Laser-assisted Optical Rotational Cell Analyzer Research Articles

Acute and long-term effects of hyperbaric oxygen therapy on hemorheological parameters in patients with various disorders.

Inhalation of 100% oxygen in a hyperbaric chamber has been accepted as a useful treatment for patients with various pathologies who suffer from hypoxia. The oxidative effects of hyperbaric oxygen (HBO) on RBCs have been investigated in animals but there is not enough data on hemorheological parameters in patients following HBO treatment (HBOT).In this study, we investigated the effect of HBO on hemorheological and haematological parameters during treatment. Red blood cell (RBC) deformability and aggregation, blood and plasma viscosity and superoxide dismutase activity were investigated in patients who underwent HBOT. Hematological parameters were determined by an electronic hematology analyzer. A Laser-assisted Optical Rotational Cell Analyzer (LORCA) was used to measure RBC deformability. RBC aggregation was measured for cells in autologous plasma and for cells resuspended in PBS containing Dextran70 (3% ) by using a Myrenne Aggregometer. A Wells-Brookfield cone/plate rotational viscometer was used for viscosity measurements. According to our results, a significant decrement of the hematocrit and the RBC count was observed after the 20th session of HBOT compared to the baseline, but none of the hemorheological parameters changed significantly. Our results showed that HBOT did not cause any significant changes in hemorheological parameters, thereby not representing any problems for the patients.

Effects of Wearing Compression Stockings on the Physical Performance of T2DM Men with MetS.

Metabolic syndrome (MetS) and type 2 diabetes mellitus (T2DM) are associated with macro- and microcirculatory complications that reduce physical performance. Wearing compression garments to potentially optimize hemodynamics has been discussed. This study investigates the effects of wearing compression stockings on physical performance-related variables in type 2 diabetic men with metabolic syndrome (n=9, 57±12 years, BMI: 36±4 kg/m(2)). Participants served as their own controls in a randomized 3*3 crossover study wearing below-knee stockings with either compression (24 or 30 mmHg ankle pressure) or no compression. Venous pooling and lower limb oxygenation profiles were determined with near-infrared spectroscopy and arterial oxygen saturation was determined using a pulse oxymeter. Measurements were performed in the supine lying position, during standing, following 10 tiptoe exercises and after submaximal intensity cycling. In addition, lactate and erythrocyte deformability were analyzed in capillary blood pre- and post-exercise. Erythrocyte deformability was analyzed using a laser-assisted optical rotational red cell analyzer. No significant differences in any variables when wearing different compression or regular stockings were evident at any point of measurement. This study did not reveal any beneficial effects of wearing compression stockings at rest and during acute bouts of moderately intense exercise in this particular patient group.

Diagnostic Power of Laser Assisted Optical Rotational Cell Analyzer (LoRRca MaxSis) Evaluated in 118 Patients Affected By Hereditary Hemolytic Anemias

