D.J. Curtis

A new biomarker quantifies differences in clot microstructure in patients with venous thromboembolism

This study compared patients with venous thromboembolism (VTE) to non‐VTE patients using a biomarker of clot microstructure (df) and clot formation time (TGP). df was the only marker that identified a significant difference (P < 0·001) between the VTE (n = 60) and non‐VTE cohorts (n = 69). The ‘abnormal’ clot microstructures in the VTE patients suggests either inadequate response to anticoagulant therapy or the presence of a procoagulant state not detected by other markers of coagulation (i.e., International Normalized Ratio). Furthermore, elevated values of df in first time VTE patients who later develop a secondary event indicates that df may identify those at risk of recurrence.

A study of microstructural templating in fibrin–thrombin gel networks by spectral and viscoelastic analysis

We report a study of the microstructural templating role of incipient fibrin–thrombin gels by analysis of rheological and confocal microscope measurements. Fractal analysis based on the spectral dimension is used, for the first time, to characterise fibrin gel microstructure in terms of the internal connectivity of gel networks. A significant correlation is found between the fractal characteristics of the incipient gel network and its eventual mature form, confirming that incipient gel microstructure templates ensuing gel development. We report an analytical basis for the study of this templating effect which reveals two different regimes of microstructural development. The first involves low thrombin concentration, in which increasing concentration decreases the gel formation time and alters the fractal characteristics of both incipient and mature gels. In the second regime, involving higher thrombin concentrations, the incipient gel formation time and the fractal characteristics of incipient and mature gels show little variation. The network formation is discussed in terms of computer simulations of incipient fractal networks by the activation-limited aggregation of clusters of rod-like particles. The significance of the work is discussed in terms of biomaterials design for applications involving controlled drug release and wound healing, and improved predictions of blood clot susceptibility to lysis.

Effects of unidirectional flow shear stresses on the formation, fractal microstructure and rigidity of incipient whole blood clots and fibrin gels

Abstract Incipient clot formation in whole blood and fibrin gels was studied by the rheometric techniques of controlled stress parallel superposition (CSPS) and small amplitude oscillatory shear (SAOS). The effects of unidirectional shear stress on incipient clot microstructure, formation kinetics and elasticity are reported in terms of the fractal dimension (df) of the fibrin network, the gel network formation time (TGP) and the shear elastic modulus, respectively. The results of this first haemorheological application of CSPS reveal the marked sensitivity of incipient clot microstructure to physiologically relevant levels of shear stress, these being an order of magnitude lower than have previously been studied by SAOS. CSPS tests revealed that exposure of forming clots to increasing levels of shear stress produces a corresponding elevation in df, consistent with the formation of tighter, more compact clot microstructures under unidirectional flow. A corresponding increase in shear elasticity was recorded. The scaling relationship established between shear elasticity and df for fibrin clots and whole blood confirms the fibrin network as the dominant microstructural component of the incipient clot in terms of its response to imposed stress. Supplementary studies of fibrin clot formation by rheometry and microscopy revealed the substantial additional network mass required to increase df and provide evidence to support the hypothesis that microstructural changes in blood clotted under unidirectional shear may be attributed to flow enhanced thrombin generation and activation. CSPS also identified a threshold value of unidirectional shear stress above which no incipient clot formation could be detected. CSPS was shown to be a valuable haemorheological tool for the study of the effects of physiological and pathological levels of shear on clot properties.

In-situ synthesis of magnetic iron-oxide nanoparticle-nanofibre composites using electrospinning

