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Biomedical subjects

M Malmsten

Publications and source records attributed to M Malmsten.

At least 37 records · Page 2Linked to original sources
PubMed2000

Formation of model lipid bilayers at the silica-water interface by co-adsorption with non-ionic dodecyl maltoside surfactant.

This present article describes a new and simple method for preparing model lipid bilayers. Stable and reproducible surface layers were produced at silica surfaces by co- adsorbing lipid with surfactant at the silica surface from mixed micellar solutions. The adsorption was followed in situ by use of ellipsometry. The mixed micellar solution consisted of a lipid (L-alpha-dioleoyllecithin) and a non-ionic sugar-based surfactant (n-dodecyl-beta-maltoside). The latter showed, by itself, no affinity for the surface and could, therefore, easily be rinsed off the surface after the adsorption step. By first adsorbing from solutions with high lipid and surfactant concentrations and then, in succession, rinsing and re-adsorbing from solutions with lower lipid-surfactant concentrations, a dense-packed lipid bilayer was produced at the silica surface. The same result can be achieved in a one-step process where the rinsing, after adsorption from the concentrated solution, is performed very slowly. The thickness of the adsorbed lecithin bilayer after this treatment found was to be about 44 +/- 3 A, having a mean refractive index of 1.480 +/- 0.004. The calculated surface excess of lipids on silica was about 4.2 mg m(-2), giving an average area per lipid molecule in the two layers of 62 +/- 3 A2. The physical characteristic of the adsorbed bilayer is in good agreement with previously reported data on bulk and surface supported lipid bilayers. However, in contrast to previous investigations, we found no support for the presence of a thicker multi-molecular water layer located between the lipid layer and the solid substrate.

Adsorption↗
PubMed1999

Orthokinetic Aggregation in Two Dimensions of Monodisperse and Bidisperse Colloidal Systems.

Orthokinetic aggregation of colloids trapped at the air-liquid interface was studied by direct imaging in a couette cell. This method allowed us to follow the temporal evolution of both the cluster-mass distribution and the cluster structure at a shear rate where Brownian aggregation is suppressed. The interactions between the monodisperse latex particles floating at the air-liquid interface were controlled either by varying the electrolyte concentration or by creating a bidisperse system through the addition of small particles. The results show that the clusters in all of the systems are characterized by a high fractal dimension, indicating that the clusters are rearranged and densified by the shear. Kinetic analysis suggests that aggregation of monodisperse systems mainly proceeds through homogeneous aggregation, i.e., large clusters sticking to other large clusters. The bidisperse system, finally, with a size ratio around 10, favored a more heterogeneous aggregation among small and large clusters throughout the aggregation process; a slightly lower fractal dimension was observed compared to the strongly aggregated monodisperse system. Copyright 1999 Academic Press.

Journal Article↗
PubMed1999

Confocal Microscopy Studies of Trypsin Immobilization on Porous Glycidyl Methacrylate Beads.

The immobilization of trypsin on porous glycidyl methacrylate (GMA-GDMA) beads has been investigated. In particular, the distribution within the beads of trypsin and of dextran used for hydrophilizing the bead surface prior to protein immobilization was investigated with confocal microscopy. For the system investigated, the fluorescence intensity profiles obtained when using borate buffer as an ambient solution displayed a distinct minimum at the center of the beads, irrespective of the observation depth. However, by reduction of the refractive index difference between the solution and the beads through the addition of glucose to the aqueous solution, artifacts relating to optical length differences could be reduced. For both low molecular weight fluorescein isothiocyanate (FITC), FITC-labeled trypsin, and FITC-labeled dextran, an essentially homogeneous distribution throughout the beads was observed. This simple "contrast matching" method seems therefore to be an interesting tool when investigating the distribution of immobilized protein in porous chromatography media. Copyright 1999 Academic Press.

Journal Article↗
PubMed1999

Surface characterisation of freeze-dried protein/carbohydrate mixtures.

