Biomedical subjects

H Elwing

Publications and source records attributed to H Elwing.

At least 19 recordsLinked to original sources

PubMed2001

Variations in coupled water, viscoelastic properties, and film thickness of a Mefp-1 protein film during adsorption and cross-linking: a quartz crystal microbalance with dissipation monitoring, ellipsometry, and surface plasmon resonance study.

We have measured the time-resolved adsorption kinetics of the mussel adhesive protein (Mefp-1) on a nonpolar, methyl-terminated (thiolated) gold surface, using three independent techniques: quartz crystal microbalance with dissipation monitoring (QCM-D), surface plasmon resonance, and ellipsometry. The QCM-D and ellipsometry data shows that, after adsorption to saturation of Mefp-1, cross-linking of the protein layer using NaIO4 transforms it from an extended (approximately 20 nm), water-rich, and hydrogel-like state to a much thinner (approximately 5 nm), compact, and less water-rich state. Furthermore, we show how quantitative data about the thickness, shear elastic modulus, and shear viscosity of the protein film can be obtained with the QCM-D technique, even beyond the Sauerbrey regime, if frequency (f) and energy dissipation (D) measurements measured at multiple harmonics are combined with theoretical simulations using a Voight-based viscoelastic model. The modeling result was confirmed by substituting H2O for D2O. As expected, the D2O substitution does not influence the actual adsorption behavior, but resulted in expected differences in the estimated effective density and shear viscosity. These results provide new insight and understanding about the adsorption kinetics and crosslinking behavior of Mefp-1. They also demonstrate how the above three techniques complement each other for biomolecule adsorption studies.

Adsorption↗

PubMed2001

In vitro study of monocyte viability during the initial adhesion to albumin- and fibrinogen-coated surfaces.

Surface adherent monocytes and macrophages play a central role in the inflammatory response to biomaterials. In the present study the adhesion, viability and apoptotic changes in material surface adherent monocytes during the first hours of cell-surface interactions in vitro were studied, using tissue culture polystyrene surfaces coated with human albumin and fibrinogen. Human peripheral blood monocytes were enriched by a two-step gradient centrifugation and resuspended (1 x 10(6)/ml) in RPMI with 10% fetal bovine serum. The cells were added to polystyrene surfaces coated with human fibrinogen or albumin and incubated in 37 degrees C (5% CO2, 100% humidity) for 30 min, 1, 2, 3 and 24 h. The adherent cells were stained for early apoptotic changes (exposed phosphatidylserine) and cell death using Annexin-V-fluorescein and propidium iodide staining, respectively. A bi-phasic adhesion was observed on the fibrinogen coated surface, having the highest number of adherent cells after 30 min and 24 h, while the cell number was markedly reduced after 1-3 h. The number of adherent cells on albumin was relatively low after all short time incubations but had reached a high level after 24 h. The number of adherent dead cells was highest after I h on both albumin (approximately 30%) and fibrinogen (approximately 15%). In the 24 h cultures, the viability of adherent cells was high on both surfaces (95-100%). Viable cells staining positive for early apoptotic changes could only be clearly observed on the albumin coated surface, after 30 min of cell-material surface interaction. Cell death, including apoptotic death, thus seems to play an important role during the initial interactions between monocytes and a foreign surface.

Albumins↗

PubMed1999

Effect of ionic strength on initial interactions of Escherichia coli with surfaces, studied on-line by a novel quartz crystal microbalance technique.

