Biomedical subjects

W B Tsai

Publications and source records attributed to W B Tsai.

10 recordsLinked to original sources

PubMed2004

Platelet adhesion to radiofrequency glow-discharge-deposited fluorocarbon polymers preadsorbed with selectively depleted plasmas show the primary role of fibrinogen.

Fluorocarbon radio-frequency glow-discharge (RFGD) treatment has previously been shown to cause decreased platelet adhesion despite the presence of adsorbed fibrinogen on the surfaces. In this study platelet adhesion to fluorocarbon RFGD-treated surfaces preadsorbed with human plasma was further examined. A series of plasma deposited fluorocarbon thin films were made by varying the C3F6/CH4 ratio in the monomer feed. The surfaces were preadsorbed with plasma, serum, or plasma selectively depleted of fibronectin, vitronectin, or Von Willebrand factor, and platelet adhesion was measured. We also measured fibrinogen adsorption to the surfaces from plasma, monoclonal antibody binding to adsorbed fibrinogen and SDS elutability of the adsorbed fibrinogen. The antibodies used bind to the three putative platelet binding sites on fibrinogen, namely, M1 antibody binds to the dodecapeptide at the C-terminus of the gamma chain, gamma (402-411), R1 antibody binds to a sequence in the Aalpha chain (87-100) which includes RGDF at Aalpha (95-98) and R2 antibody binds a sequence in the Aalpha chain (566-580) which includes RGDS at Aalpha (572-575). Fibrinogen was found to play a decisive role in mediating platelet adhesion to the fluorocarbon surfaces contacting plasma. Few platelets adhered to the fluorocarbon surfaces preadsorbed with serum, while preadsorption with plasma selectively-depleted of either fibronectin, vitronectin, or von Willebrand factor did not decrease platelet adhesion significantly. Replenishment of exogenous fibrinogen to serum restored platelet adhesion, while replenishment of the other proteins had no effect. Platelet adhesion to the fluorocarbon surfaces was lower than to PET or the methane glow-discharge-treated PET. However, there was no apparent correlation between platelet adhesion and the amount of fibrinogen adsorption or monoclonal antibody binding to surface-bound fibrinogen.

Adsorption↗

PubMed2004

Fyn kinase-tubulin interaction during meiosis of rat eggs.

Prior to fertilization, the spindle of vertebrate eggs must remain stable and well organized during the second meiotic meta-phase arrest (MII). In a previous study we have determined that the completion of meiosis is a Src family kinase (SFK)-dependent event. In the current study we have used the SFK inhibitors, SU6656 and PP2, and demonstrated that inhibition of SFKs caused the formation of a disorganized spindle. The observation that proper organization of an MII spindle is an SFK-dependent process, combined with our previous finding that Fyn kinase is localized at the microtubules (MTs), prompted us to examine the potential role of Fyn in MT signaling. Our results show an association between Fyn and tubulin, the ability of Fyn to phosphorylate tubulin in vitro and stimulation of meiosis completion by injection of a constitutively active form of Fyn (CAF). We suggested that SFKs mediate significant functions during the organization of the MII spindle. In view of CAF injection experiments, and of the pronounced concentration of Fyn kinase at the spindle, we propose that Fyn may play an important role in some aspects of the spindle functions, possibly those involving the MTs.

Animals↗

PubMed2001

Co-adsorbed fibrinogen and von Willebrand factor augment platelet procoagulant activity and spreading.

Previously we observed that platelets adherent to surfaces preadsorbed with blood plasma exhibited 1.3 to 2.4 times greater procoagulant activity than platelets on surfaces adsorbed with fibrinogen (Fg) only. These observations suggested that the adhesion proteins adsorbed from plasma may activate platelets in a cooperative, or synergistic manner. In the present study, polystyrene surfaces adsorbed with both Fg and vWF induced up to three times greater procoagulant activity than surfaces adsorbed with Fg or vWF only. The amounts of Fg and vWF adsorbed from binary mixtures that resulted in increased procoagulant activity were found to be similar to the amounts that adsorbed to PS from 100% plasma. The effect of adsorbed adhesion proteins on platelet spreading was also investigated. The proportion of fully spread platelets increased, depending on the adhesion protein preadsorbed to the surface, in the following order: vWF < Fg < Fn < (vWF + Fg) < Vn < plasma.

Blood Platelets↗

PubMed2000

Platelet adhesion and procoagulant activity induced by contact with radiofrequency glow discharge polymers: roles of adsorbed fibrinogen and vWF.

