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Biomedical subjects

R Rudolph

Publications and source records attributed to R Rudolph.

At least 163 records · Page 9Linked to original sources
PubMed1984

Growth dynamics of cultured myofibroblasts from human breast cancer and nonmalignant contracting tissues.

Myofibroblasts were successfully grown in tissue culture from the connective tissue stroma of three human breast adenocarcinomas. These cells had slower growth kinetics than fibroblasts from normal human dermis, as did myofibroblasts from two granulating wounds. Electron microscopy of breast cancer slices and tissue cultures of these specimens confirmed the presence of myofibroblasts in both. In early passages, the specificity of carcinoma-derived myofibroblast growth kinetics is preserved. The exact role of myofibroblasts in breast cancer, whether helping or hindering tumor growth, remains undetermined.

Breast↗
PubMed1984

The reticuloendothelial system in scrapie pathogenesis.

A single injection of 1 mg dextran sulphate 500 per mouse considerably prolonged incubation times and produced survivors, even when given 3 days after intravenous or intraperitoneal scrapie infection. Since this compound could be detected in mononuclear phagocytes of spleen and lymph nodes for up to 7 months, it is suggested that it impairs a particular step in the infectious process in these cells of the lymphoreticular system. Blockage of the reticuloendothelial system by trypan blue and silica did not alter the pathogenesis.

Animals↗
PubMed1984

Grass pollen allergoid. A 4-year follow-up study.

In an open comparative study with 31 grass pollen-allergic patients, two allergen extracts ('allergoid' extract/semi-depot extract) were examined. In the 1st year of treatment the allergoid group showed better results in relation to symptom score, drug consumption and IgG response, however, in the course of further treatment this difference was not evident anymore. Treatment was successful in both groups. A significant correlation between total antigen dose and IgG response was found. The better clinical results for the allergoid group in the first season after the 1st and 4th year of observation do indicate a better long-term therapeutic effect compared to the semi-depot group. This finding probably is due to the much higher antigen dose (4-fold) in the allergoid group.

Allergens↗
PubMed1984

Comparative growth dynamics and morphology between cultured myofibroblasts from granulating wounds and dermal fibroblasts.

Rat myofibroblasts from granulating wound biopsies (RGW) were successfully cultured and compared in terms of growth and morphology with fibroblasts from uninjured rat dermis (RD). Populations of early passage (P-3) RGW myofibroblasts grew significantly more slowly than RD fibroblasts. Logarithmic growth was nearly the same in late passage (P-30) populations of both cell types. Early passage RGW myofibroblasts were similar to those in vivo, as shown by well-defined microfilament bundles. RD fibroblasts contained less well defined microfilaments. Both late passage RGW myofibroblasts and RD fibroblasts displayed evidence of morphologic dedifferentiation. These data show that morphologic features of myofibroblasts and fibroblasts in vivo are maintained in vitro. Evidence is presented that cultured animal myofibroblasts maintain differentiation in early passage, whereas late passage cells suggest that these differences disappear with time.

Animals↗
PubMed1983

The reconstitution of denatured phosphoglycerate mutase.

The reconstitution of the tetrameric enzyme yeast phosphoglycerate mutase after denaturation in guanidine hydrochloride has been studied. Denaturation is almost completely reversible at enzyme concentrations greater than 10 micrograms/ml. Cross-linking by glutaraldehyde has been used to monitor the reassociation of the subunits; the kinetics of this process has been analyzed in terms of a model involving an equilibrium between monomer and dimer followed by a bimolecular association of two dimers to give a tetramer. Reactivation is found to parallel the appearance of tetramer. Structural changes during reconstitution have been measured by circular dichroism and fluorescence. Both methods reveal complex kinetics indicating the rapid formation of structured monomers (half-time less than 10 s), followed by slow subunit association. For comparison, preliminary reconstitution experiments were performed on the dimeric phosphoglycerate mutase from rabbit muscle.

Animals↗
PubMed1983

Limited proteolysis as a tool to study the kinetics of protein folding: conformational rearrangements in acid-dissociated lactic dehydrogenase as determined by pepsin digestion.

