Reduced Erythrocyte Deformability Research Articles

Altered phosphorylation of cytoskeleton proteins in sickle red blood cells: The role of protein kinase C, Rac GTPases, and reactive oxygen species

The small Rho GTPases Rac1 and Rac2 regulate actin structures and mediate reactive oxygen species (ROS) production via NADPH oxidase in a variety of cells. We have demonstrated that deficiency of Rac1 and Rac2 GTPases in mice disrupts the normal hexagonal organization of the RBC cytoskeleton and reduces erythrocyte deformability. This is associated with increased phosphorylation of adducin at Ser-724, (corresponding to Ser-726 in human erythrocytes), a domain target of protein kinase C (PKC). PKC phosphorylates adducin and leads to decreased F-actin capping and dissociation of spectrin from actin, implicating a significant role of such phosphorylation in cytoskeletal remodeling. We evaluated adducin phosphorylation in erythrocytes from patients with sickle cell disease and found it consistently increased at Ser-726. In addition, ROS concentration is elevated in sickle erythrocytes by 150–250% compared to erythrocytes from normal control individuals. Here, we review previous studies demonstrating that altered phosphorylation of erythrocyte cytoskeletal proteins and increased ROS production result in disruption of cytoskeleton stability in healthy and sickle cell erythrocytes. We discuss in particular the known and potential roles of protein kinase C and the Rac GTPases in these two processes.

Reduced erythrocyte deformability in active and untreated Behçet’s disease patients

Although the etiology of Behcet's disease is not clarified yet, the vascular endothelial damage and thrombosis are prominent features of the disease. The underlying mechanisms of increased risk of thrombosis in Behcet's disease are not completely understood. It is stated that the changes in blood rheology such as erythrocyte deformability take part in the thrombosis. There are limited numbers of studies with conflicting results about the erythrocyte deformability in Behcet's disease. The previous studies were not limited to the untreated patients with active disease. Blood samples were filtered immediately by "Imugard- leukocyte removal filter". After plasma and erythrocytes were separated by centrifugation, 10% erythrocyte suspensions were prepared with isotonic Tris NaCl buffer solution. These erythrocyte suspensions were filtered under gravitational force by "Nucleopore" to determine the filtration time. The filtration times of the erythrocyte and buffer solutions were referred as t1 and t2 respectively. Finally, deformability indices were obtained by dividing the t1 by t2. A decrease in the deformability of the erythrocytes would result in an increase in the filtration time of the erythrocyte suspension and deformability indices. The median deformability indices were 3.090 and 2.170 for patients and control subjects respectively and the difference was statistically significant (P = 0.000). We suggest that the decrease in erythrocyte deformability in Behcet's disease may be related to oxidative damage of erythrocytes and subsequently takes part in the development of thrombosis in Behcet's disease.

Sickle Erythrocytes Have Increased Adducin Phosphorylation and Increased ROS Production Mediated by Signaling Pathways Involving Protein Kinase C and Rac GTPases.