A novel quartz crystal microbalance (QCM) technique was used to study the adhesion of nonfimbriated and fimbriated Escherichia coli mutant strains to hydrophilic and hydrophobic surfaces at different ionic strengths. This technique enabled us to measure both frequency shifts (Deltaf), i.e., the increase in mass on the surface, and dissipation shifts (DeltaD), i.e., the viscoelastic energy losses on the surface. Changes in the parameters measured by the extended QCM technique reflect the dynamic character of the adhesion process. We were able to show clear differences in the viscoelastic behavior of fimbriated and nonfimbriated cells attached to surfaces. The interactions between bacterial cells and quartz crystal surfaces at various ionic strengths followed different trends, depending on the cell surface structures in direct contact with the surface. While Deltaf and DeltaD per attached cell increased for nonfimbriated cells with increasing ionic strengths (particularly on hydrophobic surfaces), the adhesion of the fimbriated strain caused only low-level frequency and dissipation shifts on both kinds of surfaces at all ionic strengths tested. We propose that nonfimbriated cells may get better contact with increasing ionic strengths due to an increased area of contact between the cell and the surface, whereas fimbriated cells seem to have a flexible contact with the surface at all ionic strengths tested. The area of contact between fimbriated cells and the surface does not increase with increasing ionic strengths, but on hydrophobic surfaces each contact point seems to contribute relatively more to the total energy loss. Independent of ionic strength, attached cells undergo time-dependent interactions with the surface leading to increased contact area and viscoelastic losses per cell, which may be due to the establishment of a more intimate contact between the cell and the surface. Hence, the extended QCM technique provides new qualitative information about the direct contact of bacterial cells to surfaces and the adhesion mechanisms involved.

Bacterial Adhesion↗

PubMed1998

Desensitization of formyl peptide receptors is abolished in calcium ionophore-primed neutrophils: an association of the ligand-receptor complex to the cytoskeleton is not required for a rapid termination of the NADPH-oxidase response.

Binding of ligands to N-formyl peptide chemoattractant receptors exposed on human neutrophils generates signals in the cells that induce an activation of the superoxide anion producing NADPH-oxidase. Ligand binding is followed by a rapid association of the ligand-receptor complex with the cytoskeleton, a process leading to desensitization of the cells with respect to NADPH-oxidase activation. We show that neutrophils that have experienced an intracellular calcium rise obtained through interaction with the calcium-specific ionophore ionomycin are "primed" with respect to the FMLP-induced production of superoxide anions. Mobilization of FMLP receptors from intracellular pools is one well-known mechanism behind the primed response. Based on our finding that ionomycin-treated neutrophils could not be desensitized, we suggest that the lack of association between the ligand-receptor complex and the cytoskeleton is an additional priming mechanism. Since in vivo-exudated neutrophils, which also had mobilized intracellular organelles, could be desensitized, we suggest that the abolished desensitization in ionomycin-treated neutrophils is not due to an inability of newly recruited receptors to couple to the cytoskeleton. We show that a rapid termination of FMLP-induced superoxide anion production is obtained in both desensitizable and nondesensitizable neutrophils, suggesting that the desensitization phenomenon is of limited importance in the oxidase termination process.

Adult↗

PubMed1998

Protein absorption and ellipsometry in biomaterial research.

Ellipsometry is an optical surface-sensitive method for the investigation of various aspects of protein adsorption mainly at reflecting metal surfaces and ceramic surfaces. One interesting feature of the method is the possibility of detailed and accurate determination of real-time adsorption kinetics of proteins without labelling of the protein. It is also possible to detect protein adsorption with the use of antibodies that adsorb onto the antigen-coated surfaces and to detect antibodies by their adsorption behaviour with regard to antigen-coated surfaces. Compared to other solid phase methods such as enzyme linked immunosorbent assay (ELISA), immunofluorescence and radioimmunoassay, ellipsometry has the advantage of not involving any labelling of the reactant and it is a relatively inexpensive method to maintain. This review is a current report of 15 years of contributions in biomaterials and biochemical research. Special consideration has been given to biologically related surface phenomena such as protein conformation changes, protein displacement effects, early events in blood clotting and complement activation. Among the technical achievements given prominence are the wettability gradient method and the rational use of silicon as an experimental surface. It is clear that ellipsometry and related methods such as reflectometry and surface plasmon resonance (Biacore) are now being increasingly used in biomaterial research as well as in other areas of research.

Absorption↗

PubMed1997

Adsorption to silica nanoparticles of human carbonic anhydrase II and truncated forms induce a molten-globule-like structure.

