Blood group, serum protein and red cell enzyme groups of Amerindian populations in Colombia

Red cell samples from persons belonging to four Amerindian linguistic groups in Colombia were investigated for genetic variants in eight blood group systems: for three of the groups investigations were extended to ten red cell enzyme and four serum protein systems. The groups studied are the Noanama (including six Empera) of the Rio Siquirisua and Rio Docampado on the Pacific lowlands and the Cofan, Ingano and Siona Indians of the Upper Rio Putumayo and its tributaries to the east of the Andes. Only blood group O was present among two of the groups and the same groups were 100% Kp(b +), k in the Kell system. Di(a +) frequencies were high in three groups and there was marked variation between groups for the MNS, Rh, P, Lewis and Duffy systems. Polymorphism in all the three linguistic groups studied for serum proteins and red cell enzymes was present only in the red cell acid phosphatase, phosphoglucomutase (locus-1) and haptoglobin systems. 6-phosphogluconate dehydrogenase was polymorphic in the Noanama, and caeruloplasmin was polymorphic in the Ingano linguistic group. In addition two persons belonging to the Cofan linguistic group revealed the presence of an “atypical” component in the lactate dehydrogenase system. No variation was found in the other six red cell enzyme and two serum protein systems. Comparison with published data on red cell enzyme and serum protein groups for other South American Amerindian populations shows the Colombian populations studied here most closely resemble the Cayapo of Brazil.     

Identification, characterization and manipulation of Babesia-bovis-infected red blood cells using microfluidics technology

Nowadays numerous microfluidic systems are being developed to address a variety of clinical problems. Latest advances in microfluidic technology are promising to revolutionize the detection of pathogens in vivo through the development of integrated lab-on-chip devices. Such microfabricated systems will undertake all steps in sample analysis from collection and preparation to molecular detection. Micro total analysis systems are suitable candidates for point of care diagnostics due to small size, low cost production and enabled portability. The work here presented aimed the use of microfluidic platforms to identify and manipulate bovine red blood cells infected by the protozoan parasite Babesia bovis. A microfabricated device based on impedance spectroscopy was used for single cell discrimination and its sensitivity and applicability as a diagnostic method for bovine babesiosis was studied. Furthermore, manipulation and sorting of normal and infected red blood cells was performed on a dielectrophoresis based microfabricated cell cytometer. Single cell analysis of normal and B. bovis infected red blood cells was performed by electrorotation and dielectric parameters such as permittivities and conductivities of the cellular membrane and cytoplasm were determined.     

In vivo survival of selected murine carrier red blood cells after separation by density gradients or aqueous polymer two-phase systems.

In order to explore possibilities of using erythrocytes as carrier systems for delivery of pharmacological agents, we have studied the in vivo survival of murine carrier red blood cell populations enriched in young or old cells. Hypotonic-isotonic dialysis has been used to modify the cells as carrier systems and Percoll/albumin density gradients or counter-current distribution in aqueous polymer two-phase systems to separate them according to age. Hypotonic-isotonic dialysis produces a decrease in the red blood cell populations in vivo survival rate (from 9.5 to 7.8 days). Among the cells modified as carriers, the enriched young red blood cell populations show a higher in vivo survival (half-life 6.5-7.4 days) than populations made up of predominantly old red blood cells (half-life 4.7-6.2 days). Half-life of young or old circulating red blood cells was approximately one day longer when these cells were separated by counter-current distribution rather than by Percoll density gradients. Based on these results, hypotonic-isotonic dialysis of whole and enriched young or old red blood cell populations, with higher or lower survival rates, can be considered as a useful tool for modification of these cells as carriers. The final outcome of such changes can be translated into better control of plasma drug delivery during therapy.     

Methylene blue-mediated hexose monophosphate shunt stimulation in human red blood cells in vitro: independence from intracellular oxidative injury

The red blood cell hexose monophosphate shunt (HMS) and proteolytic responses to several concentrations of Methylene Blue or sodium nitrite were measured. The results suggested two distinct mechanisms for activation of the HMS: (1) nitrite treatment increased HMS activity in response to oxidative challenge to red cell protein; (2) Methylene Blue treatment activated HMS without injurious oxidative challenge. Nitrite-treated cells actively degraded protein, whereas Methylene Blue-treated red cells did not activate proteolytic systems that degrade oxidized red cell protein. These observations are relevant to proposed in vitro systems for evaluation of drug hemolytic toxicity potential on the basis of HMS stimulation capacity.     

