CiteULike: Tag fibrinogen

Forced Unfolding of Coiled-Coils in Fibrinogen by Single-Molecule AFM.

AE Brown — 2007-04-24T05:39:52-00:00

Biophys J, Vol. 92, No. 5. (1 March 2007)

Fibrinogen is a blood plasma protein that, after activation by thrombin, assembles into fibrin fibers that form the elastic network of blood clots. We used atomic force microscopy to study the forced unfolding of engineered linear oligomers of fibrinogen, and we show that forced extension of the oligomers produces sawtooth patterns with a peak-to-peak length consistent with the independent unfolding of the coiled-coils in a cooperative two-state manner. In contrast with force plateaus seen for myosin coiled-coils that suggested rapid refolding of myosin, Monte Carlo simulations of fibrinogen unfolding confirm that fibrinogen refolding is negligible on experimental timescales. The distinct behavior of fibrinogen seems to be due to its topologically complex coiled-coils and an interaction between fibrinogen's alphaC-domains and its central region.

Binding strength and activation state of single fibrinogen-integrin pairs on living cells

Rustem Litvinov — 2008-03-26T17:51:03-00:00

Proceedings of the National Academy of Sciences, Vol. 99, No. 11. (28 May 2002), pp. 7426-7431.

Integrin activation states determine the ability of these receptors to mediate cell-matrix and cell-cell interactions. The prototypic example of this phenomenon is the platelet integrin, [alpha]IIb[beta]3. In unstimulated platelets, [alpha]IIb[beta]3 is inactive, whereas exposing platelets to an agonist such as ADP or thrombin enables [alpha]IIb[beta]3 to bind ligands such as fibrinogen and von Willebrand factor. To study the regulation of integrin activation states at the level of single molecules, we developed a model system based on laser tweezers, enabling us to determine the rupture forces required to separate single ligand-receptor pairs by using either purified proteins or intact living cells. Here, we show that rupture forces of individual fibrinogen molecules and either purified [alpha]IIb[beta]3 or [alpha]IIb[beta]3 on the surface of living platelets were 60 to 150 pN with a peak yield strength of 80-100 pN. Platelet stimulation using either ADP or the thrombin receptor-activating peptide enhanced the accessibility but not the adhesion strength of single [alpha]IIb[beta]3 molecules, indicating that there are only two states of [alpha]IIb[beta]3 activation. Thus, we found it possible to use laser tweezers to measure the regulation of forces between individual ligand-receptor pairs on living cells. This methodology can be applied to the study of other regulated cell membrane receptors using the ligand-receptor yield strength as a direct measure of receptor activation/inactivation state. 10.1073/pnas.112194999

Association of Fibrinogen and Fibrinogen Gene beta148 and beta854 Polymorphisms with Coronary Heart Disease.

Ai-Jun Sun — 2008-07-10T01:42:25-00:00

Cardiology, Vol. 111, No. 3. (25 April 2008), pp. 167-170.

Background: The aim of this study was to observe the association between fibrinogen C148T, G854A polymorphisms and plasma fibrinogen levels in a large cohort of Chinese patients with coronary heart disease (CHD). Methods: Fibrinogen gene beta148/beta854 polymorphisms were screened and plasma fibrinogen levels and lipids were measured in patients with angiographically confirmed CHD (n = 836) and in controls without CHD (n = 418). Results: Age, sex, smoking, hypertension, diabetes mellitus, blood lipids and fibrinogen levels were related to CHD (all p < 0. 05) while the frequencies of fibrinogen beta148 and beta854 alleles, gene and genotypes between the two groups were similar (all p > 0.05). Conclusion: This study demonstrates that the plasma fibrinogen level, but not fibrinogen beta148 and beta854 genotypes, was associated with CHD in the Chinese population.

Beta-fibrinogen haplotypes and the risk for cardiovascular disease in a dialysis cohort.

Y Liu — 2008-07-11T13:22:46-00:00

American journal of kidney diseases : the official journal of the National Kidney Foundation, Vol. 46, No. 1. (July 2005), pp. 78-85.

BACKGROUND: Elevated plasma fibrinogen levels are common in dialysis patients and may be related to an elevated risk for cardiovascular disease (CVD). We tested the hypothesis that genetic variation in the beta-fibrinogen ( FGB ) gene, shown to explain 1% to 5% of fibrinogen level variation in the general population, has an important role in elevated fibrinogen levels and excess CVD risk in dialysis patients. METHODS: Plasma fibrinogen was measured in 735 dialysis patients a median of 3 months from the start of dialysis therapy by using an automated clot-rate assay. Seven polymorphisms of the FGB gene were determined. Haplotype analysis was conducted using the Phase program to estimate haplotypes, with stratification for race. CVD events were ascertained from medical records. RESULTS: During a median follow-up of 2.1 years, 279 CVD events occurred. Genotype frequencies were in Hardy-Weinberg equilibrium. Four common haplotypes identified were not associated with fibrinogen levels or CVD risk in the entire cohort or after stratification by race. The -455A allele, known to increase gene expression in vitro, was marginally associated with fibrinogen levels only in patients without diabetes (regression coefficient [beta], 20 mg/dL [for +1 copy of the A allele; P = 0.06]), adjusted for age, sex, race, smoking, baseline dialysis modality, comorbidity, and history of diabetes and CVD. Post hoc analysis showed that -249C-->T (defining haplotype 3) was associated with greater fibrinogen levels and CVD risk among patients without diabetes and current smokers. CONCLUSION: The FGB gene likely does not have an important role in determining the variation in elevated plasma fibrinogen levels or excess CVD risk in dialysis patients.