The potential hemocompatibility of radiofrequency glow discharge (RFGD) polymers made by copolymerization of mixtures of hexafluoropropene and ethylene (C(3)F(6)/C(2)H(4)) or acrylic acid and 1,7-octadiene was investigated using in vitro assays for platelet adhesion and platelet catalyzed thrombin generation. Thrombin generation rate normalized to platelet number was used as a measurement of platelet activation (procoagulant activity). RFGD polymers produced by copolymerization of acrylic acid and 1, 7-octadiene contained varying amounts of carboxylic acid species as determined by electron spectroscopy for chemical analysis (ESCA). These polymers induced little variation in platelet adhesion, thrombin generation, or platelet activation. RFGD polymerization of C(3)F(6) and C(2)H(4) resulted in polymers with varying proportions of fluorinated species, as determined by ESCA. Fibrinogen adsorption from plasma was maximal on a polymer made with 25% C(3)F(6) (75% C(2)H(4)) in the feed. However von Willebrand factor (vWF) adsorption was greater on polymers made with increased %C(3)F(6) in the feed. Platelet adhesion decreased with increasing %C(3)F(6) in the feed. Thrombin generation was lowest for platelets adherent to polymers made from both C(3)F(6) and C(2)H(4). Therefore, procoagulant activity of platelets increased for polymers made with increased %C(3)F(6) in the feed, similar to the trend in vWF adsorption. These findings suggest that increased incorporation of fluorinated species into RFGD polymers leads to decreased platelet adhesion and increased platelet activation (which is possibly due to increased vWF adsorption).

Adsorption↗

PubMed2000

The effect of adsorbed fibrinogen, fibronectin, von Willebrand factor and vitronectin on the procoagulant state of adherent platelets.

Procoagulant (activated) platelets provide a site for assembly of the prothrombinase complex which can rapidly convert prothrombin into thrombin (a potent inducer of clot formation). Previously, we reported that adhesion of platelets to surfaces preadsorbed with blood plasma caused them to become procoagulant. In the present study we investigated the effect of adsorbed adhesion proteins (fibrinogen (Fg), fibronectin (Fn), von Willebrand factor (vWF) and vitronectin (Vn)) on the procoagulant activity of adherent platelets. Adsorbed Fn, vWF and Fg promoted platelet adhesion in the following order: Fn < vWF = Fg. However, these proteins promoted platelet activation (thrombin generation per adherent platelet) in the following order: Fg < Fn < vWF. Adsorption with a series of dilutions of normal plasma, serum, and plasmas deficient in or depleted of von Willebrand factor (de-vWF), fibronectin (de-Fn), vitronectin (de-Vn), or both vitronectin and fibronectin (de-VnFn) resulted in varied platelet adhesion, but little difference in platelet activation. However, preadsorption with dilute de-vWF plasma induced lower procoagulant activity than normal plasma. Preadsorption with normal plasma resulted in higher levels of platelet activation than preadsorption with Fg, suggesting that adsorption of plasma proteins other than Fg caused the high levels of activation observed for plasma preadsorbed surfaces.

Adsorption↗

PubMed1999

Template-imprinted nanostructured surfaces for protein recognition.

Synthetic materials capable of selectively recognizing proteins are important in separations, biosensors and the development of biomedical materials. The technique of molecular imprinting creates specific recognition sites in polymers by using template molecules. Molecular recognition is attributed to binding sites that complement molecules in size, shape and chemical functionality. But attempts to imprint proteins have met with only limited success. Here we report a method for imprinting surfaces with protein-recognition sites. We use radio-frequency glow-discharge plasma deposition to form polymeric thin films around proteins coated with disaccharide molecules. The disaccharides become covalently attached to the polymer film, creating polysaccharide-like cavities that exhibit highly selective recognition for a variety of template proteins, including albumin, immunoglobulin G, lysozyme, ribonuclease and streptavidin. Direct imaging of template recognition is achieved by patterning a surface at the micrometre scale with imprinted regions.

Adsorption↗

PubMed1999

Human plasma fibrinogen adsorption and platelet adhesion to polystyrene.

The purpose of this study was to further investigate the role of fibrinogen adsorbed from plasma in mediating platelet adhesion to polymeric biomaterials. Polystyrene was used as a model hydrophobic polymer; i.e., we expected that the role of fibrinogen in platelet adhesion to polystyrene would be representative of other hydrophobic polymers. Platelet adhesion was compared to both the amount and conformation of adsorbed fibrinogen. The strategy was to compare platelet adhesion to surfaces preadsorbed with normal, afibrinogenemic, and fibrinogen-replenished afibrinogenemic plasmas. Platelet adhesion was determined by the lactate dehydrogenase (LDH) method, which was found to be closely correlated with adhesion of 111In-labeled platelets. Fibrinogen adsorption from afibrinogenemic plasma to polystyrene (Immulon I(R)) was low and <10 ng/cm2. Platelet adhesion was absent on surfaces preadsorbed with afibrinogenemic plasma when the residual fibrinogen was low enough (<60 microg/mL). Platelet adhesion was restored on polystyrene preadsorbed with fibrinogen-replenished afibrinogenemic plasma. Addition of even small, subnormal concentrations of fibrinogen to afibrinogenemic plasma greatly increased platelet adhesion. In addition, surface-bound fibrinogen's ability to mediate platelet adhesion was different, depending on the plasma concentration from which fibrinogen was adsorbed. These differences correlated with changes in the binding of a monoclonal antibody that binds to the Aalpha chain RGDS (572-575), suggesting alteration in the conformation or orientation of the adsorbed fibrinogen. Platelet adhesion to polystyrene preadsorbed with blood plasma thus appears to be a strongly bivariate function of adsorbed fibrinogen, responsive to both low amounts and altered states of the adsorbed molecule.

