Hemocompatibility Of Biomaterials Research Articles

Formation of viscoelastic protein layers on polymeric surfaces relevant to platelet adhesion

The hemocompatibility of biomaterials is highly dependent on the adhesion and activation of platelets. Surface-adsorbed fibrinogen has a dominant role in promoting platelet adhesion to artificial surfaces by binding glycoprotein IIb-IIIa (GPIIb-IIIa), the major platelet membrane receptor. Using quartz crystal microbalance with dissipation monitoring (QCM-D), we have investigated the material-dependent binding kinetics of purified GPIIb-IIIa to polymer-adsorbed fibrinogen. The following ranking of polymer-adsorbed mass (fibrinogen and GPIIb-IIIa) to test polymers could be established: poly[desaminotyrosyl-tyrosine ethyl (DTE) carbonate]/poly(lactide-co-glycolide)>poly[DTE co-5% poly(ethylene glycol) carbonate]. The QCM-D fibrinogen adsorption data were confirmed using an immunofluorescence assay. A synthetic RGD-containing peptide, but not a control peptide, inhibited GPIIb-IIIa binding to polymer-adsorbed fibrinogen, demonstrating the specificity of binding. Importantly, the binding efficiency of purified GPIIb-IIIa to polymer-adsorbed fibrinogen correlated with increased platelet adhesion in an in vitro model. Theoretical simulations using a Voight-based model provided quantitative data on the thickness and viscoelastic properties of the polymer-adsorbed protein layers. The precision of the modeling technique was limited with respect to the shear moduli values, leading to large variations. However, the other modeling parameters showed reproducible results. The thickness of both protein layers was polymer-dependent and ranged from 5 to 35 nm and the viscosity from 0.001 to 0.005 kg/ms, whereas the protein layer densities showed little differences between the test polymers. These results suggest that material-dependent changes in the thickness and viscoelastic properties of adsorbed fibrinogen-GPIIb-IIIa layers are crucial factors in the binding behavior of platelets to biomaterials.

Prevention of Deep Vein Thrombosis and Pulmonary Embolism

Venous thromboembolism comprises deep vein thrombosis (DVT) and pulmonary embolism (PE) and strikes more than 1 in 1000 adults per year, causing discomfort, suffering, and occasionally death. DVT is defined as blood clots in the pelvic, leg, or major upper-extremity veins. These clots can break off from the veins, travel through the heart, and lodge in the lung arteries, causing potentially deadly PE. Although about 300 000 new cases are diagnosed yearly in the United States, probably 3 to 4 times as many cases occur without obvious symptoms and are never detected. This illness is often “silent” and can mimic other common conditions such as heart attack, pneumonia, and anxiety. Its aftermath spans a wide spectrum, from inconsequential to fatal. Awareness of DVT and PE is the best way to prevent this condition. Medical professionals have recognized DVT for almost 2 centuries, but until recently, only about half of Americans were informed about the disease. Without knowledge of DVT as a medical problem, the public could not engage healthcare providers to discuss lifestyle changes and more intensive measures that usually succeed in preventing this illness. Historically, many prominent public figures have been afflicted with DVT but received little attention. However, …

Analysis of immobilized L-cysteine on polymers.

Recently, we reported that L-cysteine attached to polymeric biomaterials, without prior nitrosation, enhances the hemocompatibility of biomaterials via exploiting endogenous nitric oxide (NO). As part of the polymer optimization process to further enhance platelet inhibition, a kinetic model is being developed to predict the release rate of NO. A key model parameter is the immobilized concentration of L-cysteine. This article demonstrates how several chemiluminescence-based assays, previously utilized for measuring thiols in solution, were successfully adapted to quantify immobilized L-cysteine. The assays showed that the immobilized L-cysteine on the modified PET sample is within the range of 4.1 to 6.5 nmol/cm(2). An advantage of using the more successful chemiluminescence-based assay is that it can accurately measure molar concentrations of any thiol-containing compound with a detection limit in the pmol range. The major disadvantage is that L-cysteine must first be broken off of the polymer and released into solution prior to measurement, therefore leaving the sample unable to be reused. Other thiol-measuring techniques, such as fluorescence microscopy and X-ray photoelectron spectroscopy (XPS), were used to provide qualitative and semiquantitative analysis to substantiate the polymer development.