Coordinated increase in albumin, fibrinogen, and muscle protein synthesis during hemodialysis: role of cytokines.

DS Raj — 2008-04-16T13:36:00-00:00

American journal of physiology. Endocrinology and metabolism, Vol. 286, No. 4. (April 2004)

Serum albumin, fibrinogen levels, and lean body mass are important predictors of outcome in end-stage renal disease (ESRD). We estimated the fractional synthesis rates of albumin (FSR-A), fibrinogen (FSR-F), and muscle protein (FSR-M) in nine ESRD patients and eight controls, using primed constant infusion of l-[ring-(13)C(6)]phenylalanine. Cytokine profile and arteriovenous balance of amino acids were also measured. ESRD patients were studied before (Pre-HD) and during hemodialysis (HD). Plasma IL-6, IL-10, and C-reactive protein increased significantly during HD. Despite a decrease in the delivery of amino acids to the leg, the outflow of the amino acids increased during HD. The net balance of amino acids became more negative during HD, indicating release from the muscle. HD increased leg muscle protein synthesis (45%) and catabolism (108%) but decreased whole body proteolysis (15%). FSR-A during HD (9.7 +/- 0.9%/day) was higher than pre-HD (6.5 +/- 0.9%/day) and controls (5.8 +/- 0.5%/day, P < 0.01). FSR-F increased during HD (19.7 +/- 2.6%/day vs. 11.8 +/- 0.6%/day, P < 0.01), but it was not significantly different from that of controls (14.4 +/- 1.4%/day). FSR-M intradialysis (1.77 +/- 0.19%/day) was higher than pre-HD (1.21 +/- 0.25%/day) and controls (1.30 +/- 0.32%/day, P < 0.001). Pre-HD FSR-A, FSR-F, and FSR-M values were comparable to those of controls. There was a significant and positive correlation between plasma IL-6 and the FSRs. Thus, in ESRD patients without metabolic acidosis, the fractional synthesis rates of albumin, fibrinogen, and muscle protein are not decreased pre-HD. However, HD increases the synthesis of albumin, fibrinogen, and muscle protein. The coordinated increase in the FSRs is facilitated by constant delivery of amino acids derived from the muscle catabolism and intradialytic increase in IL-6.

Fibrinogen and coronary heart disease: test of causality by 'Mendelian randomization'.

B Keavney — 2008-07-10T01:34:30-00:00

International journal of epidemiology, Vol. 35, No. 4. (August 2006), pp. 935-943.

BACKGROUND: Blood concentrations of fibrinogen have been associated with coronary heart disease risk in epidemiological studies, but it is uncertain whether this association is causal or reflects residual confounding by other risk factors. We investigated the relationship between the single nucleotide polymorphism at position -148 in the beta-fibrinogen gene promoter (beta - 148C/T), blood fibrinogen levels, and risk of myocardial infarction (MI) in sufficiently large numbers of coronary disease cases to reliably address this question. METHODS: Genotyping and measurement of blood fibrinogen concentration were carried out in 4,685 cases of confirmed MI and 3,460 controls with no history of coronary disease. A meta-analysis of ISIS and 19 other studies of beta-fibrinogen genotypes involving a total of 12,220 coronary disease cases and 18,716 controls was conducted. RESULTS: Among the ISIS controls, mean plasma fibrinogen concentrations with the C/C, C/T and T/T genotypes were 3.34 (SE 0.015), 3.48 (0.022), and 3.60 (0.064) g/l, respectively, corresponding to an increase of 0.14 (0.024) g/l per T allele (trend P < 0.0001). In the case-control comparison, 0.14 g/l higher usual plasma fibrinogen concentration was associated with an age-adjusted and sex-adjusted risk ratio for MI of 1.17 [95% confidence interval (95% CI) 1.14-1.19; P < 0.0001]. But, after further adjustment for smoking, body mass index, and plasma apolipoprotein B/A(1) ratio, this risk ratio fell to 1.03 (95% CI 1.00-1.05; P = 0.05). Moreover, fibrinogen genotype was not significantly associated with MI incidence: risk ratio of 1.06 (95% CI 0.96-1.16) per higher-fibrinogen allele in ISIS alone and of 1.00 (95% CI 0.95-1.04) per allele in the meta-analysis. CONCLUSIONS: Genotypes that produce lifelong differences in fibrinogen concentrations do not materially influence coronary disease incidence. As these genotype-dependent differences in fibrinogen were allocated randomly at conception (Mendelian randomization), this association is not likely to be confounded by other factors. Consequently, these genetic results provide strong evidence that long-term differences in fibrinogen concentrations are not a major determinant of coronary disease risk.

