In vivo characterization of cytotoxic intracellular edema by multicomponent analysis of transverse magnetization decay curves

The precise measurement of low numbers of leukocyte below 0.1 WBC/microliter in filtered red cell or platelet suspensions meet both aims: to check the compliance with previously determined requirements and to evaluate the performances of novel filtering material (5 log depletion or more), justified by more and more important clinical use. The reliability of results, obtained with the chosen method, is ensured by applying of validation protocol, including training of technologist, assessment of the analytical range and the detection limit, assessment of precision and accuracy. The flow cytometry (FC) and Nageotte Chamber (NC) method are the both techniques which are currently used in routine Quality Control (QC) and validated by multicenter studies. Recent developments are made for increasing the sensibility of these counting methods, thanks to higher concentration or volume of the sample to be analysed. Among the experimental techniques, requiring more advances before implementing in QC program, quantitative PCR must become essential as reference method for evaluating the efficiency of filtration, in the future.     

Monoclonal antibodies recognizing epitopes on the extracellular face and intracellular N-terminus of the human erythrocyte anion transporter (band 3) and their application to the analysis of South East Asian ovalocytes

This report describes the production and characterization of 13 rodent monoclonal antibodies to the human erythrocyte anion transport protein AE1 (syn. band 3). Eleven antibodies (4 murine and 7 rat) recognize epitopes dependent on the integrity of the third extracellular loop of the protein. Two antibodies (1 murine and 1 rat) recognize epitopes on the N-terminal cytoplasmic domain. Quantitative binding studies using radioiodinated IgG and Fab fragments of antibodies to extracellular epitopes on AE1 ranged from 77,000 to 313,000 (IgG) and from 241,000 to 772,000 (Fab) molecules bound at saturation. The results indicate that the epitopes recognized by different antibodies vary in their accessibility and suggest that there is heterogeneity in the organization of individual AE1 molecules in the red blood cell membrane. Quantitative binding studies on South East Asian ovalocytes using several antibodies to AE1 and an anti-Wrb show a marked reduction in the number of antibody molecules bound at saturation. These results are consistent with the existence of highly cooperative interactions between transmembrane domains of AE1 in normal erythrocytes and the disruption of these interactions in the variant AE1 found in South East Asian ovalocytes.     

A novel phosphatidylinositol-3,4,5-trisphosphate 5-phosphatase associates with the interleukin-3 receptor

To gain insight into the intracellular signaling cascades that are activated by the binding of interleukin-3 (IL-3) to its target cells, we have embarked on the identification of proteins that are associated with the IL-3 receptor (IL-3R). In a previous study we reported that a 110-kDa serine/threonine protein kinase is constitutively associated with the IL-3R and activated following IL-3 stimulation. We now report that a phosphatidylinositol-3,4, 5-trisphosphate (PtdIns-3,4,5-P3) 5-phosphatase (5-ptase) is also constitutively associated with the IL-3R. This 5-ptase is magnesium-dependent and removes the 5-position phosphate from PtdIns-3,4,5-P3 but does not metabolize PtdIns-4,5-P2, inositol (Ins)-1,3,4,5-P4, or Ins-1,4,5-P3. This substrate specificity distinguishes it from any previously characterized 5-ptase. Interestingly, it may be bound indirectly via phosphatidylinositol 3-kinase (PI 3-kinase), another enzyme that is constitutively bound to the IL-3R. However, unlike PI 3-kinase which becomes activated following IL-3 stimulation, this receptor-associated 5-ptase activity does not increase following IL-3 stimulation, and its primary function may be to keep the principal in vivo product of PI 3-kinase, PtdIns-3,4,5-P3, at low levels in unstimulated cells, to terminate the PI 3-kinase signal following IL-3 stimulation or to metabolize PtdIns-3,4,5-P3 to a metabolically active second messenger, i.e. PtdIns-3,4-P2.     

