The role of white cells in the transmission of Yersinia enterocolitica in blood components. The mechanism for the transmission of Yersinia enterocolitica in blood components has been studied experimentally. One hypothesis is that, during a Yersinia infection in the blood donor, bacteria are phagocytosed by white cells (WBCs), but are not killed. After collection of blood from such a donor and component production, the bacteria are present in WBCs for some time, during which the unit appears sterile. Later, when the WBCs disintegrate, the bacteria are released and multiply in the unit. Aliquots of whole blood and buffy coat were inoculated with 100 colony-forming units (CFU) per mL of a Y. enterocolitica strain of type O:3 and left at room temperature for 5 hours. Some aliquots were then WBC-reduced by filtration, while others retained their WBC contents. All aliquots were kept at 4 degrees C for 6 weeks. Meat extract broth culture medium was used as a control. Growth in the range of 2000 CFU per mL was obtained in the broth control by 24 hours, whereas the whole blood and buffy coat units appeared sterile for the first days of storage. After 1 week, a trace of bacteria and, after 4 weeks, massive growth were found in the WBC-containing units but not in the WBC-reduced units. The likely explanation is that the bacteria had been phagocytosed by the WBCs and were thereby hidden and not available for bacterial culture during the first phase of storage. When the WBCs spontaneously disintegrated, bacteria were released and multiplied in the blood units.(ABSTRACT TRUNCATED AT 250 WORDS)


Prevention of growth of Yersinia enterocolitica in blood by polyester fiber filtration. The ability of polyester white cell-reduction blood filters to prevent the growth of Yersinia enterocolitica in units of donated blood was studied. Sixteen units of freshly drawn blood were inoculated with 10, 50, 100, or 150 colony-forming units (CFU) per mL of a clinical isolate of Y. enterocolitica (serotype O:3). The units were subsequently fractionated into red cell concentrate and resuspended in AS-1 or AS-3 solution. One-half of the red cell concentrates in each solution were filtered within 15 hours of phlebotomy and stored for 42 days. The remaining units served as unfiltered controls. Bacterial growth was monitored by weekly cultures and, on the last storage day, by the presence of endotoxin and the formation of methemoglobin. One hundred twelve primary cultures (560 plates) were performed. Units collected in AS-1 and filtered remained sterile when initially inoculated with 50 CFU or less. Filtered units spiked with 100 CFU or less and collected in AS-3 remained sterile throughout their shelf life. All unfiltered units supported bacterial growth and the formation of endotoxin and methemoglobin. The filtration of freshly donated blood proves to limit the growth of Y. enterocolitica in red cell components.


Removal of Yersinia enterocolitica from AS-1 red cells. The growth of Yersinia enterocolitica in AS-1 red cells was investigated so as to study the organism's proliferation kinetics and to evaluate the effect of prestorage white cell (WBC) reduction on bacterial multiplication. Twenty-four 2-unit pools of ABO-compatible whole blood were prepared and inoculated with Y. enterocolitica to final concentrations ranging from 0.3 to 132 organisms per mL. After inoculation, pools were split equally, AS-1 red cells were prepared, and 1 unit of each pair (test unit) was WBC-reduced with a WBC-reduction filter. Quantitative bacterial cultures of both WBC-reduced and control units were performed at several points throughout preparation and storage. Less than 10 percent of the inoculated organisms was recovered from blood samples taken after a 7-hour room-temperature holding period. By the end of 42 days of storage, Y. enterocolitica was recovered from unfiltered red cells in 2 of the 6 units inoculated at the lowest levels (0.3 and 0.7 organism/mL), from 8 of the 12 units inoculated at the intermediate levels (2.8, 5.2, 30.7, and 43 organisms/mL) and from 6 of the 6 units inoculated at the highest levels (98.8 and 132 organisms/mL). Positive cultures were seen as early as Day 7. In contrast, filtered units inoculated at all levels less than or equal to 98.8 organisms per mL (21/21 units) were sterile at the end of the 42-day storage period, while 2 units (2/3) inoculated at 132 organisms per mL showed growth despite filtration.(ABSTRACT TRUNCATED AT 250 WORDS)


Growth of Yersinia species in artificially contaminated blood bags. Transfusion blood bags were inoculated with less than 0.1 colony-forming units (CFU) of Yersinia enterocolitica (serotypes O:3, O:5, and O:9) and Yersinia pseudotuberculosis I per mL and stored at 4 degrees C for 41 days. During storage, samples were collected periodically and colony counts of serial dilutions were performed by plating on tryptic soy agar. After a lag-phase, the growth of Yersinia species from less than 0.1 CFU per mL to 10(9) CFU per mL was observed in 3 of 10 bags. The data showed that prolonged storage of contaminated blood bags allows Y. enterocolitica and Y. pseudotuberculosis to multiply to high levels.


[Protein synthesis during acclimation of cold-blooded animals to various temperatures]. It was shown that the intensity of protein synthesis in cells of frogs, acclimated to 5 degrees C, is maintained at a high level, which is only 1.5-2 folds lower than that in animals acclimated to 20 C. In the process of acclimation to cold the intensity of synthesis decreases rapidly and already after 5 hours comprises one half of the value, which is characteristic of "warm" frogs, and the intensity of the process decreases more rapidly than the temperature of organs. On acclimation to warmth the intensity of protein synthesis increases and is getting stabilized at the level, characteristic of "warm" amphibia in 10-15 hours. It was shown that under various temperature conditions or conditions of acclimation specific proteins were synthesized against a background of the main groups of proteins.


[Heat inactivation of tissue thromboplastin]. Molecular mechanism of thermal inactivation of thromboplastin from the human brain using 1H, 31P-NMR has been studied. It was concluded that heating causes denaturation of protein moiety, disturbs hexagonal (H11) form of packing, immobilizes the lipids and decreases the mobility of their molecular segments. The dynamic properties of lipids become more homogeneous.


[Isolation and various properties of alpha-aminocaprolactam hydrolase from Klebsiella aerogenes]. L-alpha-aminocaprolactam hydrolase possessing the L lysine amidase activity was isolated from Klebsiella aerogenes and purified. The procedure of enzymes purification included cell destruction on USDN-I, fractionation by ammonium sulfate, gel chromatography on G-200. The preparation of the purified enzyme possessed specific activity of 50 mumol of lysin per 1 mg of protein per hour. Km was 2.6 mM in case of phosphate buffer (ph 7.2) for I-alpha-aminocaprolactam. Besides L-alpha-aminocaprolactam the enzyme hydrolyses lysine amide, leucine amide tryptophanamide. Magnesium ions are necessary for manifestation of catalytic activity of the enzyme.


Effect of androsole acetate on blood pressure of stress-sensitive hypertensive rats. The basal and stress-induced blood pressure was found to decrease in rats with inherited stress sensitive hypertension (SSHR) followed by androzole acetate administration orally (30 mg/kg body weight) for eight days. The basal levels of arterial blood pressure was decreased by 17.5% and that of stress-induced animals--by 19%.


[Analysis of hepatocyte cell membrane antigens in regenerating rat liver]. Antigens of plasma membranes in hepatocytes from regenerating rat liver were studied. Immunochemical investigation with polyvalent rabbits antiserum against plasma membrane proteins in hepatocytes from regenerating and normal rat liver have shown that liver regeneration processes are accompanied by the increase of proteins number with molecular weight of--80 kDa, 62 kDa, 40 kDa and 27 kDa. It is not excluded that protein with molecular weight of 27 kDa is the tissue-specific peripheral protein. The influence of antibodies against proteins of hepatocytes plasmatic membranes on histostructure of pathologically changed liver tissue has been studied. The data obtained testify to a possibility of participation of the above mentioned proteins in the regulation of rat liver regeneration processes.


[Modulating effect of surfactant on protein biosynthesis in lung macrophages]. Effect of lung surfactant on protein biosynthesis in macrophages at their cultivation in vitro has been studied. The surfactant is shown to stimulate protein synthesis in the cells. This phenomenon is probably stipulated by the increase of the transcription activity and formation of 45-pre rRNA, life of RNA. The surfactant is supposed to regulate the function of lung macrophages in norm and under pathology.


[Antiradical properties and antioxidant activity of ecdysterone]. Antiradical properties and total antioxidative activity of ecdysterone (10(-6)-10(-3)M were found on the model systems--photochemical one and that containing phosphatidylcholine liposomes oxidated with Fe2+ and Fe3+. The antioxidative activity of ecdysterone was shown to be comparable with that of the known inhibitors of peroxide oxidation of lipids--diethylparaphenylendiamine and ethylendiamine tetraacetate.


[Acid-base status of the blood and adenine nucleotide metabolism in ruminant erythrocytes during early post-natal development]. It has been established that data of acid-base balance in cows' and newborn calves' blood closely correlate with physiological status of animals during their initial days of postnatal ontogenesis and depend on the peculiarities of metabolism in fetus and newborn organisms. Data on the acid-base balance in blood of newborn calves under diarrhea syndrome are discussed from the point of view of the influence of rehydration therapy and "Namacit" preparation on clinical status of animals.


[Significance of adenylic acid catabolites for mouse thymus regeneration]. The distribution of [8-14C]adenylic acid catabolites in mouse thymocytes in cortisone-resistant and total thymocyte population has been studied. The accumulation of labelled catabolites in a form of hypoxanthine was found preferentially in cortisone resistant thymocytes but not in total population. This accumulation considerably grows after incubation of cortisone resistant thymocytes with non-peptide mitogenic factor. The excretion of labelled hypoxanthine into medium was also observed. In order to investigate a significance of hypoxanthine accumulation cortisone resistant thymocytes were incubated in the presence of hypoxanthine range concentration and [14C]thymidine incorporation into thymocyte DNA was determined. It was found that [14C]thymidine incorporation into thymocyte DNA increases after incubation in presence of 0.5-5.0 micrograms of hypoxanthine or 0.0005-5.0 micrograms of uric acid. It has been concluded that stimulation of [14C]thymidine incorporation and thymocyte proliferation by non-peptide mitogenic factor is caused by hypoxanthine accumulation.