Increased albumin and fibrinogen synthesis in hemodialysis patients with normal nutritional status.

M Giordano — 2008-04-16T13:22:07-00:00

Journal of the American Society of Nephrology : JASN, Vol. 12, No. 2. (February 2001), pp. 349-354.

This study compared the rates of whole-body proteolysis and of albumin and fibrinogen synthesis of seven hemodialysis patients (HD) with those of seven normal matched control subjects (C). HD patients had a normal nutritional and inflammatory status and serum albumin levels >3.5 g/dl. Endogenous leucine flux, albumin and fibrinogen fractional synthesis rate (FSR), and absolute intravascular synthesis rate (ASR) of albumin and fibrinogen all were evaluated by a primed/continuous infusion of 5,5,5-D3-L-leucine. Plasma volume was determined by the Evans blue dye dilution method. Endogenous leucine flux was significantly increased in HD (2.64 +/- 0.08 micromol/kg per min) compared with C (2.17 +/- 0.07 micromol/kg per min, P: < 0.05). Serum albumin concentrations were similar in HD and C. Plasma fibrinogen levels were significantly increased in HD compared with C (P: < 0.05). Plasma volume was greater in HD than in C (P: < 0.05). As a result, total intravascular pool of both albumin (141 +/- 7 versus 114 +/- 3 g/1.73 m(2), P: < 0.05) and fibrinogen (11.7 +/- 1 versus 6.7 +/- 0.5 g/1.73 m(2), P: < 0.05) were greater in HD than in C. Albumin FSR was not statistically different in HD and C. However, albumin ASR was significantly increased in HD than in C (13.7 +/- 2 versus 10.3 +/- 1 g/1.73 m(2) per d, P: < 0.05). Similarly, FSR of fibrinogen did not differ in HD and C groups, whereas ASR of fibrinogen was significantly higher in HD than in C (3.31 +/- 0.6 versus 1.94 +/- 0.3 g/1.73 m(2) per d, P: < 0.05). In summary, normoalbuminemic HD patients have an increased intravascular pool with a greater absolute synthesis rate of both albumin and fibrinogen and an increased rate of whole-body leucine flux.

Polymorphisms of the fibrinogen-beta gene are related to 2-hour glucose level after oral glucose tolerance test in Hong Kong Chinese.

LY Wong — 2008-07-10T01:49:38-00:00

Disease markers, Vol. 24, No. 3. (2008), pp. 167-173.

Fibrinogen, an acute phase protein, is an important inflammatory marker that is associated with cardiovascular diseases. We studied the association of three common human fibrinogen-beta gene (FGB) variants, -455G>A, -249C>T, and -148C>T with glycemic parameters in 265 non-diabetic Hong Kong Chinese subjects. Both FGB variants, -455G>A and -148C>T were in complete linkage disequilibrium and were associated with higher levels of plasma fibrinogen and 2-h glucose after a 75-g oral glucose load (p<0.01). Carriers of FGB AC-haplotype, comprising the two nucleotide variants at positions -455 and -249, had higher fibrinogen level (2.64 +/- 0.65 vs 2.42 +/- 0.52 g/L, p=0.002) and 2-h glucose after a 75-g oral glucose load (5.87 +/- 1.14 vs 5.47 +/- 1.22 g/L, p=0.006). The associations were significant in men, but not women. In stepwise multiple regression analysis, AC-haplotype was independently associated with plasma fibrinogen level and 2-h glucose (p=0.002 and 0.010 respectively). This suggests that fibrinogen may play a role in the development of impaired glucose tolerance.

Diabetic nephropathy is associated with increased albumin and fibrinogen production in patients with type 2 diabetes.

P Tessari — 2008-04-16T13:21:49-00:00

Diabetologia, Vol. 49, No. 8. (August 2006), pp. 1955-1961.

AIMS/HYPOTHESIS: Hyperfibrinogenaemia and albuminuria are cardiovascular risk factors, often coexisting in diabetic and non-diabetic people. Albuminuria in turn is associated with a compensatory albumin overproduction in non-diabetic patients. It is not known whether the presence of albuminuria in patients with type 2 diabetes mellitus is associated with greater albumin and fibrinogen production rates than in normoalbuminuric patients. SUBJECTS, MATERIALS, AND METHODS: Using leucine isotope methods, we measured fractional and absolute synthesis rates (FSR, ASR) of albumin and fibrinogen in post-absorptive type 2 diabetic patients with either normal (n=11) or increased (n=10) urinary albumin excretion. RESULTS: In albuminuric patients, albumin FSR (16.2+/-1.5%/day) and ASR (20.5+/-1.9 g/day) were greater (p<0.02 and p<0.05, respectively) than in normoalbuminuric patients (FSR=11.5+/-1.1%/day; ASR=15.7+/-1.2 g/day). Fibrinogen FSR was similar between patients with normal and increased albumin excretion, but concentration, the circulating pool and ASR of fibrinogen were 40 to 50% greater (p<0.035) in patients with albuminuria. Albuminuria was positively correlated with albumin ASR, with fibrinogen concentration, the fibrinogen pool and ASR, whereas albumin synthesis was inversely correlated with calculated oncotic pressure. CONCLUSIONS/INTERPRETATION: Synthesis of albumin and fibrinogen is upregulated in type 2 diabetic patients with increased urinary albumin excretion. Albuminuria is associated with enhanced fibrinogen and albumin synthesis.