Abstract 901Although sickle hemoglobin (HbSS) polymerization in hypoxic conditions is the trigger for the sickling deformation of red blood cells (RBC) containing HbSS, subsequent changes in the cytoskeleton and the mechanisms by which reversibly sickled cells become irreversibly malformed have not been fully elucidated. Actin oligomers are a significant structural component of the erythrocyte cytoskeleton. The small Rho GTPases Rac1 and Rac2 regulate actin structures and mediate reactive oxygen species (ROS) production via NADPH oxidase in a variety of cells (Schwartz.M., J.Cell Science 2004). Deficiency of Rac1 and Rac2 GTPases in mice disrupts the normal hexagonal organization of the RBC cytoskeleton and reduces erythrocyte deformability. This is associated with increased phosphorylation of adducin (an F-actin capping protein) at Ser-726, a domain-target of protein kinase C (PKC) (Kalfa et al, Blood 2006).We evaluated the cytoskeleton phosphorylation changes in RBCs from patients with HbSS and from control subjects with hemoglobin AA (HbAA) by immunoblotting with phospho-specific antibodies. We found that phosphorylation of adducin at Ser-726 and of band 4.1 at Ser-331 were significantly increased in the erythrocyte cytoskeleton of HbSS-RBCs. This was associated with elevated PKC activity in these cells, compared with normal RBCs. ROS concentration was also elevated in HbSS-RBCs by 150-250% compared to that in HbAA-RBC, as determined by flow cytometry using 5-(and 6-)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate, a peroxide-sensitive probe. Inhibition of PKC or Rac activity in HbSS-RBCs by small molecule-inhibitors, calphostin and NSC23766 respectively, resulted in decreased ROS production in a dose-dependent fashion, while PKC activation by phorbol 12-myristate 13-acetate (PMA) increased ROS production, implicating a PKC-Rac axis in erythrocyte ROS production. Additionally, Rac inhibition resulted in dose-dependent increase of adducin phosphorylation while PKC inhibition had the opposite effect. Preincubation of HbSS RBCs with calphostin appeared to decrease the subsequent susceptibility of these cells to sickling under hypoxic conditions, suggesting a role for PKC activity in the transition from normal to sickle morphology, likely through a combination of effects on volume regulation and membrane stability of the cells. Our results imply that aberrant cytoskeleton protein phosphorylation and ROS production in HbSS-RBCs, mediated by signaling pathways involving PKC and Rac GTPases, may contribute to membrane changes induced by cell sickling. Elucidation of these pathways may identify new therapeutic targets in sickle cell disease. Disclosures:No relevant conflicts of interest to declare.

Effect of 3,4,5,6-tetrahydroxyxanthone on erythrocyte deformability in apolipoprotein E-deficient mice

Previous investigations have indicated that reduced erythrocyte deformability may be an important factor contributing to the development of atherosclerosis, and endogenous asymmetric dimethylarginine (ADMA) might be an important contributor to reduction of erythrocyte deformability in atherosclerosis. In this study, the effect of 3,4,5,6-tetrahydroxyxanthone (1), a kind of polyphenolic compound, on erythrocyte deformability in apolipoprotein E-deficient (apoE − / − ) mice was evaluated. After treatment with compound 1 (10 or 20 mg/kg per day) for 4 weeks, erythrocyte deformability, antioxidant enzymes activity, erythrocyte dimethylarginine dimethylaminohydrolase (DDAH) activity, the plasma level of ADMA and malondialdehyde (MDA) level were determined. Treatment with compound 1 (10 or 20 mg/kg) increased erythrocyte deformability, antioxidant enzymes activity concomitantly, a decrease in the plasma levels of MDA and ADMA, and an increase in erythrocyte DDAH activity. The present result suggests that the beneficial effect of 1 on the erythrocyte deformability, besides inhibiting lipid peroxidation, may be related to reduction of ADMA concentration via an increase in DDAH activity.

ERYTHROPOIETIN ENHANCES OXYGENATION IN CRITICALLY PERFUSED TISSUE THROUGH MODULATION OF NITRIC OXIDE SYNTHASE

The aim of this study was to investigate the effect of human recombinant erythropoietin (EPO) on the microcirculation and oxygenation of critically ischemic tissue and to elucidate the role of endothelial NO synthase in EPO-mediated tissue protection. Island flaps were dissected from the back skin of anesthetized male Syrian golden hamsters including a critically ischemic, hypoxic area that was perfused via a collateralized vasculature. Before ischemia, animals received an injection of epoetin beta at a dose of 5,000 U/kg body weight with (n = 7) or without (n = 7) blocking NO synthase by 30 mg/kg body weight L-NAME (Nomega-nitro-L-arginine methyl ester hydrochloride). Saline-treated animals served as control (n = 7). Ischemic tissue damage was characterized by severe hypoperfusion and inflammation, hypoxia, and accumulation of apoptotic cell nuclei after 5 h of collateralization. Erythropoietin pretreatment increased arteriolar and venular blood flow by 33% and 37%, respectively (P < 0.05), and attenuated leukocytic inflammation by approximately 75% (P < 0.05). Furthermore, partial tissue oxygen tension in the ischemic tissue increased from 8.2 to 15.8 mmHg (P < 0.05), which was paralleled by a 21% increased density of patent capillaries (P < 0.05) and a 50% reduced apoptotic cell count (P < 0.05). The improved microcirculation and oxygenation were associated with a 2.2-fold (P < 0.05) increase of endothelial NO synthase protein expression. Of interest, L-NAME completely abolished all the beneficial effects of EPO pretreatment. Our study demonstrates that, in critically ischemic and hypoxic collateralized tissue, EPO pretreatment improves tissue perfusion and oxygenation in vivo. This effect may be attributed to NO-dependent vasodilative effects and anti-inflammatory actions on the altered vascular endothelium.

