The measurement of shear modulus and membrane surface viscosity of RBC membrane with Ektacytometry: a new technique

A new technique is proposed to estimate the shear modulus (mu) and membrane surface viscosity (eta(m)) of red blood cell (RBC). Theoretical formulae for finding these two parameters are first derived based on the force balance on a RBC in a flow field of low viscosity. Different types of Ektacytometry are then used to measure relevant quantities. The obtained values (mu=6.1 x 10(-6)N/m, eta(m)=8.8 x10 (-7)Ns/m for normal RBC) are consistent with those previously found by micropipette technique and in AC electric field. The present technique is, however, much easier to operate and more advantageous in reflecting the average properties of a large quantity of RBCs, and it is much cheaper to be applied in clinical practice than any other method of measuring the two parameters. The sensitivity of the technique is demonstrated by testing RBCs treated with glutaraldehyde of different concentrations. This technique was demonstrated by the flow chamber.     

Viscoelastic properties of whole blood. Influence of fast sedimenting red blood cell aggregates

Red blood cell (RBC) aggregation is known to be of deciding influence on erythrocyte sedimentation-rate (ESR) and on whole blood viscoelastic properties. The rheological behaviour of blood collected from a control-group with normal ESR is compared to the viscoelastic behaviour of blood collected from two groups with high to very high ESR, whose individuals are suffering from chronical polyarthritis and Morbus Bechterew, respectively. The rheological properties are evaluated by means of an oscillating-flow capillary-rheometer where the viscous (eta') and elastic (eta") component of the complex viscosity (eta) is measured at a constant frequency of 2 Hz. Correcting for the varying hematocrit of the different blood samples according to an exponential equation, the viscoelastic data are found to be elevated in the groups with high ESR. For the viscous properties this is only due to the increase of the plasma viscosity. A correction for the plasma viscosity, however, shows that the viscous properties at low shear- rates (2s-1) are significantly reduced, whereas elastic properties in a range of medium shear-rates (10s-1 to 50s-1) are significantly increased (P less than 0.001, t-test of Student). This result is discussed to be due to the high packing density of the RBC in fast sedimenting aggregates. High packing density reduces the effective volume of the RBC but increases the stiffness of the aggregates.     

A finite element study of invariant-based orthotropic constitutive equations in the context of myocardial material parameter estimation

A previous study investigated a number of invariant-based orthotropic and transversely isotropic constitutive equations for their suitability to fit three-dimensional simple shear mechanics data of passive myocardial tissue. The study was based on the assumption of a homogeneous deformation. Here, we extend the previous study by performing an inverse finite element material parameter estimation. This ensures a more realistic deformation state and material parameter estimates. The constitutive relations were compared on the basis of (i) ‘goodness of fit’: how well they fit a set of six shear deformation tests and (ii) ‘variability’: how well determined the material parameters are over the range of experiments. These criteria were utilised to discuss the advantages and disadvantages of the constitutive relations. It was found that a specific form of the polyconvex type as well as the exponential Fung-type equations were most suitable for modelling the orthotropic behaviour of myocardium under simple shear.     

Age and androgen-related changes in morphological parameters, haematological indices and serum protein fraction in common vole (Microtus arvalis Pall.) growing in different photoperiods

In male voles raised under different light conditions (L:D = 24:0, L:D = 18:6, L:D = 6:18) the following results were obtained. In the young males erythropoiesis seemed to be greatest in voles growing in "winter" photoperiod. In this light condition RBC count showed a tendency to decrease with age while in other illuminations erythrocyte numbers increased. The age related changes in RBC count were very similar to those observed in common voles aging in the different natural seasonal generations. The androgen level was higher after birth and then decreased. After reaching sexual maturity (13-15 weeks of vole life) it showed a peak and then decreased progressively. The age related changes in alpha 2-globulins (considered to be a potential binding proteins for steroid hormones) and in androgen level suggested a regulating mechanism based on a balance of hormones--blood proteins. The correlations stated between androgens and the parameters studied indicated more steroid than age-related metabolic changes. The more androgens, the less beta-globulin and fibrinogen while the greater rate of body and testes weight, as well as of albumin level and of RBC count were observed. All those might affect the transmission of lipoproteins from blood to adipose tissue and less predisposition to fighting with potentially lower blood coagulation capacity and more effective anabolic (erythropoietic) processes in the more "androgenic", reproductively active male voles. A higher protein metabolism in constant light, i.e. the higher level of the majority of plasma protein fraction and small weight was suggested. The high level of androgens in "winter" photoperiod might condition the aggressiveness and prevent hibernation in natural winter conditions.     