▪Introduction: Hereditary hemolytic anemias (HAs) is a group of heterogeneous disorders mainly due to defects of red cell (RBC) membrane and metabolism. HAs of RBC membrane are classified according to the morphological appearance on a blood smear: the more common are hereditary spherocytosis (HS), elliptocytosis, followed by stomatocytosis (dehydrated, DHSt and overhydrated, OHSt). Among RBC enzyme defects associated with chronic hemolysis, pyruvate kinase (PK) deficiency is the most common, followed by glucose-6-phosphate isomerase (GPI), pyrimidin 5' nucleotidase (P5'N), and more rarely, other enzymes deficiencies. Because of the rarity and heterogeneity of these diseases, diagnosis may be often challenging despite the availability of a battery of laboratory tests. Ektacytometry, and more recently, laser-assisted optical rotational cell analyser (LoRRca MaxSis, Mechatronics Instruments, NL), able to measure RBC deformability in osmotic gradient conditions (Osmoscan), is reported to be a useful tool in the detection of RBC membrane disorders and in particular for the differential diagnosis of HSt. Few data are available in other haemolytic anemias.Methods: We evaluated the diagnostic power of laser-assisted optical rotational cell analyser in 84 cases with RBC membrane disorders (69 HS, 7 HE, 7 DHSt, 1 OHSt) and 26 enzymopathies: 19 PK deficiency (14 not splenectomised), 4 GPI deficiency and 3 P5N deficiency. Moreover, we analysed 8 congenital diserythropoietic anemia type II (CDA II), of whom 3 not splenectomised. A reference curve interval was obtained from 89 healthy controls. The evaluated parameters were: Omin (osmotic value at which the EI reaches its minimum, coinciding with the 50% hemolysis in osmotic fragility assays), EImax (maximal deformability obtained near the isotonic osmolality), and Ohyper (osmotic value at which the EI reaches half of its maximum value, representing cellular hydration status).Results: Considering membrane defects, all the 69 HS, regardless the biochemical defect, showed typical altered ektacytometry curves, with a decreased EImax (0.516±0.05 vs 0.595±0.008 for controls, p<0.0001) and right shifted Omin (167±11.4 vs 143.7±7.2, p<0.0001); the 7 HE cases displayed a trapezoidal feature and a decreased EImax (0.547±0.05 vs 0.595±0.008, p<0.001). Among the hereditary stomatocytosis, the 7 DHSt were characterized by a left-shifted curve (Fig 1a).Concerning RBC enzymopathies, we found that all the GPI deficient cases showed altered enlarged Osmoscan curves associated with significant increased Ohyper (539±36 vs 462±13.4, p<0.0001), whereas not splenectomised patients with PK deficiency showed an Osmoscan curve characterised by an increased EImax (0.601±0.01 vs 0.595±0.008), although not significant (Fig 1b). Osmoscan curve in not splenectomised CDAII falls in the low reference area of normal controls. However, Omin and EImax were signifificantly different from HS patients thus permitting a differential diagnosis (Omin 149.3 ±3.0 vs 167±11.4, p<0.01; EImax 0.578±0.02 vs 0.516±0.05, p<0.05) (Fig 1c).We separately analysed splenectomised and not splenectomised patients. Interestingly, for some diseases (PK, P5N, CDAII and OHSt) Osmoscan curve after splenectomy, falls in a defined atypical area, regardless the pathology (Fig 1d).Conclusion: The Osmoscan curves were diagnostic for all the HS and HE cases analysed, with Omin and EImax parameters significantly different from controls. DHSt cases always showed typical left-shifted curve. Interestingly, Ohyper was firstly described increased in GPI deficiency, offering a new diagnostic tool for this rare enzyme defect. The interpretation of Osmoscan requires caution in splenectomised cases as splenectomy may interfere with the RBC deformability. In conclusion, the Osmoscan analysis performed by LoRRca Maxsis represents a useful and feasible first step screening test in the diagnosis of RBC membrane disorders and other rare haemolytic anemias. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Investigation of hemorheological parameters in periodontal diseases.

Periodontal diseases are frequently associated with cardiovascular diseases (CVD). On the other hand, occurrence of CVD has also been related with increased blood viscosity. This study was planned to investigate four main hemorheological parameters contributing to blood viscosity - hematocrit, erythrocyte deformability, erythrocyte aggregation and plasma viscosity - and also some biochemical parameters (hs-CRP, fibrinogen, globulin etc.) in patients with periodontal disease. We hypothesized that poor periodontal health would be associated with deterioration of hemorheological properties. According to periodontal health status, subjects were divided into three groups as control (healthy), with plaque induced gingivitis and with chronic periodontitis. All groups included 15 males who had not received periodontal therapy in the last six months before the study, were non-smokers, had no systemic diseases and were not on any medication. Erythrocyte deformability and erythrocyte aggregation were measured with laser-assisted optical rotational cell analyzer (LORCA). Plasma viscosity was measured by a cone-plate viscometer. Data were analyzed with Kruskal-Wallis, Mann-Whitney U Test and Spearman Correlation Coefficient. Plasma viscosity (1.36 ± 0.01 mPa.s in the control group and 1.43 ± 0.02 mPa.s in the chronic periodontitis group, P < 0.01), erythrocyte aggregation tendency (aggregation index, amplitude and t½ were 58.82 ± 1.78% , 20.22 ± 0.40 au, 2.80 ± 0.25 s respectively in the control group, and 67.05 ± 1.47% , 22.19 ± 0.50 au, 1.84 ± 0.15 s in the chronic periodontitis group, P < 0.01), hs-CRP, fibrinogen and globulin levels were significantly higher, whereas HDL level was significantly lower in the chronic periodontitis group (P < 0.05) compared to the control group. All of these conditions may contribute to cardiovascular morbidity and mortality observed in people with periodontal disease, via increasing blood viscosity.

