Erythrocytes Of Humans Research Articles

Effects of diet and folate on levels of micronucleated polychromatic erythrocytes

We describe a study examining the effects of fried beef consumption on the frequency of micronuclei in RNA-positive polychromatic erythrocytes (PCEs) in humans. 5 splenectomized individuals participated in a 2-week experiment consisting of 3 phases. During the first and last phases the subjects refrained from eating cooked meat for 4–5 days. This was designed to clear their system of mutagens found in cooked-meat products. In the second phase each person ate a similar diet, but also consumed 4 quarter-pound well-done hamburgers per day for 4 days. Erythrocyte-folate levels were also measured for each donor. No association between diet phase and micronucleus frequencies was observed among the 2 subjects with normal levels of folate. However, among the 3 low-folate donors, the frequency of micronucleated PCEs appeared to be associated with diet. Micronucleus frequencies began to increase 1 day following onset of the beef phase, and started to decline 1 day after finishing this phase. These observations are in agreement with erythrocyte maturation kinetics following short-term exposure. A repeat study performed on one of the low-folate donors consisted of two beef phases separated by a vegetarian phase. Beef in one phase was fried (high in mutagenic amino-imidazoazaarenes [AIAs]) while the beef in the other phase was fried after first being microwave-cooked (low in AIAs). Significant increases in the micronucleus frequency were not observed in this experiment, suggesting that AIAs formed during frying were not solely responsible for the induction of micronuclei

Memory performance of young and old subjects related to their erythrocyte characteristics

Several papers reported in recent years on a change in the age population distribution of the circulating erythrocytes in old mice, rats, rabbits, and humans. The results indicate the presence of a chronologically younger cell population in old animals and humans. The cells are typically lower in density and larger. In some reports, the cells have higher levels of enzymatic activity. We wanted to know whether changes in the characteristics of the circulating erythrocytes in old people are related to the changes in cognitive performance often observed in the elderly. Twenty young (20–40) and 21 old (70–90) volunteers submitted to memory and blood tests. Density and size distribution, aspartate aminotransferase/glutamic oxalacetic transaminase (AST/GOT) activity, and the level of glycosylated hemoglobin (HbA 1) of erythrocytes were determined. The Wechsler Memory Scale—Revised (Wechsler, 1987) was used to determine general memory and delayed recall scores for each subject. We have confirmed that old subjects have larger and less dense cells. Erythrocyte volume was the only blood parameter examined that revealed statistically significant correlations with memory performance. The old subjects with no age-related memory impairment had significantly smaller cells than the other old subjects.

Stabilization of the adenylate energy charge in erythrocytes of rats and humans at high altitude hypoxia

1. 1. Stabilization of adenylate energy charge and control of adenylate pool were analysed in the erythrocytes of the rat and the human exposed to highly hypoxic conditions. 2. 2. Red cell energy charge was decreased in the rats exposed to a simulated altitude of 5000–8000 m, and then recovered to the normal value with the depletion of adenylate pool. 3. 3. The energy charge and the adenylate pool size of the human erythrocytes did not show any change under highly hypoxic conditions. 4. 4. Anaerobic incubation of rat erythrocytes caused a marked decrease in the energy charge, and its recovery was accompanied by the depletion of total adenylates. 5. 5. The energy charge and total adenylates of human red cells did not change under the anaerobic incubation of erythrocytes. 6. 6. These results suggest that the energy charge of rat erythrocytes can be controlled by depletion of the adenylate pool, but the adenylate degradation is not responsible for the stabilization of the energy charge in human erythrocytes.

Red blood cell glucose metabolism in human chronic fluoride toxicity

Fluoride is a well known inhibitor of many enzyme systems in vitro. The most widely studied classic example of fluoride inhibition is its potent inhibition of glycolysis, specifically its action on the enzyme enolase. Despite the plethora of in vitro studies on the effects of fluoride on the enzyme activity, there is a paucity of information concerning the in vivo metabolic lesions caused by the chronic toxic doses of fluoride in humans. The present study has been undertaken with a view to assess the changes in glucose metabolism and related enzymes in erythrocytes of humans consuming toxic doses of fluoride for prolonged periods.

Studies on the metabolic fate of azuletil sodium. V: Protein binding and uptake by erythrocytes.

