scholarly journals STUDIES ON THE EXTRACELLULAR CULTIVATION OF AN INTRACELLULAR PARASITE (AVIAN MALARIA)

1950 ◽  
Vol 92 (4) ◽  
pp. 349-366 ◽  
Author(s):  
William Trager
Keyword(s):  
Avian Malaria ◽  
Culture Media ◽  
Cell Extract ◽  
Red Cells ◽  
Red Cell ◽  
Sodium Pyruvate ◽  
Plasmodium Lophurae ◽  
Adenylic Acid ◽  
Survival And Development

The erythrocytic stages of Plasmodium lophurae were freed from their host red cells by specific hemolysis directly into a favorable medium containing an extract of duck erythrocytes. Extracellular survival and development of the parasite in vitro occurred in culture media consisting essentially of a very concentrated extract of duck red cells prepared in a special nutrient solution. Omission or dilution of the red cell extract resulted in rapid degeneration of the parasites. Their survival and development were favored by the presence in the erythrocyte extract of gelatin, yeast adenylic acid, and cozymase, and especially by the further addition of adenosinetriphosphate and sodium pyruvate. Under the best conditions yet tested, all the free parasites continued their development extracellularly during the first two days of cultivation. Merozoites formed by the extracellular segmentation of the free parasites originally present developed further into trophozoites. On the third day a majority of the free parasites were still of normal appearance, but by the fourth day more were degenerate, and very few normal parasites remained on the fifth day.

scholarly journals STUDIES ON THE EXTRACELLULAR CULTIVATION OF AN INTRACELLULAR PARASITE (AVIAN MALARIA)

1952 ◽  
Vol 96 (5) ◽  
pp. 465-476 ◽  
Author(s):  
William Trager
Keyword(s):  
Malic Acid ◽  
Avian Malaria ◽  
Culture Medium ◽  
Coenzyme A ◽  
Intracellular Parasite ◽  
Sodium Pyruvate ◽  
Plasmodium Lophurae ◽  
Survival And Development ◽  
Development In Vitro

The extracellular survival and development in vitro of the erythrocytic stages of Plasmodium lophurae were favored by the addition to the culture medium of l-malic acid and concentrates rich in coenzyme A. In a concentrated extract of duck erythrocytes supplemented with these two substances in addition to adenosinetriphosphate, sodium pyruvate, and certain other materials of like nature, only 5 to 10 per cent of the extracellular parasites had become abnormal after 3 days of cultivation.

scholarly journals FURTHER STUDIES ON THE SURVIVAL AND DEVELOPMENT IN VITRO OF A MALARIAL PARASITE

1943 ◽  
Vol 77 (5) ◽  
pp. 411-420 ◽  
Author(s):  
William Trager
Keyword(s):  
Salt Solution ◽  
Cell Extract ◽  
Malarial Parasite ◽  
Red Cell ◽  
Calcium Pantothenate ◽  
Plasmodium Lophurae ◽  
Survival And Development ◽  
Gentle Agitation ◽  
Development In Vitro

The survival of Plasmodium lophurae in vitro is favored by the presence of calcium pantothenate (0.02 mg. per ml). Survival of about 2 weeks in vitro at 40–41°C. has been obtained under the following conditions: a medium consisting of duck red cell extract in balanced salt solution with glutathione and glucose or glycogen, serum, embryo extract, and calcium pantothenate; daily replacement of about half of the medium with fresh medium; addition of fresh uninfected erythrocytes every 2nd day; gentle agitation of the preparation on a rocking machine. In some of these preparations significant increases in male gametocytes and, more rarely, in total numbers of parasites occurred during the first few days.

scholarly journals STUDIES ON CONDITIONS AFFECTING THE SURVIVAL IN VITRO OF A MALARIAL PARASITE (PLASMODIUM LOPHURAE)

1941 ◽  
Vol 74 (5) ◽  
pp. 441-462 ◽  
Author(s):  
William Trager
Keyword(s):  
Liver Extract ◽  
Cell Extract ◽  
Malarial Parasite ◽  
Red Cell ◽  
High Potassium ◽  
Plasmodium Lophurae ◽  
Parasite Plasmodium ◽  
Chick Embryo Extract ◽  
Very High

The survival of Plasmodium lophurae in vitro, at temperatures of 39.5–42°C., is favored by a balanced salt solution having a high potassium content, by aeration but not by a very high oxygen tension, by an optimal density of parasites per cubic millimeter, by frequent renewal of the suspending medium, by concentrated red cell extract, by optimal concentrations of plasma or serum, of chick embryo extract, of glucose or glycogen, and of glutathione, and probably by yeast extract and a very low concentration of liver extract. In the best preparations, as judged by infectivity, more than 40 per cent of the parasites were alive on the 3rd day, more than 20 per cent on the 4th day, perhaps 1 per cent on the 5th day, and about 0.05 per cent on the 6th day. Evidence was obtained that the parasites had multiplied during the 1st day of incubation.

