Presence Of Red Cells Research Articles

Effect of RBCs on the activation of human complement by heparin- protamine complexes

Complement activation on red cells by heparin-protamine complexes was studied by using whole human serum. C3 bound to red cells was measured by radiolabeled monoclonal antibody to C3, and fluid-phase C5a was determined by radioimmunoassay. Heparin and protamine in clinically relevant concentrations caused the binding of C3 to red cell membranes, and the measurement of C3 binding provided a sensitive indicator of complement activation produced by these complexes. Complement activation by these reagents occurred at concentration ratios of protamine and heparin at which protamine neutralized the anticoagulant effect of heparin. Heparin-protamine complexes appeared to bind to red cells and produce complement activation by the classic pathway. C5a generation with heparin-protamine complexes in serum was greatly enhanced in the presence of red cells and increased with increasing red cell concentration. This enhancement of complement activation in the presence of red cells was also seen as measured by depletion of available C3 hemolytic complement units in the fluid phase. Thus red cells seem to play an important role in activation of complement by heparin-protamine complexes.

Effect of RBCs on the Activation of Human Complement by Heparin-Protamine Complexes

Complement activation on red cells by heparin-protamine complexes was studied by using whole human serum. C3 bound to red cells was measured by radiolabeled monoclonal antibody to C3, and fluid-phase C5a was determined by radioimmunoassay. Heparin and protamine in clinically relevant concentrations caused the binding of C3 to red cell membranes, and the measurement of C3 binding provided a sensitive indicator of complement activation produced by these complexes. Complement activation by these reagents occurred at concentration ratios of protamine and heparin at which protamine neutralized the anticoagulant effect of heparin. Heparin-protamine complexes appeared to bind to red cells and produce complement activation by the classic pathway. C5a generation with heparin-protamine complexes in serum was greatly enhanced in the presence of red cells and increased with increasing red cell concentration. This enhancement of complement activation in the presence of red cells was also seen as measured by depletion of available C3 hemolytic complement units in the fluid phase. Thus red cells seem to play an important role in activation of complement by heparin-protamine complexes.

Reduction of nitroxide free radical by normal and G6PD deficient red blood cells

The electron spin resonance signal of Tempol decays in the presence of red cells. The decay is due to reduction of the oxidant, paramagnetic nitroxide group by the metabolic activity of the red cell. In normal red cells, GSH level was stable and Tempol reduction rate followed a first-order kinetics. In G6PD-deficient red cells, GSH dropped and Tempol reduction rate was slower and followed a second-order kinetics. In normal red cells, diamide reversibly oxidised GSH. First-order kinetics of Tempol reduction rate was attained after a delay time proportional to the diamide concentration and corresponding to the full regeneration of GSH. In diamide-treated G6PD deficient, and in NEM-treated, normal red cells, irreversible disappearance of GSH was followed by irreversible dose-dependent decrease in Tempol reduction rate. A correlation between GSH levels and Tempol reduction rate was observed. A correlation was also established between Tempol reduction rate and stimulation of pentosephosphate shunt activity.

Effect of erythrocytes on the function and morphology of the isolated perfused rat kidney.

We have examined the effects of erythrocytes on the function and morphology of isolated rat kidneys perfused with a physiological concentration of bovine albumin (45 g/l). (1) In kidneys perfused without red cells, renal vascular resistance (RVR) was low (4.2 +/- 0.3 mm Hg/ml/min/g), fractional sodium excretion (FeNa) was high (14.5 +/- 1.8%) and concentrating ability impaired (maximum urine osmolality 343 +/- 4 mmol/kg). The erythrocyte-free kidney also developed necrosis of the cells of the medullary thick ascending limb (mTAL). (2) Erythrocytes at a hematocrit of 4-6% did not alter RVR but prevented ischemic changes in the mTAL and reduced FeNa to 9.4 +/- 0.03%. Concentrating ability was not improved by a hematocrit of 4-6% despite the presence of a morphologically normal mTAL. (3) At a hematocrit of 40-45%, RVR was increased (to 11.2 +/- 0.4 mm Hg/ml/min/g) and FeNa was further lowered to 3.5 +/- 0.6%. Also, urinary concentrating ability was markedly improved (maximum urine osmolality 640 +/- 35 mmol/kg). (4) The isolated perfused kidney (IPK) at a hematocrit of 40-45% was able to autoregulate renal perfusate flow rate of GFR but autoregulation was incomplete. A 50% increase in perfusion pressure from 100 to 150 mm Hg increased renal perfusate flow rate and GFR 27 and 29%, respectively. Thus the IPK is not able to autoregulate as efficiently as the kidney in vivo, even in the presence of red cells at a normal hematocrit.