[Enzymatic activity of chymopsin of various origin]. A number of enzymatic properties of fish pylochymopsin and bull chymopsin have been studied. Hydrolysis of synthetic ethers of N-benzoyl-L-tyrosine and N-benzoyl-L-arginine by these chymopsins depending at the time and concentration of preparations has been studied. It was found that bull chymopsin is the most active one. It was shown that concentrations of 2 to 6 micrograms/ml of bull chymopsin and of 15 to 20 micrograms/ml of fish enzyme were optimal for synthetic substrate BTME hydrolysis. The significant trypsin activity was revealed in the both preparations on a number of synthetic amides. In contrast to the bull chymopsin the treatment of fish pylochymopsin by TPCK did not completely remove the chymotryptic activity of pylochymopsin. It was shown that tryptic activity in the both preparations was completely removed with TLCK. The time and concentration dependence of the autolysis in both chymopsins has been studied. It should be noted that this process is negligible for fish pylochymopsin in contrast to bull chymopsin. Stabilization of both proteases in aqueous solution at room temperature has been studied. Stabilization of the chymopsins in solution is achieved by the addition of various protein preparations including casein and serum albumin. The degree of stabilization by these proteins was achieved at 2% concentration.


[Study of the interaction of triethylenethiophosphamide with nucleotides by mass spectrometry with ionization by fission fragments of californium-252]. Study of interaction of the antitumor alkylating drug triethylenethiophosphoramide (thioTEPA) with nucleotides (dGMP and dCMP) suggests highly perspective employment of 252-Cf fission fragment induced desorption mass spectrometry (252-Cf PDMS) in biochemical pharmacology. Using the 252-Cf PDMS the molecular masses of the unstable, unvolatile, high-molecular substances of biological origin and the chemical adducts or complexes with drugs can be used to establish some structural-functional parameters of the above mentioned biomolecules and their derivatives in microvolumes of the incubation medium. The resulting data may be used for modelling chemotherapeutic processes of "drug-biomolecule-target" type. Using 252-Cf PDMS the complexes (dGMP (thioTEPA) n), n = 1, 2, 3 and (dCMP (thioTEPA) n), n = 1, were obtained. Some quantitative parameters and stability of these complexes were studied. Binding of dGMP with drug in the presence of dCMP was shown preferential. The data are compatible with the predictions concerning the mechanism of the antitumor property of the thioTEPA which can be manifested in the impairment structure of DNA of the malignant cells.


Molecular analysis of simian varicella virus DNA. Clinical and pathological studies indicate that simian varicella virus (SVV) infection in primates is the counterpart of human varicella zoster virus (VZV) infection. The SVV and VZV genomes are also similar in size and structure. To extend studies of SVV DNA, we analyzed virus DNA from African green monkey kidney cells infected with the Delta-herpes-virus strain of SVV. The infectivity of SVV DNA was 88 PFU/micrograms. The buoyant density of SVV DNA, determined by isopycnic banding in CsCl gradients, was 1.700 +/- 0.002 g/ml, corresponding to a G + C molar ratio of 40.8%. The size of SVV DNA, estimated by analysis of restriction endonuclease digestion products and pulsed-field gel electrophoresis was 125.1 and 124.9 kbp, respectively. Electron microscopy of SVV DNA revealed a long region of 110.0 kbp, a unique short (Us) region of 5.1 kbp, and inverted repeat regions of 7.5 kbp flanking the Us. An EcoRI map of SVV DNA revealed two fragments not previously reported; our complete Pstl map also shows some differences. Mapping of SVV DNA with an additional restriction enzyme, measurement of full-length SVV DNA molecules, and the first use of pulsed-field electrophoresis to size SVV DNA, confirm and extend Gray's recent finding that SVV DNA has the same size and molecular configuration as VZV. We also show for the first time that the density of SVV DNA is similar to that of VZV DNA and that SVV DNA is infectious.


[Stimulation of replicative DNA synthesis by eukaryotic proteins bound to single-stranded DNA]. The paper deals with the effect of the single-strand (ss) DNA-binding proteins (SSB-proteins) from the Ehrlich ascites tumor (EAT) cells and from the eggs of silkworm, as well as the mouse serum blood proteins, having preferential affinity to ss DNA, on the DNA replicative synthesis in the EAT cells permeable for the macromolecules, and, for the silkworm proteins and on the DNA replicative synthesis in the nuclei from the eggs of silkworm proteins and on the DNA replicative synthesis in the nuclei from the eggs of silkworm permeable for macromolecules. SSB-proteins of EAT to considerable extent stimulated the DNA synthesis. At the same time, the other proteins (from the silkworm and from the serum) activated the DNA synthesis in the permeable cells to the less extent. It was found that SSB-proteins from the silkworm had a 1.5-13 fold stimulating effect on the DNA replicative synthesis in the homologous system (in the permeable nuclei). If the permeability for the macromolecules of the cells and nuclei treatment with Triton X-100 may be different, it is supposed that the activation of the DNA synthesis by the exogenous proteins depends on the homologous system of the DNA replicative complex. It is possible that the effect of the serum proteins on the DNA synthesis is connected with the masking of the ss regions of DNA which inhibited DNA-polymerase alpha. Perhaps the mechanisms of the activation of the DNA replicative synthesis by the proteins in vitro with the purified DNA polymerase alpha and in vivo are of different nature and are conditioned by homology of the deoxyribonucleoproteins.


Canine distemper virus increases procoagulant activity of macrophages. Inflammatory demyelination in canine distemper has been proposed to be due to a "bystander" mechanism, in which macrophages play an important role. In the present work we studied whether infection of macrophages by canine distemper virus (CDV) results in changes of macrophage functions, including Fc receptor-dependent and -independent phagocytosis, release of reactive oxygen species (ROS), and procoagulant activity (PCA). As a source of macrophages, dog bone marrow cells were seeded in teflon bags and grown for 1-2 weeks, at which time a marked enrichment of macrophages was noted. These cells were infected with the A75/17 strain of CDV. We could not detect any significant difference between uninfected and CDV-infected macrophages with respect to Fc receptor-dependent or -independent phagocytosis or with respect to the release of ROS. However, from Day 4 p.i. to the end of our observation period (10 days p.i.), PCA was up to 10-fold higher in CDV-infected unstimulated macrophage cultures than in uninfected unstimulated cultures of the same age. Increase in PCA was not due to the inoculation procedure by itself nor to components of the inoculum other than CDV; in particular, PCA was not due to contaminating endotoxin. Thus, several important macrophage functions do not appear to be impaired by CDV infection. The marked increase of macrophage PCA expression suggests that certain macrophage functions may even be enhanced as a result of infection. Such macrophage activation might contribute to the pathogenesis of the disease.


[Kinetic characteristics of creatine kinase and hexokinase from rat skeletal muscles during dietary deficit of vitamin K and administration of pelentane]. Kinetic parameters of rat creatine kinase isozymes at different vitamin K supply and treatment with antivitamin K--pelentan have been determined. MM-isozyme (skeletal muscle) has selective sensitivity to the vitamin K deficit, while BB and MB-isozymes (brain, kidney and heart) have not. The value KM for ATP of MM-isozymes increases, while maximal activity decreases. Pelentan treatment does not lead to the change of MM-creatine kinase affinity to ATP. Soluble hexokinase of skeletal muscle in rats with vitamin K deficiency and treated with pelentan has higher affinity to glucose as compared to normal rat enzyme. It has been supposed that skeletal muscle hexokinase exists in a particular molecular form under vitamin K deficiency.


Bacteriophage phi 6 envelope elucidated by chemical cross-linking, immunodetection, and cryoelectron microscopy. Bacteriophage phi 6 is an enveloped dsRNA virus which infects the plant pathogenic Pseudomonas syringae bacterium. Using low dose cryoelectron microscopy we show that the nucleocapsid, spikeless virion, and intact virion have radii of 29, 35, and 43 nm, respectively. Thus, the membrane is 6 nm thick and the surface spikes of the receptor binding protein P3 extend 8 nm from the membrane surface. Cross-linking, immunological, and complementation evidence suggest that the spikes are formed of multimeric P3 molecules and that P3 is associated with membrane-bound protein P6. We observe that the envelope can accommodate up to 400 molecules of P3 but that the average virion contains less than one-fourth of this amount. Assembly of a very small number of P3 or truncated P3 molecules onto inactive virions restores infectivity, showing that only a few spikes are necessary for receptor binding and membrane fusion.


[Passive Ca2+ influx into vesicles of the sarcoplasmic reticulum, modified by succinic anhydride]. Treatment of the sarcoplasmic reticulum (SR) vesicles with succinic anhydride in concentration of 1-2 mM modifies about 20% of amino groups. It increases initial rate and changes the pH-dependence of the passive influx of Ca2+ into vesicles and does not affect either Ca(2+)-binding or maximal passive Ca(2+)-loading of the SR vesicles. It is supposed that this effect may be caused by modification of the Ca-channel gating behaviour as a result of replacement of positive surface amino groups by carboxyl groups.


[Study of immobilization and properties of urease for creation of a biosensor based on semiconductor structures]. Many-sided investigations of urease immobilization methods were carried out to create the biosensor devices on the base of semiconductor structures. Special attention was concentrated on the biomembrane formation by means of urease and bovine serum albumin (BSA) cross-linking by gaseous glutaraldehyde. Optimal conditions for the formation process were selected which preserve about 20% of total urease activity after the cross-linking. The properties of enzyme immobilized by the above-mentioned method have been comprehensively studied. They included the urease activity dependence on pH, ionic strength, incubation buffer capacity as well as the enzyme stability during its functioning, storing and thermoinactivation. As was shown, for immobilized ureas Km value for urea at pH 7.0 and 20 degrees C is 1.65 time less than for free enzyme. In the presence of EDTA (1 mM) the enzyme activity in the biomembrane is practically unchanged under a month storing. Biomembrane possesses good adhesion to silicon surface and its swelling level under different conditions does not exceed 35%. The conclusion is made about the prospects of the used method of biomembrane formation for biosensor technology based on semiconductor structures.


Ugandan HIV-1 V3 loop sequences closely related to the U.S./European consensus. The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope protein is an important determinant for virus neutralization and cell tropism. V3 loop sequences from uncultured lymphocytes obtained in 1990 from 22 Ugandan HIV-1-infected patients could, with the exception of two patients' sequences, be divided into two groups (A and B) on the basis the V3 loop size and sequence. The V3 loop consensus sequences from both groups showed a high degree of homology to a U.S./European consensus, a characteristic also reflected by the results of peptide serology. In the case of group B the difference in sequence was only five amino acids. In contrast, the V3-flanking regions for both groups showed greater homology to an earlier (1986/1987) Ugandan consensus. The discovery of these two new Ugandan V3 loop genotypes, which are closely related to the U.S./European consensus, has implications for the understanding of the evolution of HIV-1 and for the future design of a vaccine for use in Africa.


