Toxicity evaluation of nanocrystalline silver-impregnated coated dressing on the life cycle of worm Caenorhabditis elegans.

In recent times, however, due to the emergence of bacterial strains with resistance to conventional antibiotics, silver has again gained attention as an alternative for developing new efficient bactericides, including the use of silver nanoparticles (AgNPs). However, the improper disposal of these items after use may cause toxicological effects on organisms in the environment. To evaluate the potential environmental hazard of nanosilver-coated dressings, the nematode Caenorhabditis elegans was chosen as a test organism. The assays were conducted in 24-well plates that contain four different sizes of coated dressing to obtain different concentrations. L1 and L4 C. elegans larval stages were exposed to these nanosilver concentrations. Dressing cutouts were arranged between two layers of agar for 3 days and Escherichia coli (OP 50 strain) was added as food source for the worms. After the exposure period, growth, reproduction, fertility, silver concentration in the medium and the concentration of reactive oxygen species (ROS) in the worms were evaluated. Scanning and transmission electron microscopy analyses were performed on the coated dressings, as well as analyses of zeta potential, ionic release and antibacterial power in two bacterial strains (Pseudomonas aeruginosa and Staphylococcus aureus). It was verified the antibacterial power of the coated dressing, in both bacteria strains tested. Characterization of the coated dressing indicated heterogeneous nanoparticles, as well as distinct zeta potentials for the medium in water and saline medium (0.9% NaCl). L1 larval worms exposed to nanosilver-coated dressing showed a high ROS concentration and reductions in growth, fertility and reproduction. Worms exposed to the coated dressing during the L4 stage showed almost no response. Overall, the obtained results indicate the potential environmental hazard of nanosilver-coated dressings.

Effect of Landscape Changes on Water Quality and Health Status of Heptapterus mustelinus (Siluriformes, Heptapteridae).

Substances derived from anthropogenic activities induce changes in the physical and chemical characteristics of the aquatic environment. Physicochemical and biological studies are necessary to understand how changes in landscape affect the health of the aquatic environment. The main goal of this study was to evaluate how the landscape at different spatial scales affects (1) water quality and (2) the health status of Heptapterus mustelinus, based on several biomarkers. During the dry season, individuals were caught in three sites with different degrees of anthropogenic activity. The quality of the terrestrial environment was assessed using the Riparian Quality and Land Use Indices. The water quality condition was evaluated using a water quality index, and pesticides and pharmaceuticals were measured in water. The following biomarkers were analyzed in the fish: general health status (Condition Factor, Hepatosomatic index and energetic costs), enzymatic activity (GST, CAT, AchE), carbonyl content in proteins and histopathological responses in liver and gills. The most impacted sites by the presence of pesticides showed more alterations in the surrounding landscape; specially, changes in the riparian area. In this area, biomarkers denoted more damage than in sites with protected riparian zone. Conservation status of riparian ecosystems is crucial in the determination of rivers ecological quality. Our results demonstrate the importance of monitoring the environmental quality through an integrated analysis, using native fish to understand the effects of human activities on the biota.

Characterization of efficacy and safety of pathogen inactivated and quarantine plasma in routine use for treatment of acquired immune thrombotic thrombocytopenic purpura.

BACKGROUND: Auto-immune thrombotic thrombocytopenic purpura (TTP) is a morbid multi-organ disorder. Cardiac involvement not recognized in initial disease descriptions is a major cause of morbidity. Therapeutic plasma exchange (TPE) requires exposure to multiple plasma donors with risk of transfusion-transmitted infection (TTI). Pathogen inactivation (PI) with amotosalen-UVA, the INTERCEPT Blood System for Plasma (IBSP) is licensed to reduce TTI risk. METHODS: An open-label, retrospective study evaluated the efficacy of quarantine plasma (QP) and IBSP in TTP and defined treatment emergent cardiac abnormalities. Medical record review of sequential patient cohorts treated with QP and IBSP characterized efficacy by remission at 30 and 60 days (d) of treatment, time to remission, and volume (L/kg) of plasma required. Safety outcomes focused on cardiac adverse events (AE), relapse rates, and mortality. RESULTS: Thirty-one patients (18 IBSP and 13 QP) met study criteria for auto-immune TTP. The proportions (%) of patients in remission at 30 d (IBSP = 61·1, QP = 46·2, P = 0·570) and 60 d (IBSP = 77·8, QP = 76·9, P = 1·00) were not different. Median days to remission were less for IBSP (15·0 vs. 24·0, P = 0·003). Relapse rates (%) 60 d after remission were not different between cohorts (IBSP = 7·1, QP = 40·0, P = 0·150). ECG abnormalities before and during TPE were frequent; however, cardiac AE and mortality were not different between treatment cohorts. CONCLUSIONS: Cardiac and a spectrum of ECG findings are common in TTP. In this study, IBSP and QP had similar therapeutic profiles for TPE.