We demonstrate a facile, one-step process to form polymer scaffolds composed of magnetic iron oxide nanoparticles (MNPs) contained within electrospun nano- and micro-fibres of two biocompatible polymers, Poly(ethylene oxide) (PEO) and Poly(vinyl pyrrolidone) (PVP). This was achieved with both needle and free-surface electrospinning systems demonstrating the scalability of the composite fibre manufacture; a 228 fold increase in fibre fabrication was observed for the free-surface system. In all cases the nanoparticle-nanofibre composite scaffolds displayed morphological properties as good as or better than those previously described and fabricated using complex multi-stage techniques. Fibres produced had an average diameter (Needle-spun: 125±18nm (PEO) and 1.58±0.28μm (PVP); Free-surface electrospun: 155±31nm (PEO)) similar to that reported previously, were smooth with no bead defects. Nanoparticle-nanofibre composites were characterised using scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), dynamic light scattering (DLS) (Nanoparticle average diameter ranging from 8±3nm to 27±5nm), XRD (Phase of iron oxide nanoparticles identified as magnetite) and nuclear magnetic resonance relaxation measurements (NMR) (T1/T2: 32.44 for PEO fibres containing MNPs) were used to verify the magnetic behaviour of MNPs. This study represents a significant step forward for production rates of magnetic nanoparticle-nanofibre composite scaffolds by the electrospinning technique.

The Effects of Temperature on Clot Microstructure and Strength in Healthy Volunteers

BACKGROUND:Anesthesia, critical illness, and trauma are known to alter thermoregulation, which can potentially affect coagulation and clinical outcome. This in vitro preclinical study explores the relationship between temperature change and hemostasis using a recently validated viscoelastic technique. We hypothesize that temperature change will cause significant alterations in the microstructural properties of clot. METHODS:We used a novel viscoelastic technique to identify the gel point of the blood. The gel point identifies the transition of the blood from a viscoelastic liquid to a viscoelastic solid state. Furthermore, identification of the gel point provides 3 related biomarkers: the elastic modulus at the gel point, which is a measure of clot elasticity; the time to the gel point (TGP), which is a measure of the time required to form the clot; and the fractal dimension of the clot at the gel point, df, which quantifies the microstructure of the clot. The gel point measurements were performed in vitro on whole blood samples from 136 healthy volunteers over a temperature range of 27°C to 43°C. RESULTS:There was a significant negative correlation between increases in temperature, from 27°C to 43°C, and TGP (r = −0.641, P < 0.0005). Conversely, significant positive correlations were observed for both the elastic modulus at the gel point (r = 0.513, P = 0.0008) and df (r = 0.777, P < 0.0005) across the range of 27°C to 43°C. When temperature was reduced below 37°C, significant reductions in df and TGP occurred at ⩽32°C (Bonferroni-corrected P = 0.0093) and ⩽29°C (Bonferroni-corrected P = 0.0317), respectively. No significant changes were observed when temperature was increased to >37°C. CONCLUSIONS:This study demonstrates that the gel point technique can identify alterations in clot microstructure because of changes in temperature. This was demonstrated in slower-forming clots with less structural complexity as temperature is decreased. We also found that significant changes in clot microstructure occurred when the temperature was ⩽32°C.

An enhanced rheometer inertia correction procedure (ERIC) for the study of gelling systems using combined motor-transducer rheometers

The rheological characterisation of viscoelastic materials undergoing a sol-gel transition at the Gel Point (GP) has important applications in a wide range of industrial, biological, and clinical environments and can provide information regarding both kinetic and microstructural aspects of gelation. The most rigorous basis for identifying the GP involves exploiting the frequency dependence of the real and imaginary parts of the complex shear modulus of the critical gel (the system at the GP) measured under small amplitude oscillatory shear conditions. This approach to GP identification requires that rheological data be obtained over a range of oscillatory shear frequencies. Such measurements are limited by sample mutation considerations (at low frequencies) and, when experiments are conducted using combined motor-transducer (CMT) rheometers, by instrument inertia considerations (at high frequencies). Together, sample mutation and inertia induced artefacts can lead to significant errors in the determinatio...

The application of large amplitude oscillatory stress in a study of fully formed fibrin clots

The suitability of controlled stress large amplitude oscillatory shear (LAOStress) for the characterisation of the nonlinear viscoelastic properties of fully formed fibrin clots is investigated. Capturing the rich nonlinear viscoelastic behaviour of the fibrin network is important for understanding the structural behaviour of clots formed in blood vessels which are exposed to a wide range of shear stresses. We report, for the first time, that artefacts due to ringing exist in both the sample stress and strain waveforms of a LAOStress measurement which will lead to errors in the calculation of nonlinear viscoelastic properties. The process of smoothing the waveforms eliminates these artefacts whilst retaining essential rheological information. Furthermore, we demonstrate the potential of LAOStress for characterising the nonlinear viscoelastic properties of fibrin clots in response to incremental increases of applied stress up to the point of fracture. Alternating LAOStress and small amplitude oscillatory s...

