Polycythemic Animals Research Articles

Polycythemia decreases fatigue in tetanic contractions of canine skeletal muscle.

The effects of an acute polycythemia on muscle fatigue development were investigated in the self-perfused canine gastrocnemius in situ. Following isolation of the gastrocnemius, dogs (N = 5) were made polycythemic through a bolus injection of packed erythrocytes (hematocrit (Hct) = 90-92%) to raise systemic Hct to 63.5 +/- 0.5%. Subsequently, the gastrocnemius was stimulated, through the sciatic nerve, to perform 20 min of isotonic tetanic contractions (60 x min(-1), 200 ms, 50Hz). Control (normocythemic) animals (N = 5) underwent an identical contraction regimen. Although blood flow to the gastrocnemius was not different at any time, oxygen delivery was significantly increased during polycythemia (peak = 33.7 +/- 2.2 mL x 100 g(-1) x min-1) over control (peak = 25.1 +/- 2.1 mL x 100 g(-1)x min(-1)) at all times during contraction. Oxygen uptake by the gastrocnemius, although consistently increased, was not significantly different between the normocythemic and polycythemic conditions at any time. The rate of fatigue was significantly decreased over the first 6 min of contraction in polycythemic animals (3.5 +/- 0.6% x min(-1)) when compared with controls (5.8 +/- 0.7% x min(-1)). Subsequent fatigue development was not different between groups. As a result of the early rate differences in fatigue, however, the work production in polycythemic animals was significantly greater than in normocythemic dogs for the duration of the contraction period. We conclude that during high metabolic rate isotonic tetanic contractions, muscle fatigue development is diminished by polycythemia, but the ergogenic effect appears to be transient.

Serum erythropoietin concentrations measured by radioimmunoassay in normal, polycythemic, and anemic dogs and cats.

Serum erythropoietin (Epo) concentrations were measured by radioimmunoassay (RIA) in normal, polycythemic, and anemic dogs and cats. The serum Epo concentration in normal dogs (n = 25) ranged from 7 to 37 mU/mL (median, 20 mU/mL); and in normal cats (n = 11) ranged from 9 to 38 mU/mL (median, 18 mU/mL). Polycythemic animals (PCV > 55% in dogs, > 45% in cats) were classified as those with primary (polycythemia vera), secondary, or polycythemia of uncertain etiology. Dogs with polycythemia vera (PV, n = 8) had a median serum Epo concentration in the normal range (17 mU/mL); cats with PV (n = 7) also had a median serum Epo concentration that was within the normal range (10 mU/mL). In the category of secondary polycythemias, dogs (n = 7) (median, 30.7 mU/mL) and cats (n = 2) had normal Epo concentrations. The median serum Epo concentration was significantly decreased (P < .05) in dogs with PV compared with dogs with secondary polycythemias. The median serum Epo concentrations in dogs (n = 13) and cats (n = 5) with anemias not due to chronic renal disease were significantly increased (P < .05) compared with normal dogs and cats. In cats with anemias due to chronic renal disease (n = 5) the median serum Epo concentration was not significantly different from normal cats. The measurement of the serum EPO concentration may be useful in assessment of anemia or polycythemia but the overlap of values with the normal range in all groups evaluated limit its diagnostic use.

Evaluation of an automated system for hemoglobin measurement in animals

SUMMARY In veterinary medicine, pcv determined by centrifugation of blood in a microhematocrit tube is the most common clinical test used to initially assess and monitor anemic and polycythemic animals. In contrast, blood hemoglobin (Hb) concentration, rather than pcv, is generally determined in human patients. One automated system photometrically measures blood Hb concentration after conversion of Hb to azide methemoglobin without dilution and was found to be a simple and accurate instrument for use in human medicine. We evaluated the system for its accuracy in measuring blood Hb concentration in animals by comparing it with standard techniques and for its suitability in veterinary practice. Blood samples, anticoagulated with potassium edta, from 78 healthy animals (33 dogs, 17 cats, 13 horses, and 15 cows) and 58 dogs and 4 cats with various blood abnormalities (10 anemia, 11 polycythemia, 21 lipemia, 16 leukocytosis, and 6 icterus) were analyzed. In all species, blood Hb concentration of healthy animals determined by the system was comparable to that measured by standard cyanmethemoglobin methods (ie, an automated counter; rI = 0.987 to 0.998 and a hemoglobin kit, rI = 0.946 to 0.993). The aforementioned system also yielded similar values to those obtained by use of standard methods in anemic, polycythemic and icteric dogs and cats. Moreover, the system reads the absorbance at 2 wavelengths to correct for turbidity, and therefore, accurately measured Hb concentration in blood samples with severe lipemia (triglycerides concentration &gt; 500 mg/dl) and marked leukocytosis (&gt; 50,000 wbc/μl, whereas other standard Hb techniques are known to give falsely high results. We conclude that the automated system compares favorably to standard methods, and is a simple and accurate instrument to quickly measure Hb concentration in animals.