[Peroxide modification of membranes and isomorphic composition of cytochrome P-450 of rat liver microsomes during antioxidant deficiency]. Lipid peroxidation (LPO), physico-chemical properties of the membranes and isoformic composition of microsomal cytochrome P-450 from the rat liver were studied under conditions of antioxidant insufficiency (AOI) which was modelled by exclusion of alpha-tocopherol from the animals' ration. An insignificant accumulation of microsomal diene conjugates and schiff bases against a sharp increase of the ability to the prooxidant stimulated LPO in vitro took place. A significant decrease of membrane lipid microviscosity and a change in surface properties of microsomal membranes of rats with AOI was determined. Absence of alpha-tocopherol in the ration was accompanied by a significant change in the content of separate isoforms of cytochrome P-450 exhibited in growth of a polypeptide with m. w. 54 kDa and the lowering of proteins with m. w. 48 and 50 kDa. Less intensive quenching of tryptophan fluorescence by acrylamide was also revealed, which testified to a lower accessibility of the quencher to membrane proteins or their fluorophore sites. Modification of lipid composition and of physicochemical properties of the rat liver membrane microsomes which was observed at AOI was significantly correlated by pretreatment with the antioxidant 4-methyl-2,6-ditretbutylphenol (ionol).


Three regions of cauliflower mosaic virus strain W260 are involved in systemic infection of solanaceous hosts. We have identified regions of CaMV strain W260 involved in systemic infection of Nicotiana bigelovii and Datura stramonium by constructing chimeric viruses between W260 and CM1841, a strain that is unable to systemically infect any solanaceous host. All of the chimeric viruses systemically infected turnips, demonstrating the viability of the chimeric viruses in a host that is susceptible to both CM1841 and W260. Three regions of W260, containing primarily genes I, IV, and VI, influenced the ability of that virus to induce systemic symptoms in the solanaceous hosts. The involvement of the regions containing gene I, and to a lesser extent gene IV, were affected by environmental conditions. When infected plants were grown under conditions of low light, low temperatures (18 degrees), and short days (9.5-hr day), the source of genes I and IV no longer influenced whether a chimeric virus moved systemically. As light intensity and day length were increased, the genetic requirements became more stringent and genes I and IV, as well as gene VI, had to be derived from W260.


[The role of transaminases in realizing the protective effect of L-aspartate in hypoxia]. Activation of aspartate aminotransferase and alanine aminotransferase of mitochondria introduced to the incubation medium of pyridoxal-5'-phosphate (40 microM) is approximately 2 times higher than that of the corresponding cytoplasmic forms. At hypoxia aspartate aminotransferase activity in mitochondria and postmitochondrial supernatant tends to an increase while that of alanine aminotransferase decreases (above 2 times). The protection from hypoxic damage when using L-aspartate (100 mg/kg subcutaneously 3-5 min before hypoxia) intensifies an adaptive increase of aspartate aminotransferase activity and removes a decrease of alanine aminotransferase activity. Under these conditions stimulating effect of pyridoxal-5'-phosphate on transaminases activity in vitro weakens. A simultaneous administration of vitamin-coenzyme complex (thiamine pyrophosphate, lipoate, sodium 4-phospho-pantothenate, flavin-mononucleotide, nicotinate) intensifies these metabolic shifts and protective action of L-aspartate.


A synthetic early promoter from a baculovirus: roles of the TATA box and conserved start site CAGT sequence in basal levels of transcription. Many baculovirus early genes and insect genes transcribed by RNA polymerase II have a conserved transcription start site sequence (CAGT) located downstream of a consensus TATA box. To examine the functions and interactions of these two motifs in initiating accurately positioned basal transcription, a 43-nt synthetic promoter was synthesized from the TATA box and start site sequences of the gp64 early promoter from the Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus (OpMNPV). The synthetic promoter initiated accurately and was also transactivated by the baculovirus transcriptional activator, IE1. To determine the roles of sequences within the 43-nt synthetic promoter, a series of linker-scanning and spacing mutations were analyzed for transcriptional activity, start site selection, and transactivation. Linker-scanning mutations were examined in vivo by transient expression and reporter gene assays. To examine transcription start site selection, promoter constructs were used for in vitro transcription in nuclear extracts from uninfected Spodoptera frugiperda (Sf9) cells. In vivo and in vitro analyses show that the TATA box, and not the start site CAGT, is the primary element controlling start site selection. Substitution of the conserved start site CAGT sequence resulted in a reduction of both reporter gene activity and in vitro transcripts, although transcripts initiated accurately. Data from linker-scanning and spacing mutations indicate that the conserved start site CAGT sequences are not required for accurate initiation but sequences at the start site play an important role in initiation efficiency.


Sequence and analysis of the capsid protein of Nudaurelia capensis omega virus, an insect virus with T = 4 icosahedral symmetry. We have determined the nucleotide sequence of the RNA2 segment of the Nudaurelia capensis omega virus genome. It was found to consist of 2448 nucleotides and contained one long open reading frame (ORF) encoding the 644 residue capsid protein. The deduced amino acid sequence of this protein reveals a positively charged amino terminus, a characteristic exhibited by several other viral capsid proteins, that is thought to be important for interactions between the capsid and the genomic RNA. There are 366 and 150 bases of untranslated sequence on the 5' and 3' ends, respectively. The ORF encoding the capsid protein initiates at the second AUG from the 5' end. The 5' proximal AUG specifies a short ORF (30 codons) which terminates 1 base before the initiation codon for the coat protein. Our analysis also revealed the presence of a second, previously unidentified polypeptide associated with Nudaurelia capensis omega virus particles. The amino terminal sequence of this protein corresponds to a portion of the long ORF beginning at codon 571. The lack of an initiation codon near this sequence indicates that the small polypeptide is most likely produced as a carboxy terminal cleavage product from a 70-kDa capsid protein precursor, yielding the previously identified 62-kDa protein and the 8-kDa protein that we have observed. The putative cleavage site would be at an Asn/Phe pair, somewhat resembling known cleavage sites (Asn/Ala) in the T = 3 Nodaviridae. In addition, we have found that there is also a second polypeptide similar in size to that from Nudaurelia capensis omega virus associated with particles of Nudaurelia capensis beta virus, the type member of Tetraviridae.


A cellular promoter-based expression cassette for generating recombinant baculoviruses directing rapid expression of passenger genes in infected insects. We have developed an expression cassette which allows the generation of recombinant baculoviruses that can express passenger genes under the control of a constitutive cellular promoter derived from the cytoplasmic actin gene of the silkmoth Bombyx mori. Silkmoth tissue culture cells which were infected with a recombinant B. mori nuclear polyhedrosis virus (BmNPV) containing the gene-encoding chloramphenicol acetyltransferase (CAT) under the control of this expression cassette expressed significant CAT activity beginning 5 hr postinfection (p.i.). Cells infected with a recombinant BmNPV containing the cat gene under the control of the polyhedrin gene promoter did not express CAT activity until 20 hr p.i. Silkworm larvae were also infected with the two recombinant viruses by hemocelic injections and all larval tissues examined were found to express the cat gene. While significant actin-cassette-driven CAT expression in vivo was first seen at 24 hr p.i., expression from the polyhedrin promoter was not seen until 48 hr p.i. By 60 hr p.i., tissues of larvae infected with the recombinant virus expressing cat under polyhedrin promoter control were found to exhibit sixfold higher CAT activity than those infected with recombinant virus expressing the cat gene under the control of the actin promoter. The 24-hr temporal advantage in expression of a passenger gene in infected larvae indicates that the actin-promoter-based expression cassette or other analogous cellular promoter-based cassettes could be used for generating recombinant baculovirus insecticides which could incapacitate pest insects more quickly than viruses employing the polyhedrin or other late viral promoters for expressing insect-incapacitating proteins.


Cloning and nucleotide sequence of the 5' part of v-myb cDNA. cDNA of a subgenomic v-myb mRNA from AMV-transformed BM-2 cells was cloned. Sequencing of the 5' end of this cDNA revealed the structure of both AMV leader and the splice junction in v-myb message. The leader is a novel variant of known avian retrovirus leaders. The long open reading frame in the cloned cDNA starts with six gag-derived codons spliced to the myb-specific sequence.


Effects of site-directed mutagenesis on the presumed catalytic triad and substrate-binding pocket of grapevine fanleaf nepovirus 24-kDa proteinase. Grapevine fanleaf nepovirus (GFLV) has a bipartite plus-sense RNA genome. Its structural and functional proteins originate from polyprotein maturation by at least one virus-encoded proteinase. Here we describe the cloning of the 24-kDa proteinase cistron located between the virus-linked protein (VPg) and the RNA-dependent RNA polymerase cistron in GFLV RNA1 (nucleotides 3966 to 4622). Proteinase expressed from this clone is able to cleave GFLV polyprotein P2 in order to produce the coat protein and a 66-kDa protein which is further processed to the 38-kDa presumed movement protein. The GFLV 24-kDa proteinase sequence contains sequence similarities with other nepovirus and comovirus proteinases, particularly at the level of the conserved domains corresponding to the hypothetical catalytic triad and to the substrate-binding pocket (amino acids 192 to 200). Site-directed mutagenesis of residues His43, Glu87, and Leu197 abolished proteinase activity. Inactivation of the enzyme is also observed if the catalytic residue Cys179 was substituted by isoleucine, but replacement by a serine at the same position produced a mutant with an activity identical to that of native proteinase. All our data show that GFLV cysteine proteinase presents structure similarities to the proteinases of cowpea mosaic virus and potyviruses but is most closely related to trypsin.


Feasibility of a predeposit autologous blood donation program in colorectal cancer patients: results from a randomized clinical study. The hematologic and transfusion data of a multicenter randomized trial investigating the effect of blood transfusions on the 5-year survival were used to study the feasibility of an autologous blood donation program in colorectal cancer patients. Three hundred and ten patients were randomized for autologous blood transfusions (predeposition of 2 units) or homologous blood transfusions, and transfusion rules were standardized. The Hb level in the patients who donated blood decreased by 20.1 +/- 1.3 g/l (mean +/- SEM) preoperatively and 4.5 +/- 1.8 g/l postoperatively, and in controls 3.7 +/- 1.1 g/l and 16.5 +/- 1.9 g/l (significantly different between the two groups, both pre- and postoperatively: p less than 0.01). Because blood loss and number of transfusions were similar in both groups, this indicated that either preoperative or postoperative erythropoiesis is stronger in patients who had donated blood. Twenty-three percent of the autologous patients and 61% of the homologous patients were exposed to homologous blood. The effectiveness of the procedure differed per tumor localization. In patients with a right-sided colon carcinoma, 22% of the control patients needed homologous blood, compared to 10% of the autologous patients. In patients with other colon carcinomas, this was 52 and 16%, respectively, and in patients with a rectal carcinoma 85 and 41%. We conclude that predeposition of 2 units of blood for colorectal cancer surgery is feasible and useful to prevent homologous blood usage in a significant number of patients with left colon carcinoma or rectal carcinoma.