Cation and ATP content of ferret red cells

Ferret red cells were shown to have the following properties: 1. They have a high sodium (96 mmol/l cell) and low potassium (3.9 mmol/l cell) content. 2. The majority do not appear to have an active sodium pump in their membranes. 3. Their membranes are highly permeable to rubidium indicating that they are probably also highly permeable to potassium. 4. Their magnesium (3.01 mmol/l cell) and calcium (0.01 mmol/l cell) contents are similar to those of red cells from other species. 5. Their ATP content (0.6 mmol/l cell) is similar to that of cat and dog red cells and is sufficiently high to activate known ion transport systems.     

Occurrence of lipid receptors inferred from brain and erythrocyte spectrins binding NaOH-extracted and protease-treated neuronal and erythrocyte membranes

It was previously shown in model systems that brain spectrin binds membrane phospholipids. In the present study, we analysed binding of isolated brain spectrin and red blood cell spectrin to red blood or neuronal membranes which had been treated as follows: (1). extracted with low ionic-strength solution, (2). the above membranes extracted with 0.1 M NaOH, and (3). membranes treated as above, followed by protease treatment and re-extraction with 0.1 M NaOH. It was found that isolated, NaOH-extracted, protease-treated neuronal and red blood cell membranes bind brain and red blood cell spectrin with moderate affinities similar to those obtained in model phospholipid membrane-spectrin interaction experiments. Moreover, this binding was competitively inhibited by liposomes prepared from membrane lipids. The presented results indicate the occurrence of receptor sites for spectrins that are extraction- and protease-resistant, therefore most probably of lipidic nature, in native membranes.     

Some hereditary blood factors of the Bengali Muslim of Bangladesh (red cell enzymes, haemoglobins, and serum proteins).

In a sample of Bengali Muslems from Dacca, haptoglobin, group-specific component, haemoglobin, adenosine deaminase, adenylate kinase, 6-phosphogluconate dehydrogenase, phosphoglucomutase, acid phosphatase and several other red cell enzyme types were studied. For most serum protein and red cell enzyme systems the gene frequencies are similar to those in other populations to the west of Bangladesh, but others suggest affinity with populations to the east.     

Measurement of biophysical properties of red blood cells by resistive pulse spectroscopy: volume, shape, surface area, and deformability

This paper presents a simple, new approach to the determination of size, shape, surface area, and deformability information for cells, notably red blood cells. The results are obtained by combining experimental measurements from resistive pulse spectroscopy (an extension of electronic cell-sizing methodology) with theoretical calculations for model cell systems. Assuming constancy of surface area and approximating red cell shapes by both prolate and oblate ellipsoids of revolution, values are determined for cell shape factor and volume under a variety of conditions. For red blood cells under low-stress conditions, shape factor, volume, and surface area results are found to be consistent with those available from the literature, when the oblate model is used. The applicability of this approach for determination of red cell properties under altered conditions is demonstrated by results for cell volume, at varying osmotic pressure and mechanical shear (tensile) stress. By quantitating the change in cell shape with stress, a new numerical scale for measuring cell deformability is also obtained, and data are presented on its variation for red cells at different osmolalities, over the range of 140 to 500 mOsm.     

Red cell antigen, serum protein and red cell enzyme polymorphisms in Karkar Islanders and inhabitants of the adjacent North Coast of New Guinea

Blood samples from the Waskia and Takia populations of Karkar Island, Papua New Guinea, and other nearby mainland populations, were tested for genetic variation in blood group, serum protein and red cell enzyme systems. Polymorphic variation was present in the ABO, P, MNS, Rh, Lewis, Duffy, Kidd and Gerbich blood group systems, in the Hp and Tf serum protein systems, and in the acid phosphatase, 6-PGD, ADA, PGM, MDH, and G-6-PD enzyme systems. A small number of variants was found in other systems: there were 4 Lu(a+), 1 Kp(a+), 2 C variants in the acid phosphatase system, 6 LDH variants, 1 ADA3-1 and 1 AK2-1 sample. All samples were negative for the red cell antigens Cw, Vw, He, K, Jsa, Dia, Wra, Rd and Marriott, and no variation was observed in the PHI enzyme system. The results are discussed in relation to those obtained on other Papua New Guinea populations.     

A zone phenomenon and a progressive reaction occurring with a radioactive hemolysin,sodium erucate,I 131

Sodium erucate reacts progressively (i.e., once the reaction is started in a time which is so short that the lysin is in contact with the red cells for 30 seconds, it cannot be stopped even by being diluted 10-fold) with human red cells at pH 7. At the same time, systems containing the lysin and human red cells show a zone phenomenon, lysis occurring most readily in a certain concentration of lysin but more slowly in larger or smaller concentrations. Sodium erucate-I¹³¹ can be used to investigate both the zone phenomenon and the progressive character of the reaction. As regards the former, large concentrations of the lysin react relatively poorly with the red cell surfaces and the resistance of the red cells is relatively high. This may be due to the presence of an admixed inhibitor or to the development of an inhibitory state. The lysin is taken up and fixed by material in the red cell surface, so that the "internal phase" of lysin attached to the cell surfaces is so firmly fixed that a 10-fold dilution has no effect on it. It follows that lysis in these systems is progressive, as it is found to be.     

