Biocompatibility of Polysulfone Hemodialysis Membranes and Its Mechanisms: Involvement of Fibrinogen and Its Integrin Receptors in Activation of Platelets and Neutrophils.

Activation of blood cells during hemodialysis is considered to be a significant determinant of biocompatibility of the hemodialysis membrane because it may affect patient health adversely through microvascular inflammation and oxidative stress. This study found very different cell activation among various polysulfone (PSf) hemodialysis membranes. For example, CX-U, a conventional PSf membrane, induced marked adhesion of platelets to its surface and increased surface expression of activated CD11b and production of reactive oxygen species (ROS) by neutrophils; while NV-U, a hydrophilic polymer-immobilized PSf membrane, caused little platelet adhesion and slight CD11b expression and ROS production by neutrophils. Analysis of the molecular mechanisms of the above phenomena on CX-U and NV-U indicated that anti-integrin GPIIb/IIIa antibody blocked platelet adhesion, and that the combination of anti-CD11b (integrin α subunit of Mac-1) and anti-integrin αvß3 antibodies blocked ROS production by neutrophils. Plasma-derived fibrinogen, a major ligand of GPIIb/IIIa, Mac-1, and αvß3 on membranes, was thus analyzed and found to be more adsorbed to CX-U than to NV-U. Moreover, comparison between five PSf membranes showed that the number of adherent platelets and neutrophil ROS production increased with increasing fibrinogen adsorption. These results suggested that fibrinogen, adsorbed on membranes, induced GPIIb/IIIa-mediated platelet activation and Mac-1/αvß3-mediated neutrophil activation, depending on the amount of adsorption. In conclusion, the use of biocompatible membranes like NV-U, which show lower adsorption of fibrinogen, is expected to reduce hemodialysis-induced inflammation and oxidative stress by minimizing cell activation.

Development of a Novel Site-Specific Pegylated Interferon Beta for Antiviral Therapy of Chronic Hepatitis B Virus.

Although nucleot(s)ide analogues and pegylated interferon alpha 2a (PEG-IFN-α2a) can suppress hepatitis B virus (HBV) replication, it is difficult to achieve complete HBV elimination from hepatocytes. A novel site-specific pegylated recombinant human IFN-ß (TRK-560) was recently developed. In the present study, we evaluated the antiviral effects of TRK-560 on HBV replication in vitro and in vivo. In vitro and in vivo HBV replication models were treated with antivirals including TRK-560, and changes in HBV markers were evaluated. To analyze antiviral mechanisms, cDNA microarray analysis and an enzyme-linked immunoassay (ELISA) were performed. TRK-560 significantly suppressed the production of intracellular HBV replication intermediates and extracellular HBV surface antigen (HBsAg) (P < 0.001 and P < 0.001, respectively), and the antiviral effects of TRK-560 were enhanced in combination with nucleot(s)ide analogues, such as entecavir and tenofovir disoproxil fumarate. The reduction in HBV DNA levels by TRK-560 treatment was significantly higher than that by PEG-IFN-α2a treatment both in vitro and in vivo (P = 0.004 and P = 0.046, respectively), and intracellular HBV covalently closed circular DNA (cccDNA) reduction by TRK-560 treatment was also significantly higher than that by PEG-IFN-α2a treatment in vivo (P = 0.0495). cDNA microarrays and ELISA for CXCL10 production revealed significant differences between TRK-560 and PEG-IFN-α2a in the induction potency of interferon-stimulated genes. TRK-560 shows a stronger antiviral potency via higher induction of interferon-stimulated genes and stronger stimulation of immune cell chemotaxis than PEG-IFN-α2a. As HBsAg loss and HBV cccDNA eradication are important clinical goals, these results suggest a potential role for TRK-560 in the development of more effective treatment for chronic hepatitis B infection.

A new hydrophilic polysulfone hemodialysis membrane can prevent platelet-neutrophil interactions and successive neutrophil activation.

PURPOSE:: Microaggregates have often been observed during hemodialysis and are clearly associated with complications of hemodialysis therapy. In this study, we aimed to clarify the effects of two polysulfone membranes, with different abilities to activate blood cells, on the formation of these microaggregates; we also investigated their molecular mechanisms. METHODS:: Human whole blood was circulated through a mini-module dialyzer using the membranes in vitro; platelet-neutrophil complexes in blood were determined by flow cytometry. Isolated human neutrophils were incubated with the membranes in plasma, in the presence or absence of platelets, followed by flow cytometric analysis of intracellular reactive oxygen species and cell-surface activated CD11b on neutrophils. RESULTS:: CX-U, a conventional polysulfone membrane with remarkable cell activation, induced the formation of platelet-neutrophil complexes; however, NV-U, a new hydrophilic polysulfone membrane with slight or no cell activation, did not cause complex formation. Moreover, CX-U-induced reactive oxygen species production and the increase in activated CD11b expression on neutrophils were enhanced by platelets. On the other hand, NV-U hardly affected neutrophil activation, regardless of whether platelets were present or not. The enhancement of CX-U-induced neutrophil activations by platelets was greatly inhibited by anti-CD62P antibody. CONCLUSION:: The ability of polysulfone membranes to activate blood cells is closely related to platelet-neutrophil interaction. Therefore, a biocompatible membrane, like NV-U, can be expected to prevent microaggregate formation during hemodialysis and avoid subsequent cell activation.

Reconstitution of adhesive properties of human platelets in liposomes carrying both recombinant glycoproteins Ia/IIa and Ib alpha under flow conditions: specific synergy of receptor-ligand interactions.

Liposomes carrying both recombinant glycoprotein Ia/IIa (rGPIa/IIa) and Ib alpha (rGPIb alpha) (rGPIa/IIa-Ib alpha-liposomes) instantaneously and irreversibly adhered to the collagen surface in the presence of soluble von Willebrand factor (VWF) at high shear rates, in marked contrast with translocation of liposomes carrying rGPIb alpha alone on the VWF surface. In the absence of soluble VWF, the adhesion of rGPIa/IIa-Ib alpha-liposomes to the collagen surface decreased with increasing shear rates, similar to liposomes carrying rGPIa/IIa alone. While adhesion of liposomes with exofacial rGPIa/IIa and rGPIb alpha densities of 2.17 x 10(3) and 1.00 x 10(4) molecules per particle, respectively, was efficient at high shear rates, reduction in rGPIb alpha density to 5.27 x 10(3) molecules per particle resulted in decreased adhesion even in the presence of soluble VWF. A 50% reduction in the exofacial rGPIa/IIa density resulted in a marked decrease in the adhesive ability of the liposomes at all shear rates tested. The inhibitory effect of antibody against GPIb alpha (GUR83-35) on liposome adhesion was greater at higher shear rates. Further, the anti-GPIa antibody (Gi9) inhibited liposome adhesion more than GUR83-35 at all shear rates tested. These results suggest that the rGPIa/IIa-collagen interaction dominates the adhesion of rGPIa/IIa-Ib alpha-liposomes to the collagen surface at low shear rates, while the rGPIa/IIa-collagen and rGPIb alpha-VWF interaction complements each other, and they synergistically provide the needed functional integration required for liposome adhesion at high shear rates. This study thus has confirmed for the first time the proposed mechanisms of platelet adhesion to the collagen surface under flow conditions using the liposome system.