Quality control of white cell-reduced red cells: white cell preservation and simplified counting

This article reviews recent studies concerning memory and language disorders in Alzheimer's disease and other neurodegenerative conditions. It shows how different memory and language subcomponents may differentially be impaired in different neurodegenerative diseases and at different stages of the same disease.     

Impact of various red cell concentrate preparation methods on the efficiency of prestorage white cell filtration and on red cells during storage for 42 days

BACKGROUND: White cell (WBC) reduction prior to storage of red cell (RBC) concentrates may reduce the incidence of HLA alloimmunization and may improve the quality of stored RBCs. STUDY DESIGN AND METHODS: An integrated WBC-reduction filter system was tested after various RBC preparation procedures (from whole blood), and the influence of filtration on RBCs during storage for 42 days was investigated. Four additive system RBC preparation protocols were used. Units prepared from conventional triple blood bags were held for 4 to 6 hours at 22 degrees C, and then the RBCs were separated via a hard spin and filtration, performed immediately (Group 1) or after 18 hours' storage at 4 degrees C (Group 2). Units prepared from a top-and-bottom collection system were held at 22 degrees C for 4 to 6 or 22 to 24 hours; the centrifuged RBCs were filtered immediately after preparation (Groups 3 and 4, respectively, by holding time). WBC reduction and filtration time were analyzed. The impact of WBC filtration on pH, hemolysis rate, hemoglobin content, ATP, potassium glucose, and lactate was investigated weekly during storage for 42 days. RESULTS: Filtration reduced the mean WBC count by 3 to 4 log10, to 0.19 +/- 0.25 x 10(6), regardless of the RBC preparation method. Mean filtration times differed significantly between the groups and were longest for Group 1. Besides hemolysis and pH values, which were greater in all filtered units, no major differences were found in filtered and unfiltered RBCs during the storage interval. CONCLUSION: The efficiency of prestorage WBC filtration of RBCs was unaffected by the preparation procedure. However, the filtration time for RBCs freshly prepared in the conventional triple blood bag system without buffy-coat depletion was unacceptable. No major metabolic differences between filtered and unfiltered RBCs during 42 days of storage were found.     

Protein structure prediction by threading methods: evaluation of current techniques

This paper evaluates the results of a protein structure prediction contest. The predictions were made using threading procedures, which employ techniques for aligning sequences with 3D structures to select the correct fold of a given sequence from a set of alternatives. Nine different teams submitted 86 predictions, on a total of 21 target proteins with little or no sequence homology to proteins of known structure. The 3D structures of these proteins were newly determined by experimental methods, but not yet published or otherwise available to the predictors. The predictions, made from the amino acid sequence alone, thus represent a genuine test of the current performance of threading methods. Only a subset of all the predictions is evaluated here. It corresponds to the 44 predictions submitted for the 11 target proteins seen to adopt known folds. The predictions for the remaining 10 proteins were not analyzed, although weak similarities with known folds may also exist in these proteins. We find that threading methods are capable of identifying the correct fold in many cases, but not reliably enough as yet. Every team predicts correctly a different set of targets, with virtually all targets predicted correctly by at least one team. Also, common folds such as TIM barrels are recognized more readily than folds with only a few known examples. However, quite surprisingly, the quality of the sequence-structure alignments, corresponding to correctly recognized folds, is generally very poor, as judged by comparison with the corresponding 3D structure alignments. Thus, threading can presently not be relied upon to derive a detailed 3D model from the amino acid sequence. This raises a very intriguing question: how is fold recognition achieved? Our analysis suggests that it may be achieved because threading procedures maximize hydrophobic interactions in the protein core, and are reasonably good at recognizing local secondary structure.     