A case of 'auto-immune thrombocytopenic purpura' serologically similar to post-transfusion purpura. The report describes a HPA-1a (Zwa)-negative woman with thrombocytopenia and antibodies in serum and eluate from autologous platelets with an operational anti-HPA-1a specificity. The results of the serological investigations were similar to the findings in most patients suffering from post-transfusion purpura. However, the present patient had no history of blood transfusion prior to the unset of purpura and the thrombocytopenia had persisted 6 months before splenectomy.


Quantitation of the human component C4: definition of C4 Q0 alleles and C4A duplications. We determined the C4 plasma concentrations of 48 genotypically CA4- and C4B-defined unrelated individuals from 34 families with a total of 196 members by an enzyme-linked immunosorbent assay using Rg:1,2 (C4A) and Ch:1 (C4B) specific monoclonal antibodies. The results obtained allowed the establishment of rules for the detection of C4 Q0 alleles in the heterozygous form and of C4A gene duplications. In the present study seven homoduplications of the C4A 3 allotype were defined which had not been detected by allotyping. This procedure allows the simple, reliable, and quick determination of Rg:1,2 and Ch:1 plasma levels which are not influenced by daily rhythms of C4 production.


Haemolytic disease of the newborn due to anti-Dia and incidence of the Dia antigen in Poland. A strong complement-fixing incomplete anti-Dia was found in the serum of a Polish mother who gave birth to a newborn suffering from a severe haemolytic anaemia. The whole family was of Polish origin. 9,661 donor blood samples from different regions of Poland were tested against antiserum derived from that mother. Dia antigen was present on red cells of 45 (0.46%) individuals. All of them were of Polish origin. In some cases, family studies were undertaken.


Aluminum content of source plasma and sodium citrate anticoagulant. The concentration of aluminum was determined in samples of source plasma collected by the normal plasmapheresis procedure, which involves collection in anticoagulant and immediate freezing. Samples of sodium citrate anticoagulant used in the collection of source plasma were also tested for aluminum, as were empty source plasma containers and 0.9% sodium chloride infusion (USP). Samples of source plasma were collected from a geographic cross-section of the donor population in the USA by three different manufacturers. Aliquots of these samples were mixed with Triton X-100 and sulfuric acid and analyzed for aluminum by atomic absorption spectrometry using electrothermal atomization (graphite furnace) and Zeeman background correction. The arithmetic mean and standard deviation for the aluminum content of 28 samples of source plasma were found to be 25.5 +/- 8.4 ng Al/ml. The aluminum content of the individual samples of source plasma ranged from 12 to 48 ng Al/ml. The aluminum content of 6 samples from two manufacturers of the sodium citrate anticoagulant that is used in source plasma ranged from 410 to 2,080 ng/ml. Aluminum levels found in saline for infusion and nitric acid leachates from empty source plasma containers were less than 6.9 ng/ml. The level of aluminum expected in uncontaminated human blood has been estimated to be 10 ng Al/ml or less. Comparison of this figure with the present data indicates that the sodium citrate anticoagulant contributes significantly to the aluminum load of source plasma and, therefore, to the aluminum content of products such as albumin derived from source plasma.


pH, temperature and lactate production in human red blood cells: implications for blood storage and glycolytic control. The interaction of temperature and pH in biological systems comprises two components. Temperature change may perturb the pH of solutions, and it may change the pKa of some ionizable groups that are involved in enzyme catalysis. The pH optima of single reactions and whole pathways are therefore temperature sensitive. The pH optimum of glycolysis in human red cells has been investigated only at 37 degrees C. We have measured the effect of temperature on the pH of stored blood suspensions and on the pH optimum of glycolysis in the human red cell. The pH of the cell suspensions in a traditional storage medium was 7.25 +/- 0.2 at 4 degrees C. The pH optimum of glycolysis was high (7.8-8.5) between 15 and 35 degrees C. It can be inferred from our data that human red cells are currently stored at least 0.5 pH units below the pH optimum of glycolysis at 4 degrees C. This suggestion is supported by storage experiments which showed that glycolysis at 4 degrees C was at least 1.5-fold more active at an initial pH of 7.67 versus 7.36. Equations which describe the variation in reaction velocity with pH were fitted to the pH curves for glycolysis in order to identify the ionizable groups that contribute to the effect of pH on glycolysis. It is generally accepted that hexokinase catalyses the rate-limiting step in glycolysis in the human red cell, but none of the ionizable groups implicated correspond to that involved in the hexokinase reaction.


Comparison of five different filters for the removal of leukocytes from red cell concentrates. The leukocyte depletion capacity and performance of 5 filters designed for filtration of red cell concentrates (RCC) were compared by counting leukocytes, measuring red cell volumes and by histological examination of the filters after use. To eliminate interdonor differences, 5 buffy-coat-poor RCC were pooled (in each of 10 experiments) and subsequently split up into the original bags. The RCC were passed over the Cellselect filter, a column filled with cellulose acetate, and over flat-bed polyester filters: the Cellselect Optima, the Pall RC 50, the Leukostop and the Sepacell R-500. The filtration was shortest with the Pall RC 50 (p less than 0.001 compared to the other 4 filters). Leukocyte removal was most effective with the cellulose acetate filter (p less than 0.01 compared to the other 4 filters) followed by the Cellselect Optima polyester filter (p less than 0.02 compared to the remaining 3 filters). Residual leukocytes did not exceed 50 x 10(6) for any brand of filter. Red cell recovery was similar for all 5 filters with mean values from 86.1 to 89.2%. The leukocyte numbers, counted in Türk's solution or in propidium iodide, gave comparable values in hemocytometers applying light microscopy or fluorescent microscopy, respectively. Histological examination showed that lymphocytes were mainly removed by trapping, whereas granulocytes showed a variable pattern: adhesion in presence of platelets or trapping.


Platelet concentrates stored in synthetic medium after filtration. Platelet concentrates prepared from pooled buffy coats (BCPC) were stored in Plasma-Lyte A, a glucose-free synthetic medium, after leukocyte depletion by filtration through Pall PL 50, and compared to paired unfiltered BCPC stored in the same medium. Each pair of BCPC units was prepared from a pool of 10 buffy coats split into two identical units. Platelet and leukocyte counts per unit of BCPC were 2.70 +/- 0.19 x 10(11) and 3.8 +/- 2.8 x 10(6) (filtered BCPC), 2.59 +/- 0.27 x 10(11) and 79 +/- 56 x 10(6) (control BCPC), respectively. Filtration procedures did not affect in vitro parameters of platelet quality and function such as osmotic reversal, ATP release and aggregation in response to collagen and ADP during 15-day storage. A similar decrease of platelet membrane glycoprotein Ib and a similar rise of activation markers GMP-140, gp 53 and platelet-bound fibrinogen were observed during storage of filtered and control BCPC. Our study indicates that storage of BCPC after filtration is feasible and that a reduction in leukocyte content by filtration to mean cell counts of less than five millions per unit has probably no effect on platelet storage lesion.


First autoclave-sterilized platelet-additive solution containing glucose with a physiological pH for the preparation of plasma-poor platelet concentrates. The glucose-free platelet-additive solution (termed AR solution), developed by Adams and Rock [Transfusion 1988;28:217-220], was modified by adding glucose as an energy substrate for platelets and maltose to prevent platelet lysis and by replacing sodium gluconate with sodium phosphate for better pH maintenance. The new platelet-additive solution (termed Seto solution) contained 90 mM NaCl, 5 mM KCl, 3 mM MgCl2, 17 mM tri-sodium citrate, 4.9 mM NaH2PO4, 20.1 mM Na2HPO4, 23 mM sodium acetate, 28.8 mM maltose, and 23.5 mM glucose with a pH of 7.4. The solution was sterilized by autoclaving in plastic bags in nitrogen to prevent glucose caramelization at high pH. Plasma-poor platelet concentrates prepared by adding Seto solution to the pelleted platelet buttons were stored in a LE-2 polyolefin bag at 22 degrees C with constant agitation for 5 days. The platelets suspended in Seto solution maintained oxygen consumption at a rate of 1.1 nmol/min/10(9) platelets after 5-day storage, with glucose consumption and lactate production rates of 0.5 +/- 0.2 and 1.2 +/- 0.2 nmol/min/10(9) platelets, respectively. This resulted in a final mean pH of 7.0. Those suspended in AR solution ceased glycolysis within 3 days because residual plasma glucose had been consumed. This was associated with decreases in percent hypotonic shock response and aggregation induced by adenosine diphosphate and collagen. Lactate dehydrogenase discharge in AR solution was 5 and 8 times higher at day 3 and day 5, respectively, than that of Seto solution. Morphologically, there were no ballooned platelets after storage in Seto solution.(ABSTRACT TRUNCATED AT 250 WORDS)


Euthanasia.

Euthanasia. The principles of self-determination and individual well-being support the use of voluntary euthanasia by those who do not have moral or professional objections to it. Opponents of this posture cite the ethical wrongness of the act itself and the folly of any public or legal policy permitting euthanasia. Positive consequences of making euthanasia legally permissible respect the autonomy of competent patients desiring it, expand the population of patients who can choose the option, and release the dying patient from otherwise prolonged suffering and agony. Potentially bad consequences of permitting euthanasia include the undermining of the "moral center" of medicine by allowing physicians to kill, the weakening of society's commitment to provide optimal care for dying patients, and, of greatest concern, the "slippery slope" argument. The evaluation of the arguments leads to support for euthanasia, with its performance not incompatible with a physician's professional commitment.


Observations on ethical problems and terminal care. Progress in medical diagnosis and therapy has raised new problems with far-reaching ethical implications. Medicine must remain a profession and not become a business. Textbooks must address ethical problems in the context of health care decisions and not restrict themselves to pathophysiology and practical therapeutics alone. The relative roles of the principles of autonomy, non-maleficence, beneficence, and justice must be balanced and appropriately applied to individual situations in biomedical ethics. When therapy becomes futile and the suffering of the patient does not justify any anticipated benefit, the patient (and/or patient surrogate) may request withholding or even withdrawing life-prolonging interventions. In the persistent vegetative state, even nutritional support by an unnatural (tube) route may ethically be denied at the patient's (or surrogate's) informed decision. New areas of ethical evaluation have been raised by the desire of some individuals to prolongation of their lives at high expense to the society such that other individuals are denied services because of limitation of available resources. There has been a long-standing conflict of interest between the acceptance by physicians and/or medical institutions of money or gifts from pharmaceutical companies whose drugs they prescribe, stock, or sell. This practice increases the cost of the drugs and is, in effect, a "sick tax," which is morally wrong.


