Comparative intermolecular cross-relaxation studies of human hemoglobin in red blood cells and bovine serum albumin in solution.

Intermolecular cross-relaxation rate (CR) spectra [1/T(IS) (HDO) or 1/T(IS) (H(2) O) vs f(2) (ppm) profiles] for bovine serum albumin [BSA; molecular weight (MW), 66 kDa] solution, partially hydrolyzed BSA gel (BSA*gel) and packed human red blood cells (RBCs) with normal or unstable hemoglobin (Hb; MW, 65 kDa) were studied using f(2) irradiation ranging from - 100 to 100 ppm at γH(2) /2π of 250 Hz. The CR spectra for BSA*gel (pD 4.01, 0.

Impaired erythrocyte filterability of spontaneously hypertensive rats: investigation by nickel filtration technique.

Background: Deformability of erythrocytes plays a key role in the impairment of the microcirculation in hypertension. However, erythrocyte deformability in spontaneously hypertensive rats (SHR) during development of hypertension has not been fully investigated so far.

Methods And Results: Erythrocyte filterability (whole cell deformability) was investigated in relation to blood pressure measured by the tail-cuff method in SHR and age-matched Wistar-Kyoto rats (WKY), using a highly sensitive and reproducible nickel mesh filtration technique.

Impaired filterability of erythrocytes from patients with chronic hepatitis C and effects of eicosapentaenoic acid on the filterability.

Although erythrocyte filterability plays a key role in microcirculation, it is unknown whether the filterability of erythrocytes from patients with chronic hepatitis C (CH-C) is impaired. This study aimed to investigate erythrocyte filterability in CH-C patients in relation to medical treatment. The mean erythrocyte filterability (%) for all 24 patients with CH-C (69.

Verapamil prevents impairment in filterability of human erythrocytes exposed to oxidative stress.

Effects of oxidative stress on intact human erythrocytes were investigated using tert-butyl hydroperoxide (tBHP). Exposure of erythrocytes to tBHP caused a marked decrease in filterability in a time-dependent manner. Erythrocytes exposed to tBHP also show an increase in mean corpuscular volume and a remarkable formation of methemoglobin (met-Hb) without any appearance of hemichromes that form Heinz bodies.

A new quantitative filtration apparatus to evaluate erythrocyte deformability.

We have developed a quantitative filtration apparatus to evaluate erythrocyte deformability, using a precision metal filter which we developed. The test material is sucked into the filter through the action of decreasing negative pressure. The device automatically gives a liquid level (height)-time relation and a pressure-flow rate relation, similar to gravity-based filtration.

Saturation transfer in human red blood cells with normal and unstable hemoglobin.

Saturation transfer phenomena from irradiated protein protons to observed water protons in packed human red blood cells (RBCs) with normal or unstable hemoglobin (Hb), i.e. Hb Yokohama and Hb Koeln, were studied using intermolecular cross-relaxation rates [CR; 1/T(IS)(H(2)O)], action spectra [[1-(I(infinity)/I(0))] vs f(2) (ppm), where I(0) and I(infinity) are the longitudinal magnetization of observed water protons before and after long-time f(2)-irradiation, respectively], CR spectra [CR vs f(2) (ppm)] and CR ratio vs f(2) (ppm) with f(2)-irradiation from -100 to 100 ppm at gammaH(2)/2pi of 69 or 250 Hz.

Application of the transition state theory to water transport across cell membranes.

We have applied the transition state theory of Eyring et al. (The Theory of Rate Processes, McGraw-Hill, 1941) to water transport across cell membranes. We have then evaluated free energy (Delta F(not equal)), enthalpy (Delta H(not equal)) and entropy (Delta S(not equal)) of activation for water permeation across membranes, such as Arbacia eggs, Xenopus oocytes with or without aquaporin water channels, mammalian erythrocytes, aquaporin proteoliposomes, liposomes and collodion membrane.

Relationship of high-density lipoprotein cholesterol and red blood cell filterability: cross-sectional study of healthy subjects.

The deformability of red blood cells (RBCs) is an important rheologic factor in the maintenance of normal blood flow in the microcirculation. Contrary to the well-known relationship between hyperlipidemia and atherosclerosis, the relationship between RBC rheology and the serum lipid profile has remained controversial and obscure. Moreover, the correlation of high-density lipoprotein (HDL)-cholesterol and RBC deformability has not been fully understood.

Endothelin-1 improves the impaired filterability of red blood cells through the activation of protein kinase C.

