Melanins production from enkephalins by tyrosinase.

Leu-enkephalin and Met-enkephalin are oxidized in vitro by mushroom and sepia tyrosinase giving rise to synthetic melanins whose production is dependent on incubation time and on enzyme concentration. The Enk-melanins formed are acid-insoluble brownish or reddish pigments showing a continuous absorbance in the visible region when dissolved in basic solution. The presence of the amino acid chain makes them fully soluble in pH 7.4 0.05 M phosphate buffer and methanol.     

Use of the API rapid NFT system for identifying nonfermentative and fermentative marine bacteria.

Thirty-five American Type Culture Collection type strains of marine bacteria were used to evaluate the Rapid NFT system (API Analab Products, Plainview, N.Y.) for use in identifying heterotrophic marine bacteria. The 21 biochemical and assimilation tests on the Rapid NFT test strips were treated according to the manufacturer's protocol, which included use of AUX medium (provided with the Rapid NFT system) for preparing assimilation tests, and by substituting phenol red broth base (BBL Microbiology Systems, Cockeysville, Md.) with and without an oil overlay for the AUX medium. A seven-digit numerical profile was obtained for each NFT test strip from each of the three procedures and matched to its corresponding number in the Rapid NFT identification codebook. Also, all biochemical and assimilation test results were analyzed with SASTAXAN and SAS/GRAPH programs (SAS Institute, Inc., Cary, N.C.); similarity matrices were computed for all 35 strains. For comparison purposes, bacterial strains were grouped at a similarity level of 70%. The results indicated a low efficacy of identification for all three procedures. In addition, similarity matrix analysis showed more cohesive grouping based on results of phenol red broth base-treated strains than for the AUX medium provided by the manufacturer. However, none of the three treatments provided exclusive grouping of type strains at the genus level. Thus, the reliability of the data obtained from the NFT system and modifications thereof should be evaluated carefully when environmental isolates are characterized.     

Electromyographic changes in work-related myalgia of the trapezius muscle.

In 11 patients, all women, 21-55 years of age, with unilateral work-related myalgia of the trapezius muscle, the right and left trapezius muscles were examined simultaneously for electromyogram (EMG) signs of localized muscle fatigue. All patients were tested with 0-kg hand load for 5 min, holding the arms straight at 90 degrees of elevation in the scapular plane. Only 4 of the patients tolerated exposure to higher load levels. They were tested with 1 kg hand load for 3 min and 2 kg hand load for 2 min, with a period of rest of 30 min between the trials. The EMG mean power frequency (MPF) and root mean square (rms) were calculated. Data were normalized with the initial value as a reference and regression analyses were performed. On both sides a decrease of MPF and an increase of rms were found with increasing time and load, i.e. classical EMG signs of localized muscle fatigue. Compared with the nonaffected side smaller changes were found on the affected side, possibly due to pain inhibition, impaired microcirculation and biochemical changes along the muscle fibres. At 0-kg hand load we found no change of MPF on either side despite subjective feelings of fatigue and pain. We interpreted these findings as an indication of reduced capacity of the affected trapezius muscle to sustain static load with early development of pain-associated local fatigue.     

Hydrogen peroxide production by monoamine oxidase in isolated rat-brain mitochondria: its effect on glutathione levels and Ca2+ efflux

H2O2 production and accumulation during incubation of isolated rat-brain mitochondria with substrates of monoamine oxidase A and B were investigated. All substrates gave rise to an accumulation of H2O2 which was inhibited by malate + pyruvate or isocitrate, consistent with a need for mitochondrial NADPH to maintain glutathione in the reduced state. However, in the absence of these additions the level of reduced glutathione decreased only by about 30%, indicating that only a fraction of the mitochondrial glutathione pool was accessible to the glutathione peroxidase and glutathione reductase activities responsible for the continuous removal of H2O2 generated by monoamine oxidase. The H2O2 accumulation was also inhibited by externally added reduced glutathione or NADPH but not NADH. External NADPH was oxidized by added oxidized glutathione but not alpha-ketoglutarate + NH4+. These results suggest that the removal of H2O2 generated by monoamine oxidase proceeds by way of special fractions of glutathione peroxidase and glutathione reductase that are located in the intermembrane space of mitochondria in such a way that they can react with both intra- and extra-mitochondrial glutathione and NADPH, possibly at the contact sites between the inner and outer mitochondrial membranes. Evidence is also presented that H2O2 generated by monoamine oxidase enhances Ca2+ release from mitochondria and may thus function as a regulator of mitochondrial Ca2+ efflux.     

