Microvascular pressure and functional capillary density in extreme hemodilution with low- and high-viscosity dextran and a low-viscosity Hb-based O2 carrier

Blood losses are usually corrected initially by the restitution of volume with plasma expanders and subsequently by the restoration of oxygen-carrying capacity using either a blood transfusion or possibly, in the near future, oxygen-carrying plasma expanders. The present study was carried out to test the hypothesis that high-plasma viscosity hemodilution maintains perfused functional capillary density (FCD) by preserving capillary pressure. Microvascular pressure responses to extreme hemodilution with low- (LV) and high-viscosity (HV) plasma expanders and an exchange transfusion with a polymerized bovine cell-free Hb (PBH) solution were analyzed in the awake hamster window chamber model (n = 26). Systemic hematocrit was reduced from 50% to 11%. PBH produced a greater mean arterial blood pressure than the nonoxygen carriers. FCD was higher after a HV plasma expander (70 +/- 15%) vs. PBH (47 +/- 12%). Microvascular pressure spanning the capillary network was higher after a HV plasma expander (16-19 mmHg) compared with PBH (12-16 mmHg) and a LV plasma expander (11-14 mmHg) but lower than control (22-26 mmHg). FCD was found to be directly proportional to capillary pressure. The use of a HV plasma expander in extreme hemodilution maintained the number of perfused capillaries and tissue perfusion by comparison with a LV plasma expander due to increased mean arterial blood pressure and capillary pressure. The use of PBH increased mean arterial pressure but reduced capillary pressure due to vasoconstriction and did not maintain FCD.     

Lactate concentration differences in plasma, whole blood, capillary finger blood and erythrocytes during submaximal graded exercise in humans

The aim of the study was to investigate the distribution of lactate in plasma, whole blood, erythrocytes, and capillary finger blood, before and during submaximal exercise. Ten healthy male subjects performed submaximal graded cycle ergometer exercise for 20-25 min. Venous blood samples and capillary finger blood samples were taken before exercise and every 5th min during exercise for lactate determination. The plasma lactate concentration was significantly higher (P less than 0.001, approximately 50%) than in the erythrocytes. This difference was not altered by the venous blood lactate concentration or exercise intensity. A significant difference (P less than 0.01) in lactate concentration was also found between capillary whole blood and venous whole blood. It was concluded that direct comparisons between lactate in capillary finger blood, venous whole blood and plasma could not be made.     

Microvascular and capillary perfusion following glycocalyx degradation.

Systemic parameters and microvascular and capillary hemodynamics were studied in the hamster window chamber model before and after hyaluronan degradation by intravenous injection of Streptomyces hyaluronidase (100 units, 40-50 U/ml plasma). Glycocalyx permeation was estimated using fluorescent markers of different molecular size (40, 70, and 2,000 kDa), and electrical charge. Systemic parameters (blood pressure, heart rate, blood gases) and microhemodynamics (vascular tone, velocity, and blood flow) remained statistically unchanged after injection of hyaluronidase, compared with inactivated hyaluronidase. Conversely, capillary hemodynamics were drastically affected. Functional capillary density, the capillaries perfused with red blood cells (RBCs), decreased by 35%, capillary Hct of the remaining functional capillaries increased from 16 to 27%, and penetration of 70-kDa fluorescent marker increased. Furthermore, plasma-only perfused capillaries statistically increased 30 min after hyaluronidase. The decrease in functional capillary density accounted for an increased RBC flux in the remainder of the capillaries, since the same number of RBCs had to traverse a reduced number of capillaries. Flux balances showed a reduction from baseline of 11% for the RBC flux and 20% for the plasma flux after treatment. These discrepancies are within the margin of error of the techniques used and could be explained by accounting for RBC over-velocity compared with plasma. These findings suggest that the decrease in the glycocalyx leads to capillary perfusion impairments.     