In the present investigation freeze-drying of proteins (BSA or trypsin) together with various carbohydrates, i.e. lactose, sucrose, mannitol, alpha-cyclodextrin and dextrin, has been studied with particular emphasis on the surface composition of the freeze-dried powders. The proteins were found to be over-represented on the powder surface as compared to the bulk concentration of protein. The mechanism behind the surface accumulation is believed to be that proteins adsorb preferentially over carbohydrates to the ice/liquid interface in the frozen sample. The degree of surface accumulation depended on the carbohydrate used, and was increased in annealed samples compared to reference samples. The activity of trypsin was fairly well preserved (58-90%) in the freeze-dried powders, but depended on the carbohydrate excipient, whilst the surface composition had little effect on the activity. The activity preservation was improved when the protein concentration was raised from 1 to 10% in the solids. The surface composition of powders containing mixtures of mannitol and dextrin as excipients depended on the ratio between the two carbohydrates, with the lowest surface coverage of protein obtained in 50/50 mixtures.

Calorimetry, Differential Scanning↗
PubMed1999

Spray-drying of trypsin - surface characterisation and activity preservation.

In the present study trypsin mixed with various carbohydrates, i.e. lactose, sucrose, mannitol, alpha-cyclodextrin and dextrin, was spray-dried in order to investigate the effects of spray-drying on this enzyme, with particular emphasis on the effects of interactions between trypsin and the surface formed during spray-drying. The protein was strongly over-represented at the surface of the powder particles, the surface coverage ranging from 10 to 65%, depending on the amount of trypsin in the solids (0.2-5%). This indicates that the protein adsorbs at the air/liquid interface of the spray-droplets, and that this surface is also largely preserved after drying. The surface concentration of protein in the spray-dried powders could be controlled by adding a surfactant to the mixture before drying, since the surfactant adsorbs preferentially at the air/liquid interface of the spray droplets, thus expelling protein from the surface. In general, the residual activity of trypsin in these non-optimised formulations was 90% or higher, and in no case less than 82%. It was found that the loss of activity could partly be explained by inactivation of the protein adsorbed at the surface. For mannitol and sucrose, however, the level of inactivation was higher than could be explained by surface inactivation alone, and additional mechanisms must also be considered.

Aerosols↗
PubMed1999

Physicochemical binding properties of the proteoglycan receptor for serum lipoproteins.

Proteoheparan sulfate can be adsorbed to a methylated silica surface in a monomolecular layer via its transmembrane hydrophobic protein core domain. Due to electrostatic repulsion, its anionic polysugar side chains are stretched out into the blood substitute solution representing a co-receptor for specific lipoprotein binding through basic amino acid-rich residues within their apolipoproteins. The binding process was studied by ellipsometric techniques showing that oxLDL had a deleterious effect on heparan sulfate proteoglycan binding and conformation. Ca2+ binding to and storage on the proteoheparan sulfate/LDL compound formed a 'heterotrimeric' HS-PG/LDL/Ca2+ complex of high stability, aggregability and deposit coating. On the other hand, HDL bound to heparan sulfate proteoglycan protected against LDL docking and completely suppressed calcification of the proteoglycan/lipoprotein complex.

Adsorption↗
PubMed1999

Interfacial behaviour of 'new' poly(ethylene oxide)-containing copolymers.

Block copolymers containing poly(ethylene oxide) (PEO) have a wide applicability within biomedical applications, not the least due to anti-fouling properties of surface coatings based on these copolymers. We have investigated a number of these, and results for PEO/poly(butylene oxide) (PEO/PBO), PEO/poly(lactide) (PEO/PL), and PEO/poly(ethylene imine) (PEO/PEI) copolymers, as well as for PEO-esterified fatty acids, are presented and discussed. For the former class of polymers, the effects of molecular architecture on the adsorption properties are addressed, and experimental results obtained with ellipsometry and small-angle neutron scattering are presented. For the PEO/PL block copolymers, the effects of the PEO and PL lengths for the polymer adsorption are addressed, as are the effects of degradation of the PL moiety on both adsorption and protein rejection. For the PEO-esterified fatty acids, the effects of PEO chain length and interfacial density on the protein rejection capacity of such coatings are discussed.

Adsorption↗
PubMed1998

Tumor cell locomotion and metastatic spread.