Reduced red blood cell deformability over time is associated with a poor outcome in septic patients

BackgroundTo investigate changes in red blood cell (RBC) rheology over time in critically ill patients with sepsis and their relationship with outcome. MethodsIn this prospective, non-interventional study, RBC rheology was assessed using the Laser-assisted Optical Rotational Cell Analyzer in a convenience sample of intensive care unit (ICU) patients with (n=64) and without (n=160) sepsis. Results were compared to measures in healthy volunteers (n=20). RBC rheology was also assessed on days 1 and 3 of the ICU stay in 32 of the non-septic and 19 of the septic patients. RBC deformability was determined by the elongation index (EI) in relation to the shear stress (0.3 to 50Pa) applied to the RBC membrane. An aggregation index (AI) was assessed simultaneously with the same device. ResultsThe ICU mortality rate of the septic patients was 31%. RBC deformability was already reduced in septic patients at ICU admission, an effect that persisted during the study period and worsened in the non-survivors for the large majority of shear stresses studied (e.g., EI for 50Pa of shear stress was 0.527±0.064 in non-survivors vs. 0.566±0.034 in survivors, p<0.05). These changes were not observed in non-septic patients. The AI was more elevated in septic than in non-septic patients at ICU admission, but had no prognostic value. ConclusionsAlterations in RBC rheology, including reduced deformability and increased aggregation, occur early in septic patients and reductions in RBC deformability over time are associated with a poor outcome.

In vitro hemorheological effects of parenteral agents used in peripheral arterial disease

Peripheral arterial disease (PAD) is a frequent manifestation of systemic atherosclerosis. In PAD hemorheological parameters were defined as risk factors in a number of studies and several therapeutic agents were tried in these conditions. Our study aims to investigate and compare the in vitro hemorheological effects of various drugs generally used in the parenteral treatment of intermittent claudication and critical limb ischemia. Blood samples of healthy male volunteers were incubated with iloprost, alprostadil, pentoxifylline, sulodexide or pentosan polysulfate at calculated therapeutic serum concentration. Hematocrit (Hct) was determined by microhematocrit centrifuge. Plasma and apparent whole blood viscosities (WBV) were evaluated by capillary viscometer. Red blood cell aggregation was measured by LORCA (laserassisted optical rotational cell analyzer) aggregometer, and LORCA ektacytometer was used for measuring erythrocyte deformability at 37°C. Iloprost, alprostadil, and pentoxifylline incubation did not have any significant effect on plasma and apparent WBV. Elongation index increased in samples incubated with alprostadil at low shear stresses 0.95 and 0.53 Pa (p < 0.05). Sulodexide significantly improved WBV and Hct/WBV ratio (p < 0.05). Incubation with pentosan polysulfate resulted in higher WBV, lower Hct/WBV ratio and deterioration in the aggregation parameters (p < 0.05). Sulodexide may have beneficial effect on a macrorheological parameter; alprostadil may improve a microrheological parameter. Hemorheological alterations could be important in PAD patients with hampered vasodilator capacity.

Impact of sleeve gastrectomy on red blood cell aggregation: a 12-month follow-up study

To investigate the effects of weight loss due to laparoscopic sleeve gastrectomy (LSG) on erythrocyte aggregation and the relationship of anthropometric and plasmatic factors, such as plasma viscosity, fibrinogen and lipids, with erythrocyte aggregation. The RBC aggregation and kinetics of the red blood cell aggregation were performed by the Laser-assisted Optical Rotational Cell Analyser (LORCA). Before the LSG and 6 and 12 months after the LSG, we evaluated the aggregation index (AI), amplitude (AMP) and aggregation half-time (t1/2), plasma viscosity, fibrinogen, glucose and lipids patterns in 15 non-diabetic obese subjects. The static and kinetic parameters of aggregation in obese patients at each time point after bariatric weight loss surgery were calculated and significant differences were observed at 12 months after surgery. AI and AMP decreased from 69.81±5.12% and 27.43±2.9 a.u. at baseline to 64.91±5.94% and 22.15±4.3 a.u. 12 months after surgery, respectively. The t1/2 increased from 1.7 (1.32-2.24) s at baseline compared with 2.02 (1.68-2.42) s at 12 months after the surgery. Plasma viscosity and fibrinogen decreased from 1.50±0.093 mPa s and 3.0±0.41 g l(-1) at baseline to 1.407±0.062 mPa s and to 2.66±0.25 g l(-1) 12 months after surgery, respectively. AI correlated positively with BMI (r=0.74, P=0.001), waist circumference (r=0.68, P=0.005), fibrinogen (r=0.52, P=0.045) and plasma viscosity (r=0.76, P=0.001) and negatively with percentages of weight lost after surgery (r=-0.54, P=0.034). Multivariate analyses found that the BMI, fibrinogen and plasma viscosity independently influenced the AI. The study demonstrated that weight loss due to restrictive bariatric surgery can beneficially affect red cell aggregation parameters. The improvement of the RBC aggregation behaviours among obese subjects with weight loss due to LSG was associated with changes in plasmatic factors, especially fibrinogen.