The Double Nucleation Model for Sickle Cell Haemoglobin Polymerization: Full Integration and Comparison with Experimental Data

Sickle cell haemoglobin (HbS) polymerization reduces erythrocyte deformability, causing deleterous vaso-occlusions. The double-nucleation model states that polymers grow from HbS aggregates, the nuclei, (i) in solution (homogeneous nucleation), (ii) onto existing polymers (heterogeneous nucleation). When linearized at initial HbS concentration, this model predicts early polymerization and its characteristic delay-time (Ferrone et al. J Mol Biol 183(4):591-610, 611-631, 1985). Addressing its relevance for describing complete polymerization, we constructed the full, non-linearized model (Simulink), The MathWorks). Here, we compare the simulated outputs to experimental progress curves (n = 6-8 different [HbS], 3-6 mM range, from Ferrone's group). Within 10% from start, average root mean square (rms) deviation between simulated and experimental curves is 0.04 +/- 0.01 (25 degrees C, n = 8; mean +/- standard error). Conversely, for complete progress curves, averaged rms is 0.48 +/- 0.04. This figure is improved to 0.13 +/- 0.01 by adjusting heterogeneous pathway parameters (p < 0.01): the nucleus stability (sigma(2) micro( cc ): + 40%), and the fraction of polymer surface available for nucleation (phi), from 5e(-7), (3 mM) to 13 (6 mM). Similar results are obtained at 37 degrees C. We conclude that the physico-chemical description of heterogeneous nucleation warrants refinements in order to capture the whole HbS polymerization process.

Effects of oxidized fibrinogen on the functions of blood cells, blood clotting, and rheology

Oxidatively-modified fibrinogen induces platelet aggregation and potentiates ADP-induced platelet aggregation and production of active oxygen forms in zymosan-stimulated leukocytes. Fibrinogen induces IL-8 production in primary culture of endothelial cells from human umbilical vein; the oxidized form of fibrinogen is more active, similarly as during induction of the expression cell adhesion molecules (P-selectin and ICAM-1). Oxidized fibrinogen (10 and 20% oxidation degree) impairs microrheological properties of the blood, sharply reduces erythrocyte deformability, modifies blood viscosity, and reduces suspension stability of the blood. Oxidized fibrinogen modified blood clotting parameters and ADP-, ristocetin-, and collagen-induced platelet aggregation in whole blood. Oxidized fibrinogen disordered the formation of fibrin clot and blood clotting process. Platelet aggregation was activated in response to ADP, but not to ristocetin and collagen, the degree of activation increased in direct proportion to the degree of fibrinogen oxidation. This indicates the "dysregulatory" effect of oxidized fibrinogen on platelets. The formation of platelet complexes with polymorphonuclear leukocytes was intensified in the presence of oxidized fibrinogen; polymorphonuclear leukocyte luminol-dependent fluorescence intensity in the presence of platelets increased after incubation with oxidized fibrinogen in comparison with native fibrinogen. Hence, oxidized fibrinogen plays an important role in the development of atherosclerosis and its complications (thromboses).

Asymmetric dimethylarginine reduced erythrocyte deformability in streptozotocin-induced diabetic rats