Noncovalent aggregation as a side reaction competing with the reconstitution of oligomeric enzymes is enhanced by slow conformational changes within the partially unfolded subunits. This has been shown for lactic dehydrogenase from pig muscle after acid dissociation [G. Zettlmeissl, R. Rudolph, and R. Jaenicke (1981) Eur. J. Biochem. 121, 169-175]. The present experiments confirm previous spectroscopic evidence (from circular dichroism) applying pepsin digestion and subsequent analysis of the fragments on sodium dodecyl sulfate-polyacrylamide gradient gels. The susceptibility of certain fragmentation sites toward pepsin digestion changes with increasing incubation at acid pH, in accordance with a slow M1 leads to M2 transition of the acid-dissociated monomers. Constant pulses of pepsin at varying times after transferring native enzyme to pH 2.3 yield distinct changes in the fragmentation pattern consisting of undigested monomers (Mr = 35,000) plus 12 fragments ranging from 31,000 to 5000. Short digestion of the M2 species at low concentrations of pepsin preferentially yields 25,000 and 10,500 fragments (molar ratio pepsin:lactic dehydrogenase = 1:24). The time-dependent decrease of monomers upon incubation in 0.1 M sodium phosphate, pH 2.3, at 20 degrees C strictly parallels the formation of the two fragments. The quantitative kinetic analysis of the changes in peptide pattern yields a first-order rate constant k1 = 8 +/- 2 X 10-4 s-1. The observed increase in proteolytic susceptibility is in the time range of the above mentioned decrease in the far-ultraviolet circular dichroism, and the parallel decrease in the yield of reactivation. The results suggest that during the M1 leads to M2 transition at acid pH a specific interdomain cleavage site is becoming exposed. As taken from the molecular weight of the two main fragments the trp 225-lys 226 peptide bond is the most probable candidate for this cleavage site.

Animals↗
PubMed1983

[Esophageal ulcer caused by mexiletine].

The anti-arrhythmic drug mexiletine caused an oesophageal ulcer, confirmed by endoscopy and biopsy, in a 72-year-old woman. This side-effect has not previously been published. The clinical course was similar to that observed after oesophageal ulceration caused by other drugs. An ulcer in the piriform recess also developed in a 63-year-old man who was taking both acetylsalicylic acid and mexiletine: it cleared up after mexiletine had been discontinued.

Aged↗
PubMed1983

Physicochemical characterization of a fast refolding monomeric class I fructose-1,6-bisphosphate aldolase from Staphylococcus aureus.

The class I fructose-1,6-bisphosphate aldolase from Staphylococcus aureus is proposed as a good candidate for thermodynamic and kinetic studies on protein folding. The monomeric enzyme (molecular weight 35 000 +/- 1000) has been previously described as 'unusually heat-stable' [F. Götz et al. (1980) Eur. J. Biochem. 108, 295-301]. In the present paper we show that the enzyme is reversibly denatured at relatively low temperature (26-39 degrees C), as determined by protein fluorescence and far ultraviolet circular dichroism; the van't Hoff enthalpy of the thermal unfolding is 355 +/- 63 kJ/mol. The dichroic absorption shows that the aldolase is extensively unfolded in 6 M guanidine/HCl. Complete reactivation of the guanidine-denatured enzyme in the test solution is extremely fast (less than 10 s in the temperature range from 24.6 degrees C to 7.7 degrees C). Reactivation ought to be much slower if isomerization reactions around at least some of the ten Xaa-Pro peptide bonds were rate-limiting for reactivation.

Animals↗
PubMed1983

Standardization of the dorsal rat flap model.

In an effort to standardize use of the dorsal rat flap model originally described by McFarlane, we determined the following. The effect of delay is readily demonstrated but does not prevent necrosis entirely. Almost all flaps showed some necrosis, but delayed flaps showed approximately 15% less necrosis. Measurement of surviving flap length is simpler and as reliable as the calculations of flap area. Flap survival shows little change after fifteen days, making longer periods of observation unnecessary. The simplest landmarks for flap length are the scapular tips and the palpable hip joint. No difference in survival is demonstrated by repositioning the flap's base 1 cm caudally. A one-week delay was found to be as effective as a two- or three-week delay. These observations are offered to encourage use of the dorsal rat flap model and to lessen the cost, time, and effort needed for flap research.

Animals↗
PubMed1983

Pressure ulcer of the hand masquerading as a burn.

Pressure ulcer of the hand is unusual and can be difficult to identify if not suspected. This report concerns a patient with a hand pressure ulcer that masqueraded as a burn and defied diagnosis for a prolonged time.

Adult↗
PubMed1983

Influence of glutathione on the reactivation of enzymes containing cysteine or cystine.