LACK OF EFFECT OF CORTICOSTEROIDS ON THE IN VIVO DISAPPEARANCE OF SULFHYDRYL-INHIBITED AND ANTIBODY-COATED ERYTHROCYTES

1964 ◽  
Vol 47 (3_Suppl) ◽  
pp. S28-S36
Author(s):  
Kailash N. Agarwal
Keyword(s):  
Red Cells ◽  
List Type ◽  
Red Cell

ABSTRACT Red cells were incubated in vitro with sulfhydryl inhibitors and Rhantibody with and without prior incubation with prednisolone-hemisuccinate. These erythrocytes were labelled with Cr51 and P32 and their disappearance in vivo after autotransfusion was measured. Prior incubation with prednisolone-hemisuccinate had no effect on the rate of red cell disappearance. The disappearance of the cells was shown to take place without appreciable intravascular destruction.

scholarly journals Effect of oxalate and malonate on red cell metabolism

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1389-1393
Author(s):  
E Beutler ◽  
L Forman ◽  
C West
Keyword(s):  
Pyruvate Kinase ◽  
Cell Metabolism ◽  
Inhibitory Effect ◽  
Red Cells ◽  
Red Cell ◽  
Acid Analogue ◽  
2,3 Dpg

The addition of oxalate to blood stored in Citrate-phosphate-dextrose (CPD) produces a marked improvement in 2,3-diphosphoglycerate (2,3-DPG) preservation; an increase in 2,3-DPG levels can also be documented in short-term incubation studies. Oxalate is a potent in vitro inhibitor of red cell lactate dehydrogenase, monophosphoglycerate mutase, and pyruvate kinase (PK). In the presence of fructose 1,6-diphosphate the latter inhibitory effect is competitive with phospho(enol)pyruvate (PEP). Determination of the levels of intermediate compounds in red cells incubated with oxalate suggest the presence of inhibition at the PK step, indicating that this is the site of oxalate action. Apparent inhibition at the glyceraldehyde phosphate dehydrogenase step is apparently due to an increase in the NADH/NAD ratio. Oxalate had no effect on the in vivo viability of rabbit red cells stored in CPD preservatives for 21 days. Greater understanding of the toxicity of oxalate is required before it can be considered suitable as a component of preservative media, but appreciation of the mechanism by which it affects 2,3-DPG levels may be important in design of other blood additives. Malonate, the 3-carbon dicarboxylic acid analogue of oxalate late did not inhibit pyruvate kinase nor affect 2,3-DPG levels.

scholarly journals The relationship between in vivo generated hemoglobin skeletal protein complex and increased red cell membrane rigidity

Blood ◽  
1988 ◽  
Vol 71 (5) ◽  
pp. 1427-1431 ◽  
Author(s):  
N Fortier ◽  
LM Snyder ◽  
F Garver ◽  
C Kiefer ◽  
J McKenney ◽  
...  
Keyword(s):  
Protein Complex ◽  
Sodium Dodecyl ◽  
Red Cells ◽  
Red Cell ◽  
Red Cell Membrane ◽  
Cellular Dehydration ◽  
Thalassemia Minor ◽  
Membrane Rigidity

Abstract In vitro induced oxidative damage to normal human RBCs has previously been shown to result in increased membrane rigidity as a consequence of the generation of a protein complex between hemoglobin and spectrin. In order to determine if in vivo generated hemoglobin-spectrin complexes may play a role in increased membrane rigidity of certain pathologic red cells, we measured both these parameters in membranes prepared from hereditary xerocytosis (Hx), sickle cell disease (Sc), and red cells from thalassemia minor (beta thal). Membranes were prepared from density-fractionated red cells, and membrane deformability was measured using an ektacytometer. Hemoglobin-spectrin complex was determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel analysis, as well as by Western blot analysis using a monoclonal antibody against the beta- subunit of hemoglobin. For these three types of pathologic red cells, progressive cellular dehydration was associated with increased membrane rigidity and increased content of hemoglobin-spectrin complex. Moreover, the increase in membrane rigidity appeared to be directly related to the quantity of hemoglobin-spectrin complex associated with the membrane. Our findings imply that hemoglobin-spectrin complex is generated in vivo, and this in turn results in increased membrane rigidity of certain pathologic red cells. The data further suggest that oxidative crosslinking may play an important role in the pathophysiology of certain red cell disorders.

scholarly journals The Role of Red Cell Energy Metabolism in the Generation of Irreversibly Sickled Cells In Vitro

