Discovery of FK 506 analogues that are nontoxic to mouse pancreatic beta cells in vitro but show diabetogenic potential when administered to the rat.
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Animals↗
Biomedical subjects
R Wagner
Publications and source records attributed to R Wagner.
F-Dopa as an amino acid tracer to detect brain tumors.
A 57-yr-old woman suffering from light movement disorder of the left arm and hand was referred for 18F-Dopa PET. The PET study not only proved asymmetrically reduced dopamine uptake in the putamen (influx constant Ki right 0.0064/min, left 0.0086) but also revealed pathologically increased 18F-Dopa accumulation in the right frontal lobe. Further PET examinations demonstrated increased 11C-methionine uptake and low glucose metabolism in this right frontal region. MRI and 1H-MRSI showed a heterogeneous lesion with reduced N-acetyl-aspartate and increased choline and lactate, suggesting a mixed, low-grade glioma. In 15O-water studies, during intentional movements of one hand the respective motor areas were identified, indicating asymmetries due to the mass occupying lesion. The tumor could be removed in open surgery, thus sparing the motor areas; a mild postoperative motor deficit resolved to the presurgical state. Histology confirmed the diagnosis of a grade 2 oligo-astrocytoma. This case impressively demonstrates that 18F-Dopa can be used as an amino acid tracer for brain tumor detection in addition to its established application to assess aromatic acid decarboxylase activity.
Brain↗
[Autologous keratinocyte culture on hyaluronic acid ester membranes: an alternative in complicated wound management?].
Cultivation and transplantation of autologous keratinocytes has been used in the last 15 years to treat complicated wounds of different origin. In spite of excellent technical advancements and clinical experiences cultured keratinocyte grafting still is associated with practical limitations. Application of hyaluronic acid ester membranes as carrier substrate for the transfer of keratinocytes allows improved graft handling: reduces total time required for tissue cultivation and furthermore enhances vitality of the keratinocytes because of possible grafting at semiconfluence.
Aged↗
[Extra-articular proximal femur fracture in the elderly--dynamic hip screw or intramedullary hip screw for fracture management?].
Over a 6-year period we treated 119 pertrochanteric fractures using dynamic hip screws (DHS). During the following 3 years we stabilized 112 per-, sub- and intertrochanteric, as well as "trochanter-associated" fractures by means of intramedullary hip screws (IMHS) or gamma nails (GN). Within comparable patient groups we encountered the following complications: DHS vs IMHS/GN: secondary varus malalignment of the collum femoris with "cut out": 1.7% vs. 1.8%; secondary varus malalignment without "cut out": 0.8% vs. 1.8%; infections: 5.0% vs. 2.7%; hematomas needing revision operations: 2.5% vs. 0%; torn out plate: 1.7% vs. 0%; intraoperative fissures of the shaft: 0% vs. 1.8%; intraoperative perforations of the shaft: 0% vs. 0.9%. Thus, the rate of reoperation for complications within the DHS series was 11.8%, while the rate within the IMHS/GN series was 6.3%. For stable pertrochanteric fractures we therefore acknowledge DHS as the ideal implant in our opinion, while for all other extraarticular proximal fractures of the femur we recommend IMHS or GN.
Adolescent↗
Ion channels in the chloroplast envelope membrane.
Isolated chloroplast envelope membranes were fused with azolectin liposomes. Ion transport across the membrane of these liposomes was investigated by the patch-clamp technique and in planar bilayers. Our results show that the chloroplast envelope contains voltage-dependent anion- and cation-selective channels as well as anion- and cation-selective pores with high conductances. At least one of the high-conductance pores could be located in the chloroplast outer envelope membrane. The low-conductance chloride channel and the potassium channel showed complex gating behavior with subconductant states. Potassium channel gating was affected by monovalent and divalent cations as well as by millimolar concentrations of ATP. Low concentrations of Cs+ induced a flickering block. Voltage dependence of the open probability reveals that macroscopic currents of potassium channels are rectified with preferential potassium uptake into the chloroplast. Flux measurements and determinations of the stroma pH of intact chloroplasts confirm the presence of a potassium channel that is regulated by divalent cations (Mg2+) and by ATP. The fully open potassium channel revealed a conductance of lambda approximately equal to 100 pS in asymmetric KCl (250/20 mM KCl), and the fully open chloride channel revealed a conductance of lambda approximately equal to 60 ps in 100 mM Tris/HCl. One high-conductance channel, mainly active at holding potentials > 60 mV, was slightly selective for glutamate anions (PK+/PGlu- approximately equal to 2) and revealed fast voltage-dependent gating. This high-conductance channel had a conductance of lambda approximately equal to 540 pS (in 250/20 mM potassium glutamate) and was closed most of the time. A second type of high-conductance channel, mainly open and active at holding potentials below 30 mV, was slightly selective for cations (PGlu-/PK+ approximately equal to 2) with a conductance of lambda approximately equal to 1.14 nS (in 250/20 mM potassium glutamate).