Human carbonic anhydrase II pseudo-wild type (HCAIIpwt) and two truncated variants were adsorbed to approximately 9 nm silica nanoparticles. Ellipsometry was used as an indirect measure of protein adsorption. The structural changes of adsorbed proteins were investigated with the use of circular dichroism (CD), intrinsic fluorescence, ANS binding ability and inhibitor binding capacity. It was found that the variants that were truncated at positions 5 and 17 in the N-terminal end attain a molten-globule-like state after interaction with the silica nanoparticles. In contrast, the more stable HCAIIpwt retained most of its native structure after 24 h adsorption to silica nanoparticles. The result suggests that surface induced unfolding may give rise to intermediates similar to those for unfolding induced by, for example GuHCl. Thus, the intermediate observed has some features of the molten globule.

Adsorption↗

PubMed1997

Surface-related triggering of the neutrophil respiratory burst. Characterization of the response induced by IgG adsorbed to hydrophilic and hydrophobic glass surfaces.

Hydrophilic and hydrophobic glass surfaces precoated with human albumin, fibrinogen, or IgG were investigated with respect to their ability to activate the neutrophil NADPH-oxidase. We found that IgG-coated surfaces induced a substantial and prolonged neutrophil production of reactive oxygen species (ROS). When a hydrophilic surface was used to support protein binding, a somewhat lower neutrophil response (around 35%) was obtained, compared with the response induced by IgG on a hydrophobic surface. The production of ROS was completely eliminated when cytochalasin B was added to the measuring system, suggesting the involvement of the cell cytoskeleton in the activation process. The relation between the intra- and extracellular generation of ROS was further assessed, and we found that most of the ROS produced were released from the cells, in agreement with a model in which the activating surfaces induce a 'frustrated' phagocytic response. Serum totally inhibited 'frustrated' phagocytosis provided that the IgG molecules were sticking to a hydrophilic surface.

Albumins↗

PubMed1996

A simple chemiluminescence assay for the determination of reactive oxygen species produced by human neutrophils.

We show that phagocyte production of reactive oxygen species can be measured using a microtitre plate based chemiluminescence blotting technique. The production of reactive oxygen species is determined by their ability to catalyze the oxidation of luminol or isoluminol, resulting in light emission which is recorded on a photographic film. The method permits the determination of NADPH oxidase activity from as few as 9000 cells. It could be used to detect NADPH oxidase defects in neutrophils (e.g. from patients suffering from chronic granulomatous disease), and to screen pharmaceuticals with scavenging activity for reactive oxygen species.

Humans↗

PubMed1996

Complement activation on thiol-modified gold surfaces.

To evaluate the importance of protein adsorption and chemical composition of the solid surface on complement activation, we used mercaptoglycerol (MG) and mercaptopropionic acid (MPA) modified gold surfaces as model surfaces. The complement activation by these surfaces was determined by measuring fluid phase iC3b and C5b-9 in serum that had been in contact with the surfaces. In addition, "active" C3 deposition at the modified surfaces was measured with the use of ellipsometry, an optical technique. It was found that the MG surface caused pronounced productions of iC3b and C5b-9 in serum as well as increased C3 deposition on the surface. In contrast, the bare gold surface and the MPA surface caused very little complement activation. The MG surface seems to have a high affinity with immunoglobulin G (IgG) that may be one explanation for the high complement activation ability of the MG surface. However, complement activation at the MG surface was relatively insensitive to Mg-EGTA buffer at a high serum concentration, indicating alternative pathway activation when classical pathway activation was inhibited. Corresponding control experiments performed with Mg-EGTA serum at hydrophobic silicon surfaces precoated with IgG showed no sign of alternative pathway activation. At a lower serum concentration, classical pathway activation seemed to dominate at the MG surface and the hydrophobic silicon surface precoated with IgG. Most probably, it is the hydroxyl groups at the MG surface that are associated with alternative pathway activation.

3-Mercaptopropionic Acid↗

PubMed1994

Complement activation on solid surfaces as determined by C3 deposition and hemolytic consumption.

