Max R. Hardeman

New guidelines for hemorheological laboratory techniques

This document, supported by both the International Society for Clinical Hemorheology and the European Society for Clinical Hemorheology and Microcirculation, proposes new guidelines for hemorheolog ...

The clinical importance of erythrocyte deformability, a hemorrheological parameter.

SummaryHemorrheology, the science of the flow behavior of blood, has become increasingly important in clinical situations. The rheology of blood is dependent on its viscosity, which in turn is influenced by plasma viscosity, hematocrit, erythrocyte aggregation, and erythrocyte deformability. In recent years it has become apparent that the shape and elasticity of erythrocytes may be important in explaining the etiology of certain pathological situations. Thus, clinicians have become increasingly interested in hemorrheology in general and erythrocyte deformability in particular. In the course of time, many clinical studies have been performed, but no concise review has thus far been published. This article encompasses a review of the clinically based literature on this subject.

Parameterization of red blood cell elongation index – shear stress curves obtained by ektacytometry

Abstract Measurement of red blood cell (RBC) deformability by ektacytometry yields a set of elongation indexes (EI) measured at various shear stresses (SS) presented as SS-EI curves, or tabulated data. These are useful for detailed analysis, but may not be appropriate when a simple comparison of a global parameter between groups is required. Based on the characteristic shape of SS-EI curves, two approaches have been proposed to calculate the maximal RBC elongation index (EImax) and the shear stress required for one-half of this maximal deformation (SS1/2): (i) linear Lineweaver-Burke (LB) model; (ii) Streekstra-Bronkhorst (SB) model. Both approaches have specific assumptions and thus may be subject to the measurement conditions. Using RBC treated with various concentrations of glutaraldehyde (GA) and data obtained by ektacytometry, the two approaches have been compared for nine different ranges of SS between 0.6–75 Pa. Our results indicate that: (i) the sensitivity of both models can be affected by the SS range and limits employed; (ii) over the entire range of SS-data, a non-linear curve fitting approach to the LB model gave more consistent results than a linear approach; (iii) the LB method is better for detecting SS1/2 differences between RBC treated with 0.001–0.005% glutaraldehyde (GA) and for a 40% mixture of rigid cells but is equally sensitive to SB for 10% rigid cells; and (iv) the LB and SB methods for EImax are equivalent for 0.001% and 0.003% GA and 40% rigid, with the SB better for 0.005% GA and the LB better for 10% rigid.

Small Artery Remodeling and Erythrocyte Deformability in L-NAME-Induced Hypertension: Role of Transglutaminases

Background: Hypertension is associated with inward remodeling of small arteries and decreased erythrocyte deformability, both impairing proper tissue perfusion. We hypothesized that these alterations depend on transglutaminases, cross-linking enzymes present in the vascular wall, monocytes/macrophages and erythrocytes. Methods and Results: Wild-type (WT) mice and tissue-type transglutaminase (tTG) knockout (KO) mice received the nitric oxide inhibitor Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME) to induce hypertension. After 1 week, mesenteric arteries from hypertensive WT mice showed a smaller lumen diameter (–6.9 ± 2.0%, p = 0.024) and a larger wall-to-lumen ratio (11.8 ± 3.5%, p = 0.012) than controls, whereas inward remodeling was absent in hypertensive tTG KO mice. After 3 weeks, the wall-to-lumen ratio was increased in WT (20.8 ± 4.8%, p = 0.005) but less so in tTG KO mice (11.7 ± 4.6%, p = 0.026), and wall stress was normalized in WT but not in tTG KO mice. L-NAME did not influence expression of tTG or an alternative transglutaminase, coagulation factor XIII (FXIII). Suppression of FXIII by macrophage depletion was associated with increased tTG in the presence of L-NAME. L-NAME treatment decreased erythrocyte deformability in the WT mice (–15.3% at 30 dynes/cm2, p = 0.014) but not in the tTG KO mice. Conclusion: Transglutaminases are involved in small artery inward remodeling and erythrocyte stiffening associated with nitric oxide inhibition-related hypertension.

Laser-assisted optical rotational cell analyzer measurements reveal early changes in human RBC deformability induced by photodynamic treatment

BACKGROUND:  The ability to deform is important for circulating RBCs in vivo, and earlier studies showed that this property can objectively be measured in vitro by the LORCA. In this study it was investigated whether photodynamic treatment of human RBCs (meant to inactivate contaminating pathogens) affects deformability.

Test 1 analyser for determination of ESR. 1. Practical evaluation and comparison with the Westergren technique

Abstract Background: various modifcations of the Erythrocyte Sedimentation Rate (ESR) determination have been suggested since the original Westergren procedure that has been adopted as the gold standard by the International Council for Standardization in Haematology (ICSH). Recently, an automated method, (Alifax Test 1), based on a technique completely different from Westergren, has been introduced. Material and methods: In this comparative study, ESR of blood from 680 patients with various rheumatic diseases was determined on both Test 1 and the StaRRsed automated ESR analyser which performs measurements in accordance with ICSH specifcations. Furthermore the robustness of the new technique was evaluated. Results: Direct correlation of Test 1 and StaRRsed measurements confrmed the results of previous studies: an overall correlation coeffcient of R = 0.90. However, further statistical analysis showed that, depending on the instrument that was used, in 78 samples (i.e. 11.5%) the results could lead to different treatment suggestions. Furthermore it appeared that several procedural factors could infuence the fnal Test 1 outcome. Conclusions: Due to its sensitivity for procedural variations, Test 1 measurements should be carried out under strictly standardized conditions. Especially at the higher ESR levels the Test 1 technique is, however, not a reliable alternative for the ICSH approved ‘Westergren’ method.