Acute Polycythemia Increases the Disappearance Rate of Clottable Fibrinogen in the Newborn Dog

To explore the pathophysiology of the necrotizing enterocolitis caused by polycythemia in the newborn dog, the effect of acute polycythemia on fibrinogen disappearance rate was studied in 38 puppies (3-14 days). All pups received an exchange transfusion removing 65 ml/kg of blood and transfusing 85 ml/kg of either whole blood (control, resulting hematocrit = 37), or packed red blood cells (polycythemia, resulting hematocrit = 68). Necrotizing enterocolitis was found in 15 of 19 polycythemic and four of 19 control pups (p less than 0.01). 125I fibrinogen and Evan's blue (an albumin marker) were injected 2 h after transfusion and the concentration of clottable labeled fibrinogen and albumin tracer were measured at 1/2 and 2 h after injection. The fraction of the tracer that disappeared over the 1 1/2-h period was calculated. In the polycythemic group 45 +/- 18 SD% of the clottable fibrinogen disappeared versus only 28 +/- 15% in the control group (p less than 0.01). In the polycythemic group 36 +/- 21% of the albumin tracer disappeared versus 31 +/- 12% in the control group (NS). Thus polycythemia in the newborn dog is associated with an increased disappearance rate of clottable fibrinogen not associated with a general increase in protein disappearance rate. Thus an intravascular coagulopathy is evident in the polycythemic animals. Whether this coagulopathy is the cause of the necrotizing enterocolitis or is secondary to the necrotizing enterocolitis seen in this animal model cannot be determined from this experiment.

Physiological effects of hypervolemic polycythemia in newborn dogs.

The effect of hypervolemic polycythemia (Hct 62-77) on O2 transport and O2 consumption (Vo2) was studied in 16 unanesthetized newborn dogs (age 3-10 days). A control group of 10 newborn dogs (3-10 days old) was made hypervolemic but not polycythemic in an otherwise identical experiment. Hypervolemia was attained by infusing 33 ml/kg of either packed red blood cells (polycythemia) or whole blood (controls). In the polycythemic group as a result of the transfusion, cardiac output (CO) decreased by 50% (P less than 0.001), peripheral vascular resistance (PVR) increased by 170% (P less than 0.001), but O2 transport did not change significantly, and Vo2 decreased only slightly by 13% (P less than 0.01). In the 10 control animals there were no significant changes in CO, PVR, O2 transport, or Vo2 as a result of the transfusion. Blood lactate increased only slightly in experimental (35%, P less than 0.04) and control animals (23%, NS). The ability of the animals to increase their O2 transport and Vo2 was tested by measuring the changes induced by cold stress. Cold stress produced a 20% increase in Vo2 (P less than 0.05) in both the polycythemic and the control animals. Thus in spite of a decreased CO in the nonstressed state the polycythemic animals were still able to increase O2 transport and Vo2 in response to cold stress. These results suggest that the newborn animal is capable of regulating CO to maintain O2 transport appropriate to uptake under conditions of hypervolemic, polycythemic hyperviscosity, and environmental cold stress.

Action of hemolysate of polycythemic animals on proliferations of bonne marrow cells

Erythrocytic inhibitor from polycythemic rats depressed mitotic activity of cells of of the erythroid series in mice by 40%. The inhibitory effect lasted about 12 h. The inhibitor acted on the G2 period. The “points of application” of the inhibitor were not only blast forms of the erythron but also hematopoietic stem cells.

Effect of exogenous DNA on erythropoietin-sensitive cells

Transfusion polycythemia was reproduced in Wistar rats. Preparations of homologous and heterologous DNA were injected intraperitoneally in a dose of 5 mg. On the 1st, 3rd, 5th, 10th, 15th, and 20th days after injection of DNA the erythrocytes production was determined from the Fe59 content in these cells in the peripheral blood. Despite the sharp inhibition of erythropoiesis in the polycythemic animals, injection of the preparations of exogenous DNA restored the formation of the red blood cells. The effect from injection of heterologous DNA was less than from homologous DNA. The restoration of erythropoiesis in polycythemic animals in response to injection of DNA preparations was due to the effect of the DNA on differentiation and proliferation of erythropoietin-sensitive cells.

