Biomedical subjects
Publications and source records attributed to H König.
We report on the sequence of the Methanothermus sociabilis aladh gene, which encodes the 5'-aminolevulinic acid dehydratase. The identity of the enzyme was determined by sequence comparison and by expression in Escherichia coli.
Phorbol esters stimulate and glucocorticoid hormones down-regulate a variety of promoters such as that of the collagenase gene through the transcription factor AP-1 (Fos/Jun). We now show by genomic footprinting of the collagenase promoter that phorbol ester treatment of cells results in the binding of AP-1 to its cognate DNA binding site in vivo. The DNA-protein contacts obtained in living cells are also found in vitro using cloned DNA and purified AP-1. Although in vitro synthesized glucocorticoid receptor can disturb the DNA binding of Jun homodimers, it does not interfere with the binding of Fos-Jun heterodimers or of purified AP-1 in vitro. Consistently, fully inhibitory doses of glucocorticoid hormone cause no change in apparent occupation of the AP-1 binding site in vivo. The hormone receptor acts without itself binding to DNA.
Steroid hormones and growth promoting factors regulate the expression of a number of genes involved in cellular growth and differentiation. In many cases, cellular proliferation and differentiation-specific pathways are mutually exclusive. As an example for the mechanism of mutual exclusion, transcription factors responsible for pathway-specific gene expression interact with each other. Steroid hormone receptors inhibit the action of the transcription factor AP-1 (Fos/Jun) and vice versa. This inhibitory interaction is predominantly effected at the level of protein-protein contact without the need for the interfering transcription factor to bind to DNA. In some cases the two pathways function additively or even synergistically resulting in cooperation of the transcription factors in regulation of gene expression. The examples to be discussed in this text document how elaborate and important cross-talks between signal transduction pathways are. This is particularly demonstrated by the fine tuning and reversibility of these processes.
The diagnostic value of MRI and ultrasound in the examination of 37 patients with chronic shoulder pain is compared. All the results were controlled or by arthroscopy/arthrotomy or by arthrography. Ruptures and degenerative changes of the rotator cuff and degenerative lesions could be detected with a high sensibility. The ultrasound examination as a fast, reproducible diagnostic method has now become part of the routine diagnosis. The MRI produces exacter information about the extent of the lesions; but it will be reserved for special questions.
32 patients with chronic polyarthritis were examined by MRT. Following intra-articular application of 25 mg prednisolone, 18 patients were re-examined after two of three weeks and 7 out of 18 patients were re-examined after 8 to 16 weeks, both by MRT and clinically. After an initial double echo sequence, we carried out a dynamic series of FLASH measurements (TR 30, TE 10 ms, 70 degrees) following an intravenous bolus of 0.1 mmol gadolinium-DTPA. Single intensity-time curves were treated quantitatively in relation to the pannus, muscles and effusions. Two or three weeks after treatment the curve parameters showed a reduction in pannus vascularisation which remained constant for the 8 to 16 weeks of further observation. Contrast diffusion in the pannus is initially reduced but approaches the early measurements at the time of the second examination.
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BACKGROUND: The present study was designed to define the prevalence and characteristics of skeletal muscle alterations in patients with chronic heart failure (CHF) and their relation to exercise capacity. METHODS AND RESULTS: The ultrastructure of skeletal muscle was analyzed by ultrastructural morphometry in 57 patients with CHF and 18 healthy controls. The volume density of mitochondria (Vvm) and the surface density (Svmc) of mitochondrial cristae were evaluated as a structural correlate of oxidative capacity of skeletal muscle. Vvm and Svmc were reduced by approximately 20% in patients with severe CHF irrespective of age and etiology. The cytochrome oxidase activity in mitochondria as determined by cytochemistry and subsequent morphometry in a subset of patients (n = 10) was significantly decreased in heart failure (p less than 0.01). The capillary length density of skeletal muscle was reduced in CHF (n = 12, p less than 0.05), and the fiber type distribution was shifted to type II fibers (n = 15, p less than 0.05). Vvm and Svmc were significantly related to peak exercise VO2 (r = 0.56, p less than 0.001, n = 60) and to VO2 at anaerobic threshold (r = 0.535, p less than 0.0001, n = 60). In 16 patients with severe heart failure, Vvm was inversely related to the duration of heart failure (r = 0.545, p less than 0.03). In 11 patients who underwent repeat biopsies after 4 months, a correlation was observed between the change in Vvm and the change in peak exercise VO2 (r = 0.89, p less than 0.001). CONCLUSIONS: These findings indicate that patients with CHF develop significant ultrastructural abnormalities of skeletal muscle reflecting a depressed oxidative capacity of working muscle. It appears that these alterations of skeletal muscle contribute to the decreased exercise capacity of these patients but are, in principle, reversible by an effective treatment regimen.