Renal uptake and excretion of epidermal growth factor from plasma in the rat

The rat excretes around 2 nmol epidermal growth factor (EGF) in the urine per 24 h. The urinary EGF might be derived from plasma and/or might be synthesized in the kidneys. We have used the rat to study the renal uptake and excretion of homologous EGF from plasma. I.v. injected 125I-EGF was removed from the circulation within a few minutes. 5 min after the injection, the kidneys contained 12% of the 125I-EGF. The kidneys seemed to degrade most of the 125I-EGF which they accumulated from blood, as only 4% of the injected label was excreted as intact 125I-EGF in the urine. The amount of endogenous EGF in plasma was under the detection limit of our enzyme-linked immunosorbent assay (0.03 nmol/l) and it remained so after bilateral nephrectomy. Even if plasma EGF was 0.03 nmol/l excretion of EGF from plasma could account for less than 5% of the urinary EGF. This study shows that the kidneys are able to accumulate EGF from plasma and excrete a part of it as intact EGF in the urine. However, excretion of immunoreactive EGF from plasma can only account for a minor part of the urinary EGF.     

A review of techniques and results obtained in one laboratory by an integrated system of methods designed for routine clinical flow cytometric DNA analysis

Establishing flow cytometric DNA analysis as a clinical routine procedure requires adequate and proven guidelines, by which the data can be obtained and interpreted to directly influence management of the individual patient with a specific neoplasm. The present paper is intended as a contribution to such guidelines, of which only fragments are available today. We have previously described a system of methods, designed for routine flow cytometric DNA analysis. In the present status report our experience, based on approximately 18,000 samples (clinical and experimental) is summarised. Sample acquisition with fine-needle aspiration, storage at -80 degrees C, internal standardization by chicken (CRBC) and trout red blood cells (TRBC), staining with propidium iodide (PI), and analysis in the flow cytometer is recapitulated, with emphasis on previously unpublished aspects. The method of statistical analysis which has an integrating role is described in some detail. A lack of linearity between channel number and DNA content was determined experimentally, and the coefficient of variation (CV) was found to decrease with increasing channel number. The corrections in the algorithm of deconvolution made necessary by these findings are fundamental for estimating the end results. The zero point adjustment and procedures for changing from one batch of standards to another are described. A systematic approach to interpretation of DNA histograms is attempted and illustrated by data from clinical specimens of malignant lymphoma, breast cancer, small cell lung cancer, cancer of the oral cavity, and bladder cancer. Some problems are still unsolved and visual inspection is required to determine if the quality of the individual histogram is satisfactory. Inspection of the fluorescence/light scatter dot-plot provides additional information for the recognition of artifacts. The results stress that good quality DNA histograms with as small CVs as possible are important for interpretation of the data. It is essential that statistical methods are employed to extract the key end-point results. These are the number of subpopulations and their relative representation, and for each subpopulation the DNA index (DI) and the fractions of cells in the cell cycle phases. For the DNA data to have any rationally based impact on clinical decision making, it must be demonstrated that they have an independent prognostic value. Strategies for final evaluation are discussed. Multicenter trials on fresh material, to accrue quickly the number of patients necessary for firm conclusions, are suggested.     

Dynamics of chemiluminescence response of Syrian hamster blood neutrophils to the growth of 10 different strains of tumor cells]

The production of H2O2 by blood neutrophils of Syrian hamsters, bearing 10 different transplanted tumors was studied at different stages of tumor growth by the luminol-dependent chemiluminescence (CL) test. It was demonstrated that during the tumors growth, two types of the blood neutrophils CL reaction can be registered. The first type of reaction represents very early and significant decrease of spontaneous CL of blood neutrophils, already evident before the appearance of palpable tumor nodules in animals, bearing in vivo selected cell strains. Subsequent subcutaneous tumor appearance in the animals was followed by increased CL of neutrophils. The second type of reaction was characteristic for in vitro transformed cells never selected in vivo. In this case the increase of CL of blood neutrophils at early stages of tumor growth was followed with the decrease of this activity at the period of active tumor growth. Possible relation of this reactions to the survival and growth of different tumor cells in vivo is discussed.