Effect of blood flow on gas transport in a pulmonary capillary

The effects of blood velocity on gas transport within the alveolar region of lungs, and on the lung diffusing capacity DL have for many years been regarded as negligible. The present work reports on a preliminary, two-dimensional investigation of CO convection-diffusion phenomenon within a pulmonary capillary. Numerical simulations were performed using realistic clinical and morphological parameter values, with discrete circular red blood cells (RBCs) moving with plasma in a single capillary. Steady-state simulations with stationary blood (RBCs and plasma) were performed to validate the model by comparison with published data. Results for RBCs moving at speeds varying from 1.0 mm/s to 10 mm/s, and for capillary hematocrit (Ht) from 5% to 55%, revealed an increase of up to 60% in DL, as compared to the stationary blood case. The increase in DL is more pronounced at low Ht (less than 25%) and high RBC speed and it seems to be caused primarily by the presence of plasma. The results also indicate that capillary blood convection affects DL not only by improving the plasma mixing in the capillary bed but also by replenishing the capillary with fresh (zero concentration) plasma, providing an additional reservoir for the consumption of CO. Our findings cast doubt on the current belief that an increase in the lung diffusing capacity of humans (for instance, during exercising), with fixed hematocrit, can only be accomplished by an increase in the lung volume effectively active in the respiration process.     

Blood flow in the capillary bed

From arteries to veins, the blood has to go through the ‘capillary’ blood vessels. These blood vessels are so small that often their diameter is smaller than that of the red blood cells. Intimate interactions occur, therefore, between the blood cells and the blood vessels.

A general survey of recent works on capillary blood flow is given in this article. Some details are presented for two problems: the problem of deformation of the flexible red blood cells, their motion in the capillary blood vessels, and the pressure drop due to the red cell blood vessel interaction; and the problem of the flow of plasma ‘bolus’ between neighboring red cells.

The solution supplies many details about the microcirculation phenomenon. Taken together, a method is offered for the calculation of pressure drop in the capillary as a function of various physical parameters: the red cell volume per unit blood volume, (hematocrit), the ratio of the cell diameter to the blood vessel diameter, the ratio of the length of the blood vessel to its length, the volume of individual red cells, and a parameter relating the cell membrane elasticity, plasma viscosity and the cell velocity.     

Determination of high density lipoprotein cholesterol in venous and capillary whole blood

A procedure is presented and evaluated for separation of plasma high density lipoprotein from either capillary or venous whole blood. The lipoprotein is separated by adding 50 microliter of sample to 250 microliter of 0.15 M NaCl solution containing 99.9 g/l polyethyleneglycol 6000, 0.0374 g/l dextran sulfate (Mr 15,000) and 2.6 mM Mg2+. After gentle mixing for a few minutes and standing 10 min at room temperature, mixtures are centrifuged (1,500 g) for 10 min and cholesterol is measured on 200 microliter of supernatant by an enzymatic-colorimetric method. Comparison studies demonstrate a good correlation between high density lipoprotein cholesterol in plasma and capillary or venous whole blood. The procedure is simple, has the advantage of using either K3-EDTA-anticoagulated whole blood, without the need of centrifugation, or capillary whole blood which can also be collected away from the laboratory.     

STUDIES ON BLOOD CAPILLARIES : II. Transport of Ferritin Molecules across the Wall of Muscle Capillaries

The pathway by which intravenously injected ferritin molecules move from the blood plasma across the capillary wall has been investigated in the muscle of the rat diaphragm. At 2 min after administration, the ferritin molecules are evenly distributed in high concentration in the blood plasma of capillaries and occur within vesicles along the blood front of the endothelium. At the 10-min time point, a small number of molecules appear in the adventitia, and by 60 min they are relatively numerous in the adventitia and in phagocytic vesicles and vacuoles of adventitial macrophages. Thereafter, the amount of ferritin in the adventitia and pericapillary regions gradually increases so that at 1 day the concentration in the extracellular spaces approaches that in the blood plasma. Macrophages and, to a lesser extent, fibroblasts contain large amounts of ferritin. 4 days after administration, ferritin appears to be cleared from the blood and from the capillary walls, but it still persists in the adventitial macrophages and fibroblasts. At all time points examined, ferritin molecules within the endothelial tunic were restricted to vesicles or to occasional multivesicular or dense bodies; they were not found in intercellular junctions or within the cytoplasmic matrix. Ferritin molecules did not accumulate within or against the basement membranes. Over the time period studied, the concentration of ferritin in the blood decreased, first rapidly, then slowly, in two apparently exponential phases. Liver and spleen removed large amounts of ferritin from the blood. Diaphragms fixed at time points from 10 min to 1 day, stained for iron by the Prussian Blue method, and prepared as cleared whole mounts, showed a progressive and even accumulation of ferritin in adventitial macrophages along the entire capillary network. These findings indicate: (1) that endothelial cell vesicles are the structural equivalent of the large pore system postulated in the pore theory of capillary permeability; (2) that the basement membrane is not a structural restraint in the movement of ferritin molecules across the capillary wall; (3) that transport of ferritin occurs uniformly along the entire length of the capillary; and (4) that the adventitial macrophages monitor the capillary filtrate and partially clear it of the tracer.     