3,3''-Diindolylmethane induces apoptosis in human cancer cells

Several recent reports presented conflicting data on the action of IL-4 and IL-13 in regulating the release of proinflammatory cytokines by human monocytes. Here we show that the regulation of cytokine release by IL-4 and IL-13 could be either inhibitory or stimulatory in LPS-treated murine peritoneal macrophages. When macrophages were treated with IL-13 or IL-4, between 6 and 24 hr prior to endotoxin challenge, TNF alpha and IL-6 levels were significantly augmented. On the other hand, when the cells were cotreated with LPS plus IL-13 or IL-4, the release of TNF alpha and IL-6 was inhibited. These effects of IL-4 and IL-13 were associated with the modulation of IL-10; pretreatment resulted in a decrease, whereas cotreatment gave rise to a dramatic increase in IL-10 levels. The inhibitory effect of IL-4 and IL-13 on the release of TNF alpha was partially reversed by neutralizing anti-IL10 antibody, and the inhibition of IL-6 release was completely reversed by the antibody. These data suggest that the mechanism of action of IL-13 and IL-4 in modulating macrophage TNF alpha and IL-6 release partially involves IL-10.     

Changing clinical practice. Prospective study of the impact of continuing medical education and quality assurance programs on use of prophylaxis for venous thromboembolism

OBJECTIVE: To determine the effect of continuing medical education (CME) with and without a quality assurance component (CME+QA) on physician practices in the prevention of venous thromboembolism. METHODS: A communitywide study was performed in 15 short-stay hospitals in central Massachusetts. The study population included 3158 patients in acute-care hospitals with multiple risk factors for venous thromboembolism. Study hospitals were randomly assigned to one of two educational strategies or to a control group that received no intervention. RESULTS: The proportion of patients at high risk for venous thromboembolism who received effective methods of prophylaxis increased significantly from 29% in 1986 to 52% in 1989 (P < .001). This increase was seen in all study groups: control hospitals, 40% to 51% (P < .001); CME hospitals, 21% to 49% (P < .0001); and CME+QA hospitals, 27% to 55% (P < .0001). The increase in prophylaxis use from 1986 to 1989 was significantly greater among patients cared for in hospitals whose physicians participated in a formal CME program (an increase of 28%) than in control hospitals (an increase of 11%) (P < .001). There was no significant difference in the use of prophylaxis in hospitals whose physicians received CME+QA interventions compared with hospitals whose physicians received CME interventions alone (identical increases of 28%). CONCLUSION: A formal CME program significantly increased the frequency with which physicians prescribed prophylaxis for venous thromboembolism. We believe the key factor in our CME interventions that motivated clinicians to change their practices was the provision of hospital-specific data demonstrating a compelling need for improvement. Despite the substantial investment by hospitals in QA, traditional QA intervention appeared to provide no additional benefit. Even after extensive CME/QA interventions, prophylaxis for venous thromboembolism remained underutilized, suggesting the need to develop new approaches to changing clinical practice.     

Bioethics in the medical subspecialty literature

The high affinity growth hormone-binding protein (GHBP) circulates in human blood and represents the extracellular domain of the growth hormone (GH) receptor. It is well known that repetitive bouts of endurance type exercise result in increased integrated GH secretion. As the effects of chronic exercise on plasma GHBP levels have never been studied systematically, we investigated the effect of 2 weeks of intense endurance training on plasma GHBP as well as on plasma insulin-like growth factor (IGF)-I levels in 10 healthy, young, non-obese men. IGF-I was measured as an indicator of the effects of GH release. We also studied 10 control subjects matched for sex, age and activity, who were instructed not to change their customary activities. GHBP was determined by FPLC size exclusion chromatography and subsequent Scatchard plot analysis of the binding data; IGF-I levels were measured by RIA. The results showed that plasma IGF-I and GHBP levels were increased in the subjects who followed the training program. IGF-I and GHBP changed from 252 +/- 56 ng/ml and 912 +/- 59 pmol/l before training, to 344 +/- 61 ng/ml (p < 0.01) and 1020 +/- 48 pmol/l (p < 0.01), respectively. Another effect of the training was that the aerobic capacity of these subjects was better utilized and endurance was improved. In contrast, plasma IGF-I, GHBP, utilization of aerobic capacity and endurance did not change significantly in the control subjects. We conclude that two weeks of strenuous endurance training lead to increased plasma IGF-I and high affinity GHBP levels.     

