Dog Erythrocytes Research Articles

Role of environment in the development of "natural" hemagglutinins in Minnesota miniature swine

"Natural" hemagglutinin titers against a panel of fixed erythrocyte antigens were determined for groups of Minnesota miniature swine reared conventionally, in a specific pathogen-free facility, and in germfree isolators. Sera were assayed for hemagglutination (HA) titers by the microtiter method against 12 species of erythrocytes stabilized by treatment with pyruvic aldehyde and formaldehyde. These erythrocytes were stable for up to 2 years and gave slightly enhanced HA titers as compared to fresh, unfixed erythrocytes. Of the sera from conventional swine tested, the highest "natural" HA titers were directed towards rabbit, cat, swine dog, and burro erythrocytes (greater than 1:1,000), intermediate titers were detected against human A, B, and O, and sheep, pig, and chicken erythrocytes (1:64 to 1:1,000), whereas the lowest titers were found against ox and goat erythrocytes (less than 1:8). Titers obtained with sera from specific pathogen-free swine were 2- to 16-fold lower than those of conventional swine, but were of a similar distribution with regard to the species of erythrocyte tested. Germfree swine sera uniformly exhibited HA titers less than 1:4 against all species of erythrocytes. The majority of these hemagglutinins were immunoglobulin M class but there were some agglutinins of immunoglobulin A class and a slight amount of immunoglobulin G class. Specificity of these agglutinins was examined by absorption tests. The results are consistent with the hypothesis that natural hemagglutinins develop due to dietary or microbial antigenic stimulation, or both.

Evidence for a magnesium- and ATP-dependent calcium extrusion pump in dog erythrocytes

Intact dog erythrocytes, whose Ca 2+ permeability had been increased with A23187 still maintained intracellular Ca 2+ below electrochemical equilibrium indicating that they could extrude Ca 2+. This extrusion required no Na + gradient but apparently depended on intracellular ATP and Mg 2+ suggesting that it was mediated by an ATP-fuelled Ca 2+ pump.

Rosette formation by buffalo (Bos bubalis) T and B lymphocytes

Buffalo (Bos bubalis) lymphocytes were purified and tested for their E and EAC rosette forming capacity as a marker for the detection of T and B cells, respectively. Sheep erythrocytes were found to form 17.7 per cent of E rosette with buffalo lymphocytes. This population of lymphocytes is believed to be T cell. Erythrocytes of guinea-pig, rabbit, hamster, rat, chicken, dog and donkey formed a lower percentage of rosettes. Five to 18.5 per cent of SRBC-EAC rosettes were detected with buffalo lymphocytes which are believed to be B cells.

Action of surface-active substances on biological membranes I. Effect of chemical modification of membranes on hemolysis of erythrocytes by sodium alkyl sulfates

The hemolytic action of a homologous series of sodium alkyl sulfates (C 8 toC 15) on dog and human erythrocytes was measured. The influence of trypsin, pronase and neuraminidase treatments on the lytic resistance of human red cells was studied. A new approach evaluating the hemolytic activity of ionic detergents is expressed as a ratio of a concentration providing 50% lysis to its CMC-value. The results obtained are examined for their bearing on the use of surface-active agents as a means to study the structure organization of biological membranes.

Metabolic fate of SCE-129, a new antipseudomonal cephalosporin, after parenteral administration in rats and dogs.