Dealing with the subject "polled goats intersexuality", whose problems are discussed, we studied the pathology of 7 3-month old XX-kids: 2 pseudomales, 3 males with urethral diverticulum (1 animal with 2 U.D.), and 2 testicular pseudohermaphrodites. Four horned male kids of comparable age were used as control. Urethral diverticula, seen now and then in local breeds, i.e. in "chamois-colored" mountain goats, Saanen, and Toggenburg goats, are rather unknown and only sporadically reported abroad. In agreement with previous histological studies, the testes were hypoplastic, the spermatogonia absent. In no case ovarian tissue was found. The Leydig cells appeared to be functionally active. The selection of an experimental flock of intersex-free polled goats is proposed.
At the serum response element (SRE) of the c-fos proto-oncogene a ternary complex with two proteins, ternary complex factor (p62TCF) and serum response factor (SRF) can be formed. Its formation has previously been suggested to be necessary for efficient induction of c-fos transcription by serum in mouse NIH 3T3 fibroblasts (1) and by phorbol esters, but not by serum, in mouse BALB/c 3T3 fibroblasts (2). It is shown here by genomic dimethyl sulfate (DMS) footprinting that this ternary complex is indeed formed in NIH 3T3 cells in vivo. However, cells were found, murine F9 teratocarcinoma stem cells, in which the pattern of protection and hyperreactivity is consistent with the absence of p62TCF in the protein complex in vivo, although inducibility of the endogenous c-fos gene is not impaired. Both in NIH 3T3 cells and in F9 cells a mutated c-fos promoter that binds SRF but fails to form a ternary complex, was inducible by serum and phorbol esters to the same extent as the wild-type promoter. The data suggest that ternary complex formation is not an absolute prerequisite for the transient induction of c-fos. Ternary complex formation rather appears to enhance overall promoter efficiency. A cell type specific component determines the formation of the multicomponent transcription factor complex in vivo.
The genes (slgA) encoding the surface-layer glycoproteins of the hyperthermophilic methanogens Methano-thermus pervidus and Methanothermus sociabilis were cloned and sequenced. The nucleotide sequences of these genes differ at only nine positions, resulting in three amino acid differences. In both organisms, the transcription start site was localized by primer extension analyses. The DNA sequence at this site conforms to the promotor box B motif for promotors of archaea. 24 nucleotides upstream of the transcription start is an A + T-rich region, which closely resembles the consensus box A motif of promoters of methanogens. Ribosome binding sites are exactly complementary to the 3' end of the 16S rRNA of these methanogens. Both slgA genes encode for a precursor of the mature surface-layer protein containing 593 amino acid residues with a putative N-terminal signal sequence of 22 amino acid residues. The deduced protein sequences contain 20 sequon structures representing possible carbohydrate-binding sites. In comparison with other surface-layer proteins, these obtained from the two hyperthermophilic methanogens contain unusually high amounts of isoleucine, asparagine and cysteine residues. Predicted secondary structures have a high content of beta-sheet structure (44%) and only 7% alpha-helix structures.
The in vitro fidelity of highly purified recombinant reverse transcriptase from simian immunodeficiency virus of African green monkeys (SIVagm) was determined. By using the phi X174am16 reversion assay an overall error rate of 1/19,000 was determined. This is 2.4-fold higher than the overall accuracy of purified recombinant HIV-1 reverse transcriptase, measured in parallel. The evaluation of error frequencies from nucleotide pool bias studies suggest an even higher accuracy for the SIVagm-derived reverse transcriptase. T:dGMP mismatches were formed most frequently with an error rate of 1/155,000, followed by G:dGMP (1/230,000), A:dGMP (1/315,000), G:dAMP (1/340,000), T:dCMP (1/540,000), T:dTMP (1/790,000), and A:dCMP (1/1,050,000) mispairs. Thus, according to pool bias effects and depending on the mismatch under consideration SIVagm reverse transcriptase appears to be 2 to 20-fold more accurate than the homologous enzyme from the human immunodeficiency virus type 1. This higher accuracy is not due to a co-purifying exonuclaease activity. Like the enzyme from HIV-1, the simian monkey-derived enzyme was found to be devoid of a proofreading 3' to 5' exonuclease.
