Natural zeolite clinoptilolite increases the concentrations of sphingoid bases in the yeast Yarrowia lipolytica

In the present paper, we studied the effect of natural zeolite clinoptilolite on sphingolipid metabolism in the yeast Yarrowia lipolytica. We also investigated if zeolite addition had any impact on cell shape and size, as well as on the pH alterations during the culture growth. High performance liquid chromatography analysis of sphingoid bases obtained by acid hydrolysis of complex sphingolipids from Y. lipolytica showed that their concentrations markedly rose upon the zeolite addition. The largest increase among the identified molecular species of sphingoid bases was seen in C18 phytosphingosine, whose levels rose 6.2-fold and 22.3-fold after culturing cells for 24 and 36 hours respectively in the presence of finely ground zeolite. pH measurements of the culture medium showed a similarity between pH profiles of control and zeolite-supplemented cells, suggesting that ion-exchange capacity was not probably responsible for the observed change in sphingolipid metabolism. Scanning electron microscopy revealed that zeolite affected cell size and shape. Y. lipolytica cells grown in the absence of zeolite were oval-shaped with an average cell size of 0.7-2.7 microns, whereas when cultured with zeolite, they were round-shaped and larger, having an average cell size of 1.3-2.9 microns.     

Mechanisms responsible for delayed and immediate hemolytic transfusion reactions in a patient with anti-E + Jk(b)+ Di(b) and anti-HLA alloantibodies

Immediate hemolytic transfusion reactions (IHTR) occurred in the course of delayed hemolytic transfusion reactions (DHTR). An 84-year-old man had received a blood transfusion 20 years ago. Progressive anemia developed, because of continuous bleeding from a bladder tumor. He was transfused with concentrated red blood cells (CRC) which were Rh-E antigen negative, because he had anti-E antibodies (day 0). He received CRC on day 3, and underwent resection of bladder tumor on day 6. Although crossmatch-compatible CRCs were prepared for the operation, those were not required and were kept in a refrigerator in the ward. On day 9, when a CRC kept in the ward was transfused, he suddenly had a IHTR. In order to analyze a mechanism of IHTR, the anti-Jk(b) and anti-Di(b) antibodies, anti-HLA antibodies and the concentrations of inflammatory cytokines were measured in serum samples. The anti-Jk(b) and anti-Di(b) antibodies increased prior to IHTR experienced on day 9. The concentrations of IL-6 and IL-1beta increased from day 2, while the concentration of IL-8 increased from day 7. The anti-HLA class I antibody could be detected 2 days before IHTR. Thus, the anti-Jk(b) and anti-Di(b) antibodies induced the production of inflammatory cytokines and symptoms of DHTR and IHTR. The anti-HLA class I antibody could be produced in spite of using the filer for removing leukocytes, and may take part in the induction of IHTR. Further, blood products should be transfused soon after completing a crossmatch test in patients with anti-RBC alloantibodies.     

The effect of tourniquet release timing on perioperative blood loss in simultaneous bilateral cemented total knee arthroplasty: a prospective randomized study.

Today the use of pneumatic tourniquet is commonly accepted in total knee arthroplasty (TKA) to reduce perioperative blood loss. There are a few prospective randomised and nonrandomised studies that compare the effect of tourniquet release timing in cementless or cemented unilateral TKA. However, many of these studies show an inadequate reporting and methodology. This randomized prospective study was designed to investigate the efficiency of tourniquet release timing in preventing perioperative blood loss in a simultaneous bilateral TKA study design. To our knowledge, this is the first study of its kind, in which the effect of tourniquet release timing on perioperative blood loss was investigated in simultaneous bilateral cemented TKA to compare both techniques intraindividually. In 20 patients (40 knees) one knee was operated with tourniquet release and hemostasis before wound closure, and the other knee with tourniquet release after wound closure and pressure dressing. We found no significant difference in total blood loss between both techniques (p=0.930), but a significant difference in operating time (p=0.035). There were no postoperative complications at a follow-up of 6 month. Other studies report an increase the blood loss in early tourniquet release and an increase the risk of early postoperative complications in deflation of tourniquet after wound closure. In this study we found no significant difference in perioperative blood loss and no increase of postoperative complications. Therefore, we recommend a tourniquet release after wound closure to reduce the duration of TKA procedure and to avoid possible risks of extended anaesthesia.     