Complement activation was investigated on hydrophilic and hydrophobic glass beads incubated in serum. Very little complement activation was detected with these surfaces, as indicated by a hemolytic assay and by measurement of the amount of iC3b appearing in the solution. However, preadsorption with IgG at the hydrophobic and hydrophilic beads resulted in complement activation on both surfaces. We also investigated dependent deposition of C3 at hydrophobic and hydrophilic silicon surfaces when the complement was activated. The chemistry of those surfaces is similar to the hydrophobic and hydrophilic beads. Ellipsometry, an optical method, was used for determination of the amounts of organic material deposited at the surface. C3 deposition was observed at the IgG precoated hydrophobic surface but not at the IgG-coated hydrophilic surface. The absence of C3 deposition at the hydrophilic surface was probably due to reversible binding of IgG. However, precoating of the hydrophilic surface with a double layer of IgG and anti-IgG resulted in C3 deposition also at the hydrophilic surface. The results illustrate that methods based on measuring deposition of C3 at surfaces, such as immunofluorescence or ellipsometry, cannot exclude surface-associated complement activation that is probably due to reversible binding of the complement components of the activated molecule. On the other hand, it has previously been shown that determination of complement activation in solution with the use of a C3a assay cannot exclude surface-associated activation due to immobilization of C3a at the surface. This methodologic question is an important issue because factors such as C3a and C5a act as soluble anaphylatoxins, whereas deposited factors such as C3 act as cellular receptors.

Adsorption↗

PubMed1994

Lens-on-surface method for investigating adhesion of Staphylococcus aureus to solid surfaces incubated in blood plasma.

Adhesion of Staphylococcus aureus was investigated on flat silicon oxide surfaces that had been incubated in human plasma at different concentrations. Adhesion of bacteria did not occur at high incubation concentrations of plasma or when the surface had been incubated in egg albumin. However, significant adhesion was observed when plasma was diluted. With the use of antibody method, it was noted that the adhesion of the bacteria coincided with adsorbed fibrinogen, and possibly also with IgG. We also investigated the effect of "narrow space" on the adsorption of blood plasma and subsequent adhesion of S. aureus. In these experiments, blood plasma was incubated under a convex lens placed upside-down on the silicon oxide surface. This method creates a continuous gradient of space from the contact point of the lens and outward. After rinsing off the plasma and the lens, the surface was incubated with a suspension of S. aureus followed by quantification of the attached bacteria by means of optical methods. Adhesion of bacteria occurred in several circular zones that were easily detectable with the naked eye or by the means of simple optical methods. In addition, in these experiments, adhesion coincided with adsorbed fibrinogen or IgG at the surfaces. The increased bacterial adhesion to surfaces incubated in diluted plasma, or plasma incubated in narrow space, is a variant of the so-called "Vroman effect." With a model protein system consisting of fibrinogen and IgG and the corresponding antibodies, we demonstrate that "dilution" and "incubation in narrow space" are two phenomenologically similar methods.(ABSTRACT TRUNCATED AT 250 WORDS)

Adsorption↗

PubMed1994

Effects of odorants on pigment aggregation and cAMP in fish melanophores.

Odor perception within olfactory neuroepithelium and pigment translocation within melanophores both seem to rely on a cAMP-based second messenger system. From studies on cultured frog melanophores, Lerner et al. (Proc. Natl. Acad. Sci. USA 85:261-264, 1988) suggested that some aspect of odor perception may be mediated by a nonspecific mechanism whose signal is transduced by a cAMP-based second messenger system. In the present study, odorants (beta-ionone, benzylaldehyde, cineole, cinnamaldehyde, and octanol), which previously have been shown to stimulate formation of cAMP in the olfactory neuroepithelium, were investigated for possible pigment dispersing and cAMP-increasing effects. Pretreatment of fish melanophores with the adenylate cyclase activator forskolin (1 microM) resulted in an approximately 300% increase in cAMP and an almost complete blockage of noradrenaline-induced pigment aggregation. However, none of the tested odorants were able to increase the cAMP level and only cinnaldehyde and beta-ionone were found to have any pigment dispersing activity.