The impacts of super obesity versus morbid obesity on red blood cell aggregation and deformability among patients qualified for bariatric surgery

The aim of this study was to evaluate the effects of the obesity degree on red blood cell aggregation and deformability. We studied 56 obese patients before weight loss surgery who were divided into two groups: morbid obesity and super obesity. The aggregation and deformability of RBCs were evaluated using a Laser-assisted Optical Rotational Cell Analyzer (Mechatronics, the Netherlands). The following parameters specific to the aggregation process were estimated: aggregation index (AI), aggregation half-time (t1/2) and threshold shear rate (γthr). RBC deformability was expressed as erythrocyte elongation (EI), which was measured at 18.49 Pa and 30.2 Pa shear stresses. Super obese patients presented significantly higher AI (P < 0.05) and γthr (P < 0.05) and significantly lower t1/2 (P < 0.05) compared with morbidly obese individuals. Multivariate analyses showed that fibrinogen (β 0.46, P < 0.01 and β 0.98, P < 0.01) and hematocrit (β 0.38, P < 0.05 and β 1.01, P < 0.01) independently predicted the AI in morbidly obese and super obese patients. Fibrinogen (β -0.4, P < 0.05 and β -0.91, P < 0.05) and hematocrit (β -0.38, P < 0.05 and β -1.11, P < 0.01) were also independent predictors of the t1/2 in both obese groups. The triglyceride level (β 0.32, P < 0.05) was an independent predictor of the t1/2 in the morbidly obese group. No differences in EI were observed between obese subjects. Multivariate analyses showed that the triglyceride level independently predicted EI at 18.49 Pa (β -0.42, P < 0.05 and β -0.53, P < 0.05) and 30.2 Pa (β -0.44, P < 0.01 and β -0.49, P < 0.05) in both obese groups. This study indicated that the obesity degree of patients who qualify for bariatric surgery affects RBC aggregation properties, but it does not indicate the reasons for this difference. Further studies are needed to determine factors associated with hyperaggregation in super obesity.

Association between hemorheological alteration and clinical diagnosis of metabolic syndrome among patients qualified for bariatric surgery

This study presents the association between metabolic syndrome and hemorheological parameters among obese patients qualified for bariatric surgery. We studied 73 morbidly obese patients who were qualified for bariatric surgery. Blood and plasma viscosity measurements were performed using a cone-plate viscometer (Brookfield DV-II). RBC aggregation was measured using the Laser-assisted Optical Rotational Cell Analyser (LORCA). The following parameters for the aggregation process were estimated: aggregation index (AI), amplitude (AMP), aggregation half-time (t1/2), threshold shear rate (γthr), the fast (Tfast) and the slow (Tslow) component. Metabolic syndrome was defined according to the Adult Treatment Panel III and International Diabetes Federation criteria. The obese presented differences in all rheological properties compared to control, regardless of clinical diagnosis of metabolic syndrome, except that whole blood viscosity was higher only in the obese metabolic syndrome group. No differences among the obese with and without metabolic syndrome were observed except in hematocrit. Whole blood viscosity and corrected blood viscosity correlated positively with WHR and BMI. AI, γthr, Tslow correlated positively with BMI, WHR, total cholesterol level and low-density lipoproteins. Negative correlation presented t1/2 with BMI, WHR, fibrinogen, total cholesterol level and low-density lipoproteins. The study showed that morbid obesity is associated with hemorheological disturbances independently of clinical diagnosis of metabolic syndrome.