Transient Extensional Rheology of an Aqueous Gelatin Solution: Before and During Gelation

A technique for studying the extensional rheology of gelling systems is presented. The technique is based on capillary thinning extensional rheometry, and has been used to investigate the extensional rheology of an aqueous gelatin solution before and during gelation. We discuss the qualitative differences between the response prior to and soon after the gel point. In particular, we discuss the effect of gelation on the strain‐ and rate‐dependent extensional response. We also discuss some of the methods and techniques to overcome some of the arising obstacles and limitations.

Formulation, characterisation and flexographic printing of novel Boger fluids to assess the effects of ink elasticity on print uniformity

Model elastic inks were formulated, rheologically characterised in shear and extension, and printed via flexography to assess the impact of ink elasticity on print uniformity. Flexography is a roll-to-roll printing process with great potential in the mass production of printed electronics for which understanding layer uniformity and the influence of rheology is of critical importance. A new set of flexo-printable Boger fluids was formulated by blending polyvinyl alcohol and high molecular weight polyacrylamide to provide inks of varying elasticity. During print trials, the phenomenon of viscous fingering was observed in all prints, with those of the Newtonian ink exhibiting a continuous striping in the printing direction. Increasing elasticity significantly influenced this continuity, disrupting it and leading to a quantifiable decrease in the overall relative size of the printed finger features. As such, ink elasticity was seen to have a profound effect on flexographic printing uniformity, showing the rheological tuning of inks may be a route to obtaining specific printed features.

Fourier Transform Controlled Stress Parallel Superposition (FT-CSPS): Validation and application in processing printable functional materials

In this paper, the development of a multifrequency form of controlled stress parallel superposition rheometry is reported, along with the technique’s validation and use in model gelling systems and high-value particulate suspensions. The novel technique reported herein, termed Fourier transform controlled stress parallel superposition (FT-CSPS), facilitates measurements of the superposition shear moduli and their response to an imposed unidirectional shear stress. FT-CSPS measurements are reported in applications involving (i) the determination of the relaxation properties of incipient gel networks formed in rapidly gelling samples under bulk flow conditions and (ii) measurements of the parallel dynamic moduli of non-gelling samples that experience high rates of solvent loss. By probing the rheological properties of these rapidly evolving materials using a composite waveform comprising multiple harmonic frequencies, sample mutation artefacts (which limit the use of CSPS for such materials) have been minimised. Validation of FT-CSPS has been achieved by (i) showing coincidence of data obtained using CSPS and FT-CSPS for slowly gelling systems and (ii) continuation of the expected relation between gel strength and stress relaxation exponent beyond the range of data accessible to CSPS (limited by sample mutation considerations). This work demonstrates that the rapid acquisition of parallel superposition shear moduli is feasible and facilitates the use of CSPS-based techniques for tests involving rapidly changing materials (such as those undergoing rapid gelation or relatively rapid solvent loss).In this paper, the development of a multifrequency form of controlled stress parallel superposition rheometry is reported, along with the technique’s validation and use in model gelling systems and high-value particulate suspensions. The novel technique reported herein, termed Fourier transform controlled stress parallel superposition (FT-CSPS), facilitates measurements of the superposition shear moduli and their response to an imposed unidirectional shear stress. FT-CSPS measurements are reported in applications involving (i) the determination of the relaxation properties of incipient gel networks formed in rapidly gelling samples under bulk flow conditions and (ii) measurements of the parallel dynamic moduli of non-gelling samples that experience high rates of solvent loss. By probing the rheological properties of these rapidly evolving materials using a composite waveform comprising multiple harmonic frequencies, sample mutation artefacts (which limit the use of CSPS for such materials) have been minim...