1-Octanol↗

PubMed1994

Adsorption of fibrinogen and some other proteins from blood plasma at a variety of solid surfaces.

Enzyme linked immunosorbent assay (ELISA) was used for the estimation of protein adsorption from blood plasma at some model solid surfaces. The majority of those surfaces were made in the wells of microtiterplates of polystyrene commonly used for ELISA purposes. Three of the model surfaces were made by radio frequency plasma discharge polymerization (RFPD) of the microtiterplates of polystyrene. The monomers we used were diaminocyclohexane, hexamethylenedisiloxane, and acrylic acid. Other surfaces investigated were: unmodified polystyrene, oxidized polystyrene, hydrophilic silicon oxide, and methylized silicon oxide. Two substances, Tween and bovine serum albumin (BSA), for the prevention of unintended adsorption of ELISA conjugate were also tested and the BSA method was found to be superior for this kind of investigation. Human blood plasma at different dilutions was incubated in the surface-modified microtiterplates followed by incubation of rabbit antibodies against fibrinogen (FG), fibronectin (FN), human serum albumin (HSA), complement factor 3 (C3), and immunoglobulin G (IgG). Visualization of bound antibodies was then made by standard ELISA procedure. At low blood plasma concentrations (plasma dil 1/1000), anti-IgG and anti-HSA were detected at high levels at the majority of surfaces. At high blood plasma concentration (plasma dil 1/10), anti-FG dominated at most surfaces. ELISA activity of FN and C3 were low at most of the surfaces at both plasma concentrations. An 'optimum' plasma dilution for the detection of surface bound FG (the Vroman effect) was not found with the use of the ELISA on any of the surfaces except for the silicon oxide surface. This is in contrast to findings by others who had used isotope-labelled fibrinogen diluted in plasma. However, 'false' Vroman effects occurred if nonionic surfactant was used for the prevention of unspecific binding in the ELISA.

Adsorption↗

PubMed1994

The chemiluminescence response of neutrophils on polymer surfaces made by glow discharge plasma polymerization.

Polypropylene tubes were coated with different polymers made by glow discharge plasma polymerization. Isolated human blood neutrophils were allowed to interact with the polymer surface and the chemiluminescence response of the cells was recorded as a measure of oxidative activation. The polymers represented surfaces that differed markedly with respect to charge, hardness, and wettability. We found that all polymers stimulated the chemiluminescence response in neutrophils differently; when preincubation with human serum albumin (HSA) there was a general reduction of the chemiluminescence response particularly on one of the positively charged surface 1,2-diamino-cyclohexane (DACH). Addition of a soluble stimulus, the chemoattractant formylmethionyl-leucylphenylalanine (FMLP), to the cells caused a dramatic increase in the response on one of the hydrophobic surface hexamethylene-disiloxane (HMDSO). However, there was also a pronounced reduction in the response on polymers with acrylic acid (AA). The response was normalized after addition of HSA. Taken together, the chemiluminescence response of the neutrophils interacting with the polymer surfaces differed with regard to the type of surfaces. When HSA and FMLP were added a larger difference in the response was found. Our results showed that the activation of human neutrophil granulocytes influenced by different polymer surfaces, followed unspecific different patterns which were someway related to the specific characteristics of the polymer and from this point we came to similar conclusions made by Kaplan et al. (J. Biomater. Res. 28, 377 (1994)), that it is difficult to extrapolate any activation mechanisms from one material to another.(ABSTRACT TRUNCATED AT 250 WORDS)

Acrylates↗

PubMed1993

Local and non-local receptor signalling.

It is suggested that the physiological response caused by an excited membrane-bound receptor can be either local, i.e. localized close to the receptor site, or non-local, spreading over a large volume in the cell. A simple model is presented which shows how locality and non-locality are related to the intrinsic "efficacy" of the drug used in classical receptor theory. It is further demonstrated how the shape of the physiological response versus receptor occupancy is determined by the number of receptor sites which contribute to the physiological response at a given location in the cell.