The cytoskeletal filament proteins alpha-actinin, filamin, desmin, and filamin-desmin aggregates were adsorbed to a hydrophobic silica surface. The adsorbed amount as measured by ellipsometric methods after rinsing and equilibration was 2.7 mg/m2 for alpha-actinin and 0.4 mg/m2 for filamin plus desmin, respectively. Adsorbed layer thicknesses in physiological salt solution were about 107 nm, 89 nm, 108 nm and 93 nm for alpha-actinin, filamin, desmin, and cross-linked filamin-desmin, respectively. Ca2+ ions in a concentration of 10(-4), 10(-3), and 2.52 mmol/l had no effect on the adsorbed amount, refractive index, and adsorbed layer thickness of the individual intermediate filament proteins. Cross-linked filamin-desmin, however, reacted markedly upon the addition of these Ca2+ concentrations with a change in refractive index and adsorbed layer thickness. The layer formed by the filamin-desmin complex contracted by 2-3, 6-7, and 6-7 nm, respectively. The maximum shortening occurred at 1 pmol/l Ca2+. The Ca(2+)-dependent adsorbed layer changes of cross-linked filamin-desmin supports the contractile mechanisms in muscular tissues and forms the basis for migration and motility in nonmuscular cells. These motional events are crucially involved in peripheral organ perfusion, inflammation, and tumor invasion and metastasis.

Adsorption↗
PubMed1998

Electrostatic and Hydrophobic Effects of Oligopeptide Insertions on Protein Adsorption.

The effects of oligopeptide insertions on the adsorption of the protein ZZ, where Z is the IgG binding domain of staphylococcal Protein A, was investigated by in situ ellipsometry. In particular, the interplay between hydrophobic and electrostatic interactions as driving force for adsorption was investigated by studying the effects of oligopeptide insertions of the type Tn((AlaTrpTrpPro)n), Nn((AlaTrpTrpAspPro)n), and Pn((AlaTrpTrpLysPro)n) on the adsorption at silica, methylated silica, and diaminocyclohexane (DACH) plasma polymer surfaces. For comparison, the adsorption of the inserted peptide stretches was also investigated. It was found that the adsorption of all the peptides increases with the molecular weight at methylated silica. At silica, only the Pn peptides were found to adsorb. The net negatively charged proteins modified through peptide insertions did not adsorb at the hydrophilic and negatively charged silica, irrespective of the peptide insertion, whereas an extensive adsorption was found for the positively charged DACH surface for all the proteins investigated. For hydrophobic and negatively charged methylated silica, on the other hand, the peptide insertions were found to have a major influence on the protein interfacial behavior, and the adsorption followed the peptide stretch charge, thus increasing in the order ZZNn < ZZTn < ZZPn. These effects are discussed in terms of the relative importance of hydrophobic and electrostatic interactions as driving force for the adsorption. Copyright 1998 Academic Press.

Journal Article↗
PubMed1997

Sequential Adsorption of Human Serum Albumin (HSA), Immunoglobulin G (IgG), and Fibrinogen (Fgn) at HMDSO Plasma Polymer Surfaces

The sequential adsorption of human serum albumin (HSA), immunoglobulin G (IgG), and fibrinogen (Fgn) at hexamethyldisiloxane (HMDSO) plasma polymer surfaces was investigated with ellipsometry and total internal reflection fluorescence spectroscopy (TIRF) as a function of adsorption time, pH, and excess electrolyte concentration. HSA was found to self-exchange very slowly ( approximately hours) at pH 7.2, irrespective of adsorption time in the range 90 s to 90 min. Preadsorbed HSA was exchanged by Fgn and IgG only to a limited extent irrespectively of pH (5 </= pH </= 8) and excess electrolyte concentration (5 mM </= Cs </= 150 mM). At an excess electrolyte concentration of 150 mM, the sequential adsorption of Fgn and IgG was dramatically reduced by HSA preadsorption, irrespective of pH. At an excess electrolyte concentration of 5 mM, on the other hand, there were indications of second-layer adsorption of Fgn and IgG. Copyright 1997Academic Press

Journal Article↗
PubMed1997

Analytical partitioning of poly(ethylene glycol)-modified proteins.