Influence of endurance training and marathon running on red cell deformability in HIV patients

This investigation assessed whether endurance training altered the red cell deformability (RCD) of male HIV patients. Eight subjects (38 ± 9 years, BMI 23 ± 3.6 kg/m², highly active antiretroviral therapy) trained for a marathon run for the duration of one year. Each subject underwent an acute exercise test on a treadmill at the beginning of this study (baseline) and after four months of training. Blood samples were taken at rest (T0) and at the end (T1) of each exercise test, as well as before (T0) and after the marathon run (T1). RCD was measured using the Laser Assisted Optical Rotational Cell Analyzer (LORCA), the maximal elongation index (EImax) and shear stress at half maximal deformation (SS ½ were determined using the Lineweaver Burke model. No changes were observed in RCD and SS ½ between T0 and T1 in all measurements. EImax significantly increased between T0 baseline and T0 after four months of training (P < 0.05). Measurements prior to the marathon run revealed significantly decreased EImax and SS ½ values (P < 0.05). The intensity of training in pathophysiological diseases like HIV is dependent for the alterations in RCD and possible improvements in microcirculation.

Red blood cell deformability and aggregation in chronic venous disease patients with varicose veins

Red blood cells' (RBC) rheological properties are disturbed in chronic venous disease (CVD). The aim of the study was to compare deformability and aggregation of erythrocytes taken from the varicose vein and the antecubital vein of patients with chronic venous disease. Blood samples were taken from twelve CVD patients presenting clinical, aetiological, anatomical and pathological elements (CEAP) stages II and III. Blood was sampled from varicose veins and antecubital veins of patients (as control). Deformability and aggregation of RBC were analysed with a Laser-assisted Optical Rotational Cell Analyser (LORCA). A significant increase in deformability was found in varicose vein RBC for shear stress values 4.24, 8.23 and 15.96 Pa as compared to RBC from the antecubital vein. The aggregation index was significantly lower and aggregation halftime was significantly increased for RBC taken from antecubital veins than for RBC from varicose veins. In conclusion, RBC taken from varicose and antecubital veins of CVD patients are not entirely rheologically comparable and show different deformability and aggregation. Varicose vein RBC are more deformable and show a higher tendency for aggregation than antecubital vein RBC. Perhaps the deformability of varicose vein RBC has been increased as a compensation mechanism in subjects with CVD, due to increased resistance in their microcirculation.

P70: Molecular and physiological aspects connecting red cell eNOS, red blood cell deformability and decreased microvascular function in patients with essential hypertension

Background Arterial hypertension (HT) is a complex multifactorial condition associated with cardiovascular disease. Independent studies show that HT is associated with microcirculatory and hemorheological alterations, such as decreased red blood cell (RBC) deformability, as well as decreased endothelial function and nitric oxide (NO) bioavailability. Recently we found RBC carry an active endothelial nitric oxide synthase (eNOS), which is decreased in cardiovascular disease.Hypothesis: We hypothesized that a reduced RBC deformability may contribute to induce a hypertensive phenotype via impairment of microcirculatory conductance. Methods To evaluate hemorehological alterations in HT, patients ( n = 10) with HT and age-and risk factors-matched subjects ( n = 10) without HT (control) were recruited. High BP was defined as systolic BP > 135 mm Hg and diastolic BP > 85 mm in 24 h BP measurements. RBC deformability was measured by laser-assisted optical rotational cell analyzer (LORCA) and assessed as elongation index (EI) at 1.73 Pa. RBC redox state (ROS, GSH) was assessed by labeling with diclorodihydrofluorescein and thiol-tracker and flow cytometry. Cutaneous microvascular function as post-occlusion reactive hyperemia was measured by noninvasive laser Doppler Perfusion imaging and quantified as area under the curve (AUC). Endothelial function of the brachial artery was assessed as flow-mediated dilatation (FMD) by high resolution ultrasound. Deformability of RBC from eNOS −/− mice was evaluated and compared to wild type (WT) C57Bl/6 mice. To evaluate NO and NOS-dependent changes in RBC deformability, RBC from healthy individuals ( n = 6) were treated with NO donors, NOS inhibitors and guanylate cyclase inhibitors. Measurements of cGMP production in RBC were assessed by ELISA. A stepwise linear regression as well as a multivariate linear regression model were performed to identify independent predictors for systolic BP. Results In the HT group EI (0.300 ± 0.007 vs. 0.326 ± 0.007, p = 0.013), AUC (1657 ± 422 vs. 2313 ± 473 PU; p = 0.009). and FMD (6.2% ± 0.7% vs. 5.0% ± 0.3 % p = 0.010) was reduced in the HT group, suggesting decreased deformability, shorter lasting vasodilation of microvessels and endothelial dysfunction. In RBC from HT patients the ROS levels increased and the GSH levels decreased, indicating increased oxidative stress. Univariate inverse correlations were found between systolic BP and AUC ( r = −0.485; p r = 0.726; p R 2 = 0.607, p = 0.004). In a translational approach we analyzed deformability in eNOS −/− mice, characterized by a dysfunctional endothelium and hypertension. eNOS −/− RBC are less deformable than WT RBC, show increased ROS and decreased GSH, similarly to the HT grouP Moreover, NO donors increased EI of RBC from healthy individuals, while NOS inhibition as well as guanlyte cyclase inhibition decreased cGMP levels and EI indicating that NO/NOS/cGMP dependent pathways control deformability in RBC. Conclusion The NO/NOS/cGMP pathway controls RBC mechanoproperties, and RBC deformability is an independent predictor of increased blood pressure in patients with arterial hypertension. Disclosure Nothing to disclose.