Determination of the adrenergic response by measurement of catecholamines and activity of dopamine-beta-hydroxylase (D-beta-B) in arterial capillary blood]

Plasma dopamine-beta-hydroxylase activity and catecholamines (adrenaline and noradrenaline) level in venous blood taken from the cubital vein and in arterialized capillary blood taken from the ear lobe were measured before and after a maximal exercise on a treadmill in 14 healthy untrained volunteers. The authors have shown a good correlation between these parameters in venous blood and those in capillary blood. It is concluded that the determination of plasma dopamine-beta-hydroxylase activity and catecholamines level in arterialized capillary blood could be a valid measure of the sympathetic activity.     

Dynamical clustering of red blood cells in capillary vessels

We have modeled the dynamics of a 3-D system consisting of red blood cells (RBCs), plasma and capillary walls using a discrete-particle approach. The blood cells and capillary walls are composed of a mesh of particles interacting with harmonic forces between nearest neighbors. We employ classical mechanics to mimic the elastic properties of RBCs with a biconcave disk composed of a mesh of spring-like particles. The fluid particle method allows for modeling the plasma as a particle ensemble, where each particle represents a collective unit of fluid, which is defined by its mass, moment of inertia, translational and angular momenta. Realistic behavior of blood cells is modeled by considering RBCs and plasma flowing through capillaries of various shapes. Three types of vessels are employed: a pipe with a choking point, a curved vessel and bifurcating capillaries. There is a strong tendency to produce RBC clusters in capillaries. The choking points and other irregularities in geometry influence both the flow and RBC shapes, considerably increasing the clotting effect. We also discuss other clotting factors coming from the physical properties of blood, such as the viscosity of the plasma and the elasticity of the RBCs. Modeling has been carried out with adequate resolution by using 1 to 10 million particles. Discrete particle simulations open a new pathway for modeling the dynamics of complex, viscoelastic fluids at the microscale, where both liquid and solid phases are treated with discrete particles. Figure A snapshot from fluid particle simulation of RBCs flowing along a curved capillary. The red color corresponds to the highest velocity. We can observe aggregation of RBCs at places with the most stagnant plasma flow.     

Plug Effect of Erythrocytes in Capillary Blood Vessels

As an idealized problem of the motion of blood in small capillary blood vessels, the low Reynolds number flow of plasma (a newtonian fluid) in a circular cylindrical tube involving a series of circular disks is studied. It is assumed in this study that the suspended disks are equally spaced along the axis of the tube, and that their centers remain on the axis of the tube and that their faces are perpendicular to the tube axis. The inertial force of the fluid due to the convective acceleration is neglected on the basis of the smallness of the Reynolds number. The solution of the problem is derived for a quasi-steady flow involving infinitesimally thin disks. The numerical calculation is carried out for a set of different combinations of the interdisk distance and the ratio of the disk radius to the tube radius. The ratio of the velocity of the disk to the average velocity of the fluid is calculated. The different rates of transport of red blood cells and of plasma in capillary blood vessels are discussed. The average pressure gradient along the axis of the tube is computed, and the dependence of the effective viscosity of the blood on the hematocrit and the diameter of the capillary vessel is discussed.     