Covalently grafting proteins with varying numbers (n) of poly(ethylene glycol) molecules (PEGs) often enhances their biomedical and industrial usefulness. Partition between the phases in aqueous polymer two-phase systems can be used to rapidly characterize polymer-protein conjugates in a manner related to various enhancements. The logarithm of the partition coefficient (K) approximates linearity over the range O<n<x. However, x varies with the nature of the conjugate (e.g., protein molecular mass) and such data analysis does not facilitate the comparison of varied conjugates. The known behavior of surface localized PEGs suggests a better correlation should exist between log K and the weight fraction of polymer in PEG-protein conjugates. Data from four independent studies involving three proteins (granulocyte-macrophage colony stimulation factor, bovine serum albumin and immunoglobulin G) has been found to support this hypothesis. Although somewhat simplistic, 'weight fraction' based analysis of partition data appears robust enough to accommodate laboratory to laboratory variation in protein, polymer and phase system type. It also facilitates comparisons between partition data involving disparate polymer-protein conjugates.

Chromatography, Gel↗
PubMed1997

Competitive Protein Adsorption at Plasma Polymer Surfaces

Competitive adsorption from a ternary mixture of human serum albumin (HSA), human IgG, and human fibrinogen (Fgn) at concentrations corresponding to blood plasma diluted 1/100 was investigated with the combination of Total Internal Reflection Fluorescence spectroscopy (TIRF) and ellipsometry. As substrates, three different plasma polymer surfaces, representing different surface charge and surface energy, were prepared from hexamethyldisiloxane (PP-HMDSO), acrylic acid (PP-AA), and 1,2-diaminocyclohexane (PP-DACH). In addition, adsorption from single and binary protein systems was investigated with ellipsometry. At the hydrophobic PP-HMDSO little or no displacement of any of the proteins was observed. The adsorbed layer was dominated by HSA and IgG, although Fgn was also present to a smaller extent. On PP-DACH and PP-AA, representing positively and negatively charged hydrophilic surfaces, respectively, Fgn completely dominated the adsorbed layer while HSA was almost absent and IgG was present only at a very low level.

Journal Article↗
PubMed1997

The role of the endothelium in inflammation and tumor metastasis.

In inflammation, cells interact with extracellular matrices or neighboring cells by a spatio-temporal intervention pattern of specific cell surface receptors and adhesion molecules. Resident cells of the injured tissue communicate with circulating effector cells by cytokines and direct cell-cell contact. These cytokines stimulate expression of the adhesion molecules ICAM-1, VCAM-1, and E- and P-selectin on endothelial cell surfaces and upregulate beta 2-integrins and ICAM-1 on luminal leukocytes. White blood cells then adhere to the activated endothelial cells, migrate through the vessel wall, and penetrate areas of infection or tissue damage. The basis for a cellular immune response is formed by the interaction between T lymphocytes and antigen-presenting cells amplified by adhesion molecule LFA-1,2,3 to ICAM-1 binding.

Animals↗
PubMed1996

Poly(ethylene glycol) amphiphile adsorption and liposome partition.

Surface localized poly(ethylene glycol) (PEG) amphiphiles of type C16:0-EO151 and C18:2-EO151 were studied via ellipsometry at macroscopic, flat methylated silica (MeSi), phosphatidic acid (PA), and phosphatidylcholine (PC) surfaces. At these surfaces the amphiphiles adsorb similarly, in a non-cooperative manner, achieving a plateau (approximately 0.1 PEG chains/nm2) well below amphiphile critical micelle concentration (CMC). The resultant PEG-enriched layers were 10-15 nm thick, with a polymer concentration (approximately 0.07 g/cm3) greater than the PEG-enriched phase of many dextran, PEG aqueous two-phase systems. PEG-amphiphile adsorption (mg/m2) at hydrophobic and phospholipid flat surfaces correlated with changes in the partition (log K) of PC liposomes in such two-phase systems. PEG-amphiphile adsorption at macroscopic surfaces appears to represent a balance between hydrophobic attraction and repulsive intra-chain interactions which promote chain elongation normal to the surface.

Adsorption↗
PubMed1996

Blood-flow sensing by anionic biopolymers.