Cellular determinants of low-shear blood viscosity

Low-shear viscometry is one of the methods commonly used to estimate the degree of red blood cell (RBC) aggregation in various bloods and RBC suspensions. However, it has been previously shown that alterations in RBC morphology and mechanical behavior can affect the low-shear apparent viscosity of RBC suspensions; RBC aggregation is also sensitive to these cellular factors. This study used heat treatment (48°C, 5 min), glutaraldehyde (0.005–0.02%) and hydrogen peroxide (1 mM) to modify cell geometry and deformability. Red blood cell aggregation was assessed via a Myrenne Aggregometer (“M” and “Ml” indexes), RBC suspension viscosity was measured using a Contraves LS-30 viscometer, and RBC shape response to fluid shear stresses (i.e., deformability) was determined by ektacytometry (LORCA system). Our results indicate that low-shear apparent viscosity and related indexes may not always reflect changes of RBC aggregation if cellular properties are altered: for situations where RBC aggregation has been only moderately affected, cellular mechanical factors may be the major determinant of low-shear viscosity. These findings thus imply that in situations which may be associated alterations of RBC geometry and/or deformability, low-shear viscometry should not be the sole measurement technique used to assess RBC aggregation.     

A thermodynamic optimization analysis of a possible relation between the parameters that determine the energetics of muscle contraction in steady state.

Given the phenomenological relations for muscle's steady-state contraction and proper definitions of power p and efficiency eta, the behavior of these quantities is analysed in terms of the parameters that determine the energetics of the muscle, here denoted by s(o)and alpha. s(o)is proportional to the so-called maintenance heat, while alpha is the parameter that determines the curvature of the Hill's force-velocity curve. The dependence of the muscle's power and efficiency, averaged over the whole range of force the muscle can exert, on the parameters s(o)and alpha is studied. The average power p(avg)is a function only of alpha, and is a growing function that approaches 1/6 asymptotically as alpha goes to infinity. The average efficiency eta(avg)is a function of both alpha and s(o). With the value of s(o)fixed, the graph of the function eta(avg)(s(o), alpha) is a convex curve with a single maximum. The value and the position of this maximum point both depend on s(o). In the limit alpha-->0, s(o)-->0, eta(avg)tends to 1. The points (s(o), alpha(m)(s(o))), with alpha(m)(s(o)) the value of alpha that maximizes eta(avg)for a given s(o), are fitted by the curve alpha=s(o 1/2). This relation was experimentally found by A.V. Hill in his early studies of muscle energetics. Other experimental data are found to qualitatively satisfy the same relation. Although some dynamical microscopic models for muscle contraction, based upon Huxley's cross-bridge model, show that the same kinetic parameters control both the maintenance heat (s(o)) and the muscle's power output (alpha), we suggest that the exact relation between them has been reached due to the evolutive stresses that made individuals with equally powerful and more efficient muscles more suitable to reproduce.     

Effects of couple stresses on the blood flow through an artery with mild stenosis

An analysis of the effects of couple stresses on the blood flow through thin artery in the presence of very mild stenosis has been carried out with the help of two nondimensional parameters, alpha (the length ratio parameter) and eta (the parameter characterizing the antisymmetric property of the couple stress tensor). It is shown that an increase in the couple stress (small value of alpha and eta), increases the resistance to the flow and the wall shear stress. These characteristics are further enhanced by the presence of the stenosis.     