Increased albumin and fibrinogen synthesis rate in patients with chronic renal failure.

BH Prinsen — 2008-04-16T13:51:43-00:00

Kidney international, Vol. 64, No. 4. (October 2003), pp. 1495-1504.

BACKGROUND: Hypoalbuminemia and hyperfibrinogenemia are frequently observed in patients with chronic renal failure (CRF) and are both associated with cardiovascular diseases. The mechanisms responsible for hypoalbuminemia and hyperfibrinogenemia in CRF are unknown. METHODS: In the present study, both albumin and fibrinogen kinetics were measured in vivo in predialysis patients (N = 6), patients on peritoneal dialysis (N = 7) and control subjects (N = 8) using l-[1-13C]-valine. RESULTS: Plasma albumin concentration was significantly lower in patients on peritoneal dialysis compared to control subjects (P < 0.05). Plasma fibrinogen was significantly increased in both predialysis patients (P < 0.01) as well as patients on peritoneal dialysis (P < 0.001) in comparison to control subjects. In contrast to albumin, fibrinogen is only lost in peritoneal dialysate and not in urine. The absolute synthesis rates (ASR) of albumin and fibrinogen were increased in patients on peritoneal dialysis (ASR albumin, 125 +/- 9 mg/kg/day versus 93 +/- 9 mg/kg/day, P < 0.05; ASR fibrinogen, 45 +/- 4 mg/kg/day versus 29 +/- 3 mg/kg/day, P < 0.01) compared to control subjects. Albumin synthesis is strongly correlated with fibrinogen synthesis (r2 = 0.665, P < 0.0001, N = 21). In this study, the observed hypoalbuminemia in patients on peritoneal dialysis is likely not explained by malnutrition, inadequate dialysis, inflammation, metabolic acidosis, or insulin resistance. We speculate that peritoneal albumin loss is of relevance. CONCLUSION: Synthesis rate of albumin and fibrinogen are coordinately up-regulated. Both albumin and fibrinogen are lost in peritoneal dialysis fluid. To compensate protein loss, albumin synthesis is up-regulated, but the response, in contrast to predialysis patients, does not fully correct plasma albumin concentrations in peritoneal dialysis patients. The increase in fibrinogen synthesis introduces an independent risk factor for atherosclerosis, since plasma fibrinogen pool is enlarged.

Impact of albumin synthesis rate and the acute phase response in the dual regulation of fibrinogen levels in hemodialysis patients.

GA Kaysen — 2008-04-16T13:51:37-00:00

Kidney international, Vol. 63, No. 1. (January 2003), pp. 315-322.

BACKGROUND: Fibrinogen is a risk factor for cardiovascular disease. It also is an acute phase protein (APP) and its plasma concentration increases with inflammation. Fibrinogen synthesis correlates with albumin synthesis in nephrotic patients and in patients with an expanded plasma volume even when serum albumin is normal and there is no inflammatory disease. The relationships among albumin synthesis, the acute phase response and plasma fibrinogen levels in hemodialysis patients are unknown. METHODS: In 74 hemodialysis patients, albumin synthesis, plasma volume (PV) and acute phase proteins (APPs) C-reactive protein (CRP), alpha1 acid glycoprotein (alpha1 AG), ceruloplasmin (Cer), and interleukin 6 (IL-6) were measured in serum and fibrinogen in plasma, and the results analyzed by multiple regression analysis. CRP, IL-6, alpha1 AG, Cer and fibrinogen were measured monthly, which enabled us to determine whether changes in these APPs correlated with the levels of and variability in plasma fibrinogen over time using a longitudinal modeling approach. Length of follow-up for the 74 patients ranged from 3.25 to 67.5 months. RESULTS: Baseline fibrinogen (548.6 +/- 106. 4 mg/dL) was significantly greater than levels reported for normal adults and correlated positively with albumin synthesis (P < 0.001), age (P < 0.001) and log CRP (P = 0.002) and negatively with PV (P < 0.001). Longitudinally, fibrinogen varied positively with long-lived APPs, Cer and alpha1 AG, as well as the short-lived APP, CRP. CONCLUSION: Plasma fibrinogen concentration is high in HD patients and directly correlates with increased albumin synthesis rates and the serum levels of APPs. Fibrinogen levels also correlate negatively with PV. Fibrinogen levels vary over time in synchrony with levels of other long-lived APPs, supporting the hypothesis that fibrinogen is regulated in part as a component of the acute phase response and in part by factors that increase albumin synthesis.