A numerical model for studying the effect of plasma layer on the process of blood oxygenation in the pulmonary capillaries

A two layer model for the blood oxygenation in pulmonary capillaries is proposed. The model consists of a core of erythrocytes surrounded by a symmetrically placed plasma layer. The governing equations in the core describe the free molecular diffusion, convection, and facilitated diffusion due to the presence of haemoglobin. The corresponding equations in the plasma layer are based on the free molecular diffusion and the convective effect of the blood. According to the axial train model for the blood flow proposed by Whitmore (1967), the core will move with a uniform velocity whereas flow in the plasma layer will be fully developed. The resulting system of nonlinear partial differential equations is solved numerically. A fixed point iterative technique is used to deal with the nonlinearities. The distance traversed by the blood before getting fully oxygenated is computed. It is shown that the concentration of O2 increases continuously along the length of the capillary for a given ratio of core radius to capillary radius. It is found that the rate of oxygenation increases as the core to capillary ratio decreases. The equilibration length increases with a heterogeneous model in comparison to that in a homogeneous model. The effect of capillary diameters and core radii on the rate of oxygenation has also been examined.     

Capillary electrophoresis screening of poisonous anions extracted from biological samples

A method was developed for screening human biological samples for poisonous anions using capillary electrophoresis (CE) employing indirect UV detection. The run buffer consisted of 2.25 mM pyromellitic acid, 1.6 mM triethanolamine, 0.75 mM hexamethonium hydroxide and 6.5mM NaOH at pH 7.7. Biological samples were pretreated using solid phase extraction. The method was applied to the analysis of human blood, plasma, urine, and intestinal contents. Twenty-nine different anions were detectable at aqueous concentrations of 1 part per million (ppm) with a typical analysis time less than 20 min. Intraday migration time R.S.D. and peak area R.S.D. for blood samples were less than 1.1% and 6.3%, respectively. Interday migration time R.S.D. for plasma samples ranged from 7.5% to 10.4%. The new method produced efficient separations of various target anions extracted from complex biological matrices.     

Blood flow switching among pulmonary capillaries is decreased during high hematocrit.

Pulmonary capillary perfusion within a single alveolar wall continually switches among segments, even when large-vessel hemodynamics are constant. The mechanism is unknown. We hypothesize that the continually varying size of plasma gaps between individual red blood cells affects the likelihood of capillary segment closure and the probability of cells changing directions at the next capillary junction. We assumed that an increase in hematocrit would decrease the average distance between red blood cells, thereby decreasing the switching at each capillary junction. To test this idea, we observed 26 individual alveolar capillary networks by using videomicroscopy of excised canine lung lobes that were perfused first at normal hematocrit (31-43%) and then at increased hematocrit (51-62%). The number of switches decreased by 38% during increased hematocrit (P < 0.01). These results support the idea that a substantial part of flow switching among pulmonary capillaries is caused by the particulate nature of blood passing through a complex network of tubes with continuously varying hematocrit.     

Pre-analytical factors affecting ascorbic and uric acid quantification in human plasma

We have recently described a new capillary electrophoresis assay to measure serum ascorbic and uric acids in which a baseline separation of peaks was obtained in less than 4 min by using a 60.2 cm x 75 microm uncoated capillary with a 100 mmol/L sodium borate running buffer pH 8. Since during sample preparation AA is rapidly oxidized, we employed our new capillary electrophoresis method to analyze the pre-analytical factors affecting its stability. In particular we evaluated how the standard mix preparation, the blood collection (plasma EDTA or serum) and the plasma protein precipitation influence the results of analysis. Our data suggest that standard ascorbate must be dissolved in a solution containing cysteine and EDTA in order to avoid oxidation and that EDTA blood collection is better than serum for AA measurement. Moreover, the type and the quantity of the precipitating compound are critical parameters to obtain a complete recovery of analytes. We performed AA and UA analysis in 32 healthy volunteers with the optimized experimental conditions by using our capillary electrophoresis method and a reference CE assay. Obtained data were compared to Bland-Altman test to verify the accuracy of our CZE method.     

In-silico fragment-based identification of novel angiogenesis inhibitors

Inhibition of vascular endothelial growth factor receptor-2 (VEGFR2) kinase blocks angiogenesis, the process of generating new capillary blood vessels that are important in tumor growth. To identify small molecule inhibitors of the VEGFR2 kinase, we undertook a computer assisted fragment-based screening that used 3-D structural models of the VEGFR2 kinase, and hinge region as a fragment anchor point. Seven novel non-cytotoxic compounds were identified which limited the induction of capillary networks by human umbilical vein endothelial cells in the low micromolar range.     