Granulocytic Recovery in the Polycythemic Dog Treated with Endotoxin Postirradiation

Abstract : The experiment was designed to test the hypothesis that, in hypertransfused dogs with suppressed erythropoiesis, endotoxin-induced increased muelopoietic stimulation 24 hours after irradiation would enhance leukocytic recovery. The treatment combination of transfusion polycythemia and endotoxin was compared to a control group receiving no therapy or to the results obtained from groups of dogs receiving each treatment alone. The radiation exposure was 150 rads of total body x rays. An accentuated abortive rise in the group receiving the combined treatment, accelerated recovery in both groups receiving endotixin, and impaired leukocytic recovery in the polycythemic animals were observed. Accentuated depression of platelets and splenic enlargement inversely correlated with the platelet levels were observed in the animals receiving transfusion therapy regardless of whether they received endotoxin. It is felt that the data support the concept of interdependence between the various cell lines of hematopoiesis in an experimental model not previously described.

Gastric secretion and peptic ulcer in rats with experimentally-induced polycythemia.

Experimental polycythemia was induced in rats by repeated intravenous transfusion of homologous packed erythrocytes. Rats were bearing permanent gastric cannulas and some of them had bilateral truncal vagotomy. When polycythemia was established, 24-hour gastric secretion studies were performed. During the collection of gastric juice, rats received continuous intravenous infusion of histamine or of the anticholinergic drug propantheline bromide (Pro-Banthine). Control animals were infused by normal saline. At the end of the experiment gastric mucosa of polycythemic and control rats was studied macro- and microscopically. In all polycythemic rats in which HCl and pepsin were found in gastric secretion, severe peptic ulcers of glandular mucosa were found. Vagotomized polycythemic animals in which HCl secretion was abolished had no peptic ulcers. Severity and number of ulcers were not directly related to the amount of HCl and pepsin secreted in 24 h. The etiology of these ulcers remains unexplained.

On the Mechanism of Erythropoietin-induced Differentiation: IX. INDUCED SYNTHESIS OF 9 S RIBONUCLEIC ACID AND OF HEMOGLOBIN

Erythropoietin induces the synthesis of 9 S RNA and hemoglobin by cultured bone marrow cells derived from polycythemic rats and mice in which hemoglobin synthesis has been suppressed. Synthesis of 9 S RNA starts within 1 hour (rat cells) and 4 hours (mouse cells) of addition of erythropoietin to the medium, while hemoglobin synthesis is not initiated until 8 to 10 hours later. These findings indicate that there is another regulatory step between transcription, initiated by erythropoietin, and translation. Under conditions in which no hemoglobin synthesis occurs, bone marrow cells still synthesize a very small amount of 9 S RNA which may be a messenger for a protein other than globin. Comparison of stimulated hemoglobin synthesis by cells from normal animals with cells from polycythemic animals indicates that erythropoietin has an effect on some differentiated cells in the former population as well as on the undifferentiated cells in the latter population.

Effects of blood pressure and blood viscosity on fluorescein transit time in the cerebral microcirculation in the mouse.

Fluorescein transit time from pial arterioles to pial venules was measured in the mouse by analyzing motion pictures taken at 40 frames/sec. Increased blood pressure reduced the transit time. Increased blood viscosity lengthened the transit time. Viscosity was increased by rendering the animals polycythemic, or by infusions of high molecular weight dextran. Two different dextrans were used. The one with the higher molecular weight (500,000) produced a higher viscosity and a more prolonged transit time than the one with the lower molecular weight (250,000). These dextrans produced a prolongation of the transit time equal to that, or greater than that, observed in the polycythemic animals, even though whole blood viscosity in polycythemia exceeded that of the dextran treated mice over a wide range of physiologically pertinent shear rates applied in vitro in a rotational viscometer. Results indicate that increases of blood viscosity produce decreases in the velocity of the plasma, and that viscosity increments brought about by raising the hematocrit, may be less effective than viscosity increments brought about by raising the viscosity of the plasma.