Cell transfer by filtration: evaluation of protocols for transformational competence

Collection and washing of cells by centrifugation is a time consuming and inconvenient component of many microbiological protocols, especially when sterility must be maintained. Filtration onto disposable membrane filters efficiently replaced the centrifugation steps in standard transformational competence protocols for Escherichia coli, Haemophilus influenzae, and Saccharomyces cerevisiae. With all three protocols filtered cells were competent sooner and showed transformation efficiencies comparable to or higher than those of cells prepared by centrifugation. The procedure can be easily adapted to collect cells for many other purposes, and should be especially useful in metabolic studies and in preparing cells for electroporation, as it facilitates both thorough washing and rapid but gentle resuspension of cells without compromising sterility.     

A flow cytometric method for counting very low levels of white cells in blood and blood components

Reduction of white cells (WBCs) in blood components may reduce the risk of virus transmission and HLA alloimmunization. Filtration provides a means by which to achieve high-efficiency WBC reduction. A method has been developed using flow cytometry to quantitate the number of WBCs in WBC-reduced packed red cells or platelet concentrates. This method uses a detergent and propidium iodide (PI) solution to label the WBC nuclei and incorporates a known amount of fluorescein isothiocyanate (FITC)-labeled chicken red cells (cRBCs) into the mixture as an indicator of the volume examined. The number of observed WBCs per mL is calculated as follows: Number of PI WBC nuclei events/Number of FITC cRBC events x Number of FITC cRBCs added to mixture/Volume of blood in mixture. The method may allow the detection of WBCs at a concentration as low as 0.01 per microliters (10/mL) in a blood sample. It is an efficient method of collecting data, as it requires less than 10 minutes per sample. This flow cytometric technique is suitable for research purposes and for quality control of WBC-reduced blood components, because it is precise and can be used to quantitate WBCs in large or small numbers in a sample.     

Mn-Ce-Nb-Ox/P84 catalytic filters prepared by a novel method for simultaneous removal of particulates and NO

The Mn-Ce-Nb-O_x/P84 catalytic filter for removal of particulates and NO simultaneous was prepared by a novel method(foam coating method). The process parameters including the concentrations of PTFE emulsion, particle size of catalyst and calcination temperature for preparation of catalytic filters were analyzed. In addition, the physical properties and performance for removal of NO(NH_3-SCR) and particulates of Mn-Ce-Nb-O_x/P84 catalytic filter prepared under the optimized parameters, were also systematic studied. Results show that the process parameters had significant influences on stability and performance of catalytic filter, The Mn-Ce-Nb-O_x/P84 catalytic filter prepared by foam coating method under the optimized parameters, has satisfactory physical properties and catalytic performance for removal of NO and particulates at 140-220 ℃. The NO removal efficiency of catalytic filter can reach95.3% at 200 ℃ as the catalyst loading amount is 450 g/m~2, Moreover,the dust removal efficiency of MnGe-Nb-O_x/P84 catalytic filter reaches as high as 99.98%, and the PM2.5 removal efficiency also reaches99.98%. The anti-sulfur performance of Mn-Ce-Nb-O_x catalytic filter is also attractive, after injecting150 ppm SO_2, the NO removal efficiency still retains up to 85%. It is indicated that the foam coating method can not only make a bond of high strength between catalyst and filter, but also make the catalytic filter possessing an excellent and stable performance for removal of NO and particulates.     

Decarburlzation of silicon melt for solar cells by filtration and oxidation

In studies on the decarburization of liquid silicon, two types of carbon were identified in a silicon melt containing more carbon than the solubility. One was in the form of silicon carbide particles and the other was in the form of dissolved carbon. The silicon was filtered to remove the carbide particles. Using silica or silicon carbide as the material for the filter, the carbon content was reduced from 100 to 30 to 60 ppmw. Oxidation was done under reduced pressure to eliminate the rest of the carbon as carbon monoxide, and the final carbon content was as low as 10 ppmw. Silica was chosen as oxidizing reagent. To avoid a reverse reaction, Ar was introduced on the surface to expel extraneous CO or CO₂ gas evolved. The thermodynamics of the oxidation of carbon in silicon melts is also discussed. Formerly with the Graduate School of the University of Tokyo     