Coiled-Coil Irregularities and Instabilities in Group A Streptococcus M1 Are Required for Virulence

Case Mcnamara — 2008-03-11T19:01:39-00:00

Science, Vol. 319, No. 5868. (7 March 2008), pp. 1405-1408.

Antigenically variable M proteins are major virulence factors and immunogens of the human pathogen group A Streptococcus (GAS). Here, we report the [~]3 angstrom resolution structure of a GAS M1 fragment containing the regions responsible for eliciting type-specific, protective immunity and for binding fibrinogen, which promotes M1 proinflammatory and antiphagocytic functions. The structure revealed substantial irregularities and instabilities throughout the coiled coil of the M1 fragment. Similar structural irregularities occur in myosin and tropomyosin, explaining the patterns of cross-reactivity seen in autoimmune sequelae of GAS infection. Sequence idealization of a large segment of the M1 coiled coil enhanced stability but diminished fibrinogen binding, proinflammatory effects, and antibody cross-reactivity, whereas it left protective immunogenicity undiminished. Idealized M proteins appear to have promise as vaccine immunogens. 10.1126/science.1154470

Simulation of platelet adhesion and aggregation regulated by fibrinogen and von Willebrand factor.

D Mori — 2008-01-26T20:53:57-00:00

Thromb Haemost, Vol. 99, No. 1. (January 2008), pp. 108-115.

We propose a method to analyze platelet adhesion and aggregation computationally, taking into account the distinct properties of two plasma proteins, von Willebrand factor (vWF) and fibrinogen (Fbg). In this method, the hydrodynamic interactions between platelet particles under simple shear flow were simulated using Stokesian dynamics based on the additivity of velocities. The binding force between particles mediated by vWF and Fbg was modeled using the Voigt model. Two Voigt models with different properties were introduced to consider the distinct behaviors of vWF and Fbg. Our results qualitatively agreed with the general observation of a previous in-vitro experiment, thus demonstrating that the significant development of thrombus formation in height requires not only vWF, but also Fbg. This agreement of simulation and experimental results qualitatively validates our model and suggests that consideration of the distinct roles of vWF and Fbg is essential to investigate the physiological and pathophysiological mechanisms of thrombus formation using a computational approach.

Platelet and monoclonal antibody binding to fibrinogen adsorbed on glow-discharge-deposited polymers

David Kiaei — 2007-02-06T12:06:11-00:00

Journal of Biomedical Materials Research, Vol. 29, No. 6. (1995), pp. 729-739.

The state of fibrinogen adsorbed on untreated and glow-discharge-treated surfaces was examined by measuring platelet adhesion, monoclonal antibody (mAb) binding, the amount of fibrinogen adsorbed, and the amount of adsorbed fibrinogen which could be eluted with sodium dodecyl sulfate (SDS). Tetrafluoroethylene (TFE) glow-discharge-treated polymers have a lower surface free energy (in air) and retain a larger fraction of adsorbed fibrinogen than untreated surfaces after SDS elution. Platelet adhesion was lowest on the TFE-treated surfaces which retain the highest amounts of fibrinogen after SDS elution. Fibrinogen may undergo unfolding or spreading on the TFE-treated surfaces to minimize interfacial free energy (in water) and maximize protein-;surface interactions. When it is adsorbed on the TFE-treated surfaces, fibrinogen evidently assumes a state which somehow prevents its recognition and binding by platelet receptors. Monoclonal antibodies that bind to the three regions in fibrinogen thought to be involved in platelet adhesion were therefore used to detect changes in adsorbed fibrinogen. These regions and the antibodies which bind to them are: the COOH-terminal of the ?-chain, mAb M1; the RGD peptide sequence at A? 95-98, mAb R1; the RDG sequence at A? 572-575, mAb R2. For fibrinogen adsorbed on the untreated or TFE-treated surfaces, M1 and R2 binding was relatively high compared to background, while R1 binding was low. However, the amount of binding of each mAb to fibrinogen adsorbed on the TFE-treated surfaces was equal to or greater than fibrinogen adsorbed to the untreated surfaces. Therefore, antibody-detectable changes in the platelet binding regions of adsorbed fibrinogen that might have been caused by conformational or orientational rearrangements were not observed for the TFE-treated surfaces. The data suggest that the tight binding of fibrinogen on a surface may directly affect the ability of the fibrinogen to interact with the platelet receptors - i.e., that fibrinogen must be loosely held to facilitate maximal interaction with platelet receptors. © 1995 John Wiley & Sons, Inc.

Ellipsometry studies of the effects of surface hydrophobicity on protein adsorption

Martin Malmsten — 2007-01-12T23:43:03-00:00

Colloids and Surfaces B: Biointerfaces, Vol. 3, No. 5. (2 January 1995), pp. 297-308.