Polymerized pyridoxylated hemoglobin: efficacy as an O2 carrier.

We have prepared a polymerized pyridoxylated hemoglobin solution (Poly SFH-P) with a normal [Hb] of 14 gm/dl and a normal COP of 20 torr. Although this normal [Hb] is a significant improvement over prior products, the P50 of 16-20 torr raises a concern about the ability of Poly SFH-P to effectively transport O2 in the presence of red cells with their normal P50 of 26 torr. This study quantitatively assessed the contribution of Poly SFH-P to total O2 delivery and consumption in the clinically relevant range of hematocrits. The results document that Poly SFH-P supports life at zero hematocrit, and makes significant contributions to total O2 delivery and consumption in the presence of red cells. Poly SFH-P permits a higher plasma [Hb], and has a longer intravascular persistence than any unpolymerized hemoglobin solution. Poly SFH-P is thus an effective O2 carrier, and offers greater potential than prior products as a clinically useful red cell substitute.

Effects of ischemia on capillary density and flow velocity in nailfolds of human toes

The purpose of this study was to investigate reactive hyperemia in the capillary network of human skin in terms of the flow per capillary and the density of flow-active capillaries. Seventeen male subjects 20 to 40 years of age were seated with their right foot placed on the stage of a Leitz epi-ilumination microscope such that the nailfold capillary field in their large toes could be viewed. These vessels were video taped while flow velocity in the right posterior tibial artery was recorded via Doppler ultrasound at rest, then following a 45-sec period of arterial occlusion to the foot. Subsequent to experimentation flow velocity in single nailfold capillaries was measured via video densitometry and the number of flow-active capillaries in the field of view were counted. Following the release of arterial occulsion arterial flow velocity increased 142% above rest, the velocity in single capillaries increased by 54%, and the density of flow-active capillaries, as identified by the presence of red cells, decreased by 37%. The fact that capillary flow velocity increased to a lesser degree than arterial velocity during reactive hyperemia vis-a-vis a decrease in the number of flow-active capillaries indicates that ischemia to the foot elicits a smaller dilatory effect in vascular elements controlling blood flow to the superficial cutaneous region of the toe as compared to other regional vascular networks.

Presence of P blood group antigens on human platelets.

Except for ABH antigens, the presence of red cell (RBC) antigens on human platelets has been a source of disagreement among investigators. Because ABH antigens share precursor sequences with P-system saccharides, the authors examined human platelets for evidence of P blood group antigens. Anti-P was directly labeled with 125I and incubated with normal platelets in a one-stage radioimmunoassay (RIA). Alternatively, platelets from donors of known RBC phenotype were incubated with anti-P, anti-P1, or anti-Tja, washed, incubated with FITC-labeled goat anti-human immunoglobulin and evaluated by fluorescence flow cytometry. The results of these assays demonstrate that platelets express P blood group antigens in parallel to the donor's RBCs. The role of these antigens in platelet transfusion is not known.