In vitro effect of pentoxifylline and lisofylline on deformability and aggregation of red blood cells from healthy subjects and patients with chronic venous disease.

Purpose. The aim of the study was to assess the in vitro potency of pentoxifylline (PTX) and one of its most active metabolites lisofylline (LSF) to improve rheological properties of red blood cells (RBC) from healthy individuals and patients with chronic venous disease (CVD). Additionally, the study aimed to compare the effects of PTX and LSF on RBC deformability and aggregation. Methods. Blood samples were collected from healthy volunteers (antecubital vein) and from CVD patients (varicose and antecubital vein). Deformability and aggregation of RBC were assessed using Laser-assisted Optical Rotational Cell Analyser (LORCA). Results. PTX and LSF increased RBC elongation significantly. Additionally, RBC incubation with PTX resulted in a marked decrease in RBC aggregation. PTX reduced the tendency towards the formation of RBC aggregates and of their stability. The beneficial effect of PTX on RBC aggregation was most apparent for those cells whose aggregation tendency and aggregate stability was the greatest. Conclusions. In vitro addition of PTX or LSF effectively increased deformability of RBC from healthy donors and patients with CVD. Thus, LSF may contribute to the in vivo hemorheological effects of pentoxifylline. On the other hand, there was no significant effect of LSF on aggregation of RBC in vitro. Hence, LSF has no contribution to this particular effect of PTX. Additionally, the present study demonstrated the use of RBC with impaired deformability and aggregation for the evaluation of in vitro rheological activity of xenobiotics.

Influence of a moderate physical activity intervention on red cell deformability in patients suffering from chronic obstructive pulmonary disease (COPD)

The present study investigates whether a moderate physical activity intervention may alter red cell deformability (RCD) of patients suffering from chronic obstructive pulmonary disease (COPD). Subjects (n = 10; age: 62 ± 4; body-mass index (BMI): 25.8 ± 7.5) performed a training regimen for 10 weeks. In the beginning of the study and after the training period, COPD patients underwent a WHO cycle ergometry test. Venous blood samples were taken before (T0), immediately after (T1) and 30 min after (T2) the intervention. RCD was measured with the laser-assisted optical rotational cell analyzer (LORCA). Significant improvements of the RCD were detected. The semi-maximal shear stress increased significantly. Acute exhaustion had no effect on RCD. Thus, the training period of 10 weeks influenced RCD.

Shear stress and flow dynamics of the femoral vein among obese patients who qualify for bariatric surgery

