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Biomedical subjects

U Klein

Publications and source records attributed to U Klein.

At least 145 records · Page 8Linked to original sources
PubMed1983

Cellular Fractionation of Chlamydomonas reinhardii with Emphasis on the Isolation of the Chloroplast.

A method for cellular fractionation of Chlamydomonas reinhardii, SAG 11-32/b, and isolation of intact chloroplasts from synchronized cells of the alga is described. The procedure for cell fractionation comprises essentially four steps: (1) protoplast production with autolysine; (2) lysis of the protoplasts with digitonin; (3) aggregation of broken protoplasts; and (4) separation of organelles by differential centrifugations.Replacing the differential centrifugations (step 4) by Percoll cushion centrifugations yields intact chloroplasts. Starting with 100 milliliters of an algal culture containing 3000 micrograms chlorophyll, intact chloroplasts with 100 to 200 micrograms of chlorophyll can be isolated. Envelope integrity is about 90% (ferricyanide assay). Examination of the chloroplasts by electron microscopy and marker enzyme activities indicated some mitochondrial and cytoplasmic contamination.

Journal Article↗
PubMed1983

Photosynthetic Properties of Chloroplasts from Chlamydomonas reinhardii.

Chloroplasts isolated from synchronous cultures of the unicellular green alga Chlamydomonas reinhardii, SAG 11-32/b (-), fix CO(2) at rates between 25 and 50 micromoles per milligram chlorophyll per hour. The upper value is approximately half of the rate of the intact cell.During storage in the dark on ice, the chloroplast preparation loses 30 to 50% of its CO(2) fixing capability per hour. Under reducing conditions (+ 1 millimolar dithiothreitol), this loss of activity is about twice as fast. The same reducing conditions stimulate CO(2) fixation in the light.High concentrations of inorganic phosphate (>2 millimolar) inhibit CO(2) fixation. This inhibition is overcome by the addition of glycerate 3-phosphate. It is concluded that chloroplasts from C. reinhardii possess a higher plant type phosphate translocator. With respect to dependency upon light intensity, pH and Mg(2+) concentration, the results were similar to that reported for chloroplasts from higher plants. However, in contrast to higher plant chloroplasts, maximum CO(2) fixation is observed at the relatively low osmotic concentration of 0.12 molar mannitol in the reaction buffer.

Journal Article↗
PubMed1982

[Analysis of methods for optimizing lysozyme determination].

In order to optimize the detection of lysozyme in the clinic, studies on photometric and agar diffusion methods were performed with different preparations of micrococcus luteus (lysodeikticus). Living or living lyophilized preparations of micrococcus were suitable. The influence of temperature, test period and lysozyme concentration on the result was demonstrated. The lysoplate technique has several advantages in clinical routine tests, however an exact adjustment of the reagents and the test is necessary (living micrococcus, 3 hours incubation, 50 degrees C test temperature). The lysozyme measurements of different papers are transferable not without reserve and require a careful interpretation in respect of reproducibility and clinical relevance.

Animals↗
PubMed1982

[Heart transplantation: successful treatment of postoperative complications. Case report with 19 months' follow-up].

In March, 1981, a 37-year old patient underwent cardiac transplantation, the first to be performed in Germany since 1969. Subsequent to the successful procedure, the patient has now been followed for more than 19 months. The indication for the intervention was established on the basis of endstage coronary artery disease. The operation was performed according to the technique of Lower and Shumway. Immunosuppression, in the early postoperative phase, was carried out with antithymocyte globulin, azathioprine and corticosteroids with administration oriented on the T-lymphocyte and total leukocyte counts as well as analysis of myocardial biopsies. In the late postoperative course, dosage of immunosuppressive agents was based almost exclusively on biopsy findings. Within the course of immunosuppressive therapy for cardiac transplantation, in particular, initial experience was gained with the use of antithymocyte globulin which was given in dosages of up to 12 mg/kg daily. The antithymocyte globulin proved effective for more than six weeks postoperatively. The alterations in immunosuppressive treatment necessitated by two gastrointestinal complications (perforated gastric ulcer, perforation of the small intestine) enabled a comparative analysis of the results of therapy with cyclosporin A given orally, intravascularly and intravenously. The Patient continues to be maintained on a regimen of cyclosporin A and corticosteroids and his general condition is good. For future cardiac transplantations, in the early postoperative course, alternating treatment with antithymocyte globulin, azathioprine and corticosteroids, on the one hand, with cyclosporin A and corticosteroids, on the other hand, would seem meaningful both to minimize adverse reactions as well as to provide effective immunosuppression. The experience rendered would promise to facilitate management of patients after cardiac transplantation.

