Evaluation of the risk of aflatoxin contamination in fresh medicinal plants.

Based on experimental results of aflatoxin analysis as well as information from literature, this contribution discusses the likelihood of aflatoxin contamination in fresh medicinal plants. As cultivation and collection of medicinal plants in accordance with Good Agricultural and Collection Practice (GACP) and the local climatic conditions minimise aflatoxin contamination and, as fresh raw material is normally processed immediately, aflatoxin contamination of fresh medicinal plants from Central European countries is extremely unlikely. As a result of the risk-based approach to aflatoxin testing, 3 options are proposed depending on the origin of the material and the plant parts used: no testing, skip lot testing or routine testing.

Clinical fluoroscopic fiducial-based registration of the vertebral body in spinal neuronavigation.

We present a system involving a computer-instrumented fluoroscope for the purpose of 3D navigation and guidance using pre-operative diagnostic scans as a reference. The goal of the project is to devise a computer-assisted tool that will improve the accuracy, reduce risk, minimize the invasiveness, and shorten the time it takes to perform a variety of neurosurgical and orthopedic procedures of the spine. For this purpose we propose an apparatus that will track surgical tools and localize them with respect to the patient's 3D anatomy and pre-operative 3D diagnostic scans using intraoperative fluoroscopy for in situ registration and embedded fiducials. Preliminary studies have found a fiducial registration error (FRE) of 1.41 mm and a Target Localization Error (TLE) of 0.48 mm. The resulting system leverages equipment already commonly available in the operating room (OR), providing an important new functionality that is free of many current limitations, while keeping costs contained.

Cisplatin-digoxigenin mRNA labeling for nonradioactive detection of mRNA hybridized onto nucleic acid cDNA arrays.

We optimized a novel nonradioactive hybridization technique using a cis-platin coupled digoxigenin derivative for direct labeling of mRNA. This new mRNA reporter molecule was applied to cDNA membrane arrays to simultaneously identify expression of hundreds of genes. The sensitivity of this nonradioactive mRNA hybridization technique was comparable to radioactive cDNA labeling on house-keeping gene expression but even superior on the detection limit of the expression of low abundant genes using mRNA isolated from human diploid fibroblasts. Additional advantages are faster readout and decreased total working times because of luminescence technology and avoidance of radioactivity. Finally, no potential artifactual reverse transcription step is necessary because of direct labeling of mRNA used for hybridization on nucleic acid arrays.

Low affinity binding of interleukin-1 beta and intracellular signaling via NF-kappa B identify Fit-1 as a distant member of the interleukin-1 receptor family.

The fit-1 gene gives rise to two different mRNA isoforms, which code for soluble (Fit-1S) and membrane-bound (Fit-1M) proteins related to the type I interleukin (IL)-1 receptor. To investigate IL-1 binding, we have synthesized and purified histidine-tagged polypeptides corresponding to Fit-1S and the extracellular domain of the type I IL-1 receptor using a vaccinia expression system. Fit-1S is shown to interact with IL-1 beta, but not with IL-1 alpha. However, Fit-1S binds IL-1 beta only with low affinity in contrast to the IL-1 receptor, suggesting that IL-1 beta is not a physiological ligand of Fit-1S. Moreover, expression of the membrane-bound protein Fit-1M in transiently transfected Jurkat cells did not result in activation of the transcription factor NF-kappa B following IL-1 beta treatment. However, a chimeric protein consisting of the extracellular domain of the type I IL-1 receptor and of the transmembrane and intracellular regions of Fit-1M stimulated NF-kappa B-dependent transcription as efficiently as the full-length type I IL-1 receptor. These data indicate that Fit-1M is a signaling molecule belonging to the IL-1 receptor family.

Retinoid-dependent in vitro transcription mediated by the RXR/RAR heterodimer.

