, one with high calcium, whereas normal skeletal muscle mitochondria and heart organelles showed only one broad band on the gradient. The results support the idea that both skeletal muscle and heart are affected in dystrophic mice, strain 129/ReJ dy/dy and also that in the dystrophic mouse the process of cell necrosis is associated with cellular calcium overloading.

Animals↗
PubMed1982

Analysis of fibroblast proteins from patients with Duchenne muscular dystrophy by two-dimensional gel electrophoresis.

Duchenne muscular dystrophy (DMD), the most common and severe form of the muscular dystrophies, is an X-linked inborn error of metabolism with multiple tissue involvement. Although the major pathological changes are observed in skeletal muscle, abnormalities have also been detected in the heart, nervous system, red blood cells, lymphocytes and cultured skin fibroblasts. For many reasons, such as readily available tissue material, fewer secondary changes and the potential for prenatal diagnosis, cultured skin fibroblasts should be the tissue of choice to search for the primary defect. Several abnormalities have been reported in DMD fibroblasts, suggesting that the genetic abnormality is expressed in these cells. To search for potentially mutant protein(s) we have compared the protein composition of normal and DMD fibroblasts by two-dimensional gel electrophoresis and have now found one protein spot consistently missing in DMD cells. The nature of this protein and its relation to the DMD gene are unknown.

Autoradiography↗
PubMed1981

Concanavalin A-induced chemiluminescence in rat thymus lymphocytes. Its origin and role in mitogenesis.

1. The luminol-dependent chemiluminescence of rat thymocytes responding to concanavalin A can be resolved into glucose-dependent and glucose-independent portions. 2. The glucose-dependent portion, supported by D-glucose and D-mannose oxidation, is inhibited by catalase (200 microgram/ml), amobarbital (1 mM) and hexose analogues that block D-glucose uptake. Thus concanavalin A may activate, transiently, an NAD(P)H oxidase that utilizes reducing equivalents derived from the oxidation of exogenous glucose to give dismutation products of O2- (including H2O2) as its major products. 3. The glucose-independent portion is inhibited by eicosa-5,8,11,14-tetraynoic acid but not by indomethacin. It may therefore be associated with the conversion of hydroperoxy intermediates of arachidonic acid metabolism to hydroxy products by the lipoxygenase pathway. 4. Preincubation of thymocytes for 18 h in serum-free medium enhances the subsequent chemiluminescent response to concanavalin A severalfold and evokes the response at a lower threshold concentration. The incorporation of [3H]thymidine by preincubated cells is similarly enhanced at low doses of concanavalin A, whereas the response to optimal doses is unaltered. 5. Catalase does not inhibit the enhanced incorporation of [3H]thymidine obtained in response to concanavalin A, but instead amplifies the response to low doses in the same manner as preincubation.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1980

Chemiluminescence and immune cell activation. II. Enhancement of concanavalin A-induced chemiluminescence following in vitro preincubation of rat thymocytes; dependency on macrophage-lymphocyte interaction.

The immediate chemiluminescence (CL) response to concanavalin A (Con A) of rat thymocytes is enhanced 10 to 20-fold when the cells are preincubated in serum-free medium for 5--20 h. During this period, multiple encounters between lymphocytes and macrophages occur which morphologically appear as rosettes or grape-like cell aggregates. Additionof bone marrow-derived macrophages increases the number of cell aggregates and also the CTL response to Con A. Paradoxically, removal of macrophage-containing cell aggregates after cocultivation leaves a pure thymocyte population which strongly responds to Con A with CL. Our results confirm that macrophage-depleted "competent" lymphocytes are capable of CL and, furthermore, that "competence" is gained during cocultivation with macrophages. We are therefore convinced that measurements of macrophage and lymphocyte CL are a powerful tool for further elucidation of lymphocyte differentiation and of interactions between macrophages and lymphocytes.

Animals↗
PubMed1978

Chemiluminescence and immune cell activation. I. Early activation of rat thymocytes can be monitored by chemiluminescence measurements.

Immediately after the addition of concanavalin A, rat thymocytes respond, in the presence of luminol, with a burst of chemiluminescence (CL) that can be conveniently monitored in an ordinary liquid scintillation spectrometer. Peak CL is reached after 50 sec. Addition of catalase suppresses 65% of the CL, suggesting that H2O2 generation may be its major source. CL with different kinetic characteristics can also be generated by the calcium ionophore A23187. Our rat thymocytes contain approximately 0.1% endogenous macrophages. Bone marrow-derived rat macrophages also respond to concanavalin A or A23187 stimulation with a burst of CL. However, the kinetic properties of CL as well as the inhibition of CL by catalase in these cells differ markedly from those of thymocyte preparations, suggesting that the major portion of the CL in rat thymocytes actually originates in T lymphocytes. Because CL measurements allow the monitoring of very early events in cell activation and because of the simplicity of the technique, CL measurements may become a useful method for the study of lymphocyte activation, of macrophage-lymphocyte interactions, as well as for the rapid screening for specificity and reactivity of T lymphocyte populations.