Disposition of [(14)C]SCE-129 was studied in rats and dogs after intramuscular (i.m.) or intravenous (i.v.) injection. In rats, the plasma level of i.m. [(14)C]SCE-129 attained a peak at 15 min after dosing and then declined with a half-life of 35 min, whereas the half-life after an i.v. dose was 26 min. The area under the plasma level curve within 2 h after the i.m. dose was 85% of that after the i.v. dose. Intramuscular injection of [(14)C]SCE-129 into dogs gave a peak plasma level at 30 min and a half-life of 60 min. In both rats and dogs, the plasma levels of (14)C were closely similar to those of the unchanged antibiotic, which was weakly bound to plasma protein. The rat tissue level of i.m. [(14)C]SCE-129 was maximum at 15 min, with the highest concentration in the kidney, followed by plasma, adrenal, lung, heart, thymus, gastrointestinal wall, and liver, and the lowest in the brain. The antibiotic barely entered erythrocytes of rats and dogs. Whole-body autora-diographic studies showed that i.m. [(14)C]SCE-129 scarcely crossed the rat placenta. (14)C was detected in the milk of rats given i.m. [(14)C]SCE-129. In both rats and dogs, almost all of the dosed (14)C was excreted in urine within 24 h as the unaltered antibiotic, with only small amounts appearing in feces via bile. Thus, these findings evidenced a rapid absorption and wide distribution of i.m. SCE-129, followed by extensive renal elimination as the unaltered antibiotic.

Inhibition of glycolysis by L-sorbose in dog erythrocytes.

We have demonstrated previously that in vitro L-sorbose acts directly on dog erythrocytes to induce hemolysis. Here we report that L-sorbose depresses lactate formation in dog hemolysates from glucose, mannose and fructose but not from glucose-6-phosphate and galactose, suggesting that L-sorbose interacts with glycolysis at the level of the hexokinase.

Ultrasonic energy backscattered from blood. An experimental determination of the variation of sound energy with hematocrit.

To determine how the ultrasonic energy scattered from flowing blood depends on the hematocrit, the rms amplitude of the audio Doppler signal from a range gated, pulsed Doppler ultrasonic flowmeter was measured for hematocrits ranging from 0.01 to 80%. For all investigated frequencies (5–27 MHz), the signal power, the square of the rms amplitude, was proportional to the hematocrit until 2%, reached a plateau at 10%, and was independent of the hematocrit over the rest of the investigated range. To determine how the scattered energy depends on the volume of the erythrocyte, we measured the variation of audio signal power with cell concentration for goat blood, and then without changing any of the instrument settings, repeated this measurement for dog blood. For cell concentrations ranging from 104 to 106 cells/mm3, the larger dog erythrocytes scattered 17 times more energy than the goat erythrocytes. This observation is consistent with the hypothesis that the energy scattered by a single erythrocyte is proportional to the square of the volume of the cell.

Difference in form of sialic acid in red blood cell glycolipids of different breeds of dogs.

Hematoside from dog erythrocyte membrane was previously considered to contain a mixture of N-acetyl- and N-glycolyl- neuraminic acids. However, the hematoside preparation used in the previous study was obtained from pooled blood of several dogs, and individual variation in hematoside was not examined. In this work, hematosides of erythrocytes from 31 mongrel dogs and 108 dogs of 23 breeds were examined individually by thin-layer chromatography, and the component sialic acids were analysed by gas-liquid chromatography. Individual dogs had either NAN-hematoside or NGN-hematoside: dogs with N-glycolyl-neuraminic acid also had a trace of N-acetyl-neuraminic acid, but dogs with N-acetyl-neuraminic acid had no detectable N-glycolyl-neuraminic acid. A few mongrel dogs, some Kai dogs, Kishu dogs, Japanese spaniels and most Shiba dogs had NGN-hematoside, whereas all European dogs had NAN-hematoside and no NGN-hematoside. From pedigrees of some families, inheritance of NGN-hematoside was found to be autosomal dominant. NGN-hematoside is possibly one of dog blood group substances. The sialic acid of delipidized ghost protein of dogs with NGN-hematoside was N-glycolyl-neuraminic acid, and that of dogs with NAN-hematoside was N-acetyl-neuraminic acid. The sialic acid of plasma protein was mainly N-acetyl-neuraminic acid in all dogs.