Adsorption↗

PubMed1999

The role of fibronectin in platelet adhesion to plasma preadsorbed polystyrene.

Platelet adhesion to synthetic surfaces that come in contact with blood is mediated by the adsorption of adhesive plasma proteins, especially fibrinogen. However, the roles of other adhesive proteins, such as fibronectin, vitronectin, and von Willebrand factor in platelet adhesion are not yet clear. In this study, the role of fibronectin in platelet adhesion to surfaces was assessed using three approaches. First, platelet adhesion was measured on Immulon I preadsorbed with fibronectin-depleted plasma or fibronectin-depleted plasma replenished with increasing amount of fibronectin. Under these conditions, fibronectin adsorbed from plasma did not have any effect on platelet adhesion, while fibrinogen played a major role in mediating platelet adhesion. Since fibronectin might play a role in platelet adhesion to surfaces which adsorb little or no fibrinogen, we also used two other strategies to assess the potential role of fibronectin. One was to use platelets treated with a platelet activation inhibitor, prostaglandin E1, which prevents the activation of platelet fibrinogen receptor GP IIb/IIIa. The adhesion of prostaglandin E1-treated platelets to Immulon I preadsorbed with plasma was greatly decreased compared to that of untreated platelets, but was increased by the addition of supernormal concentrations of fibronectin to the plasma. This suggests that GP Ic/IIa, rather than GP IIb/IIIa, might be the platelet receptor which is responsible for platelet adhesion to surface-bound fibronectin. Finally, we studied the effect of fibronectin on platelet adhesion to surfaces preadsorbed with fibronectin-depleted afibrinogenemic plasma. We found that fibronectin re-addition to fibronectin-depleted afibrinogenemic plasma increased platelet adhesion. However, our most important finding was that fibronectin seems to play little or no role in mediating platelet adhesion to polystyrene surfaces preadsorbed with normal plasma.

Alprostadil↗

PubMed1998

Hemocompatibility of treated polystyrene substrates: contact activation, platelet adhesion, and procoagulant activity of adherent platelets.

Platelet adhesion to biomaterials is often used as an index of blood compatibility, but a more clinically relevant issue is whether the adherent platelets are able to promote clot formation (i.e., if they are in the procoagulant state). Platelets rapidly generate thrombin when they are in the procoagulant state and the VA/Xa complex is present. We found that adherent platelets are procoagulant by three different methods: binding of FITC-Annexin V, acceleration of thrombin generation in the presence of Xa, Va, and prothrombin; and clotting of recalcified plasma. In the clotting times studies, the effect of adherent platelets on TCPS was completely eliminated by the addition of Annexin V, which is known to bind tightly to procoagulant platelets. The degree of procoagulant activity of adherent platelets was determined by measuring thrombin generation rates in the presence of the clotting factors Va, Xa, and prothrombin and normalizing to the number of adherent platelets. Two key observations were made in these studies. First, the procoagulant activity of platelets adherent to untreated and to several types of treated polystyrenes, as well as to glass and PET, was much greater than the procoagulant activity of unstimulated bulk phase platelets. Little difference in the procoagulant activity of adherent platelets was observed among the materials tested, however. Second, the procoagulant activity of platelets prestimulated with ionophore and subsequently allowed to adhere to Plastek M was much greater than when adherent platelets were stimulated by the adhesion event only. Measured values for platelet adhesion, platelet activation, and contact activation of blood plasma are discussed in the context of their potential combined impact on blood clotting.

Annexin A5↗

PubMed1993

Primary structure and characterization of an androgen-stimulated autoantigen purified from mouse seminal-vesicle secretion.

A protein extract of mouse seminal-vesicle secretion was used to immunize mature mice (Balb/c) of both sexes. Results of Western-blot analyses for these secretory proteins indicated that only one minor protein component could be recognized by the autoantisera prepared from either autoimmunization of male mice or isoimmunization of female mice. The autoantigen was purified from seminal-vesicle secretion. The purified autoantigen retained the ability to induce autoantibody formation. The autoantigen has glycoprotein characteristics: the majority of the carbohydrate is N-linked and the remainder is O-linked. Rabbit antibodies to the autoantigen were used to isolate the corresponding cDNA from a mouse seminal-vesicle cDNA library. The primary structure deduced from the cDNA sequence was confirmed by direct amino acid sequence determination. The results indicate that the core protein consists of 131 amino acid residues. Analysis of the primary structure indicates that the autoantigen has two potential acceptor sites for the N-linked carbohydrate at Asn-12 and Asn-122, three potential phosphorylation sites for casein kinase II at Thr-55, Ser-68 and Thr-76, and three potential phosphorylation sites for protein kinase C at Thr-28, Thr-40 and Thr-124. The core protein and the carbohydrate portion together have a molecular mass of 19 kDa. Results from Western- and Northern-blot analyses for various tissues indicate that the seminal vesicle is the sole organ producing this autoantigen. Expression of this autoantigen gene was stimulated by testosterone.

Amino Acid Sequence↗