Regioselectively Modified Cellulose and Chitosan Derivatives for Mono- and Multilayer Surface Coatings of Hemocompatible Biomaterials

In this study different synthetic strategies were developed and applied to introduce solely or in combination heparin/heparansulfate-like functional groups such as N-sulfo, O-sulfo, N-acetyl, and N-carboxymethyl groups into chitosan and cellulose with highest possible regioselectivity and completeness and defined distribution along the polymer chain. Completely substituted 6-amino-6-deoxycellulose and related derivatives were prepared from tosylcellulose (DS 2.02; C6 1.0) by nucleophilic substitution with azido groups only in the 6-position at 50 °C with subsequent reduction to amino groups and completely removing tosyl groups in the 2,3-position. 2,6-Di-O-sulfocellulose was prepared using the reactivity difference between C-2, C-6 and C-3 of cellulose. The reactivity difference between amino groups and hydroxyl groups was used to prepare various N-substituted derivatives. Partially 2,6-di-O-sulfated cellulose was obtained from trimethylsilylcellulose by the insertion of sulfurtrioxide into the Si–O ether linkage. Partially 3-O-sulfocellulose was synthesized by protecting C-2 and C-6 with trifluoroacetyl groups. A copper–chitosan complex was used to synthesize 6-O-sulfochitosan with a DS of 1.0 at C-6 and various partially 6-O-desulfonated products are possible. Using the phthalimido group to increase the solubility of chitosan in DMF, the regioselectivity of 3-O-sulfo groups was improved by regioselective 6-O-desulfonation of nearly complete 3,6-O-disulfochitosan. The platelet adhesion properties of immobilized regioselectively modified water-soluble derivatives on membranes have been tested in vitro. Some regioselectively modified chitosan and cellulose derivatives are potential candidates for the surface coatings of biomaterials if the regioselective reactions are somewhat further optimized.

Cardiology patient pages. Pulmonary embolism and deep vein thrombosis.

The heart pumps oxygenated blood through the aorta to smaller arteries. After the blood supplies nutrients to vital organs, it returns through veins for reoxygenation in the lungs (Figures 1 and 2⇓). Blood clots called deep vein thrombi (DVT) often develop in the deep leg veins. Pulmonary embolism (PE) occurs when clots break off from vein walls and travel through the heart to the pulmonary arteries. The broader term venous thromboembolism (VTE) refers to DVT, PE, or to a combination of both. Figure 1. The arterial (red) and venous systems (blue) are depicted. The left ventricle (LV) ejects oxygenated blood into the aorta, which circulates blood to vital organs. Deoxygenated blood travels through the venous system to the right atrium, right ventricle, and pulmonary artery. Adapted with permission from MediClip, Clinical Cardiopulmonary Images 1997, CCP01026.TIF, Williams & Wilkins, Baltimore, Md. Figure 2. Deoxygenated blood returns to the heart and lungs for reoxygenation. SVC indicates superior vena cava; IVC, inferior vena cava; RA, right atrium; RV, right ventricle; and PA, pulmonary artery. Adapted with permission from MediClip, Clinical Cardiopulmonary Images 1997, CATHRHT.TIF, Williams & Wilkins, Baltimore, Md. VTE poses a public health threat with an estimated incidence in the United States of 250 000 to 2 million cases per year. Predisposition to VTE arises from acquired conditions, inherited disorders, or both. Many of the acquired risk factors can be modified, thus lessening the likelihood of PE or DVT. Long-haul air travel is the most talked-about risk factor for PE. Other acquired risk factors include obesity, …

Improved haemocompatibility of cysteine-modified polymers via endogenous nitric oxide