[Social diagnosis: assessment scales and evaluation of "social adjustment" of psychiatric patients]. For diagnostic and prognostic considerations, rehabilitation and outcome evaluation in patients with mental disorders it proved to be indispensable to diagnose not only psychopathology but also social impairment. This study gives a review about the different instruments assessing social adjustment, and their application in out- and inpatient settings. Part of the instruments are based on statements of relevant informants, whereas others are based on observable behaviour. Hints are given to expense, objectivity of assessment and repeated measurements.


Nurse participation in decisions regarding limitation of treatment. A vast literature exists on the ethical aspects of decisions to limit life-sustaining treatments, and much of it deals with the way decisions ought to be made. Little is known, however, about how decisions are made in actual clinical practice. Empirical studies have not investigated the decision-making process directly and, with one exception, have only focused on physician practices. Through the use of a case, this paper examines the nature of hospital cancer nurses' involvement in the decision-making process. Three practice domains are identified: assisting patients to reach a truly autonomous choice, helping families to understand and to cope with the realities of the situation, and communicating with and encouraging open communication among all those involved. In addition, the potential value of the in-between position of the nurse is noted, and nurse responsibilities are summarized.


Spiritual aspects of death and dying.

Spiritual aspects of death and dying. Dying is an event beyond our comprehension, an experience that can only be imagined. Patients with cancer have a gift denied many others: some time to prepare for the approaching end of life. This time can be used to bring old conflicts to a close, to say goodbye and seek forgiveness from others, to express love and gratitude for the gifts of a life. Physicians can help patients by being aware of the spiritual dimensions to life that many patients have. In major religious traditions, death is accepted as the natural end of the gift of life and as a point of transition to another, yet unknown, existence. For many patients, it is not death that is feared, but abandonment. The physician's awareness of the spiritual needs of patients can make care of the dying more rewarding and fulfilling for all concerned.


[Eugen Bleuler (1857-1939) and psychoanalysis]. The relationship of Eugen Bleuler with psychoanalysis can be characterized as a history of overture and distance. In previous literature this relationship has been mainly studied from a historical and biographical point of view. In this article the authors review the history of Bleuler's attitude toward psychoanalysis from an ideengeschichtlich point of view. They distinguish and argue three periods. 1896-1906: the initial period was one of overt appreciation and positive rapprochement; 1906-1911: Bleulers initial positive attitude is more and more thwarted by fundamental theoretical disagreement and by his opposition against Freud's group building strategy; from 1912 on, Bleuler clearly dissociates himself from psychoanalytic theory and movement. An interpretation of this evolution is given in terms of personal and above all of theoretical incompatibility.


Clinical trials and physicians as double agents. Inherent in the dual role of physician-researcher is a conflict of interest arising out of the competing objectives of research and medical practice. Most commentary and policy recommendations on this conflict of interest have focused on the problems that arise in negotiations for informed consent. These are not, however, the only problems presented by this conflict; they are not necessarily even the most important. In order to deal with these problems, several commentators have suggested various procedural safeguards to protect the interests of the patient-subject--for example, separating the roles of physician and researcher, or introducing third parties into the relationship in order to assist in the initial or continuing negotiations for informed consent. In my view, the necessity for special procedural protections of patient-subject interests should be a discretionary judgment of the Institutional Review Board (IRB). In determining the need for special procedural protections for any research protocol, the IRB should consider three factors. To the extent that any one of these or a combination of two or more seems to present a problem, the IRB should consider it increasingly important to recommend special procedural protections: 1. There are serious impairments of the prospective subjects' capacities to consent. 2. The risk of physical or psychological injury presented by procedures done in the interests of research exceeds the threshold of "a minor increment above minimal risk." 3. The protocol is designed to introduce, test, evaluate, or compare therapeutic, diagnostic, or prophylactic maneuvers.


[Experimentally-induced stress in dyadic interactions. Presentation of the EISI (experimentally-induced stress in dyadic interactions) experiment]. In this article we present an experimental study (EISI-experiment: experimental induced stress in dyadic interaction), which focuses on a method for analysing individual and dyadic stress-experience and coping as well as its influence on dyadic interaction. The experiment offers a new diagnostic approach to analyse interaction processes under stress. We report the concept of our experimental design and show some first results. The findings suggest that our method is useful for the examination of stress phenomena in close relationships. It allows the investigation of interactional processes and in this context especially of "dyadic coping", a form of stress-management by couples.


[Looking into the past. Psychological and gerontologic aspects of memory]. Reminiscence is characterized by a dialectical relation of past and present which is of fundamental importance for personal identity. Moreover, it shows a double aspect, implying both the private recollection of one's past and the social dimension of narration, instruction and self-disclosure. This will be illustrated by a consideration of the different forms and functions of reminiscence in general, and by the results of research on the role of reminiscence in later life. An investigation into these aspects of reminiscence may also further a therapeutic approach to the crises resulting from unresolved conflicts in the life history.


Rationing health care: its impact and implications for hematology-oncology. Rationing of health care in the United States currently exists via the covert mechanism of restricting significant segments of medical care for many of those who cannot afford it. Provision of universal health care would necessitate explicit rationing of certain interventions and technologies, even though an individual could afford them. The British and Canadian experiences provide lessons from which America can profit, and the Oregon health plan is an experiment in this direction. The progressive "graying" of America has raised the question of the need for intergenerational charity as a form of rationing. The implications of these rationing plans would result in a major restructuring of the practice of hematology-oncology.


Cancer therapy: reimbursement of new therapeutic technologies. New drugs and technologies for cancer treatment are being developed at a rate that has created a reimbursement crisis. This article discusses third-party concerns about this problem and describes generic criteria that have proven to be useful in assessing any new technology. It is equally important to discontinue funding of ineffective and obsolete therapies as it is to devise a strategy for identifying and encouraging the development of new therapy that will be both clinically useful and cost-effective. Examples are provided to show that these are not necessarily mutually exclusive goals. Off-label application of standard therapy as well as the funding of new cancer therapy are considered. High-dose chemotherapy with autologous stem-cell support for treatment of a variety of neoplasms has become a major reimbursement challenge. Other technologies such as autolymphocyte therapy and use of colony-stimulating factors are considered in detail. Finally, a process for deciding how to fund new cancer therapy is described.


Childhood malignancies and decision making. Failure to obtain "adequate" medical care for a child constitutes child neglect, which may be used as the basis for prosecution of parents, removal of the child from the home, or court-ordered medical treatment. "Adequate" care is usually construed as that which is given by a licensed physician, but, in case of dispute, courts almost never engage in choosing one medical approach over another. The principle that parents may not refuse medical care, however, is made very difficult when children have malignancies--the long-term nature of the treatment means that, if the child is left at home, court order or not, the parents may flee with their child. Removing the child from the home, however, adds that trauma to the ill child's burdens. Questions should be asked before making a request to a court to order a therapy which will prolong but not save a child's life if the parents would prefer to spare their child the side effects. Parents, however, may always refuse to permit their child to participate in research studies, no matter how promising. Adolescents are increasingly believed to be capable of medical decision making; most courts, however, would not allow an adolescent to refuse life-saving treatment.


[Use of gestures to support speech comprehension of a hearing, nonverbal severely mentally handicapped adult with autistic behavior]. In the present single case study the reactions of a severely mentally handicapped adult with autistic behaviour to simple instructions were investigated; in each case with and without gestural facilitation. This procedure was varied both with regard to persons and to situations. The correct following of requests as well as special behaviour such as uncertainty, smiling, delay of reactions and enthusiasm was registered and analysed by videotape. It could be clearly proved that the requests which were supported by gestures were more often performed correctly. On top of that it could be observed that under these circumstances the handicapped person was less uncertain, showed fewer stereotypies and was generally more interested. No influence could be noticed on the delay of reaction as well as on the person's enthusiasm. No difference could be seen between different persons and situations. Further research projects should particularly analyse how far the use of gestures does not only serve more communication but also has an influence on the range of behavioural disorders.


[Heat-induced decomposition of disaccharide Amadori compounds in quasi-water-free reaction conditions]. The thermally induced decomposition of disaccharide Amadori compounds has been compared to those of monosaccharide ones under almost water-free conditions. The structure of the synthesized maltulosyl compound has been proved to be 4C1-alpha-D-glucopyranosyl- (1----4)-2C5-beta-D-fructopyranosylglycine by 1H- and 13C-NMR spectroscopy. The decomposition of Amadori compounds has been used to study the kinetics of the browning reaction. Compared to fructosylglycine and maltotriulosylglycine, the browning of the disaccharide is faster. Curie point pyrolysis at 300 degrees C and investigation of the pyrolysate by gas chromatography/mass spectrometry have shown that the disaccharide component influences the thermal process. Furanes and furanones have been detected as predominant degradation products, the main one being 2(5H)-furanone. For the first time, we suggest a reaction pathway for the formation of these products via the Maillard reaction which includes 1,6-anhydroglucose.


Coeliac active peptides from gliadin: large-scale preparation and characterization. Larger amounts of coeliac active peptides are required for pathogenetic investigations. Therefore, a simplified preparative procedure by means of gel-permeation chromatography and reversed-phase HPLC was developed for the isolation of the peptides B3141-B3146, which are present in peptic tryptic digests of gliadin [this journal (1983) 176:85-94]. The peptides are derived from the N-terminal part of alpha-gliadins and are closely related. The amino acid sequence of B3143 is VPVPQLQPQNPSQQQPQEQVPLVQQQQFPGQQQQFPPQQPYPQPQPFPSQQPYL. B3144 has proline instead of glutamine in position 34. The previously described peptide B3142 [this journal (1984) 179:371-376] corresponds to B3144 except for the missing C-terminal leucine.