Adenosine Triphosphate↗
Effects of different growth conditions on the in vivo activity of the tandem Escherichia coli ribosomal RNA promoters P1 and P2.
We have analyzed the relative activities of the Escherichia coli ribosomal RNA promoters P1 and P2 in vivo under different physiological conditions. Promoter efficiencies were determined by quantitative comparison of the transcript-specific primer extension products obtained from total RNA preparations. Cells were analyzed at different stages of the growth cycle, at different growth rates, and under conditions of stringent control. In addition, the rRNA gene dosage was altered by transformation with plasmids containing additional rrnD or rrnB transcription units, or rRNA operons in which one of the tandem promoters (P1) had been deleted. Under conditions of amino acid starvation (stringent control) we observed the expected strong reduction in P1-directed transcription. In contrast to the previous assumption that the P2 promoter is not regulated, we simultaneously noticed a smaller but significant repression of P2-directed transcription. In strains in which the rRNA gene dosage was increased by transformation with plasmids bearing rRNA transcription units, a similar degree of repression was observed. Repression of the P1 promoter activity was increased, however, when cells contained extra rRNA operons with P2 promoters only. As demonstrated under stringent control conditions, changes in the growth cycle also affected the activity of promoters P1 and P2. A greater proportion of P2-derived transcripts was observed when cells changed from exponential to stationary growth or if cultures were grown in minimal medium. Under steady-state, slow growth conditions (minimal medium) we obtained evidence showing that the ratio of P1/P2 transcription products is much lower for cells with extra rrnB as compared to extra rrnD operons or cells lacking extra rRNA operons, implying an operon-specific regulation.(ABSTRACT TRUNCATED AT 250 WORDS)
Alleles↗
A prokaryotic potassium ion channel with two predicted transmembrane segments from Streptomyces lividans.
We report the identification, functional expression, purification, reconstitution and electrophysiological characterization of an up to now unique prokaryotic potassium ion channel (KcsA). It has a rectifying current-voltage relationship and displays subconductance states, the largest of which amounts to A approximately equal to 90 pS. The channel is blocked by Cs- ions and gating requires the presence of Mg2+ ions. The kcsA gene has been identified in the gram-positive soil bacterium Streptomyces lividans. It encodes a predicted 17.6 kDa protein with two potential membrane-spanning helices linked by a central domain which shares a high degree of similarity with the H5 segment conserved among eukaryotic ion channels. Multiple alignments of deduced amino acids suggest that the novel channel has the closest kinship to the S5, H5 and S6 regions of voltage-gated K+ channel families, mainly to the subfamily represented by the Shaker protein from Drosophila melanogaster. Moreover, KcsA is most distantly related to eukaryotic inwardly rectifying channels with two putative predicted transmembrane segments.
Amino Acid Sequence↗
Mutation detection using immobilized mismatch binding protein (MutS).
An accurate and highly sensitive mutation detection assay has been developed. The assay is based on the detection of mispaired and unpaired bases by immobilized mismatch binding protein (Escherichia coli MutS). The assay can detect most mismatches and all single base substitution mutations, as well as small addition or deletion mutations. The assay is simple to use and does not require the use of either radioactivity or gel electrophoresis.
Adenosine Triphosphatases↗
Characterization of changes in the glycosylation pattern of recombinant proteins from BHK-21 cells due to different culture conditions.
The N-glycosylation patterns of a genetically engineered human interleukin-2 variant glycoprotein (IL-Mu6), produced by BHK-21 cells from long-term suspension and microcarrier cultures in the presence and absence of fetal calf serum were compared. IL-Mu6 was used as a model protein in studying the effect of different controlled cell culture conditions on the expression of N-glycans in recombinant glycoproteins. IL-Mu6 contains a single amino acid substitution (Glu100<==>Asn) generating a potential N-glycosylation recognition site (Asn100-Xxx-Thr/Ser) in addition to the natural O-glycosylation at position Thr3. Parallel cell cultivations were carried out in two continuously perfused 2.5-liter stirred bioreactors under defined culture conditions. Major differences were found in the glycoprotein products obtained during these different cultivation conditions. Serum-free cultures resulted in a higher level of terminal sialylation and proximal alpha 1-6 fucosylation. The ratio of O- to N-glycans as well as the amount of nonglycosylated product and the antennarity of N-linked carbohydrates in the model protein exhibited major differences depending on the presence or absence of serum, the condition of growth and the cultivation procedure.