Adolescent↗
PubMed1982

[Optimization of lysozyme determination: comparative study of preparations of test cultures of Micrococcus luteus (M. lysodeikticus Fleming)].

About 6000 single measurements of the susceptibility of different preparations of Micrococcus luteus (M. lysodeikticus Fleming) cells were performed in order to assess their suitability for the determination of various muramidase concentrations. In particular, a turbidimetric method was compared with an agar-diffusion technique under varying experimental conditions. From our results it can be concluded that M. luteus cells washed with ether and acetone or living lyophilized cells are most suitable for turbidimetric determinations of lysozyme activity. The optimum conditions for turbidimetric assays comprised an initial transmission of 20% and a test time of 5 min. For the agar-diffusion technique fresh living or living lyophilized cells were most suitable. For a given incubation time of 3 h an incubation temperature of 50 degrees C yielded the best results.

Acetone↗
PubMed1981

Sensilla of the cricket palp. Fine structure and spatial organization.

On the distal segment of the maxillary palp of the cricket Gryllus bimaculatus there is an extensive multimodal sensory field comprising approximately 5000 sensilla of nine morphologically distinguishable types: three types containing pores in their walls (5% of the total), three types with a pore at the tip (65%) and three types lacking pores (30%). The approximately 27,000 axons of their sense cells join in the distal palp segment to form two nerves. On the slightly smaller sensillum field on the tip of the labial palp, the same sensillum types are found in comparable density and proportion. By analogy with sensilla of which both structure and function are known, the structure of these three groups of sensillum types suggests that they function as olfactory, contact-chemoreceptive, and/or mechanoreceptive sensors. Moreover, the positioning of the sensilla on the tip of the palp is appropriate to these function. The cuticle of the pulp tip is elastically deformable; all sensory hairs are fixed via a special, presumably flexible socket suspension. These structural features could be an adaptation to stresses encountered during tactile activity of the palp.

Animals↗
PubMed1981

Sanfilippo syndrome type C: assay for acetyl-CoA: alpha-glucosaminide N-acetyltransferase in leukocytes for detection of homozygous and heterozygous individuals.

As assay for the detection in leukocytes of homozygous and heterozygous carriers of Sanfilippo syndrome type C is described. In one family with two patients suffering from Sanfilippo C syndrome, the affected individuals had no residual activity to acetyl-CoA: alpha-glucosaminide N-acetyltransferase. The determination of the acetyl-CoA: alpha-glucosaminide N-acetyltransferase/ beta-glucuronidase ratio allows the discrimination between obligate heterozygotes and normal individuals and may be used for carrier detection.

Acetyl Coenzyme A↗
PubMed1981

Localisation of acetyl-CoA: alpha-glucosaminide N-acetyltransferase in microsomes and lysosomes of rat liver.

Subcellular fractions of rat liver were obtained by differential centrifugation. The fractions enriched in lysosomes or microsomes were further fractionated in discontinuous sucrose density gradients or continuous iso-osmotic gradients made of modified colloidal silica. The fractions were analyzed for marker enzymes of the different subcellular organelles and for acetyl-CoA: alpha-glucosaminide N-acetyltransferase. The acetyl-CoA: alpha-glucosaminide N-acetyltransferase activity showed a bimodal distribution. About one fourth of the activity was associated with lysosomes, whereas the greater part of the activity was recovered in the microsomal fraction. Plasma membrane-enriched fractions contained only trace amounts of acetyl-CoA: alpha-glucosaminide N-acetyltransferase. The lysosomal and microsomal acetyl-CoA: alpha-glucosaminide N-acetyltransferases are membrane-bound and can be solubilized with Triton X-100. The pH dependence and sensitivity to various ions was similar for the lysosomal and microsomal enzyme.

Acetyltransferases↗
PubMed1981

[Serum and urine lysozyme in chronic pyelonephritis and chronic glomerulonephritis].