Proteolytic activity of isolated lamb calpains on myofibrils under the conditions of pH, Ca2+ concentration and temperature existing in postmortem muscle. The two calcium-activated neutral proteinases (calpains I and II) and their specific inhibitor were isolated by ion exchange chromatography in DEAE-Sephacel from lamb skeletal muscle (longissimus dorsi). Their proteolytic activities were then determined using myofibrils as substrate. The Ca2+ requirements were different for each form of the enzyme: calpain I needed only 50 mumol Ca2+ for half-maximal activity, while the other isoenzyme, calpain II, needed 1,000 mumol Ca2+ for reaching 50% of its maximum activity. Both calpains showed a relevant activity in the pH range 5.5-6.5 (over 40% of maximum activity found at pH 7.5). With regard to the effect of temperature, both isoenzymes retained about 25% of their activity at 25 degrees C with a temperature reduction down to 4 degrees C. It is concluded that calpain I is an active protease under conditions similar to that prevalent in lamb meat during postmortem storage.


[Surgical treatment of esophageal cancer: Part I. European results 1980-1991]. During the past 10 years, postoperative mortality associated with resection of oesophageal carcinoma has been reduced from 30% to 11%. However, all efforts to improve longterm survival with extensive excisional procedures and adjuvant chemotherapy and radiotherapy have failed. Fifty-three of 100 patients presenting to the surgeon with an oesophageal carcinoma have resectable disease. Six of them will die from postoperative complications and 47 patients will be discharged from the hospital after an average of 3 weeks. Of these patients, 30 will survive the first, 17 th second, and 7 the fifth year. Although in may be possible to further reduce postoperative complications and mortality, the chances of improving the long-term prognosis of patients with oesophageal carcinoma seem small.


[Comparative analysis of mezlocillin/metronidazole and amoxicillin/clavulanic acid as "one shot" preventive antibiotic administration in colorectal interventions]. The prophylactic effect of intraoperative "one shot" antibiotic application in colorectal surgery was investigated. Patients were randomised and the antibiotic combination Mezlocillin/Metronidazole (group A) or Amoxicillin/Clavulamid acid (group B) was applicated in 160 patients. 111 patients were selected for the study (group A: 59; group B: 52 patients). In the postoperative course 53% (group A) and 67% (group B) of the patients developed bacterial infections. Abdominal wound healing was complicated by infection in 15% (A) and 12% (B) of the patients. A significant difference between the two groups could not be proven, 24% of all patients with documented intraoperative bacterial contamination and 10% of the patients with negative findings developed wound infections. In colorectal surgery patients are still at high risk for infectious complications. Applicated antibiotics should basically cover aerobic and anaerobic germs.


[Measurement of serum protein-bound hexose--an aid in the diagnosis and after-care of colorectal cancers?]. In a group of 80 patients with colorectal carcinoma and a control group of 103 persons Protein-bound Hexose (PHex) was measured in the serum. The level was, as reported previously by other authors, significantly elevated in the tumour group. The aim of this study was to compare the sensitivity of PHex with that of the oncofetal antigens CEA and CA 19/9. The sensitivity of PHex turned out to be clearly lower than that of CEA. It was about as high as the sensitivity of CA 19/9 in colorectal carcinoma. In view of these results there is no benefit to be expected from an introduction of the PHex determination into clinical practice.


[Injuries of the portal vein and vena cava in severe blunt abdominal trauma]. Between 1/87 and 7/90 54 severely traumatized patients underwent emergency laparotomy because of blunt abdominal trauma. In 55.5% of the cases one intraabdominal organ, in 20.4% two organs, in 16.6% three and in 7.4% four or more organs were injured. Additionally in 14.8% of these cases (n = 8) a major vascular injury (portal vein n = 5, vena cava n = 2, mesenteric root n = 1) was found intraoperatively. Injuries of the portal vein were always associated with complete rupture of the pancreas, requiring left pancreatic resection in 4 cases and a duodenum preserving resection of the pancreas head in one case. In 2 of these patients the portal vein had to be reconstructed with a prosthesis. Lethality was 9.3% for the whole group (n = 54) and 25% for the patients (n = 8) with additional vascular injuries.


[The importance of puncture cytology in diagnosis of benign and malignant breast changes]. In 500 mamma needle biopsies that were cytologically examined we found a precision of 87.4%, with 2.6% false negative, and 6.2% false positive results. In 19 cases a cytological diagnosis was not possible resulting in a sensitivity of 0.94 and a specificity of 0.86. False positive findings were due to misinterpretations of the cytological pictures, as were also false negative findings. In the latter cases sampling mistakes could also be the reason. Cytological diagnostic should only be made using triple diagnostic technique since the methods are complementary to each other in respect to their diagnostic security. Practical experience in the puncturing technique as well as an experienced cytologist are prerequisites using this method successfully as a diagnostic tool.


[Lobar restricted nodular form of pulmonary alveolar proteinosis]. A case report of a 53-year-old man with lobar restricted pulmonary alveolar proteinosis is presented. The way from an accidental X-ray examination until the final histological diagnosis of the specimen taken at operation is followed. Typical diagnostic pitfalls are discussed.


[Ender nailing versus dynamic hip screw--a comparison of early postoperative results]. Proximal fractures of femur may be alternatively treated by implantation of dynamic hip screws or Ender-nails. Estimation of the value of both measures is to be done by analysis of the early postoperative course. Full load bearing capability was obtained in 83% of all cases with Ender-nails as against 68% of all cases after implantation of hip screws. Duration of operation was clearly shorter and the need for blood transfusions was reduced by implantation of Ender-nails. The amount of specific technical complications was equal in both procedures. There were no differences in the postoperative ability to walk.


[Interdisciplinary treatment strategy of aneurysmal bone cyst]. In this report the management of an aneurysmal bone cyst in the cervicothoracic region of an 13-year-old girl is described. Aneurysmal bone cyst is a histologically benign lesion that is often extremely vascular. In the spine, it can cause extensive bone destruction and compress neural structures and this can result in high-grade tetraparesis. In the treatment a three-stage operation with radical total removal of the cyst from two posterior approaches and one anterior approach for osteosynthetic stabilisation gave an excellent result. The value of preoperative embolization to reduce tumor vascularity, allowing total excision without neurological deficits to the patient, is demonstrated.


[Holistic-psychosomatic aspects of surgical organ loss]. Surgical interventions in gynaecology are from a holistic, psychosomatic perspective equally essential interventions into the life history of the treated women. Using a review of international research literature dealing with coping-processes after hysterectomy and breast surgery, the author discusses possible psychological and psychopathological consequences of these operations. In addition the presents possibilities for improving as well the preoperative doctor-patient-dialogue as postoperative psychotherapeutic and body therapeutic strategies.


[Production of prostacyclin in human umbilical cord blood vessels. Effect of natural and synthetic estrogen]. This investigation was conducted to evaluate the effects of 17 beta-estradiol and 17 alpha-ethinylestradiol on prostacyclin PGI2-production in homogenates of human umbilical cord arteries and veins in the presence of exogenous arachidonic acid. The incubation of 17 beta-estradiol at 2 different concentrations (10(-8) and 10(-6) M) in the presence of arachidonic acid for various times (5 min up to 24 h) effected an increase in the production of PGI2 (measured as 6-keto-prostaglandin F1 alpha) of 36-47% in the artery and 16-21% in the vein over basal values (100%). Under the same experimental conditions with 17 alpha-ethinylestradiol we observed an increases of 30-40% in the artery and 1-18% in the vein compared with basal values. For both estrogens this was not significantly different compared with untreated samples. In our in vitro system, natural and synthetic estrogens had no clear effect on PGI2 production in homogenates of human umbilical vessels.


[Suitability of metric structural criteria in vaginal ultrasound assessment of the postmenopausal endometrium]. In 135 postmenopausal women ultrasonic findings of the endometrium were compared with the final histopathology. Object of this study was to evaluate the ability of metric structural criterions for a vaginosonographic screening of benign and malignant endometrial neoplasms. A sonographic assessment of the endometrium was possible in 97.8% of the patients; anatomic peculiarities such as calcified fibromyomata proved to be the main-impediments. By means of the criterions examined the sensitivity for the differentiation of inconspicuous and pathological findings was 100% with a 98.2% specificity; the rate of wrong-positive results was 1.6%. In 9.2% ultrasound was classified as conspicuous requiring further controls. According to these results the vaginosonographic assessment of the endometrium regarding its thickness and structure (homogeneity, echogeneity, median echo) can be recommended for a non-invasive screening for endometrial neoplasms.


[Cryosurgery of condylomata acuminata gigantea--a contribution to therapy]. Two cases of condylomata acuminata gigantea treated with cryosurgery are described. A clinical and histologic differentiation from Buschke-Loewenstein's tumours, sex-specific differences, possible malignant transformations and differential diagnostic diseases are described and references are made to confusions with condyloma-like, primary carcinomas. Compared to conservative surgical treatment of condylomata acuminata gigantea, cryosurgery offers the advantage of rapid necrosis of the condylomata which are repelled without any further surgical intervention and heal without scars. Therefore we consider cryosurgery a more refined technique in treatment of condylomata acuminata gigantea. In extensive involvement, however, cryosurgical measures must be repeated.


Comparison of Mycoplasma bovis strains based on SDS-PAGE and immunoblot protein patterns. Whole-cell protein patterns generated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were compared for 34 isolates of Mycoplasma bovis. A high degree of similarity between most of the strains was established with strain-to-strain differences mainly confined to quantitative variations of certain protein bands, particularly in the molecular weight regions of 64-68, 55 and 26 kD. Two of the isolates provided more deviating patterns. Hydrophobic membrane protein fractions of the strains as prepared by Triton X-114 phase partitioning were compared by SDS-PAGE, which confirmed some of the characteristic strain features found with whole-cell proteins. The immunoblot analysis revealed that up to 20 of the 50-55 discrete protein bands detected in SDS-PAGE patterns were recognized to be antigenic by rabbit and bovine hyperimmune sera. It is concluded that the same set of major antigens is present in all strains investigated, but amounts of individual constituents may be differing.


Relevance of sensitivity testings (MIC) of S. aureus to predict the antibacterial action in milk. Bacterial susceptibility testings were carried out in parallel Iso-sensitest broth (ISB) and bovine milk cultures using 16 antibacterials and 4 sensitive strains of mastitic isolates of Staphylococcus aureus. Bacterial activities were analyzed by continuous turbidity monitoring (broth cultures), continuous fluorometric monitoring of the resazurin-reducing redox activity, and by analyzing the triphenyltetrazolium (TTC)-reducing capacity at the end of the incubation period. To obtain an equipotent bacteria-suppressing activity, milk cultures required in general several times more antibiotic than the respective ISB cultures. Antibacterial activities of sulfadoxine-trimethoprim, vancomycin, novobiocin, macrolides, aminoglycosides and oxytetracycline were most effectively suppressed by milk. Aminoglycosides suffered additionally from reduction of oxygen in the incubation environment. The beta-lactams (penicillin G, oxacillin, cephalothin, ceftiofur, ampicillin, ampicillin-clavulanic acid), gentamicin and enrofloxacin showed extremely variable sensitivity results depending on the S. aureus/milk combination.