We previously showed that the deformability of human red blood cells (RBCs) is affected by intracellular signaling pathways by examining the effects of Ca2+ influx and the intracellular cAMP level on mechanically-impaired RBC filterability. In the present study, we investigated whether protein kinase C (PKC) participates in the regulation of RBC deformability by affecting membrane properties. The filterability of mechanically-stressed RBCs showed a V-shaped curve depending on the extracellular Ca2+ concentration; the maximum decrease was achieved at 20-40 microM.

Kinetic analysis of the mechanism of action of the multidrug transporter.

To clarify the mechanistic role of PGP (P-glycoprotein) in multidrug transport, we constructed a kinetic model composed of four compartments: (1) the extracellular space; (2) the space in the membrane; (3) the intracellular space; and (4) the pore-like space in the PGP molecule. The kinetics of the concentration of ADM (adriamycin) in each compartment were formulated based on the assumptions that (a) the movement of ADM between two compartments by diffusion is dependent on a dynamic distribution coefficient introduced here, (b) the uptake of ADM into the pore-like structure by the pump mechanism activated by ATP is described by enzyme kinetics, (c) the movement of ADM out of the pore-like structure to the extracellular medium through a valve-like mechanism is also expressed by enzyme kinetics. The mathematical analysis of the exact solution can explain the distinct effects of verapamil and vanadate on the accumulation and release of ADM, where verapamil inhibits the efflux by the valve-like mechanism and vanadate blocks the influx by the pump mechanism.

Production of prostaglandins E1 and E2 by adult human red blood cells.

We showed that human adult red blood cells (RBCs) produce prostaglandin E1 (PGE1) and E2 (PGE2). RBCs that were mechanically stressed in the presence of extracellular Ca2+ by being injected rapidly through a fine needle produced PGE1 and PGE2 within 30 min after this mechanical stress. The amounts of PGE1 and PGE2 produced by 1 x 10(9) mechanically stressed RBCs were approximately 50 pg and 100 pg, respectively, which were determined in the cytosolic fraction from sonicated RBCs using a competitive enzyme immunoassay method.

Sickling of nucleated erythroid precursors from patients with sickle cell anemia.

The pathophysiology of sickle cell anemia is primarily explained in terms of the oxygen-dependent polymerization of sickle hemoglobin (HbS) followed by sickling of erythrocytes. Since the rate and extent of HbS polymerization depend on its intracellular concentration, it has been generally assumed that sickling occurs primarily in mature erythrocytes with their high intracellular hemoglobin concentration. In the present study, we investigated the propensity of nucleated erythroid precursors to undergo sickling; both cultured and fresh marrow-derived erythroid precursors from patients with homozygous sickle cell anemia were studied.

Regulation of red blood cell filterability by Ca2+ influx and cAMP-mediated signaling pathways.

To investigate the mechanism of the regulation of human red blood cell deformability, we examined the deformability under mechanical stress. Washed human red blood cells were rapidly injected through a fine needle, and their filterability was measured using a nickel mesh filter. The decrease in filterability showed a V-shaped curve depending on the extracellular Ca2+ concentration; the maximum decrease was achieved at approximately 50 muM.

Quantitative characterization of P-glycoprotein-mediated transport in mdr1-gene-transfected lymphoma cells.

We have established a quantitative flow cytometry system to elucidate the causal role of P-glycoprotein in the phenomenon of multidrug resistance. We have used this method to analyze the accumulation and release of adriamycin (ADM) in intact L5178Y and L5178Y/VMDR/C.06 (L5178Y/R) cells, by determining the effect of sodium orthovanadate (Na3VO4), verapamil, bovine serum albumin (BSA) and physiologically operative materials on the cells.

Filterability of mixtures of sickle and normal erythrocytes.

We investigated the deformability of sickle (SS) cells from 25 patients and mixtures of these SS cells with blood type-compatible normal (AA) cells, using a nickel mesh filtration system, with the aim of determining optimal goals for exchange therapy. We found that for air-equilibrated SS/AA cell mixtures the fraction of dense cells (MCHC > 37 g/dl) is the determinant factor in filterability and that the dense cells contribute in a linear fashion to the loss of filtration up to 15% dense cells (y = -4.41x + 98.

Contributions of sickle hemoglobin polymer and sickle cell membranes to impaired filterability.

Sickle cell anemia is a disease of abnormal rheology caused by acute and reversible, as well as chronic and irreversible, changes in the properties and deformability of sickle erythrocytes. Deformability is determined by several factors, including intracellular sickle hemoglobin polymerization, the abnormal membrane properties of sickle cells, and the abnormal rheological properties of the soluble concentrated hemoglobin solution within dense sickle red blood cells. In this study, we used a 5-microns pore nickel mesh filter to evaluate quantitatively the effects of these factors on the filterability of erythrocytes containing sickle hemoglobin.