Identification, isolation, and characterization of a species-specific 30-kDa antigen of Oesophagostomum dentatum

In the search for a serology tool for the diagnosis of nonpatent as well as patent infections with Oesophagostomum dentatum in pigs a water-soluble, unglycosilated antigen of about 30 kDa specific for the third-stage larvae of the parasite was purified by ion-exchange chromatography. In Western blots, the antigen was first detected by antibodies at day 7 postinfection. Cross-reactivity with O. quadrispinulatum, Ascaris suum, or Trichuris suis was not detected. It is suggested that this protein is a suitable tool for the species-specific serodiagnosis of O. dentatum infection in pigs. Received: 15 June 1998 / Accepted: 28 September 1998     

Inter- and intramolecular interactions in polyelectrolyte complex formation with polyampholytes

Employing polyampholytes (inclusively polybetaines) of different chemical structure containing carboxylic groups and various basic nitrogen functions, homosymplex formation, as well as the competition between homo- and heterosymplex formation on addition of an appropriate polyelectrolyte, was investigated in dependence of pH and ionic strength by means of viscometry and turbidimetry. With most, but not with all, of the polyampholytes, the expected viscosity minimum at the isoelectric point, with its steepness depending on polyampholyte structure, was observed. Competition of homo- and heterosymplex formation at and near the isoelectric point is mainly governed by the pK values of the species involved, the level of zwitter-ion formation of the polyampholyte and the effect of non-Coulombic interactions, for example, via hydrogen bonds.     

13C NMR spectroscopic studies on polyanion-polycation complexes and their component polyelectrolytes

Position and intensities of the ¹³C NMR signals and relaxation times T₁ of several anionic and cationic polyelectrolytes in the solid state were compared with those of the appropriate polyanion-polycation complexes. At a high charge density of the components, the most significant changes of the parameters in question due to complex formation are observed for the C atoms adjacent to the charge centers, indicating a strong Coulombic interaction. At lower charge density, conformational changes of the polymer chains have also to be taken into consideration.     

Correction of EOG Artifacts in Event-Related Potentials of the EEG: Aspects of Reliability and Validity

Correction of EOG artifacts using a regression approach is evaluated in terms of reliability and validity. Transmission rates are estimated for eight EEG channels in 67 subjects. The trimmed group means of these rates are shown to provide reliable measures. Eye artifact correction based on these group means is superior to the conventional rejection in terms of reducing correlation between EOG and EEG.     

Quantitation of cytomegalovirus (hCMV) DNA and β-actin DNA by duplex real-time fluorescence PCR in solid organ (liver) transplant recipients

Even now rare human cytomegalovirus (hCMV) reactivation is still a life-threatening complication after solid organ transplantation. Although PCR techniques are regarded as the most sensitive detection methods for hCMV, their accuracy and reproducibility are limited. This is a major disadvantage with quantitative PCR assays, which are thought to provide valuable information about hCMV latency or active viral replication in transplant patients. To enhance the diagnostic safety of quantitative hCMV PCR, we developed a duplex real-time fluorescence PCR that is capable of quantifying hCMV DNA and beta-actin DNA as internal control simultaneously within one reaction. By the use of 6-carboxyfluorescein and hexa-chloro-6-carboxyfluorescein as reporter fluorophores and 4-(4'-dimethylamino-phenylazo) benzoic acid as dark quencher dye, hCMV DNA and beta-actin DNA could be quantified in parallel in a wide linear range from 10(1) to 10(7) copies, each. To test the clinical applicability of this approach, we investigated hCMV DNA kinetics in peripheral leukocytes of three hCMV antigen-positive and four antigen-negative patients after liver transplantation, as assessed by intracellular hCMV pp65 alkaline phosphate-anti-alkaline phosphate (APAAP) complex. While all APAAP-negative individuals remained PCR negative, kinetics of HCMV DNA in leukocyte DNA samples of APAAP-positive patients correlated closely with hCMV antigen tests. Here, comparison of separate and simultaneous target quantitation revealed identical results. It is of interest that, while single hCMV antigen positivity is commonly not regarded as a reliable parameter of viral reactivation, in our study a viral load greater than 10(4) copies/2x10(5) beta-actin DNA copies clearly indicated a subsequent increase in APAAP-positive leukocytes. We conclude that with the presented method the reliability of hCMV quantitation via real-time PCR can be substantially increased and may be used to monitor hCMV kinetics in vivo.