An in vitro system for testing leucocyte and leukaemic cell line adhesion to synthetic fibres

Leucocyte adhesion is an important phenomenon in antimicrobial defence, inflammation and immunological mechanisms and has been shown to be dependent upon specialized adhesion molecules. To prevent side-effects related to blood transfusion (e.g. anti-human leucocyte antigen immunization and transmission of infectious agents) leucocyte reduction of blood products is now systematically performed in various countries. The most common system used for leucoreduction is blood filtration. For further understanding of the mechanisms responsible for the interaction between leucocytes and the fibres present in filters we used a flow chamber to study the adhesion of leucocytes and leukaemic cell lines to different types of fibre. Adhesion was quantified using video-microscopy and computer image analysis. Our results demonstrate that adhesion to filter fibres was dependent on the expression of beta2-integrins CD11--CD18 and was inhibited by anti-CD18. The amount of fibres present, their spatial arrangement and the physicochemical characteristics of the fibres were important factors in leucocyte adhesion. Leucocyte adhesion was the highest to polyethylene terephthalate (PET) and polyimide fibres. Lymphocytes or lymphocytic cell lines were poorly adherent to PET fibres. The retaining capacity of leucocyte filters can be improved by taking into account the different parameters for the design of new filters     

Lack of specific binding of Shiga-like toxin (verocytotoxin) and non-specific interaction of Shiga-like toxin 2 antibody with human polymorphonuclear leucocytes.

BACKGROUND: After gastrointestinal infection with Shiga-like toxin (Stx) producing Escherichia coli, the toxin is transported from the intestine to the renal microvascular endothelium. This is the main target for Stx in humans. Previous studies indicated that polymorphonuclear leucocytes (PMN) could serve as carriers for Stx in the systemic circulation. As at a later stage we could not confirm these data, we performed new studies. METHODS: The binding of Stx1 to PMN was determined in vitro (isolated human PMN and whole blood) and in vivo (injection in mice). The specificity of binding of an antibody against Stx2 to PMN from patients with haemolytic uraemic syndrome (HUS) was determined. This was compared with binding to PMN from healthy controls, and patients after haemodialysis (HD) or on peritoneal dialysis (PD). Furthermore, PMN were incubated with Stx to study possible activation. RESULTS: No specific binding of Stx1 to PMN could be detected. After intravenous injection of the toxin in mice, it was not associated with PMN. The binding of an antibody against Stx2 to PMN was detected in both patients with HUS and patients after HD, but not in patients on PD. Stx was not able to activate PMN. CONCLUSIONS: PMN are not acting as transporter for Stx in the pathogenesis of HUS. The interaction of a Stx antibody with PMN from HUS patients is not specific as it can also be observed in patients after HD (possibly due to activation of the PMN). Therefore, binding of Stx antibody to PMN is not reliable as a diagnostic tool for HUS.     

Advances in understanding the molecular mechanisms of venous thrombosis

Venous thromboembolism (VTE) represents a leading cause of global mortality, however, the determinants that contribute to thrombus development remain incompletely understood. In this review, we discuss the role of inherited abnormalities of blood coagulation in VTE pathogenesis. In addition, we also consider recent emerging data suggesting other molecular and cellular determinants may also contribute. Specifically, we describe the role played by the inflamed endothelium, and the dysregulated responses to inflammatory stimuli that create a platform for pathological clot formation. We review the accumulating evidence that blood cells, contact pathway factors and protein disulphide isomerases all play key roles in VTE development. Finally, we discuss new insights into the role of metabolites arising from commensal gut bacteria and their potential role in facilitating VTE. This overview provides an update on these state-of-the-art developments and the opportunities they provide for new antithrombotic therapies with enhanced efficacy and improved safety profiles.     

Physiology of red and white blood cells

Blood is made up of plasma and formed elements, which are red blood cells, white blood cells and platelets. The red blood cells (erythrocytes) make up the vast majority of the cells present in the blood. Their principal function is the transport of oxygen from the lungs to the tissues and the transport of carbon dioxide from those tissues back to the lungs. This is due to the presence of haemoglobin, a protein that binds easily and reversibly with oxygen. The affinity of haemoglobin for oxygen changes under certain conditions allowing increased offloading of oxygen at the respiring tissues as required. White blood cells (leucocytes) form the body's defence against invading pathogens. They can be subdivided into granulocytes and agranulocytes, which have different mechanisms of attack against those pathogens.     

