Fluorescence polarization study on the increase of membrane fluidity of human erythrocyte ghosts induced by synthetic water-soluble polymers.

The effect of water-soluble polymers on the membrane fluidity of human erythrocyte ghosts was investigated and was compared with that of concanavalin A by means of the fluorescence polarization technique. 8-Anilino-1-naphthalene sulfonic acid sodium salt and 1,6-diphenyl-1,3,5-hexatriene were used as probe molecules. The membrane fluidity was increased by the addition of polycations with concentrations of less than 2 x 10(-3) wt% 60 min after mixing. The fluidity changes were affected by the chemical structure (hydrophobicity, charge density, etc.) of polycations. Thus, the membrane fluidity increased markedly with increasing charge density on the chain backbone of polycations. On the other hand, nonionic polymers such as poly(ethylene glycol) and poly(N-vinyl-2-pyrrolidone) changed the membrane fluidity in a biphasic manner. That is, the fluidity of human erythrocyte ghost was temporarily increased and then decrease. For example, 20 wt.% of poly(ethylene glycol) gave a maximum fluidity 15 min after mixing with erythrocyte ghosts. A similar fluidity change was observed by adding concanavalin A. Such fluidity changes were not observed when lipid bilayer vesicles were used instead of cell membranes. These results suggested that the increase of membrane fluidity resulted from the intramembraneous aggregation of membrane-bound proteins which was induced by the added polymers. Cell agglutination was also induced by the addition of a large amount of polymers. This agglutination was considered to be due to the intermembraneous aggregation of membrane-bound proteins.

Phagocytosis of latex particles by leucocytes. I. Dependence of phagocytosis on the size and surface potential of particles.

A series of latex particles, having different sizes and surface structures, were prepared and the dependence of phagocytosis of latex particles by leucocytes on the particle size and surface potential was investigated by measuring the oxygen consumption of leucocytes. Most of the phagocytic behaviour in the initial stages can be explained by susceptibility of particles to heterocoagulation i.e. attachment of small particles (latex particles) onto large particles (leucocytes) by the colloidal attractive force between the two kinds of particles. Specific behaviour for fine particles seems to be attributed to the contribution of steric stabilization by the hydrated layer on the particle surface and to the inability for the leucocytes to recognize very fine particles as foreign materials.

Mechanisms of blood coagulation induced by latex particles and the roles of blood cells.

Latex particles with highly negative or positive charges shortened the clotting time of whole blood and platelet-rich plasma and activated platelet factor 3. Platelet-poor plasma was clotted by the particles with a highly negative charge, but not by those with a positive charge, except hydrophobic particles. Blood coagulation by positively-charged particles was attributed to platelet activation. An enhancement of blood coagulation was also observed in the presence of erythrocytes, leucocytes, their cell membranes or negatively charged phospholipids, and phosphatidylserine instead of platelets. Hydrophilic and low-charged particles suppressed blood coagulation.

Platelet aggregation induced by latex particles. I. Effects of size, surface potential and hydrophobicity of particles.

Latex particles induced platelet aggregation associated with the release of ATP from the platelets. The smaller the diameter of particles having the same surface structure, the greater numbers or greater total surface area of particles were required for both reactions. The higher hydrophobic and higher negatively charged particles, having a diameter of about 0.3 micron, induced platelet aggregation most easily. Hydrophilic particles without high negative surface potential activated platelets only a little. Particle-induced platelet aggregation is not only caused by colloidal electronic force and hydrophobic interaction between platelets and latex particles but also by factors concerning cell activation.

Effect of albumin immobilization by plasma polymerization on platelet reactivity.

Albumin was immobilized on polyethylene (PE) film by the plasma polymerization method. Immobilized albumin on formaldehyde polyermized PE film was hard to desorb even when the film was dipped in blood plasma. Amounts of platelet adhesion on the film and serotonin release decreased clearly. Aggregated platelets were observed on PE film but not observed on the albumin immobilized surface. These results seemed to be due to not only immobilized albumin, but also the surface charge determined by plasma protein adsorption. That is, the slightly negatively charged surface indicated good anti-thrombogenicity.

Effects of plasticizers and plastic bags on granulocyte function during storage.