Permeability of erythrocytes to glycerol and its acylated derivatives in the camel and dog

1. 1. Glycerol and glyeerol derivatives transport was compared in human, dog and camel erythrocytes. 2. 2. The camel erythrocytes had the slowest glyeerol transport, that of the dog erythrocytes was also slow, while the quickest transport occurred in human erythrocytes. 3. 3. In glyeerol mono-, di- and triacetates the transport in camel and dog erythrocytes increased until in the triacetate the quickest transport was found. 4. 4. Addition of copper ions had no inhibiting effect on glyeerol transport in camel erythrocytes but a striking effect was seen in human erythrocytes. 5. 5. Changes in pH greatly affected transport in camel erythrocytes, a decline in pH enhanced and an increase in pH slowed the transport. Storage of camel erythrocytes did not affect the glyeerol transport in camel erythrocytes. 6. 6. It is concluded that the mechanism of glyeerol transport in camel erythrocytes is one of a non-facilitated diffusion.

A Waldenström macroglobulin that is both a cold agglutinin and a cryoglobulin because it binds N -acetylneuraminosyl residues

A purified human monoclonal IgM(kappa) (cold agglutinin MKV) has been characterized as both a cold agglutinin and a cryoglobulin. Since its reactivity with human erythrocytes but not with dog erythrocytes is reduced by treatment of the cells with ficin and its reactivity with both is abolished by treatment of the cells with neuraminidase, it has by definition Pr(2) specificity [Roelcke, D. (1974) Clin. Immunol. Immunopath. 2, 266-280]. Presumably, the membrane receptors for cold agglutinin MKV are sialic acid-containing glycoproteins in human cells and sialic acid-containing glycolipids in dog cells. Agglutination of erythrocytes is specifically inhibited by II(3)-N-acetylneuraminosyllactosylceramide (GM(3)) and N-acetylneuraminosylparagloboside but not by their N-glycolylneuraminosyl forms or by lactosylceramide or paragloboside, and the reactivity of neuraminidase-treated cells can be restored by allowing them to absorb either GM(3) or N-acetylneuraminosylparagloboside. When large amounts of ganglioside are absorbed, the cells are agglutinated not only in the cold, but also at 37 degrees , showing that the density of receptor sites on the erythrocyte surface can influence the thermal amplitude of cold agglutinins. Liberation of sialic acid from cold agglutinin MKV by treatment with neuraminidase (acylneuraminosyl hydrolase; EC 3.2.1.18) does not affect its agglutinating properties but the asialoprotein is no longer a cryoglobulin. Apparently the physical basis for its precipitation in the cold is the intermolecular immune binding of N-acetylneuraminosyl residues.

Haemolysis of dog erythrocytes by sorbose in vitro.

In vitro, L-sorbose induces in dog erythrocytes haemolysis which is dependent upon time of incubation, temperature, pH and the concentration of sorbose. The in vitro susceptibility to sorbose-induced haemolysis is different in various species.

Effect of galactose oxidase, with and without prior sialidase treatment, on the viability of erythrocytes in circulation.

Previous studies have shown that sialidase-treated mammalian erythrocytes were rapidly eliminated from circulation. In contrast, chicken asialoerythrocytes remained fully viable. This investigation was undertaken to ascertain the reason for this difference in behavior as well as to determine the extent of the similarity of the physiological mechanism for the elimination from circulation of asialoglycoproteins and mammalian asialoerythrocytes. To that end, erythrocytes from dogs, rabbits, and chickens were each subjected to the action of galactose oxidase (D-galactose:oxygen 6-oxidoreductase; EC 1.1.3.9) both before and after sialidase (acylneuraminyl hydrolase; EC 3.2.1.18) treatment. The viability of the autologously transfused erythrocytes in circulation was monitored by Na2-51CrO4 labeling. Galactose oxidase had no deleterious effect on the viability of dog or chicken erythrocytes, nor did it restore the viability of dog or rabbit asialoerythrocytes. On the other hand, desialated chicken erythrocytes, which were fully viable, were rendered nonviable upon treatment with galactose oxidase. It may be concluded therefore that (a) the physiological mechanism of elimination of mammalian asialoerythrocytes from circulation is not the same as that for plasma asialoglycoproteins and (b) the treatment of chicken asialoerythrocytes with galactose oxidase results in the oxidation at carbons 6 of the galactosyl- or N-acetylgalactosaminyl residues, thereby rendering the erythrocytes nonviable.