Animals↗
PubMed1978

Biochemical investigations in cultured skin fibroblasts from patients with Duchenne muscular dystrophy.

Cultured skin fibroblasts from patients with Duchenne muscular dystrophy were studied with a double-labeling procedure. Preliminary evaluation of the growth characteristics and average viability of dystrophic cells did not show any major abnormalities. Analysis of doubly labeled fibroblast mixture did not disclose any consistent anomaly in the distribution of subcellular fractions or in the major protein components of dystrophic fibroblasts.

Cell Count↗
PubMed1978

Searching for molecular abnormalities in genetic diseases by the use of a double labeling technique. I. Rationale, techniques, and initial evaluation.

To meet the challenge of unravelling the molecular pathology of the ever expanding number of known genetic diseases in man, new efficient investigative techniques have to be designed. A procedure is presented for detection of protein defects in genetic diseases on the basis of structural rather than functional alterations. The technique is based on double labeling of normal and diseased fibroblast proteins followed by extensive fractionation and analysis. The rationale, advantages, and limitations of the procedure are discussed and the technical aspects of its use explained.

Animals↗
PubMed1978

Searching for molecular abnormalities in genetic diseases by the use of a double labeling technique. II. Deficiency of a basic protein in fibroblasts of patients with Pompe's disease.

We used a double labeling technique to search for molecular defects in two fibroblast strains obtained from patients with Pompe's disease. Analysis of the double labeled subcellular fractions by sodium dodecyl sulfate (SDS) electrophoresis did not reveal any abnormalities except in the "mitochondrial-lysosomal" fraction. In this fraction ratio deviations indicated that in Pompe's disease there was a significant decrease in counts of a protein with molecular weight of about 29,000. After solubilization by freeze-thawing this protein was shown to have an isoelectric point of 7.9 in contrast to the alpha-glucosidase which focused at about pH 4.7. Two-stage gel studies demonstrated an estimated 90% reduction of this protein in Pompe's disease. Two-stage studies of acid alpha-glucosidase did not show any abnormal ratios of leucine incorporation. Similar although quantitatively less pronounced results were obtained in the study of skin fibroblasts from a patient with adult glycogen storage disease type II.

Adult↗
PubMed1977

Altered amounts of hemoglobin synthesis in livers of dystrophic hamsters.

In liver supernatants similar amounts of leucine are incorporated into proteins of normal and dystrophic hamsters. However, using a dual labeling technique, a protein fraction is detected which shows different levels of leucine incorporation between normal and dystrophic animals in an age-dependent fashion. The major protein in this fraction comigrates with hemoglobin or its subunits under various conditions of polyacrylamide gel electrophoresis. In animals younger than 3 days there is more synthesis of this protein fraction in dystrophic animals, while at 6 days there is considerably less synthesis of this protein. These changes are paralleled by differences in the number of erythroid foci in histological sections of the livers. There is no evidence for structurally altered hemoglobin in dystrophic hamsters. The significance of this finding and its possible relation to the dystrophy process are not known at present.

Aging↗
PubMed1976

Mitochondrial calcium overload: A general mechanism for cell-necrosis in muscle diseases.

It is suggested that the mechanism of muscle-cell necrosis in various muscle diseases is explained by an increased net influx of calcium into cells which triggers a "vicious cycle" of mitochondrial calcium overloading and energy depletion. If correct, this hypothesis may offer the basis of a more rational treatment of some muscle diseases even before their primary aetiology is known.

Calcium↗
PubMed1975

On the role of mitochondria in the hereditary cardiomyopathy of the Syrian hamster.

Conventional polarographic techniques reveal no defect of oxidative phosphorylation in cardiomyopathic hamster heart mitochondria. However, in skeletal muscle of the same animals a defect is seen consistently with NAD+-linked substrates. This mitochondrial defect is caused by calcium accumulation and a consequent loss of NAD+ and magnesium. As shown by density gradient centrifugation, the defect resides in only a small fraction of all mitochondria. It is assumed that the same genetic defect in BIO 14.6 hamsters causes both skeletal muscle and heart lesions. Calcium levels in heart mitochondria are also significantly elevated in heart mitochondria. It is postulated that a calcium-associated defect similar to that in skeletal muscle mitochondria exists also in a very small fraction of the heart organelles and is responsible for the occurrence of focal necrosis.

Animals↗