An assessment of whole blood impedance aggregometry using blood from normal subjects and haemodialysis patients

Impedance aggregometry allows the measurement of platelet responses in whole blood as well as in PRP. The variability of haematocrit values encountered when applying this technique to haemodialysis patients prompted an investigation of the effects of red cells on platelet aggregation in whole blood. Collagen induced aggregation was measured in both PRP and whole blood from haemodialysis patients and healthy controls, Platelets from haemodialysis patients were less aggregable than those from the controls when tested in PRP, but more aggregable when tested in whole blood. Blood samples with a range of haematocrit values were prepared by mixing PRP and autologous red cells, and used to study the effect of haematocrit on platelet aggregation. In blood from control subjects aggregation rate was reduced by rising haematocrit but no reduction of maximum aggregation occurred until haematocrit exceeded 40%. In contrast uraemic platelets showed increased responses in the presence of red cells. In a limited cross-over study no significant difference was found in the effect on platelet aggregation of washed erythrocytes from uraemic and non-uraemic donors. It is concluded that red cell presence influences platelet aggregation by complex mechanisms during impedance aggregometry and that this effect must be considered when interpreting results.

On the mechanism for the red-cell accumulation of mefloquine, an antimalarial drug

The accumulation of the antimalarial drug mefloquine by human red blood cells has been studied by 19F-NMR spectroscopy. The uptake process was nonlinearly dependent on the external drug concentration. Concentrations inside cells as high as 60-times greater than those in the extracellular phosphate-buffered-saline were observed. Red-cell ghosts were also found to accumulate mefloquine with high-affinity binding sites for the drug. Hemoglobin was found to bind mefloquine with low affinity, but due to the high concentration of this protein it is a significant drug compartment in the red cell. Analysis of the 19F-NMR chemical shifts and linewidths of mefloquine in the presence of red cells, red-cells ghosts and hemoglobin indicates restricted mobility of the drug in the membrane-bound state and slow exchange with the extracellular medium. This is a significant characteristic of the reaction in connection with the prophylactic activity of the drug. Exchange of the drug between hemoglobin and the red-cell membrane, however, is fast and may play an important role in the bioavailability of the drug to the parasite.

Red cell deformability, platelet aggregation, and insulin action.

Abnormalities of rheologic and hemostatic properties of blood are present in uncontrolled diabetics and play a role in the development of structural micro- and macroangiopathies. Red blood cell deformability is decreased in diabetics and shows negative correlation with fast hemoglobin and actual blood glucose levels. In uncontrolled insulin-dependent diabetics, normalization of plasma glucose by an insulin infusion improves red cell deformability in 2 h. Insulin infusion (0.2 U/kg/h) [the initial hyperglycemia being maintained (hyperglycemic clamp)] also improves red cell deformability. Deformability of normal erythrocytes is reduced by incubation in plasma from uncontrolled diabetics but is normal in plasma from diabetics controlled by a 24-h insulin infusion or in diabetic plasma with insulin added in vitro. Therefore, insulin appears to have a direct effect on erythrocyte deformability. Platelet aggregation measured in whole blood is raised in uncontrolled diabetics. Aggregation of normal platelets rises in the presence of "diabetic" red cells but not in the presence of red cells from the same patients controlled by 24-h treatment with insulin. The effect of insulin on platelet aggregation, therefore, seems to be at least partly mediated by erythrocytes.

Red Cell Substitutes

Stroma-free hemoglobin (SFH) and Fluosol-DA (FL-DA) are the two acellular oxygen carriers that represent potential red cell substitutes. Although both can support life at zero hematocrit, their effectiveness as O2 carriers at intermediate hematocrits has been unclear. In the presence of red cells, the efficacy of an acellular O2 carrier can be assessed by its contribution to O2 delivery and O2 consumption. This study evaluated the efficacy of SFH and FL-DA in the presence of red cells at hematocrits less than or equal to 10%. The results illustrate that SFH and FL-DA do contribute to O2 delivery and O2 consumption in the presence of red cells. The data indicate that SFH and FL-DA are both effective acellular O2 carriers.

The relation between dicarbocyanine dye fluorescence and the membrane potential of human red blood cells set at varying Donnan equilibria.