Pulsatile blood flow and oxygen transport past a circular cylinder

The fundamental study of blood flow past a circular cylinder filled with an oxygen source is investigated as a building block for an artificial lung. The Casson constitutive equation is used to describe the shear-thinning and yield stress properties of blood. The presence of hemoglobin is also considered. Far from the cylinder, a pulsatile blood flow in the x direction is prescribed, represented by a time periodic (sinusoidal) component superimposed on a steady velocity. The dimensionless parameters of interest for the characterization of the flow and transport are the steady Reynolds number (Re), Womersley parameter (alpha), pulsation amplitude (A), and the Schmidt number (Sc). The Hill equation is used to describe the saturation curve of hemoglobin with oxygen. Two different feed-gas mixtures were considered: pure O(2) and air. The flow and concentration fields were computed for Re=5, 10, and 40, 0< or =A< or =0.75, alpha=0.25, 0.4, and Schmidt number, Sc=1000. The Casson fluid properties result in reduced recirculations (when present) downstream of the cylinder as compared to a Newtonian fluid. These vortices oscillate in size and strength as A and alpha are varied. Hemoglobin enhances mass transport and is especially important for an air feed which is dominated by oxyhemoglobin dispersion near the cylinder. For a pure O(2) feed, oxygen transport in the plasma dominates near the cylinder. Maximum oxygen transport is achieved by operating near steady flow (small A) for both feed-gas mixtures. The time averaged Sherwood number, Sh, is found to be largely influenced by the steady Reynolds number, increasing as Re increases and decreasing with A. Little change is observed with varying alpha for the ranges investigated. The effect of pulsatility on Sh is greater at larger Re. Increasing Re aids transport, but yields a higher cylinder drag force and shear stresses on the cylinder surface which are potentially undesirable.     

Changes in the blood parameters of an air-breathing fish during different respiratory conditions.

Some of the blood parameters recorded in an air-breathing eel, Amphipnous cuchia under normal respiratory condition during non-breeding period (September-April) are haemoglobin (Hb) concentration 19.26%, haematocrit value 56.16%, RBC number 1.71 million/mm3, RBC size 18.86 X 9.70 mum, mean corpuscular haemoglobin (MCH) 113.4 ng, mean corpuscular haemoglobin concentration (MCHC) 34.2%, blood sugar 77 mg% and ascorbic acid 0.435 mg%. The higher concentration of haemoglobin (19.26%) appears to be related to its obligatory air breathing habit and habitat in a water of low oxygen content. Though a definite trend of increase in the haemoglobin and haematocrit concentration with an increase in the body weight of the fish was lacking, variations were clearly marked related to intrinsic activity of the fish connected with different respiratory conditions. Asphyxiation in a submerged but continuous flow of water (liter/h) for 5 1/2 h resulted in an increase in the above-mentioned parameters to an appreciable extent. These increases were 0.23 million/mm3 in the number of erythrocytes, 6.16% in haemoglobin concentration, 10% in haematocrit value, 20% in blood sugar and 35% in ascorbic acid content. The mean corpuscular haemoglobin showed a decline of 6.2%. Exclusive aerial breathing for 5 1/2 h also caused 7.4% increase in haemoglobin concentration, 9.4% in haematocrit value, 0.14 million/mm3 in RBC number, 20% in blood sugar level, 9% in ascorbic acid content but almost no change in mean corpuscular haemoglobin. The average surface area for diffusion of gases appeared to have reduced by 6.8 mum2 per RBC.     

Effects of swimming exercise on red blood cell rheology in trained and untrained rats.

Red blood cell (RBC) mechanical properties were investigated after swimming exercise in trained and untrained rats. A group of rats was trained for 6 wk (60 min swimming, daily), and another group was kept sedentary. Blood samples were obtained either within 5 min or 24 h after 60 min swimming in both groups. In the untrained rats, the RBC aggregation index decreased to 2.60 +/- 0.4 immediately after exercise from a control value of 6.73 +/- 0.18 (P < 0.01), whereas it increased to 13.13 +/- 0.66 after 24 h (P < 0.01). RBC transit time through 5-microm pores increased to 3.53 +/- 0.16 ms within 5 min after the exercise from a control value of 2.19 +/- 0. 07 ms (P < 0.005). A very significant enhancement (166%) in RBC lipid peroxidation was detected only after 24 h. In the trained group, the alterations in all these parameters were attenuated; there was a slight, transient impairment in RBC deformability (transit time = 2.64 +/- 0.13 ms), and lipid peroxidation was found to be unchanged. These findings suggest that training can significantly limit the hemorheological alterations related to a given bout of exercise. Whether this effect is secondary to the training-induced reduction in the degree of metabolic and/or hormonal perturbation remains to be determined.     