Photometric measurements of red blood cell aggregation: light transmission versus light reflectance

Red blood cell (RBC) aggregation is the reversible and regular clumping in the presence of certain macromolecules. This is a clinically important phenomenon, being significantly enhanced in the presence of acute phase reactants (e.g., fibrinogen). Both light reflection (LR) and light transmission (LT) from or through thin layers of RBC suspensions during the process of aggregation are accepted to reflect the time course of aggregation. It has been recognized that the time courses of LR and LT might be different from each other. We aim to compare the RBC aggregation measurements based on simultaneous recordings of LR and LT. The results indicate that LR during RBC aggregation is characterized by a faster time course compared to simultaneously recorded LT. This difference in time course of LR and LT is reflected in the calculated parameters reflecting the overall extent and kinetics of RBC aggregation. Additionally, the power of parameters calculated using LR and LT time courses in detecting a given difference in aggregation are significantly different from each other. These differences should be taken into account in selecting the appropriate calculated parameters for analyzing LR or LT time courses for the assessment of RBC aggregation.

Importance of methodological standardization for the ektacytometric measures of red blood cell deformability in sickle cell anemia.

Red blood cell (RBC) deformability is severely decreased in patients with sickle cell anemia (SCA), which plays a role in the pathophysiology of the disease. However, investigation of RBC deformability from SCA patients demands careful methodological considerations. We assessed RBC deformability by ektacytometry (LORRCA MaxSis, Mechatronics, The Netherlands) in 6 healthy individuals and 49 SCA patients and tested the effects of different heights of the RBC diffraction patterns, obtained by altering the camera gain of the LORRCA, on the result of RBC deformability measurements, expressed as Elongation Index (EI). Results indicate that the pattern of RBCs from control subjects adopts an elliptical shape under shear stress, whereas the pattern of RBCs from individuals with SCA adopts a diamond shape arising from the superposition of elliptical and circular patterns. The latter represent rigid RBCs. While the EI measures did not change with the variations of the RBC diffraction pattern heights in the control subjects, we observed a decrease of EI when the RBC diffraction pattern height is increased in the SCA group. The differences in SCA EI values measured at 5 Pa between the different diffraction pattern heights correlated with the percent of hemoglobin S and the percent of sickled RBC observed by microscopy. Our study confirms that the camera gain or aperture of the ektacytometer should be used to standardize the size of the RBC diffraction pattern height when measuring RBC deformability in sickle cell patients and underscores the potential clinical utility of this technique.

Test 1 analyser for determination of ESR. 2. Experimental evaluation and comparison with RBC aggregometry.

Abstract Background: Test 1 is a recently introduced technique claiming to determine Erythrocyte Sedimentation Rate (ESR) in 20 s. In contrast to the original Westergren procedure this new technique uses undiluted blood and operates at 37°C. It is hypothesized that Test 1 is in fact an erythrocyte aggregometer and does not measure any sedimentation. Methods: Test 1 results were compared to those obtained with StaRRsed, an automated ESR analyser based on the Westergren technique, and the results of both were correlated to various indices of red blood cell (RBC) aggregation, obtained with an aggrego - meter (LORCA). Measurements were made on blood from 75 patients with various rheumatic disorders. Furthermore, blood that was experimentally manipulated in order to affect RBC aggregation, i.e. by changing the hematocrit, by diminishing plasma protein concentration, by inducing hyperaggregation or by RBC rigidification, was tested on all three instruments. Results: Generally in patient blood, Test 1 results demonstrated a higher correlation with the various aggregation parameters than StaRRsed. Highest correlation (R = −0.8)) with both Test 1 and StaRRsed outcome were seen with I20, a RBC aggregation parameter directly related to the backscatter intensity. All experimentally induced changes in RBC aggregation paralleled closely those obtained with Test 1 while StaRRsed results followed a different course. Conclusions: The results obtained in this study strongly support the hypothesis that Test 1 measures only the RBC aggregation process and does not cover any of the indices directly linked to the sedimentation process as determined by the Westergren method.

The effects of propofol compared to high-dose fentanyl anesthesia on rheologic parameters in coronary artery surgery

Propofol has previously been found to decrease hematocrit values. Because hematocrit is an important determinant of blood viscosity, lower hematocrits may cause a decrease in blood viscosity, improving blood flow and oxygen delivery. This phenomenon may be beneficial in certain intraoperative situations. To study the influence of two anesthetic techniques on a variety of rheologic parameters, 32 patients scheduled for coronary artery bypass grafting (CABG) were divided into two groups. Group I (n = 18) was induced with high-dose fentanyl anesthesia (100 micrograms/kg), and group II (n = 16) with a combination of propofol and fentanyl anesthesia (1 to 1.5 mg/kg and 35 to 50 micrograms/kg, respectively). Maintenance anesthesia continued with infusions of the same drugs. Blood and plasma viscosity, hematocrit, erythrocyte aggregation factor, and erythrocyte deformability were measured preoperatively, intraoperatively, and up to 48 hours postoperatively. Whole blood viscosity was corrected to a standard hematocrit of 0.45. The two groups were comparable with respect to age, bypass duration, blood loss, urine output, transfusions, and fluid management. Erythrocyte deformability did not decrease during or after cardiopulmonary bypass (CPB). In both groups, hematocrit and blood and plasma were decreased significantly during and after CPB (P < 0.01) and returned to baseline levels 48 hours after surgery. After induction and before CPB, blood viscosity was only decreased in group II. However, the corrected blood viscosity was significantly elevated at all shear rates in group II compared to group I at 24 and 48 hours postoperatively (P < 0.01). In group II at these sampling times, this parameter was also significantly elevated compared to preoperative values.(ABSTRACT TRUNCATED AT 250 WORDS)