The fluorescence, F, of two dicarbocyanine dyes, diS-C3(5) and diI-C3(5), depends both on the membrane potential, E, and on the intracellular pH, pHc, or human red blood cells. Compositions of isotonic media have been devised in which the equilibrium Donnan potential, E, varies at constant pHc and in which pHc varies at constant E. Dye fluorescence measurements in these suspensions yield calibrations of +1.7 % delta F/mV for diS-C3(5) and +0.6 % delta F/mV for diI-C3 (5). While pHo does not affect F of either dye, changes in pHc of 0.1 unit at constant E cause changes of F equivalent to those induced by 2--3mV. Based on these results, a method is given for estimating changes in E from dye fluorescence in experiments in which E and pHc co-vary. The relation of F to E also depends in a complex way on the type and concentration of cells and dye, and the wavelengths employed. The equilibrium calibration of dye fluorescence, when applied to diffusion potentials induced by 1 microM valinomycin, yields a value for the permeability ratio, PK.VAL/PCl, of 20 +/- 5, in agreement with previous estimates by other methods. The calibration of F is identical both for diffusion potentials and for equilibrium potentials, implying that diC-C3(5) responds to changes in voltage independently of ionic fluxes across the red cell membrane. Changes in the absorption spectra of dye in the presence of red cells in response to changes in E show that formation of nonfluorescent dimers contributes to fluorescence quenching of diS-C3(5). In contrast, only a hydrophobic interaction of dye monomers need be considered for diI-C3(5), indicating the occurrence of a simpler mechanism of fluorescence quenching.

Depletion of 18O from C18O2 in erythrocyte suspensions. The permeability of the erythrocyte membrane to CO2.

The depletion of 18O from CO2, caused by the exchange of oxygen between CO2 and water during the hydration-dehydration cycle, is catalyzed by carbonic anhydrase. This depletion process at chemical equilibrium in the presence of erythrocytes is biphasic, exhibiting a very rapid depletion rate immediately following the addition of cells to an isotonic solution containing 18O-enriched CO2, followed by a much slower depletion rate. It is hypothesized that these depletion characteristics are caused by the diffusion of labeled CO2 into erythrocytes where depletion occurs rapidly due to the large intracellular carbonic anhydrase content. Kinetic equations which describe this hypothesis are solved and a rate constant is obtained which represents the depletion of 18O in CO2 caused by the presence of red cells. These are equilibrium experiments with no net uptake or loss of CO2 in the cells. Consequently, depletion processes are not limited in rate by bicarbonate-chloride exchange or proton distribution across the membrane. The purpose of these measurements is to determine whether the rate of 18O depletion in red cell suspensions is determined by carbonic anhydrase activity in the cell or by the diffusion process by which CO2 enters the cell. This goal is achieved by partially inhibiting carbonic anhydrase with acetazolamide. The rate constant representing 18O depletion caused by the presence of red cells is unchanged, even though up to 90% of carbonic anhydrase is inhibited. From this rate constant the permeability constant of the membrane of rat erythrocytes to CO2 at 25 degrees and pH 7.4 is determined to be (7.6 +/- 1.2) X 10(-3) cm s-1 in the presence of 3.2 mM picrate, a passive anion diffusion inhibitor intended to block HCO3 -flux across the membrane. Using no picrate and allowing HCO3-flux to introduce an error in the measurements, the permeability constant is (1.6 +/- 0.4) X 10(-2) cm s-1. The permeability constants measured by this technique include the diffusion barrier to CO2 not only of the red cell membrane but also of a portion of the intracellular medium.

Adhesion of platelets to artificial surfaces: effect of red cells.