The adsorption of some model proteins, human serum albumin (HSA), IgG, fibrinogen and lysozyme on silica surfaces was investigated with in situ ellipsometry and compared to previous results obtained for methylated silica surfaces. The adsorbed amount ([Gamma]), the adsorbed layer thickness ([delta]el) and the mean adsorbed layer refractive index (nf) were obtained by a procedure involving studies of the bare substrate at two different ambient refractive indices, as well as four-zone averaging. The surface hydrophobicity strongly influenced the adsorption properties of all the proteins studied. For HSA, IgG and fibrinogen, the adsorbed amount was significantly lower on the hydrophilic surface than on the hydrophobic one, whereas the reverse was found for lysozyme. For fibrinogen, the adsorbed layer thickness on silica was smaller than that on methylated silica, whereas the adsorbed layer was more concentrated. For IgG, on the other hand, end-on adsorption was observed on both silica and methylated silica. For lysozyme, side-on adsorption in a dense monolayer was observed on silica, whereas on methylated silica, the adsorption occurs in two to three rather dilute molecular layers. Furthermore, the build-up of the adsorbed layers was studied. For fibrinogen, qualitatively the same behaviour was observed for silica and methylated silica, i.e. as the absorbed amount increases, both [delta]el and nf initially increase, while closer to adsorption saturation, nf levels off. However, at a given [Gamma], [delta]el was lower and nf higher on silica than on methylated silica. A similar finding was obtained for lysozyme. These findings are discussed in terms of adsorbed layer structure and formation.

Adsorption of fibrinogen on tantalum oxide, titanium oxide and gold studied by the QCM-D technique

Anne Hemmersam — 2006-09-28T09:20:14-00:00

Colloids and Surfaces B: Biointerfaces, Vol. 43, No. 3-4. (10 July 2005), pp. 208-215.

The adsorption of human fibrinogen on tantalum oxide, titanium oxide and gold surfaces has been studied by quartz crystal microbalance with dissipation (QCM-D) at 37[thin space][deg] C. Two different protein concentrations have been used, one close to physiological concentration (1[thin space]mg/ml) and one significantly lower (0.033[thin space]mg/ml). To further characterize the adsorbed fibrinogen layer, the subsequent binding of both polyclonal and monoclonal antibodies of fibrinogen is studied. We found that the viscoelastic properties of the fibrinogen layer depends strongly on the initial protein concentration. The trend is generally seen for all three surfaces. The fibrinogen layer on gold and tantalum oxide is found to have the same viscoelastic properties, which are different from those found for the fibrinogen layer adsorbed on titanium oxide. The dependency of the surface chemistry on the viscoelastic properties of the fibrinogen layer is observed directly for the 0.033[thin space]mg/ml solution, and indirectly through the antibody response for the 1[thin space]mg/ml solution. From this we conclude that the orientation and/or denaturation of fibrinogen on a surface depends on the surface chemistry and the protein concentration in the solution, and that the binding of antibodies is a useful way to detect this difference.

Interpretation of Protein Adsorption: Surface-Induced Conformational Changes

P Roach — 2007-01-16T20:28:46-00:00

J. Am. Chem. Soc., Vol. 127, No. 22. (8 June 2005), pp. 8168-8173.

Abstract: Protein adhesion plays a major role in determining the biocompatibility of materials. The first stage of implant integration is the adhesion of protein followed by cell attachment. Surface modification of implants (surface chemistry and topography) to induce and control protein and cell adhesion is currently of great interest. This communication presents data on protein adsorption (bovine serum albumin and fibrinogen) onto model hydrophobic (CH3) and hydrophilic (OH) surfaces, investigated using a quartz crystal microbalance (QCM) and grazing angle infrared spectroscopy. Our data suggest that albumin undergoes adsorption via a single step whereas fibrinogen adsorption is a more complex, multistage process. Albumin has a stronger affinity toward the CH3 compared to OH terminated surface. In contrast, fibrinogen adheres more rapidly to both surfaces, having a slightly higher affinity toward the hydrophobic surface. Conformational assessment of the adsorbed proteins by grazing angle infrared spectroscopy (GA-FTIR) shows that after an initial 1 h incubation few further time-dependent changes are observed. Both proteins exhibited a less organized secondary structure upon adsorption onto a hydrophobic surface than onto a hydrophilic surface, with the effect observed greatest for albumin. This study demonstrates the ability of simple tailor-made monochemical surfaces to influence binding rates and conformation of bound proteins through protein-surface interactions. Current interest in biocompatible materials has focused on surface modifications to induce rapid healing, both of implants and for wound care products. This effect may also be of significance at the next stage of implant integration, as cell adhesion occurs through the surface protein layer.

The Density and Refractive Index of Adsorbing Protein Layers

Janos Voros — 2006-09-28T09:15:18-00:00

Biophys. J., Vol. 87, No. 1. (1 July 2004), pp. 553-561.