The effects of retinoids on gene regulation are mediated by retinoic acid receptors (RARs) and retinoid X receptors (RXRs). Here, we provide the first biochemical evidence that, in vitro, ligand governs the transcriptional activity of RXR alpha/RAR alpha by inducing conformational changes in the ligand-binding domains. Using limited proteolytic digestion we show that binding of the cognate ligand causes a conformational change in the carboxy-terminal part of the receptor. We also show that recombinant RXR alpha/RAR alpha is partially active in the absence of exogenously added ligand. Trans-activation depends critically on the ligand-dependent transcriptional activation function AF-2 of RAR alpha. Full activation by recombinant RXR alpha/RAR alpha, however, requires the addition of either all-trans RA, 9-cis RA, or other RAR-specific agonists, whereas an RAR alpha-specific antagonist abolishes trans-activation. Intriguingly, the ligand-dependent AF-2 of RXR does not contribute to the level of transcription from the RAR beta 2 promoter in vitro even when the cognate ligand (9-cis RA) is bound. Thus, the major role of RXR in trans-activation of the RAR beta 2 promoter is to serve as an auxiliary factor required for the binding of RAR which, in turn, is directly responsible for transcriptional activity.

Porphyrin studies in TCDD-exposed workers.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has been shown to inhibit uroporphyrinogen decarboxylase activity resulting in chronic hepatic porphyria. From a cross-sectional study of 170 workers in chemical industry 68 showed elevated coproporphyrin levels, interpreted as secondary coproporphyrinuria. Three persons suffered from chronic hepatic porphyria in subclinical stages. None of the workers showed an overt porphyria cutanea tarda. A low-grade zinc protoporphyrinemia was observed in three persons. Forty-three of the 170 workers were evaluable for investigating the effect of TCDD on porphyrin levels. No significant correlation was found between TCDD concentration in adipose tissue and the level of uroporphyrin and coproporphyrin. The influence of a chloracne history is described.

Site-specific methionine sulfoxide formation is the structural basis of chromatographic heterogeneity of apolipoproteins A-I, C-II, and C-III.

ApoA-I and apoC-II are eluted in two isoforms and apoC-III2 is eluted in three isoforms by reversed phase high performance liquid chromatography (HPLC). The structural basis of these nongenetic heterogeneities was unravelled using HPLC of proteolytic peptides and time-of-flight secondary ion mass spectrometry (TOF-SIMS). In apoA-I, the chromatographic microheterogeneity was caused by the formation of methionine sulfoxides (MetSO). However, only residues Met112 and Met148 were found oxidized, whereas Met86 was unaffected and also resistant towards artificial oxidation. To assess whether and to what extent amino acid substitutions in apoA-I might affect methionine sulfoxidation, the tryptic peptides of 13 different mutant apoA-I proteins from 24 heterozygous apoA-I variant carriers were analyzed by HPLC. In normal apoA-I, the ratios MetSO112/Met112 and MetSO148/Met148 were highly variable. By contrast, the relative ratio of oxidation of methionine residues 112 and 148 was constant. The amino acid changes Lys107----Met, Lys107----O, Glu139----Gly, Glu147----Val, and Pro165----Arg resulted in the preferential oxidation of Met112, and Asp103----Asn resulted in a preferential oxidation of Met148; whereas Pro3----Arg, Pro3----His, Pro4----Arg, Asp89----Glu, Ala158----Asp, Glu198----Lys, and Asp213----Gly had no impact. ApoC-II and apoC-III isoforms differed by the oxidation of the two methionine residues in these proteins. Whereas in apoC-II both methionine residues were oxidized in parallel, in apoC-III the two methionine residues differed in their susceptibility towards oxidation. We conclude that the formation of MetSO depends on the molecular microenvironment within a protein.

Apolipoprotein C-III(Lys58----Glu). Identification of an apolipoprotein C-III variant in a family with hyperalphalipoproteinemia.