Sickle cell rheology is determined by polymer fraction--not cell morphology.

Sickle cell disease pathophysiology is mediated by acute and chronic impairment of cell flexibility due to the formation of intracellular sickle hemoglobin (Hb S) polymer as cells are partially deoxygenated in the microcirculation. We have recently developed a method to measure the relationship between the formation of intracellular polymerized Hb S and cell filtration. In this study, we have used this method to examine whether sickle cell morphology, independent of Hb S polymer fraction, had an effect on cell rheology.

Rheologic and pathophysiologic significance of red cell passage through narrow pores.

To elucidate the pathophysiologic significance of red blood cell (RBC) filterability, we measured RBC rheology with our own designed nickel mesh with 3-microns pores, smaller than the previously used 5-microns pores. Vertical and cylindrical pores with no pore coincidence were regularly distributed across the filter, the pore entrances of which showed a round and rather smooth transition to the pore inside. An advantage of the nickel mesh is the repeated use (at least 100 times) of the same filter possible after ultrasonic washing.

Effect of low-osmolality contrast media on red cell filterability.

The effects of low-osmolality contrast media (CM) on red blood cell (RBC) filterability were investigated using a recently developed nickel mesh filtration method. The conventional hypertonic CM iothalamate, low-osmolality iohexol, and the recently synthesized iomeprol were studied. Among them, the osmolality of iomeprol was the lowest.

Impaired deformability of Heinz body-forming red cells.

Although a decrease in the deformability of red blood cells (RBCs) has been suspected in Heinz body-forming hemolytic anemia, it remains uncertain whether the formation of Heinz bodies themselves impairs RBC deformability or not. To elucidate this question, we used RBCs treated with phenylhydrazine and RBCs from patients with unstable hemoglobin (Hb) disease (Hb Yokohama) to investigate the effect of Heinz body formation on deformability in terms of RBC filterability through nickel mesh and viscosity of the RBC suspension. The phenylhydrazine-treated RBCs exhibited a marked decrease in deformability in a dose-dependent manner.

Augmentation by erythropoietin of the fetal-hemoglobin response to hydroxyurea in sickle cell disease.

Background: Hydroxyurea increases the production of fetal hemoglobin in patients with sickle cell anemia, inhibiting the polymerization of hemoglobin S and potentially improving vaso-occlusive manifestations and hemolysis. Recombinant erythropoietin increases the number of reticulocytes containing fetal hemoglobin in laboratory animals and in humans. We studied whether hydroxyurea and erythropoietin might have a potentiating effect on the production of fetal hemoglobin in patients with sickle cell disease.

Effects of superoxide anions on red cell deformability and membrane proteins.

The effect of superoxide anions (O2-) on red blood cells (RBC) deformability and membrane proteins was investigated using hypoxanthine-xanthine oxidase system. Exposure of RBC to O2- caused a marked decrease in RBC deformability with a concomitant increase in cell volume and shape changes. The RBC exposed to O2- also displayed pronounced degradation of membrane proteins such as band 3 protein and spectrin; new bands of low molecular weight products appeared as the original membrane proteins tended to diminish, without the appearance of high molecular weight products.

Possible role of red cell deformability and microvasculature in microcirculation.

The roles of the deformability of red blood cells (RBC) and the microvasculature in the maintenance of blood flow were investigated in terms of the pressure (P)-flow rate (Q) relationships in human RBC suspension perfusions of bullfrog hind limb. Although isotonicity for the bullfrog is approximately 215 mOsm/kgH2O, perfusions in intact hind limbs showed no change in the P-Q relationship at test solution osmolalities ranging from approximately 150 to approximately 300 mOsm/kgH2O. The deformability of RBC was examined in glutaraldehyde-fixed hind limbs.

Further investigations of red cell deformability with nickel mesh.

Although the filtration method has been widely employed in red cell deformability studies, the structural irregularity of the pores of a Nuclepore polycarbonate membrane has always been a major problem. Anegawa, T. et al.

Electron spin resonance studies of erythrocyte membrane in spinocerebellar degeneration.

Erythrocyte membrane fluidity was examined by electron spin resonance spectra using nitroxide fatty acid spin labels in spinocerebellar degeneration (SCD). Subjects with SCD, motor neuron disease (MND) and controls did not differ in fluidity of the deep site (hydrophobic region) of the erythrocyte membrane. However, the fluidity of the shallow site (hydrophilic region) in the erythrocyte membrane was significantly less fluid in SCD than in controls and MND (outer hyperfine splitting of 5-nitroxide stearic acid: SCD 54.