Binding of thyroid hormone by erythrocyte cytoplasmic proteins

Characteristics of iodothyronine-binding to dog erythrocyte cytosol proteins are described. Half-time of association of both thyroxine (T 4) and triiodothyronine (T 3) is 60 min and equilibrium is achieved at 120 min (20°C). Binding is enhanced at 37°C compared to 20°C. T 4 and T 3 binding capacities of the cytosol are 10 and 5 picomoles/mg cytosol protein, respectively. Gel filtration (G-100) reveals 3 protein fractions that dissociably bind both T 4 and T 3. Quantitative and qualitative differences distinguish the erythrocyte cytosol “receptor” proteins from those previously described in other dog tissues. The erythrocyte is a model for studying functions of cytosol “receptors” for iodothyronines.

High-pressure liquid chromatography of glycosphingo-lipids (with special reference to gangliosides)

The analysis of mixtures of gangliosides from adult human or bovine brain, supplemented with Tay-Sachs ganglioside, and haematoside from dog erythrocytes by high-pressure liquid chromatography using a moving-wire detector system is described. The complete separation of six gangliosides within 40 min has been achieved, using silica as the stationary phase and acidified chloroform-methanol-water mixtures as the eluent on a 25-cm column. Neutral glycosphingolipids, viz., the major components from normal human erythrocytes, can be completely separated on the same column, using non-aqueous and non-acidic eluents. It is shown that the methods described are useful for both analytical and (micro)-preparative purposes.

Proton-induced uptake of mammalian DNA by dog erythrocyte pink ghosts

Low pH (below 6) induces the uptake of mammalian DNA in dog erythrocyte pink ghosts. Uptake requires either Ca 2 or Mg 2 and is stimulated by ATP. These agents induce a rapid sphering of the ghosts at 37°C and sphering is required for uptake. Uptake is increased in ghosts which have been incubated 60–90 min before adding the DNA. Uptake is strongly temperature-dependent. Lowering the temperature of a suspension of ghosts taking up DNA at 37-0°C stops uptake. It is concluded that uptake depends on active membrane processes and that it may depend on the capacity of the ghosts to maintain cation exchange.

Cation specificity of propranolol‐induced changes in RBC membrane permeability: Comparative effects in human, dog and cat erythrocytes

Propranolol, in the presence of calcium, causes marked K efflux from human red blood cells (high K, low Na). The studies reported here indicate this effect of propranolol is specific for K and does not represent a nonspecific permeability increase for intracellular cations to leave the cell. Amphotericin-treated human RBC's (high Na, low K) and dog RBC's (high Na, low K) both gain K and increase in size when incubated in a K-medium containing propranolol and calcium. No effect was noted when cat RBC's (high Na, low K) were similarly treated. Propranolol, independent of added calcium, also inhibited the normally increased Na efflux observed when dog RBC's are suspended in K-medium. These species differences in response to propranolol thus may serve as a focus for elucidating the mechanism by which this drug alters normal membrane physiology. The unique drug effect on Na permeability of canine erythrocytes also may be a useful probe for the study of dog RBC volume regulation.

Genetic variants of glucose phosphate isomerase (E.C. 5.3.1.9) in canine erythrocytes.