Increased leucocyte histamine release by Entamoeba histolytica antigen in patients with amoebic abscess of the liver

Leucocytes (basophils) from non-atopic adult subjects living in an area highly endemic with Entamoeba histolytica release histamine in a dose-dependent fashion upon in vitro exposure to an antigen of axenically grown E. histolytica (histolyticin). Leucocytes of patients with acute amoebic liver abscess were significantly more sensitive to this antigen than leucocytes of control subjects, including patients that had recovered from amoebic liver abscess. By comparison Concanavalin-A induced histamine release found in patients with amoebic liver abscess and healthy controls suggest an immunological mechanism for histolyticin induced in vitro histamine release. This is also suggested by the inability of histolyticin to release histamine from leucocytes of healthy newborn infants and the significant fall in sensitivity to histolyticin following incubation of leucocytes in acid pH. Histamine and other mediators may contribute locally to the early intense inflammatory reaction observed in tissue invasion by E. histolytica.     

Reduced leucocyte cholesteryl ester transfer protein expression in acute coronary syndromes

Objective. Cholesterol ester transfer protein (CETP) plays an important role in HDL cholesterol metabolism. Leucocytes, including monocyte‐derived macrophages in the arterial wall synthesize and secrete CETP, but its role in atherosclerosis is unclear. The aim of the current study was to investigate the effect of acute coronary syndromes (ACS) on leucocyte CETP expression. Research design. Peripheral blood mononuclear cells (PBMCs) were freshly isolated from hospitalized ACS patients displaying Braunwald class IIIB unstable angina pectoris (UAP) on admission (t = 0) and at 180 days post inclusion (t = 180) for analysis of CETP expression. In addition, to prove the potential correlation between leucocyte CETP and ACS the effect of acute myocardial infarction on leucocyte CETP expression was studied in CETP transgenic mice. Results. Upon admission, UAP patients displayed ～3–6 fold (P < 0.01) lower CETP mRNA and nearly absent CETP protein expression in PBMCs, as compared to healthy age‐/sex‐matched controls. Interestingly, CETP mRNA and protein levels were significantly elevated in PBMCs isolated from UAP patients (both stabilized and refractory) at t = 180 as compared to t = 0 (P < 0.01), which was correlated with a reduced inflammatory status after medical treatment. In agreement with the data obtained in UAP patients, markedly down‐regulated leucocyte CETP mRNA expression was observed after coronary artery ligation in CETP transgenic mice, which also correlated with increased serum amyloid A levels. Conclusions. We are the first to report that episodes of UAP in humans and myocardial infarction in CETP transgenic mice are associated with reduced leucocyte CETP expression. We propose that the impairment in leucocyte CETP production is associated with an enhanced inflammatory status, which could be clinically relevant for the pathogenesis of ACS.     

Effects of cylindrospermopsin on the phagocytic cells of the common carp (Cyprinus carpio L.)

Cylindrospermopsin is a cyanotoxin with cytotoxic activity. It is released into water during and after cyanobacterial water blooms and thus poses a threat to the health of fish. There is very little information available concerning the effects of the toxin on fish immune cells. In this study, we assessed the potential impact of cylindrospermopsin on the basic functions of phagocytic cells from common carp (Cyprinus carpio L.), including phagocytosis, reactive oxygen and nitrogen species production, and the structure of microfilaments and selected cytokine expression. Phagocytic cells, isolated from fish head kidneys, were exposed to the toxin at concentrations of 0.05, 0.1, 0.5 or 1 µg ml^?1, for up to 24 h. Cytotoxicity, detected by lactate dehydrogenase release, was observed at the highest studied concentration. A decrease in phagocytic activity and changes in actin cytoskeletal structures were observed after the cell exposure to the toxin at 0.5 and 1 µg ml^?1. Moreover, at all tested concentrations, cylindrospermopsin increased the production of reactive oxygen and nitrogen species. It also evidently influenced the expression of genes of proinflammatory cytokines interleukin‐1β and tumour necrosis factor‐α and, to a minor extent, anti‐inflammatory transforming growth factor‐β, but had no effects on interleukin‐10. The results indicated that the cyanotoxin cylindrospermopsin is able to modify basic features of carp phagocytic cells, which might result in adverse consequences for fish health. Copyright © 2015 John Wiley & Sons, Ltd.