The binding to various proteins and the uptake by erythrocytes of 14C-KT1 -32 and its major metabolites (14C-M1, 14C-M2) were studied. The binding of 14C-KT1-32, 14C-M1 and 14C-M2 to total plasma, albumin, α1-acid glycoprotein(AAG) and γ-globulins were determined in humans and various animals in vitro, ex vivo and in vivo. The uptakes by erythrocytes were determined in humans, rats and dogs in vitro or in vivo.1. The binding of 14C-KT1-32, 14C-M1 and 14C-M2 to plasma and albumin were greater than 97%. AAG and γ-globulins bindings of 14C-KT1-32, 14C-M1 and 14C-M2 were less than 15%.2. Protein binding of 14C-KT1-32 to rat, dog and human plasma was characterized as reversible.3. Protein binding of 14C-KT1-32 to human plasma was not affected by the additions of cimetidine, ranitidine, famotidine and bromazepam. Protein binding of 3H-cimetidine, ranitidine, famotidine and bromazepam to human plasma was also not affected by the addition of KT1-32.4. The uptakes of 14C-KT1-32 by erythrocytes of humans and various animals were less than 41% in vivo or in vitro.

Age population distribution of erythrocytes in young and old healthy donors

Reports from several laboratories on a shorter life span of erythrocytes (E) in old animals and humans, induced the authors to search for a simple method for determining the younger age distribution of E in the blood of 20 old (over 70), as compared to 20 young (below 40), healthy donors. The following tests were performed: 1) Density Distribution of Cells (DDC), 2) Osmotic Fragility, 3) Agglutinability of E by Poly-L-lysine, 4) Analysis of Aspartate Amino Transferase (AST) activity, 5) Test for the presence of immunoglobulin on the surface of E (rosette formation on K562 cells); and 6) All the usual clinical and hematological tests were performed in order to avoid pathology. The most significant difference between the blood of the young and the old was found in the DDC. The shift of the cumulative curve indicated a younger population of cells in the blood of the elederly. The activity of AST was higher in the blood of the elderly, also indicating a younger cell population. The rosette formation was higher with the E from the blood of the elderly, indicating that the E, had more immunoglobulins on their surface than the E from the blood of the younger donors.

Different sensitivities of rat and human red cells to exogenous Ca2+

During an examination of the effects of shear and of the Ca2+ ionophore A23187 on Ca2+ entry into erythrocytes of rats and humans, we noted that rat erythrocytes were much more sensitive to Ca2+-induced hemolysis than the human cells. An examination of the effect of Ca2+ on transglutaminase, a cytosolic enzyme in the erythrocyte which cross-links membrane proteins and renders cells less deformable, demonstrated a correlation between enzyme activity and Ca2+-induced hemolysis. Both rat and human cells subjected to shear-induced Ca2+ entry exhibited increased enzyme activity and altered membrane protein SDS-PAGE patterns. Twenty micromolar A23187 with Ca2+ at concentrations above 80 microM caused hemolysis of rat erythrocytes. In contrast to human erythrocytes, under these conditions no membranes were recoverable from rat erythrocytes. At lower concentrations of Ca2+ (25 and 50 microM), however, rat erythrocytes maintained integrity, and exhibited enhanced transglutaminase activity and cross-linking of membrane proteins. The rat enzyme can be activated 30% by 10 microM Ca2+, while 50 microM Ca2+ was necessary to achieve a similar activation of the enzyme from human red blood cells. In studies of shear-stimulated Ca2+ uptake by erythrocytes the rat red cell enzyme was more readily activated. The SDS-PAGE pattern of rat red cell membranes after a 30 sec shear showed specific changes in protein banding, including the appearance of bands greater than 330 kDa. Changes in protein banding were also apparent in cytosolic proteins. This work supports the view that shear-induced Ca2+ entry activates transglutaminase that leads to cross-linking of membrane components, a loss of cell integrity, and eventual cell death.

Identification of glucose and nucleoside transport proteins in neonatal pig erythrocytes using monoclonal antibodies against band 4.5 polypeptides of adult human and pig erythrocytes.