Blood ◽  
1973 ◽  
Vol 42 (6) ◽  
pp. 835-842 ◽  
Author(s):  
Michael Jensen ◽  
Stephen B. Shohet ◽  
David G. Nathan
Keyword(s):  
Energy Metabolism ◽  
Calcium Metabolism ◽  
Red Cells ◽  
Red Cell ◽  
Hemoglobin S ◽  
Cell Energy ◽  
Incubation Buffer ◽  
Membrane Defect

Abstract An acquired membrane defect is believed to be responsible for the maintenance of the sickled shape in oxygenated irreversibly sickled cells (ISC), because the hemoglobin S in these cells is not in the aggregated, "sickled" state. In the present study, it is demonstrated that the acquisition of the membrane defect in vitro depends on cellular metabolism. Only if cellular ATP is almost completely depleted while the cells are sickled, do they become unable to resume the biconcave disk shape upon reoxygenation. If calcium is omitted from the incubation buffer, ISCs are not generated despite metabolic depletion. This suggests an action of ATP mediated through calcium metabolism similar to that which prevents membrane stiffening in normal red cells. No ISCs were produced by repeated sickling and unsickling. Thus, a membrane alteration occurring as a consequence of metabolic depletion seems to be a more important factor in the generation of ISC than sickling-unsickling induced fragmentation.

scholarly journals Reticulocyte Survival in Sickle Cell Anemia: Effect of Cyanate

Blood ◽  
1972 ◽  
Vol 40 (5) ◽  
pp. 733-739 ◽  
Author(s):  
Blanche P. Alter ◽  
Yuet Wai Kan ◽  
David G. Nathan
Keyword(s):  
Cell Survival ◽  
Sickle Cell ◽  
Fetal Hemoglobin ◽  
Red Cells ◽  
Red Cell ◽  
Hemoglobin Molecule ◽  
Red Cell Survival ◽  
And Control

Abstract Cyanate prevents sickling in vitro and apparently prolongs the survival of 51Cr-tagged sickle erythrocytes in vivo. Cautious interpretation is required because the effects of cyanate on 51Cr binding to sickle and fetal hemoglobin-containing red cells are unknown, and comparison of the effect of cyanate on sickle red cell survival to control red cell survival must be performed sequentially. We have studied the survival of sickle reticulocytes utilizing radioactive amino acids that are incorporated into hemoglobin. Two informed adult patients with sickle cell disease were studied. In each study, two 50-ml samples of blood were incubated separately with 14C- and 3H-leucine for 2 hr, after which 50 mM cyanate was added to one aliquot for 1 hr. The cells were then washed and reinfused. Frequent venous samples were obtained, and the specific activities of 14C and 3H in the hemoglobin were followed. The t ½ of the carbamylated cells was tripled, but remained below normal. This method provides a generally useful measurement of the influence of drugs bound to red cells on reticulocyte lifespan. The labels are incorporated into the hemoglobin molecule of the reticulocyte, and simultaneous comparison of the survivals of the same cohort of drug-treated and control cells is achieved.

scholarly journals Effect of oxalate and malonate on red cell metabolism

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1389-1393 ◽  
Author(s):  
E Beutler ◽  
L Forman ◽  
C West
Keyword(s):  
Pyruvate Kinase ◽  
Cell Metabolism ◽  
Inhibitory Effect ◽  
Red Cells ◽  
Red Cell ◽  
Acid Analogue ◽  
2,3 Dpg

Abstract The addition of oxalate to blood stored in Citrate-phosphate-dextrose (CPD) produces a marked improvement in 2,3-diphosphoglycerate (2,3-DPG) preservation; an increase in 2,3-DPG levels can also be documented in short-term incubation studies. Oxalate is a potent in vitro inhibitor of red cell lactate dehydrogenase, monophosphoglycerate mutase, and pyruvate kinase (PK). In the presence of fructose 1,6-diphosphate the latter inhibitory effect is competitive with phospho(enol)pyruvate (PEP). Determination of the levels of intermediate compounds in red cells incubated with oxalate suggest the presence of inhibition at the PK step, indicating that this is the site of oxalate action. Apparent inhibition at the glyceraldehyde phosphate dehydrogenase step is apparently due to an increase in the NADH/NAD ratio. Oxalate had no effect on the in vivo viability of rabbit red cells stored in CPD preservatives for 21 days. Greater understanding of the toxicity of oxalate is required before it can be considered suitable as a component of preservative media, but appreciation of the mechanism by which it affects 2,3-DPG levels may be important in design of other blood additives. Malonate, the 3-carbon dicarboxylic acid analogue of oxalate late did not inhibit pyruvate kinase nor affect 2,3-DPG levels.

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