Capillary electrophoresis method optimized with a factorial design for the determination of glutathione and amino acid status using human capillary blood

Plasma aromatic and sulfur containing amino acids are good indicators of protein anabolism/catabolism, while blood reduced and oxidized glutathione reflect oxidative status in an organism. Using a full factorial design for screening important variables (pH, concentration, temperature) we developed a capillary zone electrophoresis method permitting their measurements in the single run, without any derivatization procedures. The best separations were obtained within less than 30min employing a 10mmol/l phosphate buffer, pH 2.8, 18 degrees C, 15kV voltage. Fairly good precision with a linear relationship between peak area and concentrations (r=0.995-0.999) were obtained. The method was used to analyze human capillary blood.     

Validation of HCV-NAT Assays and Experience with NAT Application for Blood Screening in Germany

Validation of HCV-NAT assays is an important prerequisite for the use of NAT for screening plasma or blood donations. The main NAT features to be validated are specificity, detection limit and robustness. Preliminary experience in Germany obtained with different methodical and logistic approaches shows the feasibility of HCV-NAT as a screening test for blood donations.     

Neutrophil activation in systemic capillary leak syndrome (Clarkson disease)

Systemic capillary leak syndrome (SCLS; Clarkson disease) is a rare orphan disorder characterized by transient yet recurrent episodes of hypotension and peripheral oedema due to diffuse vascular leakage of fluids and proteins into soft tissues. Humoral mediators, cellular responses and genetic features accounting for the clinical phenotype of SCLS are virtually unknown. Here, we searched for factors altered in acute SCLS plasma relative to matched convalescent samples using multiplexed aptamer‐based proteomic screening. Relative amounts of 612 proteins were changed greater than twofold and 81 proteins were changed at least threefold. Among the most enriched proteins in acute SCLS plasma were neutrophil granule components including bactericidal permeability inducing protein, myeloperoxidase and matrix metalloproteinase 8. Neutrophils isolated from blood of subjects with SCLS or healthy controls responded similarly to routine pro‐inflammatory mediators. However, acute SCLS sera activated neutrophils relative to remission sera. Activated neutrophil supernatants increased permeability of endothelial cells from both controls and SCLS subjects equivalently. Our results suggest systemic neutrophil degranulation during SCLS acute flares, which may contribute to the clinical manifestations of acute vascular leak.     

The effect of different blood sampling sites and analyses on the relationship~ between exercise intensity and 4.0 mmol ·1? blood lactate concentration

The aim of the study was to examine whether the difference in lactate concentration in different blood fractions is of practical importance when using blood lactate as a test variable of aerobic endurance capacity. Ten male firefighters performed submaximally graded exercise on a cycle ergometer for 20-25 min. Venous and capillary blood samples were taken every 5 min for determination of haematocrit and lactate concentrations in plasma, venous and capillary blood. At the same time, expired air was collected in Douglas bags for determination of the oxygen consumption. A lactate concentration of 4.0 mmol.l-1 was used as the reference value to compare the oxygen consumption and exercise intensity when different types of blood specimen and sampling sites were used for lactate analysis. At this concentration the exercise intensity was 17% lower (P less than 0.01) when plasma lactate was compared to venous blood lactate, and 12% lower (P less than 0.05) when capillary blood lactate was used. Similar discrepancies were seen in oxygen consumption. The results illustrated the importance of standardizing sampling and handling of blood specimens for lactate determination to enable direct comparisons to be made among results obtained in different studies.     

A model of blood flow distribution during filtration-reabsorption processes in capillaries

A model of blood flow in a capillary was constructed, which takes into account the movement of plasma through its porous wall. The functions of changes in pressure and the rate of blood flow along the capillary were calculated. It was shown that, in the general case, the distribution of hemodynamic parameters as a result of filtration-reabsorption processes is nonlinear. Possible mechanism of tissue edema resulting from the disturbance of the filtration-reabsorption equilibrium were analyzed.