Women's occupational health: a critical review and discussion of current issues

Action to improve women's occupational health has been slowed by a notion that women's jobs are safe and that any health problems identified among women workers can be attributed to unfitness for the job or unnecessary complaining. With increasing numbers of women in the labor force, the effects of work on women's health have recently started to interest health care providers, health and safety representatives and researchers. We begin our summary of their discoveries with a discussion of women's place in the workplace and its implications for occupational health, followed by a brief review of some gender-insensitive data-gathering techniques. We have then chosen to concentrate on the following four areas: methods and data collection; directing attention to women's occupational health problems; musculoskeletal disease; mental and emotional stress. We conclude by pointing out some neglected occupational groups and health issues.     

Radical functions in vivo: a critical review of current concepts and hypotheses

Most of the basic knowledge about radical reactions comes from radiation chemical studies in vitro. In view of the rapidly increasing knowledge on radical reaction in vivo, it is important to reconcile the fundamental physico-chemical reaction characteristics of radicals with the need to explain their alleged biological effects. Severe problems in the understanding of their in vivo action remain unsolved. An example is phagocytosis, which seems to be a paradigm of a 'deleterious' radical process. The exact mechanism is not clear; so it is an open question whether the intruder is eventually killed by radicals (like OH) or by endproducts of radical reactions (like H2O2 and/or HOCl). It is even more difficult to understand signalling by radicals: owing to their chemical nature they are 'unspecifically' reacting species--they withdraw or add electrons--and thus their reactions are governed by redox-properties. Since all radicals have different redox characteristics and different molecular shapes, the usual key-and-keyhole picture for molecular interaction does not apply, as there, is no reactive site conceivable which has the property of reacting with radicals 'specifically. Our intent in this article is: (i) to briefly review some fundamental characteristics of in vitro radical reactions, (ii) to extrapolate from this to the conditions in vivo, and (iii) to discuss current hypotheses concerning the redox-regulation of cellular signalling. This leads us to the tentative conclusion that radicals per se must be tolerated by the cell and do not threaten its life, if they stay below a certain concentration limit. The main biological implication of radical-reactions seems to be that the cell derives signals from the balance of oxidative versus reductive processes and that radicals may interact with pathways of intra- and intercellular communication.     

Prestorage versus bedside white blood cell filtration of red blood cell concentrates: effects on the content of cytokines and soluble tumor necrosis factor receptors

BACKGROUND: The cytokine network has important implications for the systemic inflammatory and metabolic response in trauma and infection. The objective of this study was to investigate the influence of white blood cell (WBC) filtration on the cytokine content in red blood cell concentrates (RBCs). STUDY DESIGN AND METHODS: Tumor necrosis factor-alpha (TNF), interleukin-1-beta (IL-1), interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-10 (IL-10), and soluble TNF receptors I and II (sTNF-RI, sTNF-RII) were investigated in filtered and nonfiltered RBCs during storage. After 40 days of storage the originally nonfiltered units were filtered. RESULTS: On day 1 prestorage filtered RBCs had lower concentrations of WBCs (p < 0.001), TNF (p < 0.01), sTNF-RI (p < 0.01) and sTNF-RII (p < 0.05) compared to nonfiltered units. IL-1 concentrations increased from day 1 to day 40 (p < 0.05) in nonfiltered RBCs and were higher in nonfiltered units compared to prestorage filtered ones on day 40 (p < 0.05). An increase of IL-8 was found in nonfiltered RBCs as well as prestorage filtered units from day 1 to day 40 (p < 0.05) but the concentrations of IL-8 were higher in nonfiltered units on day 40 compared to prestorage filtered units (p < 0.05). Filtration at the end of the 40-day storage period had no influence on the concentrations of cytokines and soluble TNF receptors. CONCLUSION: The present results suggest that prestorage WBC filtration may be more efficient in reducing the cytokine content of RBCs compared to filtration at the the end of the storage period. The clinical impact of passive transfer of components of the cytokine network via RBCs, e.g., in critically ill patients, is however unclear and needs further investigations.     