Cytochalasin B and nitrobenzylthioinosine (NBMPR), which inhibit membrane transport of glucose and nucleosides, respectively, have served as photoaffinity ligands that become covalently linked at inhibitor binding sites on transporter-associated proteins. Thus, when membranes from erythrocytes of neonatal pigs with site-bound [3H]cytochalasin B or [3H]NBMPR were irradiated with uv light, two labeled membrane polypeptides (peak Mr values: 55,000 and 64,000, respectively) were identified. Treatment of the photolabeled membranes with endoglycosidase F increased the mobility of [3H]cytochalasin B- and [3H]NBMPR-labeled material (peak Mr values: 44,000 and 57,000, respectively) and limited digestion with trypsin yielded different polypeptide fragments (Mr values: 18,000-23,000 and 43,000, respectively). Identification of the photolabeled polypeptides as transporter components was established using monoclonal antibodies (MAbs) raised against partially purified preparations of band 4.5 from erythrocytes of adult pigs and humans. MAbs 65D4 and 64C7 (anti-human band 4.5), raised in this study, reacted with [3H]cytochalasin B-labeled material from membranes of human erythrocytes and bound to permeabilized erythrocytes but not to intact cells. MAb 65D4 also bound to erythrocytes of mice and neonatal pigs and to a variety of cultured cells (mouse, human, rat), including AE1 mouse lymphoma cells, which lack an NBMPR-sensitive nucleoside transporter. Also employed was MAb 11C4 (anti-pig band 4.5), which recognizes the NBMPR-binding protein of erythrocyte membranes from adult pigs. When membrane proteins from neonatal and adult pigs were subjected to electrophoretic analysis and blots were probed with different MAbs, MAb 65D4 (anti-human band 4.5) bound to material that comigrated with [3H]cytochalasin B-labeled polypeptides (band 4.5) from neonatal, but not adult, pig erythrocytes, whereas MAb 11C4 (anti-pig band 4.5) bound to material that comigrated with [3H]NBMPR-labeled band 4.5 polypeptides of erythrocytes from both neonatal and adult pigs. These results, which indicate structural differences in the cytochalasin B- and NBMPR-binding proteins of pig erythrocytes, establish the presence of both proteins in erythrocytes of neonatal pigs and suggest that only the NBMPR-binding protein is present in erythrocytes of adult pigs.

Electron spin resonance studies of erythrocytes from spontaneously hypertensive rats and humans with essential hypertension.

The purpose of the present study was to investigate erythrocyte membrane abnormalities in hypertension by means of an electron spin resonance and spin-label technique. The erythrocytes from spontaneously hypertensive rats (SHR) and humans with untreated essential hypertension were examined and compared with their normotensive counterparts, and electron spin resonance spectra were obtained for a fatty spin-label agent (5-nitroxy stearate) incorporated into the erythrocyte membranes. The value of outer hyperfine splitting (2T' parallel) was significantly higher in erythrocytes of SHR and humans with essential hypertension than in erythrocytes of normotensive controls (at 37 degrees C: SHR, 56.14 +/- 0.51 gauss [G], n = 8; Wistar-Kyoto rats, 52.22 +/- 0.86 G, n = 4, p less than 0.01; humans with essential hypertension, 56.94 +/- 0.27 G, n = 11; normotensive subjects, 55.44 +/- 0.36 G, n = 8, p less than 0.01). The order parameter (S) was also increased in the hypertensive rats and humans compared to their respective normotensive controls. When calcium was loaded to erythrocytes with calcium ionophore A23187 (0.9 microM) and CaCl2 (1.0 mM), the parameters of the spectra were increased. These changes were more prominent in the hypertensive groups than in the normotensive controls. These results revealed that the erythrocyte membranes of the hypertensive subjects tolerated different spin motions than those of the normotensive controls in the electron spin resonance study and that membrane fluidity might be decreased in hypertension. Additionally, calcium loading to erythrocytes caused the reduction of membrane fluidity. Therefore, it is suggested that an abnormality of calcium handling at the cellular level might affect physical properties of the biomembranes in hypertension.

Cationic concentrations and transmembrane fluxes in erythrocytes of humans during exercise.

The effect of exercise on the intraerythrocyte cationic concentrations and transmembrane fluxes such as the Na+-K+-adenosinetriphosphatase (ATPase) pump, the Na+-K+ cotransport, and the Na+-Li+ countertransport system was studied in 11 normal male volunteers. All subjects performed an uninterrupted incremental exercise test on a bicycle ergometer, starting at an initial work load of 20% of the subjects' maximal exercise capacity, as determined in a pretest. The work rate was increased with an additional 20% each 6 min up to a final work load of 80%. Blood samples were taken at rest, at 60 and 80% of maximal exercise capacity, and 1, 2, 3, 4, 5, and 30 min after cessation of exercise. At moderate exercise (60% of maximal exercise capacity) the intraerythrocyte potassium concentration was not changed, but at severe exercise (80% of maximal exercise capacity) it was decreased. After exercise the intraerythrocyte potassium concentration returned to base line within 2 min. Exercise did not affect the intraerythrocyte concentrations of sodium and magnesium. The activity of the Na+-K+-ATPase pump and the Na+-K+ cotransport in the erythrocytes during and after exercise was no different from the resting level. The activity of the Na+-Li+ countertransport system on the contrary tended to decrease during exercise. It is concluded that exercise is accompanied by a leakage of potassium out of the erythrocytes without major alterations in the active red cell cationic fluxes.