The possibilities of MRI and ultrasound in visualizing the structures of the shoulder joint are compared. Both methods have a good accuracy in detecting changes of the soft tissue of the shoulder. Changes of the cartilage and the bony structures are better seen in the MRI. The MRI gives a static and clear view of the shoulder joint, meanwhile the ultrasound depends on the dynamic examination. Ultrasound and MRI are both reproducible and non-invasive methods, who seem to complete one another at the examination of the shoulder.
The aim of this study was to improve the MRI diagnosis of CMP, with special reference to the early stages and accurate staging. For this purpose, the retropatellar cartilage was examined by MRI while compression was carried out, using 21 patients and five normal controls. The compression was applied by means of a specially constructed device. Changes in cartilage thickness and signal intensity were evaluated quantitatively during FLASH and FISP sequences. In all patients the results of arthroscopies were available and in 12 patients, cartilage biopsies had been obtained. CMP stage I could be distinguished from normal cartilage by reduction in cartilage thickness and signal increase from the oedematous cartilage during compression. In CMP stages II/III, abnormal protein deposition of collagen type I could be demonstrated by its compressibility. In stages III and IV, the method does not add any significant additional information.
The cell wall of Methanosarcina barkeri consists of a heteropolysaccharide (methanochondroitin), which resembles the eukaryotic chondroitin. From cell extracts of Methanosarcina barkeri four uridine diphosphate and one undecaprenyl pyrophosphate-activated intermediate(s) of the methanochondroitin were isolated. In contrast to the known biosynthetic pathways of polysaccharides from other prokaryotes and eukaryotes, nucleotide activated oligosaccharide precursors are involved in the case of the methanochondroitin. Usually, oligosaccharides are synthesized at the lipid stage.
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If a clinician is faced with acute intoxications a rapid toxicological test is essential. The toxic substances are identified by a combined use of various systems of analysis such as gas chromatography, thin-layer chromatography or UV-spectroscopy. As the result of the analysis a data pattern is obtained which can be interpreted by comparing it with substance-related reference patterns. This paper describes a computer-program which allows more efficient reference comparisons and which gives a support of the decisions. The algorithms of the comparisions are based on fuzzy-theory.
Native reverse transcriptase from simian immunodeficiency virus was purified from virus with good recovery to near homogeneity. The optimum reaction conditions of the enzyme were determined with respect to divalent cations, pH and ionic strength. The enzyme was shown to possess both RNA-dependent and DNA-dependent DNA synthesis activity. In addition, we could demonstrate an associated RNase H activity. Employing novel assay conditions, activated DNA as a heteropolymeric substrate was used more efficiently than the homopolymeric substrate poly(rA).oligo(dT) which in turn was used twofold more effectively as the template primer than poly(dC).oligo(dG). Other homopolymeric substrates, including poly(rC).oligo(dG), were also tested but were found to be poorly used by the reverse transcriptase. The Miachaelis-Menten constants were determined for each of the four nucleotides needed to elongate a natural template primer. Simultaneously, using dideoxyadenosine triphosphate as nucleotide analogue, we could show that this compound acts as a competitive inhibitor with respect to dATP, whereas it acts as a non-competitive inhibitor with respect to the other nucleotides. Gel electrophoretic analysis showed the enzyme to consist of two polypeptides with apparent molecular masses of 64 and 48 kDa. Using activity gel electrophoresis, we were able to demonstrate that both subunits exhibit DNA synthesis activity.
The following putative precursors of the pseudomurein were isolated from trichloroacetic acid extracts of Methanobacterium thermoautotrophicum: a uridine diphosphate activated derivative of glutamic acid and the uridine diphosphate activated peptides (see text). The activated glutamic acid residue and the three activated pepetides lack the glycan components N-acetylglucosamine and N-acetyltalosaminuronic acid present in the intact pseudomurein. In this case uridine diphosphate should be directly linked to the amino group of a glutamic acid residue, which represents a new mode of amino acid and peptide activation.