Reduction in tumor necrosis factor receptor affinity and cytotoxicity by glucocorticoids

Micromolar concentrations of glucocorticoids rendered L-M cells (a murine tumorigenic fibroblast line) less sensitive to the cytotoxic activity of murine TNF. The potency of different steroids paralleled their known anti-inflammatory potency, and pretreatment was more effective than post treatment. Sex steroids and mineralocorticoids were ineffective. Dexamethasone also decreased the sensitivity of MCF-7 (a human mammary carcinoma line) to the cytotoxic activity of human recombinant TNF. Pretreatment of both cell lines reduced the affinity of specific cell surface receptors for the binding of their species 125I-TNF about 3-fold while retaining the same number of binding sites. The decrease in sensitivity was not due solely to the inhibition of early TNF-induced events (such as binding, internalization or signal transduction). Dexamethasone modestly enhanced inhibition beyond that of neutralizing antiserum alone when both were added midway in the L-M killing reaction (after receptor down regulation but before the onset of complete cell death).     

Preparation of Oligonucleotides and Their Use in Molecular Weight Estimations

Abstract

Yeast RNA was used to prepare oligonucleotides employed to calibrate a G-50 Sephadex column. The oligonucleotides' preparation, isolation, desalting and characterization is described. Data obtained by chromatography of the oligonucleotides demonstrate that the molecular weights of oligonucleotides can be easily determined by interpolation using plots of elution volumes (Ve) versus molecular weights (M). Errors greater than 20% are obtained if the conventional plot of Ve-Vo/Vs versus log M is used (where Vo is the void volume of the column and Vs is the volume of the column occupied by the inert phase, the 6-50 Sephadex).     

Comparison of porcine liver tRNA preparations: Purification of tRNA and its separation from RNA-peptidyl complexes

Six fractions of soluble RNA were obtained from phenol extracts of porcine liver and were tested for their acceptance of 14 amino acids under aminoacylation conditions and for their effects on the aminoacylation of tRNA. Two of the fractions contained appreciable amounts of tRNA, and three of the fractions affected the aminoacylation of tRNA. Based on these observations a revised method of tRNA preparation was developed that includes essentially all the tRNA in one fraction but that excludes the RNA-peptidyl complexes. The revised method is rapid and convenient and provides better quality tRNA than three alternate methods to which it is compared.     

A comparison of the palaeolimnology of Peipsi and Võrtsjärv: connected shallow lakes in north-eastern Europe for the twentieth century, especially in relation to eutrophication progression and water-level fluctuations

We applied a multi-proxy palaeolimnological approach to provide insights into the natural variability and human-mediated trends of two interconnected temperate large shallow lakes, Peipsi and Võrtsjärv, during the twentieth century. The history of the lakes was assessed on the basis of age-related changes in the sediment main constituents (water, organic matter and carbonate), sub-fossil pigments, diatom assemblages and organic matter dissolved in pore water. The temporal changes in the palaeodata indicate an increase of the in-lake biological production in both lakes from about the 1960s, suggesting enhanced nutrient inputs. In subsequent decades, the gradual increase of autochthonous organic matter becomes more obvious, indicating progressive eutrophication of the lakes. Palaeolimnological indicators from the sediment record of Lake Peipsi indicate a slight recession of the lake’s eutrophication in the 1990s but not for Lake Võrtsjärv. The results of the study also suggest that after the lakes became eutrophied, the climatically induced water-level fluctuations ceased to be the main driver determining the abundance of phytoplankton. Responses of the lakes to human-induced impacts are better recorded in the sediments of Lake Peipsi than in those of Lake Võrtsjärv, which is shallower of the two and where the wave-induced resuspension of deposits markedly smooths or erases the signals of environmental changes. The results of the investigation expand the knowledge on how large shallow lakes respond to human-mediated and natural perturbations, including those in the lake catchment areas and the capability of the lakes to store the chronology and sequence of these changes.     

Development of a Novel Strategy to Isolate Lipophilic Allergens (Oleosins) from Peanuts

Background

Peanut allergy is one of the most severe class I food allergies with increasing prevalence. Especially lipophilic allergens, such as oleosins, were found to be associated with severe symptoms, but are usually underrepresented in diagnostic extracts. Therefore, this study focused on isolation, molecular characterization and assessment of the allergenicity of peanut oleosins.

Methods and Results

A comprehensive method adapted for the isolation of peanut oil bodies of high purity was developed comprising a stepwise removal of seed storage proteins from oil bodies. Further separation of the oil body constituents, including the allergens Ara h 10, Ara h 11, the presumed allergen oleosin 3 and additional oleosin variants was achieved by a single run on a preparative electrophoresis cell. Protein identification realized by N-terminal sequencing, peptide mass fingerprinting and homology search revealed the presence of oleosins, steroleosins and a caleosin. Immunoblot analysis with sera of peanut-allergic individuals illustrated the IgE-binding capacity of peanut-derived oleosins.

Conclusion

Our method is a novel way to isolate all known immunologically distinct peanut oleosins simultaneously. Moreover, we were able to provide evidence for the allergenicity of oleosins and thus identified peanut oleosins as probable candidates for component-resolved allergy diagnosis.