Amino Acid Sequence↗
Fluorescence analysis of the Escherichia coli transcription regulator H-NS reveals two distinguishable complexes dependent on binding to specific or nonspecific DNA sites.
Here we report a structural investigation of the transcription factor H-NS and its DNA interaction. H-NS has a general effect on transcription by compacting DNA; but for a number of specific genes, it is known to act directly as repressor or activator. The homodimeric protein binds to the major groove of DNA in a sequence-nonspecific manner, recognizing a curved conformation of the target DNA. H-NS consists of 136 amino acids with a single tryptophanyl residue at position 108. To overcome the apparent lack of any other structural details, we took advantage of the intrinsic fluorescence of Trp-108. Static and dynamic quenching constants obtained with the neutral quencher molecule acrylamide are consistent with a hydrophilic environment and high degree of solvent exposure for Trp-108. In addition, quenching studies in the presence of the anionic quencher iodide indicate a positively charged microenvironment for the same amino acid residue. Specific and nonspecific H-NS.DNA complexes were studied by gel retardation and fluorescence analysis. While specific H-NS.DNA complex formation is accompanied by a clear enhancement of the tryptophanyl fluorescence intensity, interaction in the presence of the nonspecific competitor DNA poly(dI-dC) decreases the fluorescence quantum yield.
Bacterial Outer Membrane Proteins↗
[Current advances in HIV pathogenesis. Results of therapy and vaccine development].
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Biomarkers↗
Leukemia patient's spirits lifted high.
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Adolescent↗
Protection against EHV-1 challenge infection in the murine model after vaccination with various formulations of recombinant glycoprotein gp14 (gB).
Four formulations of the equine herpesvirus type 1 (EHV-1) glycoprotein gp 14 (gB), were tested for their ability to protect mice against intranasal (inas) EHV-1 challenge infection. The preparations tested included (i) a truncated gp14 produced in Escherichia coli or (ii) a truncated gp14 expressed in insect cells by a recombinant baculovirus, (iii) truncated gp14 coexpressed with human immunodeficiency virus type 1 (HIV-1) gag virus-like particles (VLP) in insect cells, and (iv) a gp14-DNA vaccine under the control of the cytomegalovirus immediate early promoter. All antigen preparations and the DNA vaccine elicited a humoral and delayed-type hypersensitivity (DTH) immune response to EHV-1 after intramuscular (im) immunization. After inas immunization, only the VLP-gp14 preparation produced both a good humoral and a prominent DTH immune response; gp14 expressed by insect cells elicited high titers of EHV-1-specific antibodies, whereas gp14 produced in E. coli and the DNA vaccine elicited only low antibody titers. Glycoprotein gp14 expressed by bacteria, however, induced a strong DTH reaction after inas application. Mice were completely protected against EHV-1 challenge infection after both the im and the inas immunization with the VLP-gp14 preparation. Protection was less efficient after immunization with the E. coli and insect cell gp14s as well as after DNA vaccination. Although the transmembrane domain of EHV-1 gp14 was deleted, recombinant gp14 could be demonstrated in insect cell membranes at late times postinfection and aggregated with the VLPs. It is suggested that the transmembrane domain is not required for gp14-association with membranes in that system.
Administration, Intranasal↗
[Laboratory diagnosis of HIV infection].
Recent use of modern methods in the field of molecular biology has provided new insights into the genetic and structural nature of HIV, which are now increasingly being applied to the diagnostic work-up. For the reliable laboratory diagnosis of HIV infection, three different possibilities are available: detection of specific antibodies against viral proteins (anti HIV AB), detection of the virus itself (HIV antigen) and detection of viral nucleic acid using in vitro amplification techniques. In this overview, we compare clinical and diagnostic parameters of an HIV infection, and discuss the current state of the art of an HIV infection and the future prospects offered by new molecular biological methods such as the use of recombinant antigens or the polymerase chain reaction (PCR) for the detection of viral nucleic acid.
AIDS Serodiagnosis↗
The Escherichia coli ribosomal RNA leader nut region interacts specifically with mature 16S RNA.