The structure of the adsorbing layers of native and denatured proteins (fibrinogen,gamma -immunoglobulin, albumin, and lysozyme) was studied on hydrophilic TiO2 and hydrophobic Teflon-AF surfaces using the quartz crystal microbalance with dissipation and optical waveguide lightmode spectroscopy techniques. The density and the refractive index of the adsorbing protein layers could be determined from the complementary information provided by the two in situ instruments. The observed density and refractive index changes during the protein-adsorption process indicated the presence of conformational changes (e.g., partial unfolding) in general, especially upon contact with the hydrophobic surface. The structure of the formed layers was found to depend on the size of the proteins and on the experimental conditions. On the TiO2 surface smaller proteins formed a denser layer than larger ones and the layer of unfolded proteins was less dense than that adsorbed from the native conformation. The hydrophobic surface induced denaturation and resulted in the formation of thin compact protein films of albumin and lysozyme. A linear correlation was found between the quartz crystal microbalance measured dissipation factor and the total water content of the layer, suggesting the existence of a dissipative process that is related to the solvent molecules present inside the adsorbed protein layer. Our measurements indicated that water and solvent molecules not only influence the 3D structure of proteins in solution but also play a crucial role in their adsorption onto surfaces. 10.1529/biophysj.103.030072

The kinetics of baboon fibrinogen adsorption to polymers: <I>In vitro</I> and <I>in vivo</I> studies

TA Horbett — 2007-02-06T12:11:58-00:00

Journal of Biomedical Materials Research, Vol. 20, No. 6. (1986), pp. 739-772.

Fibrinogen adsorption on polymers from blood may mediate or potentiate thrombosis because of its involvement in both the intrinsic clotting system and the formation of platelet aggregates. While the kinetics of fibrinogen adsorption from plasma in vitro have previously been found to be very different on polar and nonpolar surfaces [T. A Horbett, ?The kinetics of adsorption of plasma proteins to a series of hydrophilic-hydrophobic copolymers,? ACS Org. Coat. Plas. Chem., 40, 642-646 (1979)] the significance of this difference with respect to thrombogenesis in vivo has not been clarified. In this study, the kinetics of deposition of baboon 125I fibrinogen from plasma in vitro or from blood in vivo on a series of polymers was measured. The polymers chosen for this study had previously been found to have a large range in surface polarity and reactivity in the in vivo baboon shunt model. The kinetics of fibrinogen adsorption in vitro were observed to be of three types, depending on the polymer: (1) high initial adsorption decreasing to a lower steady state value; (2) constant throughout the time course; (3) low initial adsorption rising steadily to a plateau value. In vivo, fibrinogen deposition kinetics were of two types: (1) low, constant deposition throughout the time course, independent of heparinization; (2) low deposition initially followed by a second phase of greatly increased deposition (probably as fibrin) which was prevented or greatly decreased by heparinizing the animals. Polymers for which fibrinogen adsorption increased to a plateau in vitro were found to have a heparin inhibitable second phase of enhanced in vivo fibrinogen deposition. These polymers also have been found in previous studies to enhance the rate of platelet destruction when used as in vivo shunts on baboons. Conversely, most polymers with high initial in vitro fibrinogen adsorption followed by a decrease had low fibrinogen deposition behavior in vivo and were also minimally destructive of platelets. The adsorption kinetics of fibrinogen to polymers from blood in vivo and in vitro and the consumption of platelets in vivo induced by the polymers all vary with polymer polarity. More polar polymers had in vitro fibrinogen kinetics characterized by a rise to a plateau, in vivo fibrinogen deposition characterized by a second stage of great increase inhibitable by heparin, and enhanced platelet consumption. The correlation of three separate indicators of surface thrombogenicity with surface polarity suggests that more polar materials may be more thrombogenic because of an influence on the way in which fibrinogen interacts with these surfaces.

Surface Tailoring for Controlled Protein Adsorption: Effect of Topography at the Nanometer Scale and Chemistry

P Roach — 2007-01-16T19:45:46-00:00

J. Am. Chem. Soc., Vol. 128, No. 12. (29 March 2006), pp. 3939-3945.

Abstract: Protein adsorption behavior is at the heart of many of today's research fields including biotechnology and materials science. With understanding of protein-surface interactions, control over the conformation and orientation of immobilized species may ultimately allow tailor-made surfaces to be generated. In this contribution protein-surface interactions have been examined with particular focus on surface curvature with and without surface chemistry effects. Silica spheres with diameters in the range 15-165 nm with both hydrophilic and hydrophobic surface chemistries have been used as model substrates. Two proteins differing in size and shape, bovine serum albumin (BSA) and bovine fibrinogen (Fg), have been used in model studies of protein binding with detailed secondary structure analysis being performed using infrared spectroscopy (IR) on surface-bound proteins. Although trends in binding affinity and saturation values were similar for both proteins, albumin is increasingly less ordered on larger substrates, while fibrinogen, in contrast, loses secondary structure to a greater extent when adsorbing onto particles with high surface curvature. These effects are compounded by surface chemistry, with both proteins becoming more denatured on hydrophobic surfaces. Both surface chemistry and topography play key roles in determining the structure of the bound proteins. A model of the binding characteristics of these two proteins onto surfaces having differing curvature and chemistry is presented. We propose that properties of an adsorbed protein layer may be guided through careful consideration of surface structure, allowing the fabrication of materials/surface coatings with tailored bioactivity.