To investigate the effect of asymmetric dimethylarginine on erythrocyte deformability in streptozotocin-induced diabetic rats, a single intraperitoneal injection of streptozotocin (STZ, 65 mg/kg) in male Sprague–Dawley rats was carried out to induce diabetes and normal erythrocytes were incubated with asymmetric dimethylarginine or aortic rings from diabetic rats in the presence of l-arginine or vitamin E. We found that erythrocyte deformability was significantly decreased in diabetic rats. The levels of asymmetric dimethylarginine in plasma and erythrocytes of diabetic rats were elevated significantly from 2-week diabetic duration to 8-week diabetic duration. Nitric oxide in erythrocytes was decreased at 8-week diabetic duration while plasma nitric oxide remained unchanged all along. The content of malondialdehyde in erythrocytes of diabetic rats was increased. After incubation of erythrocytes with asymmetric dimethylarginine (10 − 6 M) for 30 min, erythrocyte deformability and nitric oxide level in erythrocytes were decreased markedly. Reactive oxygen species and malondialdehyde production in erythrocytes were promoted by asymmetric dimethylarginine. Both l-arginine and vitamin E reversed the effects of asymmetric dimethylarginine. After incubation of erythrocytes with aortic rings from diabetic rats, erythrocyte deformability was decreased, which was attenuated by l-arginine. These results indicated that reduction of erythrocyte deformability in diabetic rats was associated with promoted oxidant stress as well as impaired nitric oxide synthesis by elevation of asymmetric dimethylarginine.

Reduced erythrocyte deformability in Krushinsky-Molodkina rats with acute hemorrhagic stroke

Acute hemorrhagic stroke in Krushinsky-Molodkina rats was used to assess the ability of erythrocytes to change their shape in a shear flow. Membrane rigidity and internal viscosity of erythrocytes were measured by laser diffractometry (i.e., obtaining a diffraction pattern from a thin layer of an erythrocyte suspension in a shear flow followed by computer processing of the image). The results testify to reduced deformability of erythrocytes under hemorrhagic stroke.

Redistribution of intestinal microcirculatory oxygenation during acute hemodilution in pigs.

Acute normovolemic hemodilution (ANH) compromizes intestinal microcirculatory oxygenation; however, the underlying mechanisms are incompletely understood. We hypothesized that contributors herein include redistribution of oxygen away from the intestines and shunting of oxygen within the intestines. The latter may be due to the impaired ability of erythrocytes to off-load oxygen within the microcirculation, thus yielding low tissue/plasma Po(2) but elevated microcirculatory hemoglobin oxygen (HbO(2)) saturations. Alternatively, oxygen shunting may also be due to reduced erythrocyte deformability, hindering the ability of erythrocytes to enter capillaries. Anesthetized pigs underwent ANH (20, 40, 60, and 90 ml/kg hydroxyethyl starch; ANH group: n = 10; controls: n = 5). We measured systemic and mesenteric perfusion. Microvascular intestinal oxygenation was measured independently by remission spectrophotometry [microcirculatory HbO(2) saturation (muHbO(2))] and palladium-porphyrin phosphorescence quenching [microcirculatory oxygen pressure in plasma/tissue (muPo(2))]. Microcirculatory oxygen shunting was assessed as the disparity between mucosal and mesenteric venous HbO(2) saturation (HbO(2)-gap). Erythrocyte deformability was measured as shear stress-induced cell elongation (LORCA difractometer). ANH reduced hemoglobin concentration from 8.1 to 2.2 g/dl. Relative mesenteric perfusion decreased (decreased mesenteric/systemic perfusion fraction). A paralleled reduction occurred in mucosal muHbO(2) (68 +/- 2 to 41 +/- 3%) and muPo(2) (28 +/- 1 to 17 +/- 1 Torr). Thus the proposed constellation indicative for oxygen off-load deficits (sustained muHbO(2) at decreased muPo(2)) did not develop. A twofold increase in the HbO(2)-gap indicated increasing intestinal microcirculatory oxygen shunting. Significant impairment in erythrocyte deformability developed during ANH. We conclude that reduced intestinal oxygenation during ANH is, in addition to redistribution of oxygen delivery away from the intestines, associated with oxygen shunting within the intestines. This shunting appears to be not primarily caused by oxygen off-load deficit but rather by oxygen/erythrocytes bypassing capillaries, wherein a potential contributor is impaired erythrocyte deformability.