Adhesion of platelets to several polymer- and protein-coated glass surfaces has been studied in vitro. The apparatus consists of a cylindrical probe rotating in a test tube containing the platelet medium and allows close control of fluid shear and mass transport. Suspensions of washed pig platelets constitute the basic platelet medium, and can be modified by adding back red cells and plasma proteins. Adhesion is measured via 51Cr-labeling of platelets. In the absence of red cells, identical low levels of adhesion were seen on all surfaces and saturation was reached within 2 min. In the presence of red cells, adhesion was greater. Saturation on all surfaces except fibrinogen and collagen again occurred within 2 min. The adhesion levels on polymer surfaces and glass were indistinguishable, while those on albumin were lower and those on fibrinogen were higher. Collagen was the most reactive surface. It did not equilibrate within 15 min., and kinetic data indicated a platelet diffusivity strongly dependent on hematocrit. These effects were attributed to rotational and translational motion of the red cells causing increased diffusion and surface-platelet collision energy.

ラット胎存肺サファクタント産生に関する実験的研究

Perinatal anoxia was experimentally produced in fetal rats to examine the effect of anoxia on pulmonary lecithin levels. The right uterus of adult Sprague-Dawley rats was exposed by laparatomy on the 19th day of pregnancy, and the right uterine artery was compressed for 2 minutes and then released for 1 minute. This procedure was repeated five times. Thus the rat fetuses in the right uterine horn experienced anoxia for a total of 10 minutes, while the littermates in the left horn were free from anoxia. The fetuses were delivered by cesarean section 24 hours after these treatments, and the lecithin content of the lungs was measured.Pulmonary lecithin levels of the anoxic group were found to be significantly lower than those of the control group. Furthermore, it was found that body weight was significantly less and lung wet weight was significantly greater in the anoxic group. The lungs of the anoxic group showed partial atelectasis, interstitial edema, congestion and the presence of red cells and desquamated epithelial cells in the alveolar lumen.A second experiment was conducted to examine the effects of CDP-ch oline and prednisolone upon the pulmonary lecithin levels of the anoxic fetuses. The fetuses in both uterine horns were exposed to anoxia by the same method used in the first experiment. The rats were divided into three groups. The fetuses in the right horn of the first group were injected intraperitoneally with CDP-choline (200mg/kg), the fetuses in the right horn of the second group with CDP-choline (200 mg/kg) and prednisolone (60mg/kg), and the fetuses in the right horn of the third group with prednisolone (30 mg/kg). The fetuses in the left horn of each group were injected with an amount of saline solution equivalent to the amount of medication received by the fetuses of the right horn, and served as the control group.It was found that when CDP-ch oline was administered in this way (group one), there was no acceleration in the production of lecithin. But the newborn rats which had been injected with prednisolone (group three) showed a significantly higher titer of pulmonary lecithin than did the control group. As for group two, injection with a large dose of prednisolone did not result in an increase of lecithin production, but there was a significant weight loss.It can be conc luded from these results that an anoxic episode during the perinatal period inhibits production of pulmonary surfactant, and is one of the causes of intrauterine growth retardation. The effects of anoxia cannot be corrected by a single administration of CDP-choline. An adequate prednisolone injection can accelerate pulmonary lecithin production, as already reported by other investigators. An overdose of prednisolone reduces the pulmonary lecithin levels, and retards weight gains of rat fetuses significantly.

Clinical features in enteritis due to vibrio parahemolyticus

Clinical features of a case of enteritis due to Vibrio parahemolyticus are presented. Superficial ulcerations of colonic mucosa were observed directly by sigmoidoscopy. Previously such damage had not been documented but inferred from the presence of red cells and leukocytes in the stools. The literature pertinent to V. parahemolyticus and its recent presence in the United States is reviewed.

Unexpected temperature dependence of potassium release from the isolated perfused rat liver.

At temperatures from 15–25° C perfused rat livers take up potassium from the perfusion medium or release small amounts of the cation. Both increasing or decreasing temperature enhance the K+-release in the absence as well as in the presence of red cells. Two possible explanations of this phenomenon, hypoxia and phase-transition of the plasma membrane, are discussed.

Hereditary nephritis. Early clinical, functional, and morphological studies.