The influence of the plasticizers, di-(2-ethylhexyl)phthalate (DEHP) and tri-(2-ethylhexyl)trimellitate (TOTM), on granulocyte function was examined. Polyvinyl chloride (PVC) bags with DEHP (DEHP-PVC) leaked DEHP into plasma, but TOTM did not dissolve in plasma under the same conditions. Glow discharge treatment inhibited the leakage of DEHP from DEHP-PVC bags. Depending on the amount of DEHP added into granulocyte suspension, chemotaxis and bactericidal activity decreased, but cell counts and phagocytosis were not affected. During storage for 24 h at 22 degrees C, granulocyte function decreased greatly in DEHP-PVC, but was well maintained in the bags which did not leak plasticizers, TOTM-PVC and glow-discharged DEHP-PVC.

Physical and chemical changes of ACD-preserved blood: a comparison of blood in glass bottles and plastic bags.

ACD blood was preserved in glass bottles with or without aeration and in plastic bags in air or nitrogen gas at 4.5 degrees C. The blood was examined for physical and chemical changes of erythrocyte membrane resistance, hemoglobin in the plasma, the viscosity of the blood, pH of plasma, and ATP and 2,3-DPG content of erythrocytes. The blood preserved in plastic bags showed less changes than blood in glass bottles. The presence of air or nitrogen gas in blood seems to increase the pH perhaps by elimination of carbon dioxide (CO2) which in turn causes the different rates of glycolysis in the erythrocytes.

Effects of autoclave sterilization on the physical properties of storage bags and granulocyte function.

Autoclave sterilization altered the leaching of plasticizer, CO2 gas permeability, surface area and the surface wettability of bag films. These changes affected granulocyte cell counts and functions during storage. Four types of polyvinyl chloride bags, with di-(2-ethylhexyl)phthalate (DEHP) or tri-(2-ethylhexyl)trimellitate (TOTM) as plasticizer, with or without treatment by glow discharge (H2), were sterilized with ethylene oxide (EO) or autoclaving (AC). The greatest amounts of plasticizer leached from DEHP-EO bags. TOTM plasticizer did not leach into plasma. CO2 gas permeability was greater with TOTM than DEHP. AC sterilization decreased the surface area of bags. Wettability of film surfaces was greatest with H2-TOTM-EO. After storage in these bags for 24 and 48 h at 22 degrees C, the granulocyte cell counts and functions were greatest in H2-TOTM-EO bags with the nonleaching plasticizer, higher CO2 gas permeability and higher wettable surface due to glow-discharge treatment. The H2-TOTM-EO bag was useful as a granulocyte storage container.

Change of oxygen affinity of hemoglobin in different conditions of blood preservation.

Changes of oxygen affinity of hemoglobin were studied in blood preserved in different conditions. Changes in hemoglobin function were least in CPD blood in plastic bags, and more extensive in ACD blood in plastic bags, ACD packed red blood cells in plastic bags and ACD blood in glass bottles, respectively. These decreases are consistent with the changes of organic phosphates levels during storage found in earlier studies. When old erythrocytes were incubated with inosine, pyruvate and phosphate, defective oxygen transport function was completely restored even after 7 weeks storage in ACD.

Leukocyte-poor platelet concentrates at the bedside by filtration through Sepacell-PL.

We developed a new filter, 'Sepacell-PL', consisting of a blood administration set for the preparation of leukocyte-poor platelet concentrates at the patient's bedside. This filter is packed with 1.8 microns polyester fibers coated with a new polymer, and has a volume of 14 ml. Sepacell-PL removed more than 99% of the leukocytes while approximately 93% of platelets were recovered from 10 units of platelet concentrates which contained 5 x 10(8) leukocytes and 3 x 10(11) platelets. The functions of pre- and postfiltrated platelets were unchanged. The filtration procedure is very simple, without priming with saline to remove air and wet the filter. Moreover, platelet concentrates can be passed through the filter at a flow rate of 4 ml/min, or at a higher flow rate of 54 ml/min by gravity.

Prevention of leakage of Di-(2-ethylhexyl)phthalate from blood bags by glow discharge treatment and its effect on aggregability of stored platelets.

Platelets storage in glow discharge treated PVC bags was studied. The amount of leaked di-(2-ethylhexyl) phthalate (DEHP) was 150-200 micrograms/ml/day in the nontreated PVC bags, but only 20-40 micrograms/ml in the treated bags after 48 h. The adhesion of silicone to PVC was much improved, and consequently, uniform coating with silicone became feasible. The decrease of the ability of platelets to aggregate was accelerated by DEHP. When stored platelets were resuspended in fresh plasma, the ability to aggregate was gradually restored. However, the degree of restoration of the ability of platelets which had been incubated with DEHP was low. When platelets were stored in the glow discharge treated and then silicone-coated PVC bags, their adhesion on the surface and the decrease of their function were prevented.