Clinical evaluation in organ transplant patients of a polymerase chain reaction test for CMV DNA applied on white blood cells and serum

A polymerase chain reaction (PCR) test for CMV DNA was evaluated for clinical usefulness. Leukocytes and serum were sampled from 36 patients who had recently undergone organ transplantation. Clinical symptoms, virus culture, and IgG and IgM antibodies were used to identify, in retrospect, patients with CMV disease certified beyond all doubt, with probable disease, with asymptomatic infection, or without infection. PCR tests for CMV DNA in leukocytes (BC-PCR) and serum (SE-PCR) were then evaluated. BC-PCR was positive in all patients with certified CMV disease but also in 31% of the samples from patients without infection. SE-PCR was positive in 11/13 patients with certified disease and was concordant with CMV culture in 192/231 tests. Of the 39 discordant cases, 27 had a positive SE-PCR with a negative culture. The effect of ganciclovir treatment could not be predicted by any test. In conclusion, a negative BC-PCR is strong evidence against CMV disease and a positive SE-PCR strongly suggests CMV disease, but the opposite results are of little clinical help.     

Dyslipidemia: clinical approaches, evaluation of methods and strategies for standardization

Disorders in lipid metabolism (dyslipidemia) can result to the chronic heart disease. The low density lipoprotein (LDL) is a critical subfraction of total cholesterol present in serum because it is directly linked to coronary heart disease (CHD). The growing awareness of the risks of CHD stipulates the need for more accurate and precise measurement of LDL cholesterol. Current approaches in diagnosing and monitoring CHD is largely dependent on calculated LDL (CLDL) value due to the inherent complexity of ultracentrifugation method. While friedwald's calculated formula may provide comparable values with ultracentrifugation method, it may provide a result which is different. This difference may be of clinical significance. The lipoprotein electrophoresis may be useful in measuring LDL cholesterol, in the diagnosis of type III hyperlipidemia (broad beta band) and when the triglyceride level exceeds 400 mg/dl. The result that compares the CLDL with that obtained by the electrophoresis showed a significant difference (P > or = 0.000) for LDL and insignificant difference (P = 0.068) for high density lipoprotein (HDL) cholesterol.     

Animal and plant cell technology: a critical evaluation of the technology/society interface

The paper describes the energetic feed evaluation systems for ruminants, pigs, poultry and horses presently used in Germany. During the last ten years the "Ausschuss für Bedarfsnormen" (AfB; Committee of Nutrient Requirements) of the "Gesellschaft für Ern?hrungsphysiologie" (GfE, Society of Nutritional Physiology) introduced new recommendations for energy requirements of domestic animals including national and international references. The energetic requirements were factorially deduced (demand for maintenance and various performances) under consideration of partial efficiency of utilized metabolizable energy (ME) for various performances (k-values). Except for lactating cows (Net Energy Lactation; NEL) the energy requirements of other ruminants as well as of pigs and poultry are given in ME. At present, the energy requirement of horses is still expressed as digestible energy (DE). Besides several energy requirements reported there are equations to determine and to calculate the energy content of feeds (GfE, 1986-1998) and corresponding tables with include the nutritive values of feedstuffs for various species. The energy content of mixed feeds may be estimated by specific equations.     

Identification of mutants of pyruvate kinase from red blood cells by means of trypsinization, electrophoresis, kinetic properties and immunological methods

Enzymopathies of pyruvate kinase (PK) are characterized by polymorphism. 11 different mutants of PK were detected in 12 analysed cases. The frequency of double heterozygotes of the PK deficiency is very high. The mutant forms have been identified by electrophoretic, kinetic and immunologic methods in red blood cells of patients and members of their families. The effect of trypsin has greatly increased the sensitivity of the differentiation procedures. Differences in activity and quality of the proteolytic systems in liver and kidney in comparison to red blood cells are responsible for inactivation and degradation of PK mutants in both organs. Homozygotes and double heterozygotes show clinic manifestations. The rigidity of red blood cells increases with the severity of the nonspherocytic haemolytic anaemia.     