The early migration of Elaphostrongylus rangiferi in goats. The migration of nematodes of the genus Elaphostrongylus (Metastrongyloidea: Protostrongylidae) from the gut into the tissues of ruminants has not been described. Detailed histologic studies were performed on five goat kids that had received oral doses of 1,000-1,500 infective larvae of Elaphostrongylus rangiferi Mitskevich, 1960 and were subsequently autopsied at days 2, 4, 6, 10, and 14 post inoculation. The main migratory route of E. rangiferi larvae seemed to be haematogenous. The larvae penetrated the venules of the abomasum and followed the blood stream via the liver to the lungs. In the lungs, the larvae entered pulmonary venules and passed into the arterial circulation reaching the central nervous system (CNS) and other organs between days 6 and 10 post inoculation. Some of the larvae that had lodged in tissues outside the CNS probably migrated into it along the spinal nerves. A marked eosinophilia was present from day 8 post inoculation.


Evaluation of the API Coryne test system for identification of Actinomyces pyogenes. The present study was designed to evaluate the accuracy of the API Coryne test system for identification of Actinomyces pyogenes. The test system correctly identified 36 of 42 A. pyogenes and 4 of 5 comparatively studied Arcanobacterium haemolyticum-cultures. The biochemical profiles of the remaining 6 A. pyogenes- and 1 A. haemolyticum-cultures were not included in the analytical profile index. None of the cultures were misidentified. According to the API database (ATB Plus V 1.5.4.) the unidentified cultures could be correctly identified as A. pyogenes and A. haemolyticum respectively. A greater repertoire of A. pyogenes specific biochemical profiles incorporated into the analytical profile index would improve the applicability of this test system for veterinary diagnostics.


Evaluation of the effect of 3 probiotics on experimental Escherichia coli enterotoxaemia in weaned piglets. In this report the efficacy of 3 different probiotics (Bacillus cereus "toyoi", Lactobacillus spp. and Streptococcus faecium) was investigated. They were supplemented in the food of recently weaned piglets that were orally infected with E. coli O141 K85ab. Supplementation could not prevent mortality and clinical symptoms nor reduce the faecal excretion of hemolytic E. coli. The possible explanations for the unsatisfying results are various.


Opsonic activity in mammary secretion and serum of gilts during the lactation period. Opsonic activity was studied in mammary secretion and serum during lactation in four healthy gilts. The opsonic activity was determined as peak chemiluminescence and time to peak chemiluminescence in a luminol enhanced chemiluminescence assay. The peak chemiluminescence was significantly (p less than 0.05) higher in mammary secretion around parturition than later in lactation. No changes in the opsonic activity were seen in serum during lactation. Overall, the peak chemiluminescence and time to peak chemiluminescence were significantly higher (p less than 0.001) and shorter (p less than 0.05) respectively, in serum than in mammary secretion. The present study indicates that the opsonic activity is highest in mammary secretion around parturition but decreases later in lactation, and that this change is confined to the mammary gland.


Cytotoxicity of cyanobacterium Microcystis aeruginosa. Cytotoxic effects of crude extracts and fractions of the purification steps towards Microcystin-LR (MCYST-LR) were investigated in vitro. Cytotoxicity was evaluated by measure of lactate dehydrogenase liberation of Chang liver cells and by hemolysis. Crude extracts of strain PCC 7806 damaged the cells within a few minutes. In contrast, MCYST-LR did not show any detectable cytotoxic effects. The cytotoxic activity could be related to a heat-labile substance with a molecular weight of about 30,000 Da.


Haematology of experimental babesiosis and ehrlichiosis in steroid immunosuppressed horses. An investigation was carried out to study the haematology of steroid immunosuppressed horses experimentally infected with Babesia equi and Ehrlichia equi, separately or simultaneously. Horses infected with both pathogens showed less marked changes in their haematology than those inoculated with either pathogen separately. This appeared to result from early elimination of the more pathogenic Babesia as Ehrlichia spread through the granulocytes. The apparent suppression of Babesia by Ehrlichia is of field clinical importance and merits further investigation for its apparent useful potentials in the control of babesiosis in endemic areas.


Preparation and characterization of monoclonal antibodies against Mycoplasma bovis. Monoclonal antibodies (mabs) against Mycoplasma (M.) bovis were prepared for use in diagnosis of bovine mastitis. From the original 32 hybridomas actively secreting mabs against M. bovis, 6 stable lines were cloned. Two of them, Mb 5D8 and Mb 4F6, recognized M. bovis antigens of estimated molecular weights of 33 and 26 kDa, respectively. They showed no cross-reaction to other bovine mycoplasmas, thus rendering them useful for specific detection of this pathogen. All mabs investigated cross-reacted with M. agalactiae which is known to be closely related to M. bovis, but does not occur in cattle. Two other mabs, Mb 5D4 and Mb 1F6, exhibited further cross-reactions to a number of bovine mycoplasma species. Finally, mabs Mb 5D5 and Mb 2G5 reacted with all mycoplasmas tested. The possibility that they recognized constituents of the broth culture medium is discussed.


The isolation and identification of Leptospira interrogans serovar bratislava from a pig in Germany. During a study on the improvement of selective media for the isolation of leptospires from clinical material, leptospires were isolated from the kidneys of a pig. Test material was cultured in EMJH-medium containing 0.4% rabbit serum and 3 inhibitory substances (Rifampicin 10 micrograms/ml, Amphotericin B 2 micrograms/ml, 5-Fluorouracil 100 micrograms/ml). The cultured leptospira-strain (Berlin 406) was identified to serogroup level using the cross agglutination method and it was further typed to serovar level by MAB's, factor analysis and restriction endonuclease analysis (REA). MAB typing of Berlin 406 gave an agglutination pattern profile typical of serovar bratislava. Factor serum identification clearly identified this strain as serovar bratislava. On REA examination, Berlin 406 gave a profile identical to that of the serovar bratislava type strain Jez Bratislava i.e. genotype B1. This is the first report of a confirmed isolate of Leptospira interrogans serovar bratislava from domestic animals in Germany. There exist a number of possibly important implications as this agent is being incriminated in the cause of clinical disease not only in pigs, but also in horses and dogs in other countries.


The overwintering of Fasciola hepatica eggs under semi-arid and temperate Mediterranean climate (Madrid, Spain). In this experimental work, conducted over two consecutive years (1988 and 1989), we have analyzed the overwintering possibilities of Fasciola hepatica eggs deposited in winter in a semi-arid and temperate Mediterranean area (Madrid, Spain). Eggs did not survive for more than a week in faeces at environmental moisture conditions. In contrast, over a 75% of the eggs maintained in permanently wet faeces survived winter temperatures, showing no high mortality until important thermic increment from June. The success of overwintering was higher in eggs arrested in wet faeces than included in water (55% "versus" 20% of hatching in spring), and it was unrelated to the deposition time (excretion in early or late winter). The hatching period of overwintered eggs started in May, at the same time as those excreted in early spring. Therefore, overwintering of eggs will have an effect on the level but not on the start of the spring contamination with miracidia. The dry winters of this semi-arid area greatly restrict the overwintering eggs contamination to soils next to streams, ponds or irrigation channels.


Relationship between group B streptococcal serotypes and cell surface hydrophobicity. Cell surface hydrophobicities of streptococci of serological group B were determined by the adherence of the bacteria to hexadecane droplets. A significant adherence to hexadecane was observed with the group B streptococcal type reference strains Ib, V, Ic, R and X, but not with those of serotype Ia, II, III and IV. Cultivation of the bacteria in microcapsule-inducing media reduced the hexadecane adherence properties. The adherence to hexadecane was not related to fibrinogen binding properties of the cultures. Screening a large number of group B streptococci isolated from humans and bovines revealed that those with polysaccharide type antigen alone were generally hydrophilic, those with protein antigen alone or with protein antigen in combination with polysaccharide antigen were mostly hydrophobic. Cultivation of the bacteria under microaerobic conditions or after a single mouse passage enhanced microcapsule production and correspondingly reduced the hexadecane adherence values. Treatment of the bacteria by guanidinium chloride or by neuraminidase enhanced the hexadecane adherence. The hydrophobic component on group B streptococcal surface appeared to be partly inactivated by heat or proteolytic treatment of the bacteria.


Characterisation of Newcastle disease viruses isolated in India. Eleven Newcastle disease virus (NDV) isolates obtained from outbreaks of disease in chickens (9) and Japanese quail (2) in Tamil Nadu, India were characterised in pathogenicity tests, antigenically, using mouse monoclonal antibodies (MAbs), and other established tests devised to distinguish between different strains. All 11 isolates were shown to be highly virulent for chickens. In indirect immunoperoxidase tests used to assess the ability of a panel of 28 MAbs to bind to infected cell cultures, 10 of the isolates showed an identical reaction pattern, the other isolate (No. 4) failed to react with one MAb which bound to cells infected with the other isolates. Isolates 9 was unstable at pH 3 while the other 10 were stable. All other properties were shared by the 11 isolates.


Mechanism of impaired growth hormone secretion in patients with Cushing's disease. The function of the growth hormone-releasing hormone (GHRH)-growth hormone (GH) axis in Cushing's disease was studied by monitoring (a) the GH responses to GHRH loading and L-dopa loading, (b) the GHRH response to L-dopa loading, and (c) the daytime profiles of plasma GH concentration. GH release following GHRH and L-dopa was blunted in patients as compared to that in age-matched control subjects. However, GHRH release following L-dopa was similar in patients and controls. The plasma GH levels in four patients measured every 20 min by a highly sensitive immunoradiometric assay for GH showed pulsatile GH secretion at low levels during the observation period. These results indicate that GHRH release from the hypothalamus is preserved in patients with Cushing's disease, and support the hypothesis that glucocorticoid inhibits GH secretion by altering the hypothalamic somatostatin tone.