In 130 patients with chronic pyelonephritis and 215 patients with chronic glomerulonephritis the serum lysozyme content was established and in 114 and 186, respectively, the enzyme content of their urine tests. Moreover the lysozyme measurement in the serum of 28 patients undergoing haemodialysis was performed. A collective of 50 healthy persons served as comparative group. The lysozyme estimation was performed by means of the agar-diffusion technique after Ossermann and Lawlor in own modification. The average serum lysozyme levels of the patients with pyelonephritis (mean =7.3 micrograms/ml) as well as of the patients with glomerulonephritis (mean = 5.7 micrograms/ml) were significantly increased in contrast to the control group (mean = 4.5 micrograms/ml). Differences could be recognized between the various forms of glomerulonephritis. 34.2% of the patients with pyelonephritis and 37.1% of the patients with glomerulonephritis showed a lysozymuria. In functional restrictions of the kidneys as well as in active forms of the diseases increased concentrations in serum and urine could be established.

Adult↗
PubMed1980

Metabolism of sulfated glycosaminoglycans in rat hepatocytes. Synthesis of heparan sulfate and distribution into cellular and extracellular pools.

Primary cultures of rat hepatocytes grown in a serum-free medium supplemented with [35S]sulfate synthesize 35S-labelled glycosaminoglycans at an almost constant rate for 58 h. Approx. 57% of the newly synthesized 35S-labelled glycosaminoglycans remain within the hepatocytes, approx. 30% become associated with the cell surface and only 13% are secreted into the medium. The amount of cell-surface-associated 35S-labelled glycosaminoglycans became constant within 36 h, whereas no equilibrium was reached in the intra- and extracellular pool. During a 24 h chase more than 50% of the intracellular and cell-surface-associated 35S-labelled glycosaminoglycans disappears, more than 80% of this material is degraded and radioactivity is recovered as inorganic sulfate. A minor part is released into the medium in a macromolecular form. Heparan sulfate accounts for more than 95% of the 35S-labelled glycosaminoglycans in each of the three pools. It is distinguished from heparan sulfates from other sources by the presence of unsubstituted glucosamine residues. In all three pools, heparan sulfate chains of mean molecular weights between 24 000 and 30 000 are part of an alkali labile proteoglycan. Intra- and extracellularly, however, part of the heparan sulfate appears to have little, if any, protein attached. Hepatocytes contain heparan sulfate-degrading endoglycosidase activity, which may contribute to the variation of molecular weights observed for the heparan sulfate.

Animals↗
PubMed1980

Characterization of dermatan sulfate in mucopolysaccharidosis VI. Evidence for the absence of hyaluronidase-like enzymes in human skin fibroblasts.

Dermatan sulfate-chondroitin sulfate copolymers with a high content of dermatan sulfate are stored in cultured human skin fibroblasts from patients affected with mucopolysaccharidosis VI (Maroteaux-Lamy disease). Characterization of the storage material provided evidence that hyaluronidase-like enzymes are not present in these fibroblasts. This is based on the following observations: (i) dermatan sulfate chains stored intracellularly show no reduction of molecular size as compared with intact chains isolated from the extra-cellular space; (ii) the stored dermatan sulfate chains lack reducible end groups generated by endoglycosidases; (iii) homogenates of human skin fibroblasts do not degrade hyaluronate and (iv) the stored dermatan sulfate chains are degraded by testes hyaluronidase.

Cells, Cultured↗
PubMed1980

A 3H-labelled trisaccharide from heparin as substrate for acetyl-CoA: 2-amino-2-deoxy-alpha-D-glucoside N-acetyltransferase.

The tetrasaccharide fraction obtained by gel chromatography after treatment of commercially available heparin with nitrous acid was reduced with NaB3H4 and then hydrolysed with 2M trifluoracetic acid at 70 degrees for 3 days. By gel chromatography and electrophoresis, the 3H-labelled trisaccharide 1 bearing and unsubstituted 2-amino-2-deoxy-D-glucosyl group in the non-reducing position was obtained (18% from the 3H-labelled tetrasaccharide). By sequential, enzymic degradation, the structure alpha-D-GlcN-(1 leads to 4)-beta-D-GlcA-(1 leads to 4)-[1-3H]aManol was obtained for 1, which is a substrate for acetyl-CoA: 2-amino-2-deoxy-alpha-D-glucoside N-acetyltransferase, an enzyme that is deficient in the Sanfilippo C syndrome. In human-skin fibroblasts, the pH optimum of acetyl transfer onto 1 was between pH 5.5 and 7.0, and dependent on the buffer. An apparent Km for 1 of 0.14mM was found.