Refolding of dimeric porcine cytosolic or mitochondrial malate dehydrogenases and of tetrameric pig heart and skeletal muscle lactate dehydrogenases (containing 5-7 cysteine residues), as well as reformation of the four cystine cross-bridges of bovine pancreatic ribonuclease, were studied in the presence of reduced and oxidized glutathione (GSH and GSSG). At the intracellular GSH level (5 mM) reduced ribonuclease can be reoxidized by 0.01-0.5 mM GSSG (pH 7.4) both at 20 degrees C and 37 degrees C. In this physiological range of GSSG concentrations and pH, the dehydrogenases show at least partial reactivation. With GSSG concentrations greater than 5 mM, reactivation is found to be completely inhibited for all the enzymes given. The results show that at the intracellular level of GSH and GSSG, thiol groups in reduced, unfolded ribonuclease are oxidized to form intramolecular cystine cross-bridges, while thiol groups of typical cysteine enzymes, such as lactate and malate dehydrogenase, remain in their reduced state during refolding. The rate of reactivation of lactate dehydrogenase (porcine muscle) is not affected by GSSG. In the case of ribonuclease, increasing concentrations of GSSG increase the rate of reactivation: At 20 degrees C, the halftime of the correct disulfide bond formation varies from approximately equal to 80 h in the presence of 0.01 mM GSSG to approximately equal to 10 h in the presence of 0.25 mM GSSG. A further increase in the rate of reactivation at higher GSSG concentrations is accompanied by a decrease in yield. Reactivation of ribonuclease is also observed at the low glutathione level found in blood plasma (5-25 microM GSH).

Animals↗
PubMed1983

Dimers of porcine skeletal muscle lactate dehydrogenase produced by limited proteolysis during reassociation are enzymatically active in the presence of stabilizing salt.

Lactate dehydrogenase from pig skeletal muscle is known to be a "dimer of dimers" stabilized in its tetrameric state by an N-terminal sequence of 20 aminoacid residues. Limited proteolysis of dimeric intermediates of association with thermolysin prevents association of the enzyme to its tetrameric structure. The resulting stable "dimers" are still capable of binding to a Procion Green dinucleotide affinity column. This may indicate that the dimeric intermediates contain the dinucleotide fold of the native enzyme. Since this structural feature is preserved after thermolysin treatment, affinity chromatography may be applied to separate the "dimer" on a preparative scale. Endgroup analysis and peptide mapping of the proteolytic dimer indicates that the reaction product consists of intact chains lacking the "N-terminal arm", apart from "nicked subunits" with fragments of 18 000 and 12 000 molecular weight. According to its hydrodynamic and conformational characteristics (sedimentation velocity and circular dichroism), the "dimer" does not differ significantly from the native enzyme in the backbone structure of its subunits. Its sedimentation properties resemble those of other dimeric dehydrogenases. The spectral data are similar to those observed for the dimeric intermediates that form during reconstitution after acid denaturation. Under standard test conditions, the proteolytic "dimer" (as well as the dimeric intermediate on the pathway of folding) do not show enzymatic activity. However, in the presence of "structure-making ions" like 2 M ammonium sulfate, about 40% of the native catalytic function is restored in the dimeric state. As shown by activity transport measurements in the ultracentrifuge, both the native tetramer and the proteolytic "dimer" maintain their quaternary structure in this solvent.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1982

Rate-determining folding and association reactions on the reconstitution pathway of porcine skeletal muscle lactic dehydrogenase after denaturation by guanidine hydrochloride.

Reactivation of tetrameric porcine skeletal muscle lactic dehydrogenase after dissociation and extensive unfolding of the monomers by 6 M guanidine hydrochloride (Gdn . HCl) is characterized by sigmoidal kinetics, indicating a complex mechanism involving rate-limiting folding and association steps. For analysis of the association reactions, chemical cross-linking with glutaraldehyde may be used [Hermann, R., Jaenicke, R., & Rudolph, R. (1981) Biochemistry 20, 2195-2201]. The data clearly show that the formation of a dimeric intermediate is determined by a first-order folding reaction of the monomers with k1 = (8.0 +/- 0.1) x 10(-4) s-1. The rate constant of the association of dimers to tetramers which represents the second rate-limiting step on the pathway of reconstitution after guanidine denaturation, was then determined by reactivation and cross-linking experiments after dissociation in 0.1 M H3PO4 containing 1 M Na2SO4. The rate constant for the dimer association (which is the only rate-limiting step after acid dissociation) was k2 = (3.0 +/- 0.5) x 10(4) M-1 s-1. On the basis of the given two rate constants, the complete reassociation pattern of porcine lactic dehydrogenase after dissociation and denaturation in 6 M Gdn . HCl can be described by the kinetic model (formula: see text).

Cross-Linking Reagents↗