All ribosomal RNAs are preceded by leader sequences not present in the final ribosome particles. The highly conserved leader sequences of bacterial rRNAs are known to be important for the folding and assembly of functional ribosomes. Very likely transient binding of the leader to mature parts of the 16S RNA occurs during transcription. To better understand the mechanistic details of these functions we have performed a secondary structural analysis of E. coli ribosomal RNA leader transcripts by chemical modification and enzymatic hydrolysis studies. The data were combined with results from thermodynamic stability calculations to yield a generalized structural model. The same secondary structure of the leader core, comprising the nut-like sequences up to the mature 5' end of the 16S RNA, was deduced, irrespective if transcripts started at promoter P1 or 120 nucleotides downstream at P2. Employing gelshift and cross-linking studies we were able to demonstrate that a part of the leader core, namely the nut-like sequence elements bind directly to specific regions within the mature 16S RNA. The sites of RNA-RNA cross-linking could be localized by sequencing. They map in the 16S RNA 5' domain at nucleotide positions G27 to G42, C48, G68, G117 and G126. The results may explain the recently observed scaffolding function of the leader RNA during ribosome biogenesis.
Base Sequence↗
Circulating forms of ICAM-3 (cICAM-3). Elevated levels in autoimmune diseases and lack of association with cICAM-1.
The intercellular adhesion molecule-3 (ICAM-3) has been identified as the third LFA-1 ligand in addition to ICAM-1 and ICAM-2. In this report we have identified circulating forms of ICAM-3 (cICAM-3) in human serum. Using a sandwich ELISA with two monoclonal anti-ICAM-3 Abs, we detected cICAM-3 in concentrations between 40 and 360 ng/ml in all of 112 healthy controls. An analysis of patient sera from 10 different immune-mediated diseases revealed a distinct pattern of expression. Significantly elevated cICAM-3 levels were found in rheumatoid arthritis, systemic lupus erythematosus, Guillain-Barré syndrome, and multiple sclerosis, but not in type I diabetes, Grave's disease, chronic autoimmune thyroiditis, ulcerative colitis, or Crohn's disease. cICAM-3 levels were significantly higher in systemic lupus erythematosus patients with active compared with nonactive disease. Despite their binding to the same integrin receptor, serum levels of cICAM-3 did not correlate with cICAM-1 concentrations in either normal persons or in patients. The majority of patients had either elevated cICAM-3 or cICAM-1 levels but not both. In conclusion, a circulating form of ICAM-3 is present in human sera. cICAM-3 expression is elevated in certain immune-mediated diseases but occurs independently of cICAM-1.
Adolescent↗
A procedure for the rapid preparation of guanosine tetraphosphate (ppGpp) from Escherichia coli ribosomes.
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Adenosine Triphosphate↗
Floor of mouth carcinoma. The management of the clinically negative neck.
OBJECTIVES: Examine the management of the clinically negative neck and evaluate the role of elective neck dissection in patients with squamous carcinoma of the floor of the mouth. DESIGN: Retrospective analysis of a cohort of patients with squamous carcinoma of the floor of the mouth and N0 stage disease of the neck who were treated between 1973 and 1992. The mean follow-up was 6 years. PATIENTS: The cohort consisted of 129 patients. Excluded from analysis were patients without evidence of disease but less than 3 years of follow-up and those with uncertain resection margins. INTERVENTION: Resection of the floor of the mouth lesion with or without marginal mandibulectomy. Elective lymphadenectomy was performed in 26 (23%) of the 129 patients. OUTCOME MEASURE: Estimates were obtained of survival according to mode of therapy, classification of treatment modality, determinate cure, locoregional failure, salvage, and occult disease by clinical stage. RESULTS: Occult disease was detected in 23% of the patients who underwent elective neck dissection. Recurrence in the neck occurred in 36% of 103 patients who received follow-up but did not undergo elective neck dissection. The determinate survival at 3 years was 100% for patients with occult disease who underwent elective neck dissection. Overall, 96% of the patients who were treated with elective neck dissection were cured; 85% of the patients who received no initial treatment of the neck were cured; and 59% of the patients with failure in the neck were salvaged. CONCLUSIONS: A more aggressive approach to the neck with N0 disease may be warranted. Selective neck dissection allows early removal of occult metastases with acceptable morbidity. In elective dissection for clinically and histologically negative necks, the high rate of survival may result from the removal of metastatic carcinoma that was missed in the histopathologic sampling process.
Adult↗