Using 23Na-NMR techniques we could show that the polyanion proteoheparan sulfate integrated into the membrane of endothelial cells may serve as "flow sensor'. Based on its viscoelastic properties, heparan sulfate proteoglycan is present as a random coil under "no flow' conditions, whereby most of its polyanionic sites undergo intramolecular hydrogen bonding. With increasing flow the macromolecule becomes unfolded into a filamentous structure. Additional anionic binding sites to which Na+ ions from the blood bind are released by this shear stress-dependent conformational change. The Na+ binding triggers the signal transduction chain for a vasodilatory vessel reaction. Decrease in flow effects, for reasons of the intramolecular elastic recoil forces of the macromolecules, an entropic coiling, the release of Na+ ions and thus an interruption of the signal chain. Proteoheparan sulfate adsorbed onto a hydrophobic surface in physiological Krebs solution at pH 7.3 demonstrated clearly its characteristic as a Na+ sensor. While Ca2+ ions modulated the adsorption (promotion with increasing Ca2+ concentrations) by changing the conformation of the sensor molecule, the adsorbed amount was determined preferably by the Na+ concentration. K+ and Mg2+ ions showed slightly desorbing properties with increasing concentrations. Thus, it may be concluded that Na+ ions play the role as "first messenger' in flow-dependent vasodilation.

Biopolymers↗
PubMed1996

Effects of Hydrophilization and Immobilization on the Interfacial Behavior of Immunoglobulins.

The adsorption and immobilization of rabbit anti-human immunoglobulin (rabbit IgG), as well as the effects of rinsing with buffer and addition of bovine serum albumin (BSA) or human IgG on the amount and reactivity of bound rabbit IgG, were investigated with ellipsometry, total internal reflection fluorescence spectroscopy (TIRF), and enzyme immuno assay (EIA). It was found that although rabbit IgG readily adsorbs at hydrophobic hexamethyldisiloxane (HMDSO) plasma polymer surfaces, a substantial fraction of the adsorbed protein molecules is desorbed upon rinsing with buffer. BSA was found to adsorb readily at the surfaces obtained after rinsing, although also this protein desorbed to a large extent (about 60%) upon further rinsing with buffer. The adsorption of BSA causes a further reduction in the amount of rabbit IgG adsorbed. Immobilization of rabbit IgG to acrylic acid (AA) plasma polymer surfaces, achieved by covalent coupling via a strongly adsorbed PEG-PEI copolymer, was found to overcome the problem of the desorption of rabbit IgG upon rinsing with buffer or addition of BSA. Furthermore, nonspecific adsorption was virtually absent after immobilization. However, covalently bound rabbit IgG reacted strongly with human IgG, as observed by ellipsometry, TIRF, and EIA. The immobilization of rabbit IgG to hydrophilized surfaces was found to facilitate the interpretation of EIA results.

Journal Article↗
PubMed1996

Anionic biopolymers as blood flow sensors.

The finding of flow-dependent vasodilation rests on the basic observation that with an increase in blood flow the vessels become wider, with a decrease the vascular smooth muscle cells contract. Proteoheparan sulphate could be the sensor macromolecule at the endothelial cell membrane-blood interface, that reacts on the shear stress generated by the flowing blood, and that informs and regulates the vascular smooth muscle cells via a signal transduction chain. This anionic biopolyelectrolyte possesses viscoelastic and specific ion binding properties which allow a change of its configuration in dependence on shear stress and electrostatic charge density. The blood flow sensor undergoes a conformational transition from a random coil to an extended filamentous state with increasing flow, whereby Na+ ions from the blood are bound. Owing to the intramolecular elastic recoil forces of proteoheparan sulphate the slowing of a flow rate causes an entropic coiling, the expulsion of Na+ ions and thus an interruption of the signal chain. Under physiological conditions, the conformation and Na+ binding proved to be extremely Ca(2+)-sensitive while K+ and Mg2+ ions play a minor role for the susceptibility of the sensor. Via counterion migration of the bound Na+ ions along the sensor glycosaminoglycan side chains and following Na+ passage through an unspecific ion channel in the endothelial cell membrane, the signal transduction chain leads to a membrane depolarization with Ca2+ influx into the cells. This stimulates the EDRF/NO production and release from the endothelial cells. The consequence is vasodilation.

Biosensing Techniques↗