Physiological and biochemical responses of Eichhornia crassipes exposed to Cr (III).

The effect of exposure of Eichhornia crassipes to Cr (III) was assessed by measuring changes in photosynthetic pigments, malondialdehyde, superoxide dismutase, glutathione reductase, catalase, and guaiacol peroxidase activities, as well as Cr concentration in tissues. Cr concentration in roots was significantly higher than in aerial parts and increased with Cr concentration in water. Photosynthetic pigments increased significantly, whereas the activities of antioxidant enzymes varied differently in plant tissues. Low Cr concentrations induced a rapid response of E. crassipes during short-term exposure, implying that the antioxidant system conferred redox homeostasis. Results showed that Cr (III) was more toxic at the two highest concentrations and long-term exposure, while it was not harmful but beneficial at the two lowest concentrations and short-term exposure. This work concludes that E. crassipes was able to grow under Cr (III) stress by protecting itself with an increase in the activity of its antioxidant system.

[Ethics and transfusion--seminar report]. / Éthique et transfusion sanguine--Échos d'un séminaire.

This paper brings together the abstracts and proceedings of a seminar held on the topic of "ethics and transfusion", October 15, 2013 at the National Institute of Blood Transfusion, Paris.

[Pathogen inactivation in platelet concentrates: the French experience]. / Inactivation des pathogènes des concentrés plaquettaires: expérience française.

The transfusion of platelet concentrates is increasing in oncohematology patients due to chemotherapy and hematopoietic stem cell grafts. The transmission of pathogenic agents, viruses, parasites and especially bacteria with platelet concentrates stored at room temperature (20-24°C) is associated with a septic risk, partly prevented by bacterial detection. Photochemical inactivation of platelet concentrates, using a technique associating amotosalen and UVA, has been used for five years in a French region for the whole population and a large spectrum of patients, with efficacy and safety. Universal implementation of pathogen inactivation in labile blood products is a major and key step to improve safety against infection in transfusion.

[Photochemical inactivation of pathogens in platelets and plasma: five years of clinical use in routine and hemovigilance. Towards a change of paradigm in transfusion safety]. / Inactivation photochimique des pathogènes des plaquettes et du plasma: cinq ans d'utilisation clinique de routine et d'hémovigilance. Vers un changement de paradigme de la sécurité en transfusion.

The transfusion of labile blood products is vital and essential for patients in absence of alternative treatment. Patients and doctors have always feared transfusion-transmitted infections by blood, blood components and blood-derived drugs. Photochemical inactivation of platelet concentrates and plasma, using a technique associating amotosalen and UVA, has been used for five years in a French region for the whole population and a large spectrum of patients, with efficacy and safety. It would seem wise to introduce labile blood products, submitted to pathogen inactivation by a technique already approved by a regulatory agency and not to wait for a perfect system including red blood cells concentrates. Universal implementation of pathogen inactivation in labile blood products is a major and key step to improve safety against infection in transfusion.

Mechanisms underlying FeCl3-induced arterial thrombosis.

BACKGROUND: The FeCl(3)-induced vascular injury model is widely used to study thrombogenesis in vivo, but the processes leading to vascular injury and thrombosis are poorly defined. OBJECTIVES: The aim of our study was to better characterize the mechanisms of FeCl(3)-induced vascular injury and thrombus formation, in order to evaluate the pathophysiological relevance of this model. METHODS: FeCl(3) was applied at different concentrations (from 7.5% to 20%) and for different time periods (up to 5 min) to mouse carotid or mesenteric arteries. RESULTS: Under all the conditions tested, ultrastructural analysis revealed that FeCl(3) diffused through the vessel wall, resulting in endothelial cell denudation without exposure of the inner layers. Hence, only the basement membrane components were exposed to circulating blood cells and might have contributed to thrombus formation. Shortly after FeCl(3) application, numerous ferric ion-filled spherical bodies appeared on the endothelial cells. Interestingly, platelets could adhere to these spheres and form aggregates. Immunogold labeling revealed important amounts of tissue factor at their surface, suggesting that these spheres may play a role in thrombin generation. In vitro experiments indicated that FeCl(3) altered the ability of adhesive proteins, including collagen, fibrinogen and von Willebrand factor, to support platelet adhesion. Finally, real-time intravital microscopy showed no protection against thrombosis in GPVI-immunodepleted and ß(1)(-/-) mice, suggesting that GPVI and ß(1) integrins, known to be involved in initial platelet adhesion and activation, do not play a critical role in FeCl(3)-induced thrombus formation. CONCLUSION: This model should be used cautiously, in particular to study the earliest stage of thrombus formation.