Platelet-erythrocyte interactions enhance αIIbβ3 integrin receptor activation and P-selectin expression during platelet recruitment: down-regulation by aspirin ex vivo

Activated platelets release biologically active compounds, which then recruit additional platelets into an evolving thrombus. We studied activation of alpha(IIb)beta(3) and exposure of P-selectin on platelets recruited by releasates obtained from collagen-treated platelets and evaluated modifications in prothrombotic effects of releasates induced by platelet-erythrocyte interactions and aspirin treatment. Releasates from collagen-stimulated platelets induced alpha(IIb)beta(3) activation and P-selectin exposure (monitored by flow cytometry using fluorescein isothiocyanate-PAC-1 and phycoerythrin-CD62 antibodies). These responses were markedly amplified by releasates from combined platelet-erythrocyte suspensions. This finding demonstrates a novel mechanism(s) by which erythrocytes intensify platelet aggregability and mediate increased platelet recruitment. Because P-selectin and alpha(IIb)beta(3) are potential sites for platelet-leukocyte interactions, erythrocytes may also modulate leukocyte recruitment. Following aspirin ingestion both the recruiting capacity of platelet releasates and erythrocyte-induced amplification of platelet recruitment were down-regulated. These events represent an additional antithrombotic property of aspirin. We also examined the possibility that arachidonic acid, or eicosanoids derived therefrom, can induce a prothrombotic activity of erythrocytes. The TXA(2)-analog U46 619 and free arachidonate, but not PGI(2) or 12-HETE, induced increases in cytosolic Ca(++) and promoted phosphatidylserine (PS) exposure on a subpopulation of erythrocytes. PS exposure and increases in erythrocyte [Ca(++)](i) are associated with enhanced procoagulant activity, increased endothelial adhesion, and reduced erythrocyte deformability. Our findings, therefore, suggest that TXA(2) and arachidonic acid, derived from activated platelets, induce a prothrombotic phenotype on erythrocytes in proximity. We conclude that by these mechanisms, erythrocytes can actively contribute to platelet-driven thrombogenesis and microvascular occlusion.

Prohemorrhagic potential of dipyrone, ibuprofen, ketorolac, and aspirin: mechanisms associated with blood flow and erythrocyte deformability.

Dipyrone, ibuprofen, ketorolac, and aspirin were tested in a well-defined perfusion system (shear rates: 300/s, 800/s, and 1,800/s). Whole blood samples were treated with the drugs at analgesic doses and platelet interaction with damaged subendothelium was measured. All the drugs fully inhibited platelet cyclooxygenase, as assessed by classic aggregometry. Perfusion studies showed that there was a general tendency to reduce the percentage of large aggregates (thrombus; %T), to increase the percentage of adhered platelets (adhesion; %A), and to reduce the height of thrombi with respect to control. Aspirin significantly increased %A and reduced %T at all shear rates tested, whereas dipyrone had the same effect at 800/s, and ketorolac and ibuprofen at 1,800/s. In addition, aspirin significantly reduced erythrocyte deformability with respect to the other drugs. In conclusion, under our experimental conditions, aspirin showed the most remarkable effects on platelet function, closely followed by dipyrone. The effects of ketorolac were moderate, whereas ibuprofen had a minor impact on platelet function.

Hemorheological changes and overproduction of cytokines from immune cells in mild to moderate dementia of the Alzheimer’s type: adverse effects on cerebromicrovascular system.

An association between hemorheological alterations (i.e., whole-blood and plasma hyperviscosity, reduced erythrocyte deformability, increased red cell aggregation, hyperfibrinogenemia and increased acute-phase protein levels) and the mild stage of senile dementia of the Alzheimer’s type (DAT) was suggested in the present study. In particular, hyperfibrinogenemia and the increase of erytrhocyte aggregation were correlated with the increased generation and release of TNF-α and IFN-γ (spontaneous release and IL-2-modulated release) from natural killer (NK) lymphocytes (CD16+, CD56+, CD3- cells) of patients with DAT; whereas a normal cytokine release from NK cells was found in healthy old subjects and in patients with vascular dementia (VaD). The in vitro and in vivo administration of the hemorheologic drug pentoxifylline (PTX) significantly reduced spontaneous and IL-2-modulated cytokine overproduction from NK cells (in vitro effects with 500 U/ml and 1000 U/ml/NK cells) and improved all the hemorheological parameters. Taken together, these data suggest that disturbances of cerebrovascular flow and of hemorheology could be considered a negative component related to the pathogenesis and progression of DAT neurodegeneration. The association between hemorheological changes and alterations of TNF-α and IFN-γ release from NK may indicate a potential immunorheologic mechanism associated with cerebrovascular damage in DAT and could suggest the use of vascular protective drugs as support of the main pharmacological and non-pharmacological therapy of AD.