Effect of nickel on red blood cell parameters and on serum vitamin B12, folate and homocysteine concentrations during pregnancy with and without anemia

BackgroundResearch to date suggests that nickel affects not only the metabolism of vitamin B12 but also folates and thus may affect hematopoiesis processes.ObjectiveThe aim of the study was to examine the relationship of nickel (Ni) status to red blood cell (RBC) parameters and serum vitamin B12, folate and homocysteine concentrations in the course of normal pregnancy and in pregnant women with anemia.MethodsThe study included fifty-three pregnant women recruited to the study from the Lower Silesia region of Poland, 17 % of whom developed anemia. Nickel concentration was determined in urine, whole blood and food samples by atomic absorption spectrometry. At the same time as the food and urine samples were taken, blood was also collected for the determination of RBC parameters and serum vitamin B12, homocysteine and folate concentrations.ResultsThe median reported Ni intake, and the urinary and whole blood nickel contents for the studied pregnant women for the first trimester were respectively – 162.46 μg/day, 3.98 μg/L and 3.32 μg/L; for the second trimester – 110.48 μg/day, 6.86 μg/L and 1.04 μg/L; and for the third trimester – 132.20 μg/day, 3.41 μg/L and 0.70 μg/L. With regard to Ni concentration in whole blood (p = 0.0204) and in urine (p = 0.0003), the differences in the values for individual trimesters were statistically significant. The whole blood Ni level was significantly higher (9.28 vs 3.62 μg/L, p = 0.0114), while the concentration of homosysteine was significantly lower (4.09 vs 5.04 μmol/L, p = 0.0165) in pregnant women with anemia compared to those without anemia. The whole blood Ni concentration was negatively correlated with almost all RBC parameters in non-anemic pregnant women.ConclusionsNi status changes with the development of normal pregnancy, and in the case of anemia, an increase in Ni concentration in whole blood is observed. The demonstrated correlations between the Ni status in pregnant women and RBC parameters as well as serum vitamin B12 and folate concentrations suggest that nickel is associated with the methionine–folate cycle, iron homeostasis and bacterial synthesis of vitamin B12 in humans.     

On the application of a constitutive equation for whole human blood

In consideration of the pulsatile blood flow in a conduit, the constitutive equation for the whole human blood of F. J. Walburn and D. J. Schneck (Biorheology, Vol. 13, 1976, pp. 201-210) is utilized. Governing equations are solved numerically yielding the velocity and the shear stress distributions. These results are discussed and compared with the Newtonian fluid, Casson's fluid, and Bingham fluid applications.     

Effects of blood storage on rheology and damage in low-stress shear flow

Data are presented on the rheological and hemolytic behavior of whole human blood as it ages while stored at 4 degrees C (as in blood banking practice) up to 26 days. The viscometric properties of steady shear viscosity eta and oscillatory (complex) viscosity eta * = eta' - i eta" reported over ranges of shear rate gamma and radian frequency omega of 33 less than gamma less than 4130 s-1 and 1.5 less than omega less than 48 s -1; data on autologous plasma are given for reference. The Cox-Merz relation, eta (gamma) = [eta *(omega)] omega = gamma, is found to be a good approximation, with eta greater than or equal to [eta *], over the range studied. Release of hemoglobin (Hgb) and lactate dehydrogenase (LDH) into the plasma during shearing is tracked as a function of time for 30 min, and its sensitivity to gamma magnitude is measured. Bloods from four different donors are studied, with primary attention given to one (SSR). For all bloods, the release of both Hgb and LDH increases with storage age, but differences in such aging characteristics between different bloods can be substantial (even when rheological properties are identical). A post-shear incubation at 4 degrees C for one day shows no enhancement of plasma Hgb and LDH levels beyond those expected from normal aging after the shearing experience, demonstrating the absence of significant delayed-action effects as a consequence of shearing trauma.     

Myocardial material parameter estimation: a comparison of invariant based orthotropic constitutive equations

This study investigated a number of invariant based orthotropic and transversely isotropic constitutive equations for their suitability to fit three-dimensional simple shear mechanics data of passive myocardial tissue.