[Occurrence of severe, persistent thrombocytopenia following allogeneic bone marrow transplantation, attributable to anti-HPA-1 allo-immunisation of the host]. / Survenue d'une thrombopénie sévère et durable, au décours d'une greffe de moelle allogénique, imputable à l'existence chez l'hôte d'une allo-immunisation anti-HPA-1.

A 56 year-old, multiparous woman suffering from a myeloproliferative syndrome, who had received multiple red blood cell and platelet transfusions, was the recipient of an allograft of peripheral blood stem cells derived from her HLA-A, B, DR, DQ and DP and ABO identical sister, following myeloablative conditioning. The persistence of severe, isolated thrombopenia resistant to platelet transfusions led to the discovery of anti-HLA class I allo-immunisation. As HLA compatible platelet transfusions did not result in satisfactory platelet increments, we then discovered the simultaneous presence of anti-HPA-1a allo-immunisation. Genotyping of the HPA-1 systems of the patient (HPA-1B/B) and her sister (HPA-1A/B) enabled us to elucidate the mechanism underlying the persistent thrombopenia and the inefficacy of transfusion. In fact, only transfusion of HPA-1B/B platelets (HLA compatible or incompatible) proved to be efficacious. To reduce the level of anti-HPA-1a antibodies, we performed plasmapheresis sessions and used an anti-CD20 monoclonal antibody. It was only on achieving total haematopoietic chimerism, through rapid interruption of the immunosuppression, that we obtained spontaneous normalisation of the platelet count. The present case emphasises the necessity, before undertaking any allograft of haematopoietic stem cells - even if the latter come from a strictly HLA identical member of the family - of performing a search for eventual anti-HPA allo-immunisation.

Proplatelet formation deficit and megakaryocyte death contribute to thrombocytopenia in Myh9 knockout mice.

BACKGROUND: Inactivation of the mouse Myh9 gene (Myh9Δ) or its mutation in MYH9-related diseases leads to macrothrombocytopenia. Paradoxically, previous studies using in vitro differentiated megakaryocytes showed an increased capacity for proplatelet formation when myosin was absent or inhibited. METHODS: To explore the origin of the thrombocytopenia induced by myosin deficiency, we studied proplatelet formation using bone marrow explants of wild-type (WT) and Myh9Δ mouse where megakaryocytes have matured in their native environment. RESULTS AND DISCUSSION: A dramatic decrease in the number and complexity of proplatelets was observed in megakaryocytes from Myh9Δ mice, while inhibition of myosin activity by blebbistatin increased proplatelet formation from WT mature megakaryocytes. Moreover, Myh9Δ megakaryocytes had a smaller size than the WT cells. These data indicate that myosin deficiency acts negatively on proplatelet formation, probably by impairing in situ megakaryocyte maturation, while myosin activity is dispensable at the latest stage of proplatelet formation. In addition, ultrastructural examination of Myh9Δ bone marrow revealed an increased proportion of megakaryocytes exhibiting signs of non-apoptotic cell death as compared with the WT mice. CONCLUSION: These data indicate that thrombocytopenia in Myh9Δ mice results from defective development of megakaryocyte size, impaired proplatelet formation and increased cell death.

Immature myeloid dendritic cells capture and remove activated platelets from preformed aggregates.

BACKGROUND: Immature dendritic cells (DCs) patrol the circulation, where they can uptake antigens. It has been reported that mature monocyte-derived DCs have the ability to interact with an activated platelet monolayer under high shear conditions (1500s(-1) ). OBJECTIVES: In this study, we investigated whether platelets can recruit immature myeloid DCs (CD1c(+) ) directly isolated from blood (BDCs) and if so, which receptors are involved. RESULTS: Using flow cytometry and electron microscopy, we showed that BDCs interact with activated but not resting platelets in suspension. Interaction was also observed after perfusing BDCs under low flow conditions (150 s(-1) ) through collagen-coated microcapillaries in which platelets had adhered and formed aggregates. No such interaction could be detected at higher shear rates. Whereas initial transient attachment required the exposure of P-selectin on activated platelets and PSGL-1 on BDCs, subsequent stationary adhesion was dependent on α(IIb) ß(3) and α(M) ß(2) integrins on platelets and BDCs, respectively. Moreover, during their transient interaction, BDCs preferentially removed platelets located at the outer margin of the thrombus in a P-selectin- and integrin-dependent manner. CONCLUSION: This study provides evidence for an interaction between activated platelets and immature myeloid DCs only under low shear conditions. This could be of importance for BDC maturation and antigen uptake during normal hemostasis or in the context of atherothrombosis at sites of reduced blood flow.