The aim of this study was to evaluate the effects of obesity on wall shear stress and its relationship to erythrocyte aggregation. We studied 35 morbidly obese patients who were qualified for bariatric surgery. The control group consisted of 20 non-obese people. Blood rheological measurements were performed using the Laser-assisted Optical Rotational Cell Analyzer (Mechatronics, the Netherlands) and a cone-plate viscometer (Brookfield DV-II). The venous flow dynamics were assessed using a duplex ultrasound. The shear rate was estimated from the measured blood flow velocity and the diameter of the femoral vein. Venous wall shear stress was calculated from the whole blood viscosity and the shear rate. The shear rate (P < 0.005) and the venous wall shear stress (P < 0.05) were significantly lower in obese patients compared with the controls. The aggregation index (P < 0.001), syllectogram amplitude - AMP (P < 0.05) and Tslow (P < 0.001) were significantly higher in the obese patients; the aggregation half-time (P < 0.001) and Tfast (P < 0.001) were decreased compared with the control group. Multivariate regression analyses found waist circumference (β -0.31, P < 0.05), thigh circumference (β 0.33, P < 0.05) and Tslow (β -0.47, P < 0.005) to be variables that independently influenced the shear rate. Nevertheless, the AMP (β 0.34, P < 0.05) and Tslow (β -0.47, P < 0.01) were independent predictors that influenced the wall shear stress. This study indicates that there is a relationship between wall shear stress in the femoral vein and the rheological impairment of the RBC among obese patients, but further studies are necessary to confirm this suggestion.

Relationship between hemorheology and endothelial dysfunction in renal transplant patients receiving calcineurin inhibitors

Calcineurin inhibitors, mainly cyclosporin A (CsA), are associated with endothelial dysfunction in renal transplant recipients (RTRs). Hemorheological disturbances including decreased erythrocyte deformability (ED), increased plasma viscosity and erythrocyte aggregation (EA) have also been reported in CsA-treated RTRs. The aim of this study was to investigate the relationship between hemorheological factors and endothelial dysfunction in CsA- and tacrolimus (Tc)-treated RTRs. Thirty-one RTRs and 16 healthy subjects were recruited. The RTR group received either CsA (n = 16) or Tc (n = 15). Endothelial function was evaluated by flow-mediated dilation (FMD) of the brachial artery. ED and EA were measured with laser-assisted optical rotational cell analyzer, and plasma viscosity by a cone-plate viscometer. FMD of the CsA group was significantly lower than that of controls (6.3% ± 5.1% vs. 11.9% ± 5.6%, p = 0.024), whereas, there was no significant difference between the Tc group (8.8% ± 5.4%) and controls. At shear stresses ranging between 0.95 and 30 Pa, EDs of the CsA group were significantly lower compared with controls. In the Tc group, the decrease in ED was significant at shear stresses ranging between 0.53 and 5.33 Pa. ED indices did not correlate with FMD in any of the groups. The degree of endothelial dysfunction and reduction in ED were more remarkable in patients on CsA therapy. Hemorheological factors were not likely to be associated with endothelial dysfunction in RTRs.

Reversing a red blood cell storage (RBCS) lesion: S-Nitrosothiol replenishment in aged RBCS

Transfusion of RBCs has been linked to disappointing patient outcomes. Specifically, a correlation between transfused RBC storage duration and increased incidence of mortality and morbidity has been observed in some clinical settings. Chemical changes (e.g., decreases in pH and 2,3-diphosphoglycerate levels) are well documented to produce “storage lesions” in banked RBCs. Recent work has revealed novel defects that progress over the typical 42-day storage of banked RBCs. Of note, loss of the S-nitrosothiol (SNO) moieties bound to the β93 cysteine of hemoglobin and to other, membrane-resident protein occurs rapidly (hours) after collection. This SNO loss and the attendant impairment in the nitric oxide (NO)-derived vasoactivity of RBCs may contribute to problems associated with transfusions of aged RBCs. We report on strategies to replenish the stored-RBC SNO/NO content via exposure to NO donors (e.g., N-diazeniumdiolates). We examine the RBC SNO/NO yield and distribution, measured via photolysis-chemiluminescence, as a function of incubation conditions (e.g., aerobic or anaerobic, NO-treatment duration) and of NO donor-specific NO release kinetics. For instance, bolus release of NO from the N-diazeniumdiolate PROLI/NO during longer anaerobic incubation favored heme-bound NO rather than SNO. Furthermore, we report on the efficacy of these treatments in improving RBC function with focus on enhanced deformability and decreased RBC adhesion to endothelium as assayed via laser-assisted optical rotational cell analyzer (LORCA) and in vitro flow chamber assays, respectively. Benefits associated with replenished SNO/NO content have the potential to impact clinical outcomes for a wide population of transfused patients.