IgG from Adult Atopic Dermatitis (AD) Patients Induces Thymic IL-22 Production and CLA Expression on CD4+ T Cells: Possible Epigenetic Implications Mediated by miRNA

Atopic dermatitis (AD) is a common relapsing inflammatory skin disorder characterized by immune-mediated inflammation and epidermal barrier dysfunction. The pathogenesis of AD is multifactorial and has not been fully elucidated to date. This study aimed to evaluate whether serum IgG from adult AD patients could modulate the thymic maturation of IL-22-producing T cells and CLA+ T cells of non-atopic infants. Given that miRNAs regulate immune response genes, we evaluated whether miRNA expression is also altered in cultured thymocytes. Thymocytes were cultured with purified IgG from AD patients or control conditions (mock, Intravenous-IgG (IVIg), non-atopic IgG, or atopic non-AD IgG). Using flow cytometry analysis, we assessed the expression of CLA and intracellular levels of IL-4, IFN-γ, and IL-22 on double-positive T cells (DP T), CD4 T cells, or CD8 T cells. We also investigated the frequency of IgG isotypes and their direct interaction with the thymic T cells membrane. The miRNA profiles were evaluated by the Illumina small RNA-seq approach. MiRNA target gene prediction and enrichment analyses were performed using bioinformatics. Increased frequencies of IL-22 and CLA+ producing CD4+ T cells cultured with IgG of AD patients was seen in non-atopic infant thymocytes compared to all control conditions. No alterations were observed in the frequency of IgG isotypes among evaluated IgG pools. Evidence for a direct interaction between IgG and thymic DP T, CD4 T, and CD8 T cells is presented. The small RNA-seq analysis identified ten mature miRNAs that were modulated by AD IgG compared to mock condition (miR-181b-5p, hsa-miR-130b-3p, hsa-miR-26a-5p, hsa-miR-4497, has-miR-146a, hsa-let-7i-5p, hsa-miR-342-3p, has-miR-148a-3p, has-miR-92a and has-miR-4492). The prediction of the targetome of the seven dysregulated miRNAs between AD and mock control revealed 122 putative targets, and functional and pathway enrichment analyses were performed. Our results enhance our understanding of the mechanism by which IgG can collaborate in thymic T cells in the setting of infant AD.     

Cellulose acetate reverse osmosis membranes: Optimization of preparation parameters

Asymmetric cellulose acetate based membranes usually employed in reverse osmosis as well as in separations in aqueous systems can possibly be applied in the so‐called salinity process of energy generation. For these applications, membranes with a relatively high water permeability (sometimes also called water flux) and low salt permeability (or high salt rejection) are required. In this study the authors present the optimization of such membranes, which concerns the preparation parameters. The membranes studied were prepared from a solution whose composition were previously optimized.⁴ The authors concluded that the optimum preparation parameters are as follows: thickness of the liquid film of 100 μm; 30 s allowed for evaporation of the solvent; and temperature of coagulation bath of 0–4°C and 80–85°C as annealing temperature. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 134–139, 2007     

Anticoagulant, Antioxidant and Antitumor Activities of Heterofucans from the Seaweed Dictyopteris delicatula

In the present study, six families of sulfated polysaccharides were obtained from seaweed Dictyopteris delicatula by proteolytic digestion, followed by acetone fractionation and molecular sieving on Sephadex G-100. Chemical analyses demonstrated that all polysaccharides contain heterofucans composed mainly of fucose, xylose, glucose, galactose, uronic acid, and sulfate. The fucans F0.5v and F0.7v at 1.0 mg/mL showed high ferric chelating activity (～45%), whereas fucans F1.3v (0.5 mg/mL) showed considerable reducing power, about 53.2% of the activity of vitamin C. The fucan F1.5v presented the most prominent anticoagulant activity. The best antiproliferative activity was found with fucans F1.3v and F0.7v. However, F1.3v activity was much higher than F0.7v inhibiting almost 100% of HeLa cell proliferation. These fucans have been selected for further studies on structural characterization as well as in vivo experiments, which are already in progress.