Apolipoprotein C-III is a major protein constituent of triglyceride rich lipoproteins and HDL. It occurs in plasma in three isoforms differing by their sialic acid content. Apo C-III putatively inhibits lipolysis and the apo E mediated hepatic uptake of remnants from triglyceride rich particles. We identified a heterozygous carrier of an apolipoprotein C-III variant by the presence of additional bands after isoelectric focusing (IEF) of VLDL. Structural analysis of the variant protein by HPLC, time-of-flight secondary ion mass spectrometry, and automated gas phase sequencing revealed a lysine to glutamic acid replacement in position 58. The underlying A to G exchange was verified by direct sequencing subsequent to amplification by polymerase chain reaction of exon 4 of the apo C-III gene. Family studies revealed vertical transmission of this defect. The two variant carriers exhibited plasma concentrations of HDL cholesterol and apo A-I above the 95th percentiles of sex matched controls whereas the unaffected father and sister showed normal values. The plasma concentrations of apo C-III in the two variant carriers were decreased by 30-40% compared with those of the two unaffected family members and to random controls. Using two-dimensional immunoelectrophoresis as well as IEF and subsequent scanning densitometry, we found that the low serum concentration of apo C-III was a consequence of diminished concentrations of the variant apo C-III isoproteins in both VLDL (15% of normal) and HDL (25% of normal). Apo C-III(Lys58----Glu) heterozygotes possessed unusual HDL as demonstrated by nondenaturing gradient gel electrophoresis. They consisted mainly of HDL2b and contained a proportion of atypically large particles, enriched in apo E, with a Stokes diameter of 13-18 nm and resembling HDLc. In conclusion, heterozygosity for a structural apo C-III variant--apo C-III(Lys58----Glu)--was identified in two hyperalphalipoproteinemic subjects characterized by the presence of low plasma apo C-III concentrations and atypically large HDL.

[Barrett's syndrome (author's transl)]. / Barrett-Syndrom. Ein endoskopisch nicht so seltener Befund.

Barrett's syndrome is a rare serious ulcerative inflammation of the middle or distal parts of esophagus, characterized by heterotopic gastric mucosa within the esophagus. In 4 cases, diagnosis of Barrett's syndrome suspected by means of x-ray examination was confirmed endoscopically and by taking biopsies under vision. The etiology of the disease is not clear. Alcoholism is common. In our patients, conservative treatment has been used without a great deal of benefit. Only by resection of the parts of esophagus lined by columnar epithelium a beneficial result can be expected, this surgical procedure certainly being a high risk.

Factors influencing survival of patients with permanent cardiac pacemakers.

With the advent of permanent cardiac pacing, technology has produced power sources that will last two, four, and even 20 years. The cost of these devices is proportional to their complexity and expected life. Therefore, the selection of an appropriate pulse generator should be based on the patient's prognosis. An actuarial analysis of 319 consecutive patients receiving permanent cardiac pacemakers was performed in order to determine whether prepacing factors influenced prognosis. The survival probability at 5 years was not influenced by sex, race, type of conduction defect, or antecedent disease. Survival rate decreased only slightly for each decade of life, but even in the ninth decade there was an acceptable prognosis.

[Size comparison of poly (A)-RNA from polysomes and from nuclei of the yeast Saccharomyces cerevisiae (author's transl)]. / Grössenvergleich der Poly (A)-RNA aud Polysomen und aus Kernen bei der Hefe Saccharomyces cerevisiae

The size of poly (A)-RNA from polysomes and cell nuclei of the yeast Saccharomyces cerevisiae was investigated. Pulslabelled cells ([14C] adenin) were cracked by the French press; polysomes and nuclei were separately isolated and the RNA was finally extracted with phenol. The separation of poly (A)-containing and poly (A)-lacking fractions was achieved by oligo (dT) cellulose. These fractions were characterized by sedimentation analysis. The main portion of polysomal poly (A)-RNA sedimented with a rate of 8 to 14S, whereas the poly (A)-RNA of nuclei exhibited a sedimentation rate of 12 to 17S. Thus nuclear poly (A)-RNA is about 20-30% larger than polysomal poly (A)-RNA.

[Characterization of messenger ribonucleoproteins from yeasts (author's transl)]. / Charakterisierung von Messenger-Ribonukleoproteiden aus Hefezellen

A method to produce in 15 min stable sphaeroplasts from yeast of the early log-phase is described. This method was used to isolate polysomes, after specific radioactive labelling of mRNA. The mRNP *-particles, isolated from polysomes, have sedimentation-coefficients from 10 to 80S and a density of 1.4 g/cm3, the polysomes a density of 1.58 g/cm3 and the ribosomes a density of 1.6. g/cm3.