Alterations in circadian rhythm of serum thyrotropin in critically ill patients. To evaluate the 24-h pattern of serum thyrotropin (TSH) in critically ill patients, we measured serum concentrations of TSH in blood samples collected every 2 h for 24 h from nine patients (six with malignancy, two with liver cirrhosis, one with chronic renal failure), who had subnormal levels of both triiodothyronine (T3) and thyroxine (T4), in the absence of history, symptoms or signs of thyroid disease. Analysis of the data, performed using a second-order inferential statistical methodology for rhythmometry (cosinor method), demonstrated that critically ill patients still had daily oscillations of serum TSH which significantly adapted to the function approximating the circadian rhythms (R2 = 74.3%). However, the mean level (mesor) in the rhythm of the patients was found to be significantly lower than that of healthy subjects (0.96 vs 2.18 mU/l); the amplitude of rhythmical daily variations also was lower in patients than in healthy subjects (0.23 vs 0.56 mU/l), even though the amplitude/mesor ratio was similar (23% vs 26%). Lastly, the highest level in the TSH rhythm of the patients was found to be in the late afternoon, in contrast to healthy subjects, who had a TSH surge after midnight. Although these alterations are consistent with the existence of a dysregulation at suprahypophyseal level in critically ill patients, it remains to be established whether the state of low T3 and T4 may be ascribed to anomalous circadian rhythm of TSH.


Evaluation of the degradation of desamino1,D-arginine8-vasopressin by nasal mucosa. The nasal route is a convenient and simple way to administer peptides to humans. Absorption of the drug, however, requires passage of the substance through the nasal mucosa. This is a possible site of enzymatic degradation of the peptide. It is shown that rabbit nasal mucosa homogenates rapidly degrade the synthetic anti-diuretic hormone analogue desamino1,D-arginine8-vasopressin in vitro. The metabolite formed has been identified as des-(amino,arginine8,glycineamide9)- vasopressin, which is stable under the prevalent in vitro incubation conditions. It is proposed that this process is catalyzed by intracellular post-proline cleavage enzyme. Reversed phase chromatography in combination with immunological detection has been used to study the possible presence of this metabolite in the circulation after intranasal administration to humans. The metabolite des-(amino,arginine8,glycineamide9)-vasopressin could not be detected in plasma following intranasal administration, possibly indicating a paracellular absorption of desamino1,D-arginine8-vasopressin or absence of this enzymatic activity in humans.


Long-term stability of low insulin responses to glucose in non-diabetic subjects. We investigated the stability of the insulin response to glucose in healthy subjects by making retrospective comparisons of insulin responses after two 60 min glucose infusion tests performed many years apart. The subjects (N = 49) were divided into two lower and two higher quartiles as assessed by the incremental 0-10 min insulin area during the initial glucose infusion test. Ages were initially 32.3 +/- 2.8 years in lower quartiles and 26.6 +/- 1.1 in higher quartiles and body mass indexes 21.6 +/- 0.6 kg/m2 and 21.8 +/- 0.5, respectively. The interval between the first and second glucose infusion tests was 8.1 +/- 2.8 years for lower quartiles and 10.4 +/- 1.3 for higher quartiles. In lower quartiles, the 0-10 min insulin area at first testing was 157.1 +/- 15.9 mU/l x 10 min and at follow-up 202.2 +/- 26.6 (+29%, NS). In higher quartiles, the insulin area decreased from 654.8 +/- 70.6 mU/l x 10 min at first testing to 489.8 +/- 53.6 at follow-up (-25%, p less than 0.05). The 0-60 min glucose area did not change significantly between glucose infusion tests in lower quartiles (+5%), but did increase by 12% (p less than 0.005) in higher quartiles. Only one subject of the lowest quartile at first testing changed to higher quartiles at follow-up. Predictable "regression toward the mean" at follow-up was moderate, hence the individual insulin response to glucose was relatively stable with time. This finding is compatible with the hypothesis that genetic factors are of major importance for the insulin response to glucose.


Relationship between plasma growth hormone concentration and cellular sodium transport in acromegaly. We investigated the relationship between mean plasma growth hormone (GH) concentration and cellular sodium transport in untreated and treated acromegaly. Seventeen patients (age 55 +/- 3 years) with active acromegaly were studied with respect to plasma GH (mean of 24 h GH profile) and erythrocyte electrolyte content as well as transmembrane sodium transport. The patients were reinvestigated two weeks after successful surgery (N = 14) and again after one year (N = 13). Erythrocyte electrolytes were analyzed by flame photometry and sodium influx and efflux rate constant determined by in vitro incubation using a modified Keyne's formula. In patients with active acromegaly there was a significant positive correlation between IGF-1 and cellular sodium transport, while GH tended to show a negative relationship to the same parameter. After successful treatment, both IGF-1 and GH disclosed a positive relationship to cellular sodium transport. After one year, a significant increase in erythrocyte sodium content was seen in the patients compared to the preoperative situation. In conclusion, if this is a generalized phenomenon the results are compatible with a sodium-retaining effect of GH via stimulation of transmembrane sodium transport. In active acromegaly this may be counteracted by a sodium transport inhibitor giving the reverse relationship between GH and cellular sodium transport.


Immunoglobulin G of patients with circumscribed pretibial myxedema of Graves' disease stimulates proteoglycan synthesis in human skin fibroblasts in culture. Hyaluronic acid and proteoglycan accumulate in the affected skin of Graves' disease patients with pretibial myxedema (PTM). We aimed to determine whether an autoantibody IgG circulating in PTM patients stimulates proteoglycan synthesis in human skin fibroblasts, resulting in PTM. IgGs were purified from 14 normal subjects, 11 Graves' disease patients with PTM. 5 Graves' disease patients with active ophthalmopathy and 15 Graves' disease patients with neither PTM nor ophthalmopathy. Human skin fibroblasts were incubated with the IgGs and labeled with [35S]sulfate. The medium and cell layer were separated and the proteoglycan was extracted. The 35S radioactivity in the proteoglycan fraction was measured. Compared with normal IgGs or with those of Graves' disease without PTM or ophthalmopathy, PTM IgGs significantly increased the incorporation of the 35S into the proteoglycan. The effect of PTM IgG was dose-dependent. As PTM IgG did not alter degradation of the 35S labeled proteoglycan, an increase in 35S incorporation reflects increased synthesis. The effect was mediated through a mechanism other than adenylate cyclase activation. The present study demonstrates the presence of an autoantibody in PTM IgG that stimulates proteoglycan production through human skin fibroblasts. This is not correlated with the thyroid stimulating antibody activity. It is suggested that the activity of this antibody leads to the development of PTM.


Serum and plasma epidermal growth factor in thyroid disorders. Epidermal growth factor (EGF) is an important mitogen and its secretion in neonatal animals has been shown to be affected by thyroid hormone levels. EGF in blood of humans is found in both platelets (as reflected in its serum level) and in plasma; its origin in plasma remains unclear. Serum and plasma EGF were studied in a group of patients with thyroid disorders. Twenty hyperthyroid subjects (3M, 17F) aged 37.3 +/- 14.9 years and 10 hypothyroid patients (3M, 7F) aged 58.3 +/- 18.6 years were studied before and after euthyroidism was restored. Before treatment, serum EGF in the hyperthyroid patients was elevated compared to normal controls (501 +/- 376 vs 270 +/- 154 pmol/l, p less than 0.001). After treatment of hyperthyroidism, serum EGF returned to the normal levels (232 +/- 176 pmol/l). In contrast, serum EGF was not significantly different in the hypothyroid subjects either before or after treatment (151 +/- 194 and 237 +/- 153 pmol/l respectively). A significant correlation (r = 0.461, p less than 0.001) between serum EGF and serum-free thyroxine index (FTI) was found when all samples from both untreated and treated hyper- and hypothyroid patients were examined. Multiple regression analysis revealed that both serum FTI and platelet count independently affected the serum EGF levels. Similarly, plasma EGF was also elevated in untreated hyperthyroid patients with a median of 26.4 pmol/l (range less than 16.6-88.0), whereas all normal controls and hypothyroid subjects had unmeasurable levels.(ABSTRACT TRUNCATED AT 250 WORDS)


Cerebrospinal fluid and blood concentrations of luteinizing hormone, follicle stimulating hormone and prolactin following castration of adult male rats. Comparative levels of LH, FSH, and PRL in the serum and cerebrospinal fluid (CSF) of adult male rats were studied at different periods following castration. Intact and sham-operated animals served as controls. Blood and CSF were collected at 1, 3, 7, 14, 21, 28, 35 and 46 days following castration. The CSF was collected via cisterna-magna puncture, while the blood was collected from abdominal aorta. Serum gonadotropins increased progressively beginning day 1 post-castration to reach maximum by day 35 or 46 post-castration. Sham operation and castration did not affect mean CSF, LH and FSH levels compared to intact controls. Analysis of the temporal pattern of serum and CSF gonadotropin levels following castration revealed significant positive correlation between CSF and serum LH (r = 0.58) and FSH (r = 0.64) levels respectively. The data suggest that CSF gonadotropins may be derived from systemic circulation. Serum PRL levels were not affected by castration, but CSF PRL levels were significantly reduced at days 28, 35 and 46 post-castration compared to intact controls. CSF PRL levels showed negative correlation with serum LH and FSH levels but failed to show a correlation with serum PRL levels. Hypothalamic norepinephrine turnover rate increased at days 28, 35 and 46 post-castration. Hypothalamic dopamine contents and turnover rates were reduced at days 21 and 28 post-castration. It is suggested that CSF PRL may have a role in the regulation of serum gonadotropins.


Primary culture of prepubertal human testicular cells isolated from testes collected at necropsy. The aim of the present work was to develop a method for maintaining prepubertal human testicular cells in culture. Seven pairs of testes of boys who died of causes unrelated to endocrine or metabolic diseases were obtained at necropsy. Histology of the testes was normal. Testes were digested with collagenase and dispersed cells were seeded in multi-well dishes in the presence of 5% bovine fetal serum. After the first day, cells were cultured for five days in serum-free medium in the presence or absence of 918 pmol/l insulin. At the end of culture, microscopic examination showed healthy looking cells with characteristics compatible with pre-Sertoli cells; peritubular cells were identified by immunocytochemistry. In the presence of insulin, cells were able to secrete either testosterone or estradiol into the medium, as well as to reveal aromatase activity. In order to study the effect of the time elapsed between death and beginning of cultures, steroidogenic activity was related to this post mortem time. It was found that, in the presence of insulin, cells obtained from testes with less than 24 h of post mortem time secreted testosterone (64 +/- 7.2 pmol/10(6) cells.24 h, mean +/- SD) while cells obtained from testes with more than 24 h of post mortem time did not secrete testosterone. With long post mortem times, aromatase activity under insulin increased from non-detectable to 35 pmol/10(6) cells.24 h. Time course studies showed that cells with capacity to secrete testosterone increase this secretion gradually up to day 10 of culture, while those with detectable aromatase activity showed increments in this activity during the first week of culture.(ABSTRACT TRUNCATED AT 250 WORDS)