Improvement of erythrocyte deformability by cholesterol-lowering therapy with pravastatin in hypercholesterolemic patients

Erythrocyte deformation is an important regulatory factor of the microcirculation. The present study was designed to examine whether erythrocyte deformability is altered in hypercholesterolemic patients and, if so, whether cholesterol-lowering therapy affects this parameter in these patients. The erythrocyte deformability of 37 hypercholesterolemic patients was evaluated before and after 1 year of therapy with pravastatin, an inhibitor of hepatic hydroxymethyl glutaryl coenzyme A reductase, under various shear stresses (4.7, 9.5, 23.6, 47.3, 118.1, and 236.2 dyne/cm 2) using laser diffractometry. At study entry, erythrocyte deformability under 4.7 and 9.5 dyne/cm 2 shear stress, which is actually observed in human vessels, was reduced compared with that in 20 age-matched normocholesterolemic subjects and was inversely correlated with serum cholesterol and low-density lipoprotein (LDL) cholesterol. Pravastatin therapy for 1 year, which reduced serum cholesterol from 288 ± 28 to 223 ± 20 mg/dL, significantly improved erythrocyte deformability by approximately 20%. There was a significant relation between the improvement of erythrocyte deformability and the reduction of serum cholesterol or LDL cholesterol. The results suggest that erythrocyte deformability is reduced in hypercholesterolemic patients, and that long-term cholesterol-lowering therapy can improve reduced erythrocyte deformability, which may contribute to the improvement of organ perfusion.

Reduced erythrocyte deformability related to activated lipid peroxidation during the early postburn period

The role of lipid peroxidation in reduced red cell deformability has been examined after thermal injury of rats (full skin thickness over 15–20 per cent of total body surface). An increased concentration of erythrocyte malonyldialdehyde (MDA) and blood thiobarbituric acid (TBA) reactive product levels during the first 3 days after thermal trauma was established. There was a remarkable decrease in red cell superoxide dismutase (SOD) activity after burns and an increase in glucose-6-phosphate dehydrogenase (G-6-PD) on day 3 postburn. The deformability of erythrocytes was diminished significantly during the first 3 days after burns. There was a significant negative correlation between the deformability and the amount of MDA accumulated in erythrocytes (r= −0.92). Both the accumulation of MDA and the reduction of SOD activity in erythrocytes were suppressed by alpha-tocopherol treatment, which also prevented the decrease in erythrocyte deformability. These results suggest that the activation of the peroxidative process is a possible mechanism for the decreased deformability of erythrocytes during the early stage after thermal injury.

Correction of erythrocyte deformability defect in ALX-induced diabetic rabbits after treatment with aminoguanidine.

To test whether treatment with aminoguanidine, a drug known to prevent cross-linking between glycated proteins, is effective in improving reduced erythrocyte deformability in diabetes, we studied a group (n = 6) of ALX-induced long-term (12.7 +/- 2.2 mo of hyperglycemia) diabetic New Zealand white rabbits before and after 20 wk of treatment with aminoguanidine (100 mg.kg-1 x day-1). The key findings were as follows: 1) at 12 wk of treatment with aminoguanidine, mean erythrocyte deformability normalized and remained within the normal reference range throughout the period of aminoguanidine administration; 2) 10 wk after discontinuing aminoguanidine in a subset of diabetic rabbits, mean erythrocyte deformability deteriorated by approximately 50%; 3) blood glucose and total GHb did not vary significantly during treatment with aminoguanidine nor after its discontinuation.

Effects of alloxan-induced diabetes on hemorheology in rabbits.

