Biosynthesis of chondroitin sulfate side chains and hyaluronate. Time-course studies with cartilage slices of calf ribs under different conditions.

The time course of double labeling with 35SO24- and [3H]glucosamine was followed in a semi-in vitro system of cartilage slices from calf ribs whose chondroitin sulfate peptide pool consists of (A) less than 1% of very short under sulfated side chains of less than 10 disaccharide units length, (B) 3--5% of short under sulfated longer side chains (16 to 25 disaccharide units), (C) 3--5% of short, slightly oversulfated side chains (16--23 dissacharide units, very probably containing some dermatan sulfate), (D) the bulk material (74--82% of total uronate) of longest, slightly undersulfated or equally sulfated side chains (22--42 disaccharide units). After 10 min incubation rapid chain elongation with [3H]glucosamine and prelabeling with 35SO24- of endogenous acceptors are apparent. Chains of type A exhibit highest specific radioactivities. During 30--60 min incubation it is mainly chains of type B that show highest specific radioactivities, after 90 min chains of type C. On the after hand, chains of type D always incorporated the highest total amount of both precursors. Preincubation of slices for 40 min at 37 degres C strongly enhances labeling rates of all types A and B. After 10 min preincubation followed by 35SO24- labeling for 60 min, a decrease of radioactivity of type A and a distinct increase with type B are observed during the post incubation period. After pulse chase experiments type B exhibits highest specific radioactivities. The data make it evident that undersulfated short chondroitin sulfate side chains form very rapidly in a well organised manner and grow, by elongation and proceeding sulfation processes, to longer higher sulfated chains. The labeling of the hyaluronate pool is about half of that of the chondroitin sulfate pool after a lag phase of 10 min. The latter increases linearly after 35--45 min incubation time. However, after preincubation and chase experiments the hyaluronate pool is more highly labeled. The data indicate different precursor pools of both biosynthesis mechanisms, probably located in different cell compartments and/or different cartilage cells.

Disorganization of glycolytic and gluconeogenic pathways in skeletal muscle of aged persons studied by histometric and enzymatic methods.

Histometric data obtained by the point counting method, and the enzyme patterns of glycolysis, gluconeogenesis, fatty degradation and energy transfer have been determined in the same muscle specimens of m. vastus lateralis from 12 untrained patients between the ages of 4 and 78 years who suffered no disturbance of the neuromuscular system. Activities of 18 enzymes have been related to pure muscle weight corrected for fatty and connective tissue content, as well as to single fibre weight. A comparable muscle enzyme pattern was found in persons of around 20 years old and around 70 years old when expressed per gram of single fibre weight. However, in terms of grams of pure muscle weight, a significant activity decrease with age was obtained for 6-phosphofructokinase, triosephosphate dehydrogenase and phosphoenolpyruvate carboxykinase, whereas activity of hexose diphosphatase increased with age as also did 3-hydroxyacyl-CoA dehydrogenase activity. Five other cytoplasmic enzyme activities involved in glycolysis and energy transfer did not change significantly with age, nor did lysosomal acid phosphatase. The mitochondrial enzyme activities of gluconeogenesis (for example, pyruvate carboxylase, malic enzyme) were diminished to a lesser extent as also the auxiliary enzymes glutamic-oxaloacetic transminase and glutamic-pyruvic transaminase; glutamate dehydrogenase activity remained unchanged. The findings indicate a distinct disorganization of cytoplasmic glycolysis and gluconeogenesis pathways in presenile human skeletal muscle, confirming the histometric data already described. They cannot be explained by changes with age in numerical or areal ratio of type I and type II fibres.

Inhibitory and stimulating effects of various types and series of prostaglandins on in vitro biosynthesis of proteochondroitin-4,6-sulfate and protein and anaerobic glycolysis in calf rib cartilage.

The effect of various types of prostaglandins (PGs) have been studied in a semi-in-vitro system with cartilage slices of calf ribs. 0.1 mmol/1 PG B1, D2, E1, E2, F1 alpha, F2 alpha inhibit biosynthesis of Ch-4,6-S protein to a higher extent than 10 mumol/l; 1 and 2 series PG E and F (but not B) inhibit similarly, PG A2 inhibits twice as much. With biosynthesis of total protein 2-series PG A, B, E, F act more inhibitorily than 1-series PG. 10 mumol/l PG A2, E1, E2 produce cAMP-like effects, e.g. acceleration of biosynthesis of Ch-4,6-S protein and total protein as well as of anaerobic glycolysis; PG F2 alpha stimulates the first two anabolic processes, PG B1 only the second one. PG A1 stimulates Ch-4,6-S protein biosynthesis and anaerobic glycolysis, a cGMP-like effect. 20 mumol/l diBu-cAMP or cAMP (together with 0.1 mmol/l theophylline) produce stimulating and inhibitory effects on these three anabolic processes; both compounds produce additively positive or additively negative effects in connection with the inhibitory PG effects on these three anabolic processes.

[Diagnosis of cerebrospinal fluid: the use of semiquantitative rapid tests. Investigation of the adsorbance of proteins to glass and plastic tubes (author's transl)]. / Liquordiagnostik: Untersuchungen mit Schnelldiagnostica Untersuchungen zur Adsorption von Proteinen in Glas- und Kunststoffröhrchen.

The semiquantitative rapid tests used in the present study proved to be sufficiently sensitive for the estimation of erythrocytes in CSF samples, but they were inadequate with respect to the estimation of less than 100 microliter mononuclear and less than 10/microliter segmental neutrophilic leucocytes, of bilirubin, and albumin. Plastic tubes and cuvettes (e.g. of polystyrol) did not adsorb IgG, IgA, albumin or prealbumin from CSF samples, but adsorbed up to 80% of each purified single protein.

Hyaluronate-proteoglycan complex: evidence for separate biosynthesis mechanisms of the macromolecules.

In cartilage cells biosynthesis of hyaluronate and chondroitin-4-,-6-sulfate from proteoglycan takes place via two different distinct precursor pools; the synthesis of hyaluronate appears to require the unaffected formation of nucleotides and nucleic acids, whereas that of proteoglycan is very sensitive to the modulation of protein biosynthesis.

[Cerebrospinal fluid cytology using prestained slides (author's transl)]. / Liquorzytologie mit farbbeschichteten Objektträgern.

The vital staining method using prestained slides permits satisfactory characterization and differentiation of benign and malignant cells present in CSF using 10 and 20 mul of CSF. In comparison the sedimentation technique of Sayk and the cytocentrifuge with subsequent Pappenheim staining require larger volumes of CSF. They have the disadvantage of possible loss of cells and of cell denaturation.

Flattening and/or expanding of pH gradients in isoelectric focusing gels exemplified with PhastSystem.

A simple method of flattening and/or expanding of pH gradients in isoelectric focusing is described for any pH interval desired: to modify pH gradients near one electrode a paper strip soaked with carrier ampholytes is applied onto the gel close to the opposite electrode. In order to flatten central parts of pH intervals paper strips are applied onto the gel at both electrodes. Conditions and criteria (e.g. amount and pH intervals of carrier ampholytes, width and localization of the paper strip, separation period) for optimization are presented with PhastSystem using ready-made gels with three different pH intervals and pI marker proteins (Pharmacia). Examples utilizing erythrocyte lysates are presented.

Biochemical and autoradiographical distribution of hyaluronic acid in calf rib cartilage.

In calf rib cartilage, about one half of total hyaluronate is soluble with guanidinium hydrochloride, the other half only after collagenase treatment. Evidence is presented for its pericellular and intracellular distribution.