Prevention by antioxidants of the hemolysis of erythrocytes of cattle, pigs and humans treated with t-butyl hydroperoxide

1. Urate, 3-ribosylurate, ascorbate, glutathione and plasma protected bovine, porcine and human erythrocytes from hemolysis caused by t-butyl hydroperoxide (t-BHP). 2. Urate partially protected porcine erythrocytes from hemolysis by t-BHP when it was added 15 min after the addition of the t-BHP, but it did not protect when added 30 min after the t-BHP. 3. Glutathione and ascorbate protected oxyhemoglobin from oxidation to methemoglobin by t-BHP; 3-ribosylurate gave only slight protection. 4. Urate stimulated the formation of methemoglobin from oxyhemoglobin during treatment with t-BHP.

A study of zinc distribution in erythrocytes of normal humans.

The assignment of zinc bound to carbonic anhydrase isoenzymes (CA-I and CA-II) and Cu2 Zn2 superoxide dismutase (SOD1) was investigated in the hemolysates from 21 normal male subjects. Sufficient care was taken to remove leukocytes and platelets. The following values of zinc distribution were obtained: total zinc, 113.8 +/- 22.7 (mean +/- S.E.) micrograms X 100 ml-1; CA-I-derived zinc, 866.6 +/- 26.2 micrograms X 100 ml-1; CA-II-derived zinc, 99.9 +/- 3.9 micrograms X 100 m-1; SOD1-derived zinc, 60.3 +/- 1.9 micrograms X 100 ml-1; the other zinc, 87.0 +/- 12.6 micrograms X 100 ml-1. Namely, 7.6% of the zinc in human erythrocytes is not bound to the carbonic anhydrases and Cu2Zn2 superoxide dismutase, but present in available form or attached to other enzymes.

Variation in erythrocyte enzyme activity among primate species

1. 1. Significant differences between species in the mean level of enzyme activity, ranging from 50 to 400%, were detected during a study of 10 enzymes from erythrocytes of humans and five non-human primates. 2. 2. The activity differences were consistent with variation, between species, at the structural loci for specific enzymes rather than differences in mean cell age, hemoglobin content or other pleiotropic effects. 3. 3. For seven of the eight loci at which direct comparisons among the six species could be made, the number of species-specific activity differences was greater than the number of variants observed by electrophoresis. 4. 4. Thus not only do activity measurements detect previously “hidden” variation among species but they also provide information regarding genetic differences which may be of potential physiological and/or selective relevance.

The effect of ascorbic acid on nitrite-induced methemoglobin formation in rats, sheep, and normal human erythrocytes

Interspecies differences were demonstrated with respect to the occurrence of sodium nitrite-induced methemoglobin (METHB) in the erythrocytes of rats, dorset sheep, and normal humans, with the rats displaying approximately 25–33% of the sensitivity of sheep and human erythrocytes. Ascorbic acid incubation along with the nitrite was able to significantly reduce METHB formation in a dose-dependent manner in both rats and humans but not in sheep.

The effect of ascorbic acid on copper-induced oxidative changes in the erythrocytes of rats, sheep, and normal humans

Rats, sheep, and normal humans displayed a comparable sensitivity to copper acetate (3 m M)-induced changes in reduced glutathione (GSH) levels it vitro. However, the humans erythrocytes were more sensitive than either animal to methemoglobin (METHB) formation with the rat being least sensitive. Ascorbic acid incubation markedly enhanced the occurrence of copper acetate-induced increases in METHB and decreases in GSH in the sheep and humans. However, ascorbic acid incubation reduced the occurrence of copper acetate-induced increases in METHB, while not effecting changes in GSH in rats.

The Chemical Removal of Embryophoric Blocks from Eggs of Taenia ovis and Taenia hydatigena Prior to In vitro Cultivation