Animals↗

PubMed1993

Conformational epitopes of C3 reflecting its mode of binding to an artificial polymer surface.

The aim of the study was to investigate the incompletely understood mechanisms of complement (C) activation and binding on artificial biomaterials. Polystyrene in the form of microtitre plates was used as target for C binding, detectable by ELISA using monoclonal anti-C3 antibodies specific for conformational epitopes expressed by bound C3 and C3 fragments. C3 binding in whole blood/plasma/serum is maximal at low dilutions and occurs predominantly by C activation. At higher dilutions, C3 binding occurs at approximately 1/3 of maximal levels and is solely an effect of adsorption. C3 adsorption in the lower serum dilution range, occurs at low but clearly detectable levels. Comparative epitope analysis between C3 fragments, actively bound to polystyrene in the presence of serum, and of iC3b bound to sheep erythrocytes, clearly indicates that C3 binding/activation on polystyrene takes place as a C3 convertase-mediated reaction, which in serum/plasma is followed by a secondary factor I-dependent degradation of the bound C3b into iC3b. The neo-epitope analysis of serum-contacting polystyrene revealed that the adsorbed C3, throughout the entire serum dilution range tested, deposits in a state closely similar to that observed for purified C3 at a high packing density. Polystyrene surfaces with adsorbed purified C3 expressing this epitope profile were found to mediate APW dependent deposition of C3b in pig serum, presumably by forming a hybrid convertase with porcine Bb. These data therefore suggest that adsorbed C3 on serum-contacting polystyrene surfaces may initiate complement activation via the APW.

Adsorption↗

PubMed1993

Modification of the complement binding properties of polystyrene: effects of end-point heparin attachment.

In recent years, conjugation of heparin to biomaterials has been shown to improve its biocompatibility. The purpose of the present work was to compare complement activation and binding of C3 to unmodified and heparin-treated polystyrene surfaces of microtitre plates. When polystyrene was incubated with human serum, C3 was deposited on the surface by both adsorption and binding dependent on activation of the classical (CPW) and alternative (APW) pathways. After end-point attachment of heparin, significant C3 deposition, although at reduced levels, occurred only by CPW-mediated mechanisms, while adsorption and APW-mediated binding were strongly reduced. Generally, the modified surface bound lower amounts of protein, e.g. serum albumin and IgG, than the unmodified. By contrast, it had increased affinity for C1q which leads to binding of C1 and activation of complement via the CPW. Nevertheless, the net effect of the surface modification on the complement reaction was an overall reduction of C3 binding due to obliteration of APW. This can be related to an enhanced factor H/I-dependent down-regulation of C3b and to the lowered protein-adsorbing property of the surface, both of which have inhibitory effects on APW and on the C3 shunt-dependent activation of the complement system.

Complement Activation↗

PubMed1992

Antisera binding onto metals immersed in human plasma in vitro.

Titanium (Ti), silicon (Si), silver (Ag), vanadium (V), gold (Au), and chromium (Cr) surfaces were made hydrophilic by radiofrequency plasma treatment. The binding of antifibrinogen (a-fib) and anti-high-molecular-weight kininogen (a-HMWK) after incubation of surfaces in 10% human blood plasma was investigated with ellipsometry. Ti and Si surfaces bound a-HMWK but no detectable amounts of a-fib, whereas the other metals bound both types of antisera. Protein adsorption in a fluid gradient produced under a convex lens indicated that the small deposition of a-fib on Ti and Si surfaces is probably a result of protein displacement. This displacement was not evident at lens positions simulating low plasma concentrations where large amounts of a-fib adsorbed. Surfaces washed sequentially in trichloroethane, acetone, and ethanol produced more hydrophobic surfaces and resulted in slower rates of a-fib displacement. These studies would indicate that some metals form complexes with specific plasma proteins or, alternatively, that different metals may bind equal amounts of plasma proteins which express differing antigenicities toward their surroundings.

Adsorption↗