Rheological blood behaviour is not related to gender in morbidly obese subjects

To the Editor. We have read with interest the recently published article by Wiewiora et al. [7] regarding hemorheological differences related to gender in morbidly obese subjects. In this study authors found several hemorheological differences between male and female. In this sense, males showed higher blood viscosity and higher erythrocyte deformability than females and native blood viscosity correlated with several erythrocyte aggregation parameters evaluated with the Laser-Assisted Optical Rotational Cell Analyser (LORCA) [3]. In studies with small sample sizes (18 men and 20 women) there is always the risk that differences in certain variables may appear by chance or by imbalances in the groups. Moreover, it was not discussed why the erythrocyte deformability should be higher in males. Therefore we aimed to evaluate the rheological profile in a larger sample of morbidly obese subjects selected for bariatric surgery, to ascertain whether there are rheological differences in morbidly obese related to gender. Before statistical testing, all continuous variables were checked for normal distribution. Those who were markedly skewed, were log transformed to improve normality. Analytical tests were performed according to previous studies of our group [4, 5] and in accordance with the ethical guidelines for Clinical Hemorheology and Microcirculation [1]. Results are expressed in Tables 1 and 2 as mean ± sd. P values <0.05 were considered as statistically significant. All analyses were performed using the Statistical Package of Social Sciences (SPSS, version 11.0) for Windows. The results of the present study revealed no differences in erythrocyte deformability, at any of the shear stresses tested when males and females were compared, in line with the homogeneity of erythrocyte indices in both groups. In this sense, Wiewiora’s study did not include any information on erythrocyte indices [7], although these indices are of paramount importance when evaluating erythrocyte deformability [2, 6].

Deformability of Red Blood Cells and Correlation with ATP Content during Storage as Leukocyte-Depleted Whole Blood

Background: Storage duration of red cells has been associated with increased morbidity and mortality following transfusion. This association has been attributed to the loss of deformability of stored red cells leading to deterioration of microvascular perfusion. ATP content is considered a critical determinant of the deformability of stored red cells. Methods: ATP content and deformability were determined after storage for up to 49 days in 40 leukocyte-depleted whole blood units. Red cell deformability was determined using a laser-assisted optical rotational cell analyzer (LORCA®) employing shear stress (SS) ranging from 0.3 to 30 Pa. Deformability was expressed as the elongation index (EI). EI was correlated with ATP content. Results: ATP content decreased from 3.5 to 1.7 mmol/g hemoglobin. EI increased from 0.03 to 0.05 at an SS of 0.3 Pa, and decreased from 0.62 to 0.59 at an SS of 30 Pa. Correlation coefficient (r) of ATP vs. EI at 0.3 Pa ranged from –0.17 to +0.15 during storage. At 30 Pa, r ranged from –0.03 to +0.45. Correlation increased with storage irrespective of SS, and increased with SS irrespective of storage. Conclusions: ATP content is not a valid surrogate marker for red cell deformability and may not reflect in vivo survival of stored red cells.

Impaired Red Blood Cell Deformability in Patients with Coronary Artery Disease and Diabetes Mellitus

Patients with diabetes mellitus (DM) have an increased cardiovascular morbidity and mortality. There is increasing evidence that diabetes mellitus is associated with pathological hemorheological alterations, which might contribute to impaired coronary blood flow in coronary artery disease (CAD). We hypothesize that red blood cell (RBC) deformability is impaired in diabetic patients with CAD in comparison to nondiabetic patients with CAD. RBC deformability was measured in 21 patients with CAD and type 2 diabetes mellitus (CAD + DM) and 24 patients with CAD (CAD - DM). RBC deformability was measured by the Laser-assisted optical rotational cell analyzer by determining the elongation index (EI). RBC deformability was reduced in patients with CAD + DM in comparison to patients with CAD - DM (EI @ 1.12 Pa 0.236 ± 0.008 vs. 0.260 ± 0.005, p=0.007). Inverse univariate correlations were found between the EI @ 1.12 Pa and plasma glucose concentration (r= - 0.57; p<0.001) and HbA1c (r= - 0.45; p=0.002). Multivariate linear regression analysis identified plasma glucose concentration as the independent predictor of RBC deformability (β= - 0.58; p=0.007) thereby indicating that increased glucose concentrations determine RBC deformability in diabetic patients with CAD. In patients with CAD, diabetes mellitus leads to an impairment of RBC deformability which might contribute to increased morbidity of diabetic patients with CAD.