Interaction of fibrinogen with solid surfaces of varying charge and hydrophobic--hydrophilic balance : I. Adsorption isotherms

A Schmitt — 2007-01-06T15:34:13-00:00

Journal of Colloid and Interface Science, Vol. 92, No. 1. (March 1983), pp. 25-34.

Adsorption of fibrinogen to a series of polyelectrolyte complex surfaces of varying charge density (bulk ion-exchange capacity ranging from +1.21 to -1.33 meq g-1) and hydrophilicity (water content from 4.5 to 60 wt%) was studied using an iodine-labeling technique under static conditions. In the high concentration region of the isotherms (c > 0.5 mg ml-1) pseudoplateaux were observed with adsorption values in the range expected for a close-packed monomolecular layer. The surface of a material of +0.8 meq g-1 ion-exchange capacity showed a particularly high value. At low concentrations (c -2mg ml-1), the data were found to fit a Freundlich-type relation with identical slopes for all surfaces. Free energies of adsorption estimated from these data were of the order of 7 kcal mole-1. These values are indicative of nonspecific relatively weak bonding perhaps of the hydrophobic interaction type. It is concluded that fixed surface charge of the order of magnitude of that used in the present work has little effect on the equilibrium (isotherm) aspects of protein adsorption.

Visualisation of human plasma fibrinogen adsorbed on titanium implant surfaces with different roughness

Paola Cacciafesta — 2007-01-13T10:30:20-00:00

Surface Science, Vol. 491, No. 3. (1 October 2001), pp. 405-420.

Direct visualisation of adsorbed human plasma proteins on biomaterial surfaces may help the understanding of the performance of implants. The aim of this study was the visualisation of human plasma fibrinogen (HPF) adsorbed on different titanium implants with characterised surface properties. Seven types of titanium samples were used: mechanically polished (P); mechanically ground (G); sand blasted with alumina (B); sand blasted with alumina and etched in HF/HNO3 (BN); sand blasted with alumina and etched in HCl/H2SO4 (SLA); mechanically polished and etched in HCl/H2SO4 (PSLA); and plasma sprayed (TPS). The surface morphology and roughness of these surfaces were analysed by atomic force microscopy (AFM). The surface chemical composition of the implants was analysed by X-ray photoelectron spectroscopy (XPS) and time of flight secondary ion mass spectrometry (ToF-SIMS). The seven processing methods influenced the roughness and produced distinct morphologies of the implant surfaces. The main chemical constituent of all surfaces was TiO2, with some samples showing the presence of other elements. HPF adsorbed on the titanium surfaces was visualised by AFM phase imaging. Visualisation of the adsorbed HPF was successfully achieved on samples P and G, but not on the other five samples. Differences in the dimensions and in the phase contrast of HPF molecules adsorbed on P and G surfaces were observed and discussed. No correlation between surface topography and morphology of the adsorbed proteins was observed on surfaces P and G. Possible reasons for not detecting HPF with AFM on five of the seven sample types were attributed to the surface physico-chemical properties of these samples

Effect of Surface Hydrophobicity on Adsorption and Relaxation Kinetics of Albumin and Fibrinogen: Single-Species and Competitive Behavior

CF Wertz — 2007-01-06T20:47:06-00:00

Langmuir, Vol. 17, No. 10. (15 May 2001), pp. 3006-3016.

Abstract: This work compares the spreading and relaxation rates of albumin and fibrinogen, inferred from single-component and competitive adsorption kinetic experiments, on model surfaces of varying hydrophobicity. Kinetics from the single-component studies revealed a constant spreading rate, where the adsorbed protein footprint grew linearly in time for at least 15 min. This spreading rate increased with substrate hydrophobicity (ranging from 0.02 to 0.16 nm2/molecule/s for albumin and from 0.04 to 0.26 nm2/molecule/s for fibrinogen), resulting in a larger extent of footprint growth and a lower ultimate coverage on hydrophobic surfaces when compared with hydrophilic surfaces at the same adsorption conditions. Competitive adsorption studies were in qualitative agreement with the single-component experiments but were able to probe longer spreading time scales. Although spreading appeared to occur initially at a constant rate in the competitive experiments, after 2 h the spreading rate had slowed dramatically and the spreading process had begun to level off.

Fibrinogen and bleeding: old molecule--new ideas.

VG Nielsen — 2008-06-08T11:32:58-00:00

Anesthesia and analgesia, Vol. 105, No. 4. (October 2007), pp. 902-903.