Plasma viscosity (PV), apparent whole blood viscosity (WBV), relative blood viscosity (RV) and erythrocyte deformability (filterability) (EDF) were determined in 13 New Zealand White (NZW) rabbits with alloxan induced-diabetes (AID) and 8 normal NZW rabbits, matched for age, sex and weight. AID rabbits were divided into two groups depending on the duration of hyperglycemia (long-term, greater than 6.0 months (n = 7), and short-term, less than or equal to 3.0 months of hyperglycemia, n = 6). Comparing long-term AID rabbits to normal animals, we found significant increases in WBV (P less than 0.001, 0.005 for high and low rates of shear, respectively), and a marked reduction in EDF (P less than 0.001). There was no significant difference in PV between long-term AID and normal rabbits. Conversely, PV was significantly increased in rabbits with short-term diabetes (P less than 0.01) while there was a concurrent significant increase in WBV measured at high and low rates of shear (P less than 0.001, 0.001, respectively). No difference was detected in EDF between normal and short-term AID rabbits. Furthermore, in long-term AID rabbits there was a strongly positive correlation between RV and reduced erythrocyte deformability (r = 0.94, P = 0.006) while WBV strongly correlated with PV (r = 0.92, P = 0.004) in the short-term AID subgroup. We conclude from these data: (1) elevated blood viscosity in long term AID rabbits is associated with reduced erythrocyte filterability; and (2) elevated WBV in short-term AID rabbits is associated with increased PV.

Control of the erythrocyte free Ca2+ concentration in essential hypertension.

Since Ca2+ ions seem to directly participate in the control of erythrocyte membrane structure and deformability and because cell Ca2+ metabolism has been repeatedly proposed to be modified in hypertension, the intracellular calcium ion concentration ([Ca2+]i) was investigated in red blood cells from hypertensive and normotensive subjects. [Ca2+]i was measured by using the fluorescent Ca2+ chelator fura-2. Red blood cell [Ca2+]i was increased in hypertensive compared with normotensive subjects in the whole population and further increased when hypertensive were compared with age-matched normotensive subjects. An inverse relation between age and [Ca2+]i was observed when calculated with blood pressure adjusted. In hypertensive patients, high [Ca2+]i values were associated with a reduced erythrocyte deformability. The initial rate of 45Ca2+ uptake did not differ between the two blood pressure groups. Similarly, when the extracellular Ca2+ concentration was elevated from 1 to 2 mmol/l, [Ca2+]i increased by 16 +/- 4% (p less than 0.03) in red blood cells from both groups, thus maintaining a significant difference between hypertensive and normotensive subjects. Under these conditions, the addition of 10(-7) mol/l nicardipine, a dihydropyridine Ca2+ antagonist, decreased [Ca2+]i by 15 +/- 4% (p less than 0.05) and 7 +/- 5% in erythrocytes from hypertensive and normotensive subjects, respectively, thereby reducing the difference in [Ca2+]i observed between these two groups. This nicardipine effect was positively correlated to the initial [Ca2+]i. In the presence of 5 mumol/l W7, a calmodulin antagonist, [Ca2+]i increased significantly only in erythrocytes from hypertensive patients (26 +/- 6%, p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Reduced Erythrocyte Deformability as a Possible Contributing Factor to Pulmonary Hypertension and Ascites in Broiler Chickens

Deformability of erythrocytes and hematological parameters in white leghorn and broiler chickens were measured at age 7, 14, 21, 28, and 35 days. For deformability testing, a simple vertical apparatus containing a polycarbonate membrane with 5-microns pores was used. This technique assesses erythrocyte deformability by measuring the filtration time of an erythrocyte suspension through the pores. There was a significant difference in filtration time between the leghorns and broilers at all sampling times. These results indicate that reduced erythrocyte deformability in broilers may be one of the predisposing factors that increase resistance to blood flow and alter the rheology of blood in the microcirculation in the lung. Increased resistance to flow may result in pulmonary hypertension, heart failure, and ascites in broiler chickens.

Reduced Erythrocyte Deformability in Anemic Rats with Adjuvant Arthritis

Erythrocytes in anemic rats with adjuvant arthritis (AA) were less deformable compared with normal rats. Swelling of the spleen was noticed in anemic rats with AA. The treatment of anemic rats with recombinant human erythropoietin (r-HuEPO; 30 and 100 U/kg, i.v., for 5 days) resulted in a normalization of both the anemia and erythrocyte deformability. It is suggested that erythrocytes with reduced deformability may be sequestered by endothelial slits of the spleen, which may play a causative role in the anemia in rats with AA.