stimulation of SOD activity by an as yet unknown mechanism. Because we found the difference in reticulocytosis between infected (2.7%) and uninfected mice (0.4%) to be small, and because the SOD activity in the infected mice showed no correlation with the reticulocyte level, the observed differences in SOD activity between the red blood cells from the two groups of mice should not arise from the reticulocytes alone. No variation in SOD level was observed in the young and mature erythrocytes in humans (Michelson, 1977. In Frontiers in physicochemical biology, B. Pullman (ed.). Academic Press, New York, p. 351). Finally, enzyme activity and protein-stained patterns of extracted SOD from red cells of uninfected and infected mice and SOD from isolated parasites appeared identical. The parasite may be able to use this enzyme for combating toxicity of oxygen or oxidant stress during the conversion of ingested hemoglobin (containing heme (Fe II)) to yield hemozoin pigment (containing hemin (Fe III)). Supertimulation of SOD activity by an as yet unnown mechanism. Becaus we found the diference in reticulocyt sis between inf cted .7%) and uninfected mice (0.4%) to be small, d because the SOD activity n he fect d oxide ca arise from the reduction of oxygen bound to hemoglobin during the oxidation of Fe (II) of heme to Fe (III) of hemin (Rotilio et al., 1976. In Superoxide and superoxide dismu ase, J. M. McCord and I. Fridovich (eds.). Academic Press, New York, pp. 239-244). Disulfiram can potentiate oxygen toxicity in rats by interacting with SOD (Forman et al., 1976, J. Pharmacol. Exp. Therapeut. 212: 452-455). Perhaps the antimalarial action of a bioconverted product of disulfiram, diethyldithiocarbamate, on P. falciparum is related to its inhibition of the parasite's SOD activity (Scheibel et al., 1979, Proc. Natl. Acad. Sci. USA 76: 5303-5307). We found that diethyldithiocarbamate at concentration as low as 0.2 mM inhibited the SOD extracted from the isolated P. berghei and also that from infected red blood cells from 20 mice. We thank UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (Project no. 780001) for financial support.

Variance in the enzymatic properties of the chloride-activated arginine aminopeptidase from erythrocytes of healthy humans

The use of a strictly identical procedure for the purification of the human erythrocyte Cl −-dependent arginine aminopeptidase (EC 3.4.11.6) separately from 10 healthy adults resulted in enzymes which showed considerable variance in substrate specificity, values of the inhibition constant of p-chloromercuribenzoate, reaction velocities, and degree of substrate inhibition, although the purified enzymes were immunologically identical. These differences were considered to reflect changes in enzyme conformation. The present data were regarded as new because other (polymorphic) peptidases of human erythrocytes previously described differ from the present Cl −-dependent arginine aminopeptidase by virtue of substrate specificity.

In vitro binding of Trypanosoma congolense to erythrocytes.

Trypanosoma congolense Broden, an intravascular parasite, binds to vessel walls and erythrocytes of infected hosts. In an attempt to characterize T. congolense adhesion to host cells, an in vitro assay was devised. It was shown in the in vitro experiments that T. congolense binds to bovine, sheep, and goat erythrocytes, but not always to erythrocytes of rats, mice, rabbits, horses or humans. Only the anterior part of live trypanosomes adheres to erythrocytes, and the attachment site on the trypanosomes is destroyed by trypsin and chymotrypsin-trypanosomes did not adhere to bovine erythrocytes that had been incubated with neuraminidase, sodium periodate and poly-L-lysine. The foregoing experiments suggest that the surface of T. congolense contains a protein-associated site which binds to sialic acid of some host cells. This surface site is most likely responsible for attachment to blood vessels in vivo.

The H, HI, I and i blood group antigens in butanol extracts of rabbit tissues.

Aqueous and butanol phase extracts of rabbit erythrocytes and the tissues gastric mucosa, liver, lung and kidney were examined by inhibition of agglutination for the presence of H, HI, I and i blood group substances. Of the four specificities, only I substance was recovered from erythrocytes and only in the aqueous phase extracts. Both aqueous and butanol extracts of gastric mucosae contained H substance, but HI and I were present only in the aqueous extracts of this tissue. Extracts of rabbit lungs only had H and I activity and only in the aqueous phase. No H, HI, I or i substance was detected in any of the extracts of liver and kidney. Reduced levels of H substance were found in aqueous extracts of gastric mucosae of 'A-like' rabbits. Three H specificities were demonstrated in the butanol phase extracts of gastric mucosae of rabbits. These specificities were similar to those previously shown in extracts of gastric mucosae and erythrocytes of humans.

Glycolytic intermediates and glutathione levels of erythrocyte: Comparisons between erythrocytes of humans and thoroughbred horses

1. 1. 2,3-DPG, ATP, ADP, AMP and GSH content of red cells of horses are compared with the levels in human red blood cells. 2. 2. Horses showed lower values for these intermediates in terms of numbers of erythrocytes. 3. 3. If the values are considered in terms of MCV and MCH, the amount of 2,3 DPG and GSH from horses are more than those from human cells whilst the adenosine phosphates remain at lower values.