A novel method for improving the haemocompatibility of biomedical materials through endogenous nitric oxide (NO) is presented. l-cysteine was covalently immobilized onto two biomedical polymers: polyurethane (PU) and polyethylene terephthalate (PET). The l-cysteine content on the polymers was approximately 5–8 nmol/cm 2 as quantified via a chemiluminescence-based assay. The haemocompatibility of the modified polymers was evaluated in terms of the number of adhered platelets when exposed to a platelet suspension labeled with Cr 51. Platelet adherence on the l-cysteine-modified polymers was reduced more than 50% as compared to the control (glycine-modified polymers) when the platelet suspension contained plasma constituents. No difference in platelet adhesion was observed in the absence of plasma constituents. Further experiments demonstrated that NO was easily transferred to the l-cysteine-modified polymers from S-nitroso-albumin in PBS buffer. The NO was then released from the polymer. NO transfer or release was not observed for the control. The results suggest that l-cysteine-modified polymers are effective in reducing platelet adhesion via the transfer of NO from endogenous S-nitrosoproteins in plasma to the polymer followed by the subsequent release of NO. Thus, exploiting endogenous NO is a viable option for improving the haemocompatibility of biomaterials.

The Haemocompatibility of Biomaterials In Vitro: Investigations on the Mechanism of the Whole-blood Clot Formation Test

Twenty years ago Imai & Nose introduced a whole-blood clotting test for the estimation of haemocompatibility of biomaterials in vitro In our paper a modification of this assay is described and the mechanism of clot formation further elucidated. It was found that neither the inhibition of platelet function nor the removal of platelets from blood significantly changed the clot formation rate on glass and polyvinyl chloride in comparison to the rate tor whole blood. Scanning electron microscopy demonstrated that platelets were not involved in clot formation near the blood/biomaterial interface. Thus, it was concluded that the system of contact activation of the coagulation cascade dominates during clot formation under static conditions. The latter conclusion was supported by the fact that preadsorption of human serum albumin or human fibrinogen onto the glass plates used, decreased the clot formation rate in the same manner.

Cellular morphology and distribution on a stretching blood-material interface.

In order to investigate the interactions of cellular elements and protein on constantly deforming (fatiguing) blood contact surfaces, a series of ex vivo canine arteriovenous shunt experiments were conducted. While fresh blood was flowing through Silastic tubing shunts, portions of the tubing were stretched 20 to 60% at a frequency of 20 to 90 cycles per minute for 10 to 90 min. The surfaces of the tubing that were stretched were compared with control tubing surfaces taken from the arterial side of the test segment using scanning electron microscopy and interference phase contrast microscopy. Approximately the same number of platelets were deposited on the stretched as on the unstretched portions of the tubing in the ten minute experiments. On the control portions of the tubing, the platelets were deposited singly and uniformly in what appeared to be a fairly inactivated state. On the stretched tubing, more pseudopod extension and aggregation was observed. In these preliminary experiments, no differences were noted as a function of frequency of stretch. As the blood contact time and the percent stretch were increased, only nonuniform, scattered aggregations of platelets, and platelets mingled with fibrin were seen. Significant numbers of spread white blood cells were observed on many of the segments of Silastic tubing stretched 20% for as short a time as 15 min. Granulocytes have occasionally been reported on less hemocompatible biomaterials after exposure to canine blood. This helps to confirm that substrate stretching of 20-60% had an adverse effect on the blood compatibility of the Siliastic tubing.

Adsorption-desorption processes of proteins at solid/blood interfaces

Many attempts have been made to find one common physicochemical determinator of the hemocompatibility of biomaterials. In terms of general laws, a universal mechanism of hemocompatibility has not yet been found. We propose a pragmatic approach to analyze the changes in the adsorption-desorption processes of plasma proteins as functions of controlled variations of specific surface properties. We use a radioisotopic method to measure the kinetics of formation of an irreversible protein layer. The character of the reversible adsorption was estimated via kinetic parameters of surface-induced complement activation. For commercial polyetherurethane alterations of surface morphology, in particular the domain form, size, and distribution, was shown to change the type of protein adsorption. The existence of four types of protein adsorption on the surfaces was established.