A number of orthotropic laws based on Green strain components and one microstructurally based law have previously been investigated to fit experimental measurements of stress-strain behaviour. Here we extend this investigation to include several recently proposed functional forms, i.e. invariant based orthotropic and transversely isotropic constitutive relations.

These laws were compared on the basis of (i) ‘goodness of fit’: how well they fit a set of six shear deformation tests, (ii) ‘variability’: how well determined the material parameters are over the range of experiments. These criteria were utilised to discuss the advantages and disadvantages of the constitutive laws.

It was found that a specific form of the polyconvex type as well as the exponential Fung-type law from the previous study were most suitable for modelling the orthotropic behaviour of myocardium under simple shear.     

Deformability of red blood cells from different species studied by resistive pulse shape analysis technique

The Cell Transit Analyzer (CTA) is now being used widely in clinical hemorheology. Most of the data obtained by CTA are limited to human blood, although the CTA has an important potential to be used in experimental studies on animal models. However, behavior of red blood cells (RBC) from various species might be different in CTA. Eight parameters reflecting different aspects of cell passage through pores with 5 μm diameter and 15 μm length were determined or human, guinea pig, dog, rabbit, rat, mouse and sheep RBC, together with instrument precision and biological variation. These parameters have a wide range when measured in different species and correlate with cell volume. Sensitivity of these parameters to the glutaraldehyde-induced alterations in RBC deformability was not same for different laboratory mammals. The main reason for this difference seems to be related to the cell size and thus sensitivity might be significantly limited if 5 μm pore-size filters are used to test the smaller RBC. The results of this study may help in designing experimental studies on laboratory mammals using the CTA.     

The effects of cholesterol oxidation products in sickle and normal red blood cell membranes.

The oxysterol content in normal and sickle red blood cell (RBC) membranes was assessed using thin-layer chromatography and capillary gas chromatography/mass spectrometry. Several more oxysterols were present in sickle RBCs compared to normal RBCs. Sickle RBC membranes had a higher concentration of 5 alpha,6 alpha-epoxycholesterol, 5 alpha-cholestane-3 beta,5,6 beta-triol, 7-ketocholesterol and 19-hydroxycholesterol than normal RBC membranes. The increased oxysterols in sickle RBC may be an effect of the increased oxidative stress which occurs in sickle RBC membranes. Physical characteristics of normal and sickle RBC membrane ghosts with and without inserted oxysterols were examined by Fourier transform infrared spectroscopy. The data are consistent with a greater sterol content in sickle cells compared to normal RBC membranes, and a possible oxysterol-cholesterol synergism.     

Discrimination between red blood cell and platelet components of blood clots by MR microscopy

Magnetic resonance imaging (MRI) of pulmonary emboli obtained ex vivo, verified by immunohistochemistry, showed that platelet layers display brighter signal intensity than areas containing predominantly red blood cells (RBC) in T (1)-weighted MRI. These results were surprising since platelets do not contain paramagnetic haemoglobin that would enhance magnetic relaxation. Our assumption was that the fibrin meshwork areas with entrapped RBC retain abundant extracellular space filled with serum, whereas platelets regroup into tight aggregates lacking serum, essentially mimicking solid tissue structure, rich with cellular proteins that enhance T (1)-relaxation. Our hypothesis was examined by MRI and NMR relaxometry of in vitro RBC suspensions and sedimented platelets, as well as by MRI of model clots and pulmonary emboli obtained ex vivo. Pure sedimented platelets exhibited shorter proton spin lattice relaxation times (T (1) = 874 +/- 310 ms) than those of venous blood of a healthy male with 40% haematocrit (T (1) = 1277 +/- 66 ms). T (1)-values of RBC samples containing high haematocrit (>/=80%) resembled T (1) of platelet samples. In T (1)-weighted spin-echo MRI echo time and repetition time (TE/TR = 10/120 ms) the ratio of signal intensities between a non-retracted whole blood clot (with a haematocrit of 35%) and a pure platelet clot was 3.0, and the ratio between a retracted whole blood clot with an estimated haematocrit of about 58% and a pure platelet clot was 2.6. We conclude that T (1)-weighted MRI can discriminate between platelet layers of thrombi and RBC-rich areas of thrombi that are not compacted to a haematocrit level of >/=80%.