Effects of buffers and pH on rabbit red blood cell hexokinase

The kinetic properties of rabbit red blood cell hexokinase in different buffer systems have been studied. At pH 8.0 the reaction velocity (v) is about 30% higher in glycylglycine compared to Tris, Tea, Hepes or ammonium acetate buffers. The enzyme stability, heat-dependence and spectral properties of the enzyme are also affected by the buffer utilized. None of the following kinetic properties of red blood cell hexokinase varies with pH in the range 6.8-8.5: Km of glucose; Km of ATP and Ki of glucose 6-phosphate.     

A calcium-activated potassium channel present in foetal red cells of the sheep but absent from reticulocytes and mature red cells.

Red cells of adult sheep, like those of other ruminants, lack the calcium-activated potassium channel which is present in the membrane of human red cells. Since the activities of other transport systems in the sheep red cell are known to decrease during maturation of the cell or during development of the animal it was investigated whether the K+ channel is present in red cells from younger animals or in reticulocytes. Using the divalent cation ionophore A23187 to increase the intracellular Ca of intact cells, it was found that the K+-selective channel is present in foetal red cells from the foetus or newborn animal but not in reticulocytes. The presence of the channel showed no dependence on the K+ genotype of the sheep and was not associated with either "high K+"- or "low K+"-type Na+ pump. No Ca2+-dependent change in K+ permeability was found in red cells from either newborn or adult donkeys suggesting that its presence in the red cells of the foetus may not be general. The role of the K+ channel in the mammalian red cell and the relationship between the K+ channel and the Na+ pump are discussed.     

Future needs and expected trends in peripheral blood cell analysis: erythrocyte histograms

Erythrocyte histograms should be analysed in accordance with the ICSH recommended protocol to determine: How many red cell populations are present and the proportion of the total red cells which are included in each population. The central tendency (mode, median & mean) and dispersion (size ratio, geometric standard deviation & coefficient of variation) for each population. The proportion of microcytic or macrocytic cells in each population. Examples are given to show how the patient's diagnosis is assisted by careful analysis of the histograms and the effect of treatment monitored by sequential testing. Histograms can be generated from blood films or from aperture-impedance and light-scatter volume measurements. Volume measurements on these systems are influenced by the red cell shape and internal refractive index. Mean cell hemoglobin concentration affects both shape in the aperture-impedance orifice and internal refractive index in light-scatter systems. There is poor agreement between volume histograms obtained on the two measuring systems and the histograms available on current automated instruments provide no more useful information than can be obtained from the blood film. Automated instruments need to produce histograms with fewer artefacts and the histograms should then be examined in accordance with the ICSH protocol. This approach should maximise the diagnostic value of the complete blood count.     

Photocontrol of Growth, and of Anthocyanin and Chlorogenic Acid Production in Cultured Callus Tissues of Haplopappus gracilis

Cultures of dark-grown Haplopappus callus (strain AI) were exposedto continuous blue, green, red, far-red, and white light for33 days at energy levels of approximately 10 J m^-2s^-1. Growthwas suppressed in all but far-red. Blue had the greatest suppressiveeffect, green the least; red and white were about equally effective.Mean cell generation times were increased from 8–8 days(dark control) to 12.5 days in red light and 20.5 days in blue.There was a slight increase in mean wet weight per cell in bluelight but a slight decrease in red, whereas there was almosta twofold increase in mean dry weight per cell in blue and littlechange in red. In contrast, far-red stimulated growth; the meancell generation time was reduced to 6–5 days and therewas little change in wet or dry weight per cell. Anthocyanin synthesis was promoted by all wavebands except far-red.Blue had the greatest effect, then white, red, and green inthat order. In blue light the pigments accumulated rapidly,but only during the early stages of culture. The maximum amountper cell was attained after 7 days and thereafter the valuesdeclined. In red, however, the pigments accumulated relativelyslowly, and the maximum cell content was not attained until22 days; the amount attained was less than half that attainedin blue light. Initially, the ratio of cyanidine-3-glucosideto cyanidine-3-rutinoside exceeded 5.0 in blue light, but theratio fell to almost unity with time. This probably reflecteda rapid initial synthesis of the glucoside accompanied by asteady conversion to the rutinoside. Blue light was also more effective than red in acceleratingchlorogenic acid production. The response to blue light occurredafter the initial rise in anthocyanins and continued for therest of the culture period. The data are discussed in relation to similar high-energy photoresponsesreported for intact systems.     