Extract: The accumulation of literature on hereditary nephritis with or without deafness attests to the increased awareness of the existence of this syndrone. It is the purpose of his report to show that there are distinguishing histologic findings in young paitents with minimal disease and normal renal function. Eleven paitents were studies; 4 were females and 7 were males. Releveant family histories and clinical laboratory findings are shown in table I. In all paitents, hematuria, gross or microscopic, was present. Exacerbations of the hematuria occurred as a part of the exaggerated response of these paitents to upper respiratory infection. Renal functional data are shown in table II. These data were generally within normal limits for our laboratory. Table III outlines the morphologic findings observed on renal biopsy. In renal biopsies obtained from 10 patients, there was observed persitence of fetal-like glomeruli characterized by the circumferential crowding of visceral epithelium and an apparent decrease in the number of patent capillaries. The most striking finding in the tubules was the presence of red cell and blood cell casts seen mainly in the distal and collecting tubules. Foam cells were an infrequent finding but, when presnt, were unassociated with interstitial fibrosis. Examination of kidney specimens under the electron microscope reveated the presence of at least ferential crowing of visceral epithelial cells, particularly at the surface, facing Bowman's capsule. These epithelial cells were characterized by large nuclei, small amounts of cytoplasm, apparently poorly developed introacytoplasmic organelles, and fiequent villous-like cytoplasmic projections on the surface. In families from whom past histories are unavailable, recognition of distinct renal biopsy characteristics would help identify this syndrome in the early stages of disease. At this stage, the combination of morphologic features considered to be characteristic would include the presence of a) fetal-like glomeruli, which were seen by electorn microcscopy in 7 paitents and by light microscopy in 10 patients; blood cell casts; and d) foam cells in the absence of interstitial fibrosis. The fetal-like glomeruli present in our paitents did not correspond to any of the normal stages of glomerular development observed under electron microscopy by other investiagators, but rather, appeared to represent dysgenesis. Most patients from whom data are reported were more than 2 years of age, at which time fetal gomeruli are exceptional. Of interest was the observaton that almost identical histology was found to exist among the siblings. Speculation: The fetal-like glomeruli observed on renal biopsy might be the result of either a developmental abnormality or dysgensis. The changes obeserved in all glomeruli studied by electron microscopy suggest that even those glomeruli that appear to be morphologically unchanged under light microscopy were involved in or by the pathologic process. It is suggestd that there is a common rather than a divers etiology for ranal disease in children. Longgitudinal follow-up with repeated renal functionl studies and biopsies will be needed to delineate the natural history.

Vascular injury due to topical application of cardiovascular contrast medium in the hamster cheek pouch.

The microcirculatory toxicity of water-soluble cardiovascular contrast agents has received increasing attention in recent years (1). This interest has been stimulated by attempts to visualize smaller and smaller vessels with selective and magnification technics requiring the delivery of highly concentrated radiopaque contrast material for detailed visualization. The effect of intra-arterial and intravenous injection of these compounds in various microcirculation preparations has been reviewed by Wiedeman (2). Read et al. (3) showed experimentally that obstructive circulatory phenomena produced by the administration of angiographic media were dependent upon the presence of red cells in the perfusate. In addition, these workers disclosed that the viscosity of blood and red cell suspensions increased when their osmolality was raised in a glass capillary tube model (4). Although other factors have been considered in the production of intravascular sludging following the injection of radiopaque media, the phen...

On virus induced bacterial agglutination

By replacing red cells with different Colitypes in an analysis of the Hirst-test, it could be shown that influenza viruses, as well as one type of ECHO-virus, were capable of agglutinating bacteria. The agglutination titers of the same viruses against different Coli-types and ECHO-type 7 were dissimilar. It is concluded that, for the demonstration of agglutinating abilities, the presence of red cells is not necessary, since these can be replaced by certain bacteria. Further studies are in progress to elucitate the relationship between the original Hirst-test and the phenomenon described. The descripted observation on antagonismus have to be continued by research with other viruses and bacteriae. These kind of studies will be of theoretical and practical significance.