Inventory management.

A critical aspect of blood transfusion is the timely provision of high quality blood products. This task remains a significant challenge for many blood services and blood systems reflecting the difficulty of balancing the recruitment of sufficient donors, the optimal utilization of the donor's gift, the increasing safety related restrictions on blood donation, a growing menu of specialized blood products and an ever-growing imperative to increase the efficiency of blood product provision from a cost perspective. As our industry now faces questions about our standard practices including whether or not the age of blood has a negative impact on recipients, it is timely to take a look at our collective inventory management practices. This International Forum represents an effort to get a snap shot of inventory management practices around the world, and to understand the range of different products provided for patients. In addition to sharing current inventory management practices, this Forum is intended to foster an exchange of ideas around where we see our field moving with respect to various issues including specialty products, new technologies, and reducing recipient risk from blood transfusion products.

The platelet glycoprotein GPIbbeta intracellular domain participates in von Willebrand factor induced-filopodia formation independently of the Ser 166 phosphorylation site.

SUMMARY BACKGROUND: Circulating platelets are initially recruited at the site of vessel injury by von Willebrand factor (VWF) immobilized on collagen fibers. This process, mediated by the GPIb-V-IX complex, is accompanied by specific intracellular signaling leading to reorganization of the platelet actin cytoskeleton and extension of filopodia. OBJECTIVES/METHODS: To evaluate the GPIbalpha and GPIbbeta intracellular domains contribution to this signaling, we generated Chinese hamster ovary (CHO) cells expressing a GPIb-IX complex with mutant forms of the two subunits and we measured their ability to extend filopodia upon adhesion on a VWF matrix. RESULTS: Complete intracellular deletion or elimination of the filamin or the 14-3-3zeta binding sites in GPIbalpha did not prevent filopodia extension. In contrast, deletion of the juxtamembrane (Leu(150)-Arg(160)) or central (Ala(159)-Pro(170)) intracellular segment of GPIbbeta resulted in a 21% and 23% reduction in the number of cells extending filopodia, respectively. This occurred without decreasing adhesion efficiency or GPIb-IX association with filamin A or 14-3-3zeta. Alanine scanning mutagenesis of the Leu(150)-Pro(170) segment identified Arg(164), Leu(165), Leu(167), Thr(168) and Pro(170) as important residues for efficient filopodia formation. Surprisingly, mutation of the Ser(166) PKA phosphorylation site did not alter adhesion and shape change. A role for the GPIbbeta subunit was reinforced by the decreased capacity to extend filopodia upon adhesion on VWF of platelets from knock-in mice expressing a GPIbbeta intracellular deletion mutant. CONCLUSIONS: Altogether, our results strongly support participation of GPIbbeta and its intracellular region in GPIb-dependent platelet activation and shape change triggered by a VWF matrix.

Arterial thrombosis: relevance of a model with two levels of severity assessed by histologic, ultrastructural and functional characterization.

BACKGROUND: We previously described a model of laser-induced thrombosis in mesenteric arterioles with superficial and deep levels of injury producing a transient thrombus resolving within 2 min and a larger almost occlusive thrombus, respectively. Both types of lesion were sensitive to platelet GPIIb-IIIa and P2Y(12) inhibition, whereas only deep injuries were sensitive to thrombin blockade. OBJECTIVE: The aim of the present study was to use histologic methods and electron and intravital microscopy to characterize the lesions and thrombi and to extend our knowledge of the sensitivity of this model to genetic and pharmacologic inhibition. RESULTS: A superficial injury was found to detach the endothelial cells and expose a collagen III- and IV-rich subendothelium where platelets could adhere. Tissue factor and fibrin were not detected. Deeper penetration of the external elastic lamina occurred in deep injuries, with exposure of collagen I, III and IV. Here the thrombus was composed of platelets exhibiting a decreasing gradient of degranulation from the deepest lesion area to the surface. Fibrin was found close to the most activated platelets. Consistently, glycoprotein VI (GPVI)-collagen and GPIb-von Willebrand factor (VWF) interactions were found to be critical in superficial injuries. After deep lesion, thrombus formation was modestly reduced in GPVI-immunodepleted mice and still strongly inhibited in VWF(-/-) mice. Combined hirudin infusion and GPVI depletion further inhibited thrombosis after deep injury. CONCLUSIONS: This study confirms the feasibility of inducing arterial thrombosis with distinct levels of severity and establishes the central roles of collagen and VWF in thrombus formation after superficial injury. Collagen, VWF and thrombin all appear to contribute to thrombosis after deep arterial lesion.