Acetyltransferases↗
PubMed1980

Phosphorylated oligosaccharides in lysosomal enzymes: identification of alpha-N-acetylglucosamine(1)phospho(6)mannose diester groups.

In human fibroblasts, the recognition of lysosomal enzymes by cell surface receptors is mediated by mannose 6-phosphate residues located on oligosaccharides that can be cleaved by endo-beta-N-acetylglucosaminidase H. About half of these oligosaccharides, as isolated from beta-hexosaminidase and cathepsin D secreted by human skin fibroblasts, are anionic. Most of these are resistant to alkaline phosphatase. The resistance is due to alpha-N-acetylglucosamine residues linked to mannose 6-phosphate by a phosphodiester bond. The major phosphorylated oligosaccharides contain one and two and possibly three phosphate groups blocked by N-acetylglucosamine. Besides the blocked phosphate groups these oligosaccharides contain a common inner core consisting of Man alpha 1,6-(Man alpha 1,3)Man alpha 1,6(Man alpha 1,3)Man beta GlcNAc and either one or two alpha 1,2-linked mannose residues.

Acetylglucosamine↗
PubMed1980

[The technetium-99-m-pyrophosphate scintigram in the assessment of the size of acute myocardial infarction: comparison with CK-(MB) curves mortality (author's transl)].

While the technetium-99m-pyrophosphate scintigram enables diagnostic proof of acute myocardial infarction, its use in the evaluation of the extent of infarction has not been clearly established. This study, in 30 patients with acute myocardial infarction was, thus, undertaken to assess the relationship between the findings of the technetium-99m-pyrophosphate scintigram, with respect to infarct area and uptake pattern, the infarct size, as determined from the total CK an CK-MB curves, and the mortality. The scintigraphically determined infarct areas ranged between 1.7 and 29.3 cm2; the 20.2 plus or minus 6.0 cm2 average for anterior wall infarction (n=18, range 7.4 to 29.3 cm2) was significantly greater (p smaller than 0.005) than the 8.3 plus or minus 5.3 cm2 average for inferior infarction (n=12, range 1.7 to 15.8 cm2). Correlation coefficients between the scintigraphically and enzymatically determined infarct sizes for the entire patient population ranged from 0.71 to 0.80. Anterior infarctions correlated more closely (0.66 to 0.84) than inferior infarctions (0.46 to 0.66). The technetium-99m-pyrophosphate uptake pattern was focal in 20 patients and ring-shaped (doughnut) in ten. Infarcts with focal uptake patterns were significantly smaller than those displaying a doughnut pattern (12.2 plus or minus 6.4 vs. 24 plus or minus 4.0 cm2, p smaller than 0.005). The infarct weight calculated from the CK-MC curve with application of individually determined disappearance rate for those infarcts displaying a focal uptake pattern was 34 plus or minus 29 grams while that associated with a doughnut uptake pattern was significantly greater at 86 plus or minus 25 grams (p smaller than 0.005). During the 18 -month observation period there were six deaths; the average scintigraphic infarct area of 22.8 plus or minus 3.6 cm2 in those who died was significantly greater (p smaller than 0.005) than that of the 13.8 plus or minus 8.2 cm2 of the survivors. Of the non-survivors, five had a doughnut uptake pattern and one displayed focal uptake. In the 24 survivors, a focal uptake pattern was found in 19 and a doughnut pattern in five. Conversely, 19 of the 20 patients with a focal uptake pattern survived while only five of the ten patients with a doughnut pattern were alive after 18 months. Thus, comparison with the enzymatically determined infarct weight as well as the mortality indicate that the technetium-99m-pyrophosphate scintigram yields clinically relevant data with regard to infarct size. Since no patient with an infarct area of less than 17 cm2 died within the 18-month observation period, designation of scintigraphically determined infarct size as small (smaller than 17 cm2) and large (larger than 17 cm2) was enabled. The corresponding cut-off point between large and small infarctions as determined enzymatically has been designated at 65 grams. Accordingly, agreement was found in 70% (14 of 20 patients) with scintigraphically small infarcts and in 80% (8 of 10 patients) with scintigraphically large infarcts...

Acute Disease↗