Evidence for a critical role for IL-2 in CD40-mediated activation of naive B cells by primary CD4 T cells

The interaction of CD40 on B cells with the CD40 ligand (CD40L) on preactivated CD4 T cells is critical for the initiation of T-dependent Ab responses. It is believed that signals via CD40 synergize with cytokines (e.g., IL-4 and IL-5) to drive B cell activation. However, primary T cells preactivated via CD3 alone cannot induce B cell proliferation; we have shown previously that costimulation of T cells via CD3 and CD28 stabilizes the expression of the CD40L, which we propose contributes to their capacity to act as competent helper-effector cells. Here we show that an additional, critical reason why CD3-stimulated CD40L-bearing T cells are incompetent helper cells is because they secrete insufficient IL-2. In contrast, CD28/CD3-activated T cells induce B cells to become IL-2 responsive via a combination of CD40L and IL-2-mediated signals, and these two stimuli subsequently drive B cell proliferation and IgM secretion. We therefore propose that T cells must first encounter Ag in conjunction with CD80/86 on APCs. This leads to the stable expression of CD40L and maximal secretion of IL-2, which together render primary T cells competent to activate B cells in an IL-2-dependent fashion.     

Economic costs of diabetes mellitus: critical review and cost-benefit evaluation of proposed strategies for its reduction

A multistage protozoan parasitic disease was used as a cachexia model to study the effects of daily administration of bovine growth hormone (GH) on endocrine and body composition changes of young calves from the onset of the acute phase response (APR). Male calves averaging 127.5 +/- 2.0 kg body weight were assigned to control, ad libitum fed, noninfected (C); ad libitum fed, infected (250,000 oocysts Sarcocystis cruzi, per os, I); noninfected, pair-fed (PF) to matched I-treatment calves and these respective same treatments in calves injected daily with GH (USDA-bGH-B1), 12.5 mg/calf/day, im) designated as CGH, IGH and PFGH. GH injections were initiated on Day 20 postinfection (PI), 3 to 4 d before the onset of clinical signs of APR, and continued to Day 56 PI, at which time animals were euthanized for tissue collections. Abrupt increases in rectal temperature commensurate with up to 70% reduction in voluntary feed intake were observed in I and IGH beginning 23-25 d PI. For the trial period between Days 20 and 56 PI, average daily carcass protein gains were 123, 52, 109, 124, 48, and 67 g/d and average daily carcass fat gains were 85, 11, 43, 71, -23, and 29 g/d for C, I, PF, CGH, IGH, and PFGH, respectively. Effects of GH were significant for fat accretion and plasma urea depression. Rectus femoris was highly refractory to catabolic effects of infection while psoas major was significantly catabolized during infection. Plasma concentrations of insulin-like growth factor-I (IGF-I) increased significantly in all GH-treated calves between Day 20 and 23 PI. Plasma IGF-I declined well below Day 20 values in all infected calves from the onset of the APR through the end of the study. The decrease in plasma IGF-I concentrations in I and IG was highly correlated with the magnitude of the fever response. Hepatic mRNA for GH receptor and IGF-I was decreased in infected calves. Hepatic microsomal membrane binding of 125I-GH did not differ between groups. The data suggest that effects of GH and parasitism on tissue metabolism during disease may vary among different specific tissue pools. The data demonstrate that daily GH administration in young calves does not prevent lean tissue losses and may accelerate fat depletion associated with cachectic parasitism. Furthermore, the onset of APR overrode the capacity for GH to maintain elevated plasma concentrations of IGF-I, an effect not readily explained through changes of GH-receptor binding.