Genetic polymorphism of glucose phosphate isomerase (GPI) was found in the erythrocytes of dogs of six Japanese breeds by using starch gel electrophoresis. Analysis of parentage records of dogs revealed that the phenotypic variation of erythrocyte glucose phosphate isomerase was controlled by one autosomal locus with two codominant alleles, GPIA and GPIB. The allele GPIB was observed in the following breeds: San'in-Shiba, Shinshu-Shiba, Shikoku, Kai and Kishu, but not in Hokkaidoes and Akitas. All the dogs belonging to 25 European breeds, 5 oriental or China-origin (except Japan) breeds examined in this experiments had the genotype constitution GPIA/GPIA, whereas one Dalmation dog was heterozygous GPIA/GPIB.

De novo synthesis of alanine by the perfused rat hindlimb.

Due to the disproportionately large production of alanine by muscle, it has been suggested that part of the alanine released by muscle is synthesized de novo by the transamination of glucose-derived pyruvate. This glucose-alanine conversion was quantitated in the isolated rat hindlimb perfused with a solution of bicarbonate buffer containing 2% albumin, 2.4% dextran, 2.5-15.9 mM glucose, 32-34% dog erythrocytes, and 0.05 muCi/ml [14C]glucose. Measurement of labeled alanine production allowed quantitation of de novo alanine synthesis. De novo derived alanine accounted for an average of 33% of the total alanine released by the perfused tissue (perfusate glucose concentration 8.3 mM), concurrently 2.7% of the glucose taken up by the limb was converted to alanine. By increasing the glucose concentration perfusing the muscle, both the rate of glucose uptake and de novo alanine release were increased. Addition of insulin to the perfusate (700 muU/ml) resulted in a significant increase in the rate of glucose uptake and de novo alanine production, but the rate of total alanine release was significantly decreased by the hormone. It was concluded that de novo alanine production accounts for a sizeable portion of the total alanine released by muscle, nevertheless a comparatively small fraction of the glucose carbons are actually transformed to alanine.

Effect of exogenous ATP on sodium transport in mammalian red cells.

The effect of exogenous adenosine triphosphate (ATP) and other nucleotides on the transport of Na in various mammalian red cells has been studied. While they have no effect on the transport of Na in human and cat red cells, in dog red cells adenosine and its mono-, di- and triphosphorylated forms were found to increase Na-influx. Of these, ATP has the most striking effect, causing a more than 8-fold increase at a concentration of 0.6 mM and exerting this effect at a dose range of 10(-5) to 10(-3) M. The effect of ATP is rapid (less than 5 minutes) and can be reversed by washing or the addition of calcium or magnesium. In contrast to the adenosine series other phosphorylated nucleotides (GTP, CTP, UDP, GDP and cAMP) have no effect. The well known volume dependent Na-transport in these cells is reversed in the presence of 0.6 mM ATP. It is suggested that ATP acts on passive cation movements either by chelation of membrane charge or by a direct interaction with membrane proteins and may be involved in the volume regulation of cation transport in the dog erythrocyte.

Effects of oxygen supply on electrical and secretory responses of humorally stimulated acinar cells in isolated rat pancreas.

A technique is described for perfusing isolated rat pancreas with a modified Krebs-Henseleit solution. This preparation made feasible the simultaneous measurement of transmembrane potential, effective membrane resistance, rate of flow of pancreatic juice and amylase output. The hyperpolarizing effect of cholecystokinin-pancreozymin (CCK-PZ) was confirmed by perfusion with a solution containing 5 mU CCK-PZ/ml. The hyperpolarizing efffect of CCK-PZ was enhanced when the pancreas was perfused with an oxygenated solution containing dog erythrocytes. This effect was inhibited by ouabain and anaerobic conditions. It is suggested that the oxidative metabolism supplies energy for a mechanism which is responsible for the hyperpolarizing effect of CCK-PZ. The amylase output and the rate of flow of pancreatic juice were enhanced concurrently under conditions that augumented the hyperpolarizing effect, and they were inhibited under conditions that suppressed the effect. Cellular events in stimulus-secretion coupling in the pancreatic acinar cell are discussed by correlating the electrophysiological responses and the secretory responses under various conditions.