Genetic studies of blood markers in Przewalski's horses

Ninety-six Przewalski's horses (Equus przewalskii) were blood typed using systems of inherited blood variants known to be highly effective for parentage testing of domestic horses (E. caballus). Sixteen red cell antigenic factors detected using sera prepared by alloimmunization of domestic horses were shown to be inherited in six systems (A, C, D, P, Q, and U) and in the same patterns as domestic horses. Family data confirmed autosomal, codominant inheritance at five loci of serum protein variants (Al, Tf, Xk, Pi, and Es) and three loci of red cell proteins (PGM, PHI, and Hb). One serum protein locus (Gc) and two red cell protein loci (PGD and CA) appeared to be monomorphic. Despite the narrow genetic base and high inbreeding coefficients of captive Przewalski's horses, average heterozygosity calculated over 18 loci was estimated to be 0.320 +/- 0.05, which was similar to that found in five breeds of domestic horses.     

Carbonic anhydrase histochemistry: Evidence for non-enzymatic reaction and artifact production

Summary The fine structural localization of histochemically demonstrable carbonic anhydrase (CA) has been studied in tissues of the rat with particular regard to the red blood cell and associated reactions. A variety of fixation methods, including immersion in warm solution for periods up to one week, were used. All methods showed a precipitate along the red cell plasma membrane, with less reaction inside the cell. Secondly, the pinocytotic vesicles of capillary endothelium, pericytes, and smooth muscle cells accumulated precipitate; this staining was independent of neighboring red cell activity. A third finding was filling of the extracellular space, accessible pinocytotic vesicles, and mesaxons of small nerves by dense material. None of these staining patterns appear to be mediated by enzyme activity; the first may represent CA localization via direct staining, but the others are probably artifacts typical of many systems which employ a simple heavy metal salt.     

Effect of surface modification of rat erythrocytes of different ages on their partitioning behavior in charge-sensitive two-polymer aqueous phases

Partitioning differences between cells in two-polymer aqueous phase systems originate from subtle differences between the surface properties of cells. Because of the exponential relation between the parameters affecting the partition ratio (P) and the P itself, differences in membrane components suspected of effecting the differential partitioning of closely related cell populations cannot be directly established by conventional chemical assay techniques. In order to study the chemical nature of the components responsible for the age-related changes in surface properties of rat red cells we have devised an approach which uses a combination of isotopic labeling of erythrocyte subpopulations of distinct cell age with different enzyme and/or chemical treatments followed by countercurrent distribution in charge-sensitive two-polymer aqueous phase systems. These studies show that: neuraminidase-susceptible sialic acid is not responsible for the cell age-related surface differences detected by partitioning; the component(s) responsible for the cell age-related surface differences can be extracted (from aldehyde-fixed red cells) with ethanol or cleaved with dilute sulfuric acid. Our data are consistent with the hypothesis that ganglioside-linked sialic acid is the chemical moiety responsible for the cell charge-associated surface differences among rat red blood cells of different ages.     

Population study of electrophoretic polymorphisms of red cell enzymes and plasma proteins in Caucasian Canadians

Blood samples from a random series of Canadian Caucasians were phenotyped for 28 red cell enzyme systems and eight plasma protein systems. Polymorphism was found in 17 and rare variants in 11 of the systems. Allele frequencies are presented for these; distribution of phenotypes is in accordance with the Hardy--Weinberg equilibrium theory. In a complementary study of families there was no evidence of de novo mutation in any of the 36 systems and they allowed a minimum estimate of the frequency of null alleles in the ADA, C2, and GPT systems.     

Contribution of hydrogen peroxide to the cytotoxicity of green tea and red wines

Green tea and red wine are claimed to have health benefits because of their high content of polyphenolic compounds, but they have also been reported as mutagenic in some test systems. In this paper, we show that a commonly used cell culture medium, Dulbecco's modified Eagle's medium (DMEM), catalyses oxidation of green tea and red wines to generate H(2)O(2). The level of H(2)O(2) produced from green tea accounted for all of the cytotoxic effects of this beverage on PCl2 cells. By contrast, H(2)O(2) was only responsible for part of the cytotoxicity of the red wines examined. Our data illustrate the danger of extrapolating from cell culture studies to predict the effects of complex beverages in vivo.     

The use of an agglutination micromethod for erythrocyte phenotyping. Importance of improved blood transfusion safety

The authors describe an easy, sensitive and inexpensive micromethod for red blood cell typing within the Rhesus, Kell and Lewis systems.