Studies on the Regulation of Hemopoietic Spleen Colonies

Abstract The in vivo intrasplenic cloning of haemopoietic cells was used as an experimental system for the study of some aspects concerned with the regulation of erythropoiesis. Experiments on the effects of polycythemia on the formation of erythroid clones demonstrated that the "feedback" suppression of such clones was inhibited if the polycythemic animals were kept under hypoxic conditions. Overloading with oxygen is, thus, an essential factor in the polycythemia-induced suppression of erythroid clones. Polycythemic animals which were transferred to hypoxic conditions for only 48 hours before the termination of the experiment showed the formation of erythroid clones. Analysis of clone formation following such short exposures to hypoxia suggests that the endogenous erythropoietin which had been functioning in this situation acted not upon the single clone-forming cell, but rather on small cell populations which derived from the clone-forming cells in the absence of erythropoietin. There is, therefore, an early, erythropoietin-independent phase of replication of the clone-forming cells. The application of erythopoietin only during this "early" phase of replication resulted in the formation of erythroid clones, which, following the discontinuation of erythropoietin, disappeared from the spleen. Similar results were obtained when the action of endogenous erythropoietin was terminated by subjecting animals to polycythemic conditions a few days after the injection of bone marrow. It appears that in both these cases the reduction in the number of spleen colonies following discontinuation of erythropoietin activity is due to the total maturation of the erythroid colonies, ending in the evacuation from the spleen of the mature red blood cells. The differentiation of the cloned cells is, thus, causally related to the cessation of the replicating effect of erythropoietin.

Polycythemic mice produced by hypoxia in silicone rubber membrane enclosures: a new technique.

SummaryUtilizing enclosures covered with silicone rubber membrane, a new, safe, and simple method for producing polycythemia by hypoxia is presented. The polycythemic animals are excellent for use in erythropoietin assays.

The Course of Plasmodium berghei Infection in the Polycythemic Mouse

The erythrocytic forms of P. berghei are reported to have a predilection for infecting young red cells in rodents. In an attempt to clarify the role of immature red cells, P. berghei infections were studied in the polycythemic mouse in which erythropoiesis was markedly inhibited. The infectivity of the parasite is directly related to the availability of immature red cells. The infection can be completely blocked even in the presence of a large population of mature red cells. The malaria parasite of rodents, Plasmodium berghei, is reported to preferentially invade immature erythrocytes (Galliard, 1948; Ramakrishnan, 1950; Baldi, 1950; Corradetti et al., 1951; Sergent, 1958). During the course of infection in the mouse the total number of red blood cells falls rapidly as reticulocytosis greatly increases (Singer, 1954). The appearance of parasites in reticulocytes may result from the anemic condition with the increased availability of young red cells. Hsii and Geiman (1952) reported the potentiation of P. berghei virulence in mice which were anemic from Eperythrozoan coccoides infection. The mice exhibited a marked reticulocytosis. Xirradiation of normal mice before and after the inoculation of a large number of parasites has certain effects on the course of P. berghei, which are apparently related more to inhibition of erythropoiesis than to a direct effect on the parasite (Singer, 1953). The infectivity and virulence of P. berghei has been compared in normal, anemic, and polycythemic mice in an attempt to better clarify the role of young red cells. Polycythemic mice can be prepared so that erythropoiesis is essentially halted as judged by the absence of reticulocytes in the peripheral blood (Jacobson et al., 1957; Filmanowicz and Gurney, 1961). This model permits the regulation of the availability of young red cells to the parasite under physiological conditions. MATERIALS AND METHODS Male CFW strain mice are maintained in this laboratory with Plasmodium berghei infections. Received for publication 15 September 1965. * This paper is contribution number 50 from the Army Research Program on Malaria. The work was supported by U. S. Army Contract No. DA49-193-MD-2545. Directed by Dr. John Arold. Hypertransfused mice were prepared in the following manner: 5 to 7 weeks old virgin male mice, weighing 15 to 20 g, were given daily intraperitoneal injections of washed homologous red cells. Red cells were collected from the jugular vein of stock mice in a heparinized glass beaker. Plasma was removed and the cells were washed three times with 0.85% NaCl. One-half ml of washed packed red cells was injected daily for 5 days. On day 6 a hematocrit and a reticulocyte count were prepared from the tail vein of each animal. Reticulocyte counts showed complete absence of young red cells peripherally. Hematocrits ranged from 70 to 80%. On day 7 a standard inoculum of approximately 30,000 parasitized red cells was injected intraperitoneally. This is approximately 300 times the minimum number of parasitized red cells required to produce an infection (Fabiani et al., 1951). A second group of hypertransfused mice were continually given IP red cell injections during the 10-day period following the inoculation of parasites. The red cells used in this experiment were taken from stock polycythemic animals which showed less than one reticulocyte per 1,000 red cells. The third and fourth groups of polycythemic animals were given 500 R or 750 R whole body X-irradiation 1 day following the final intraperitoneal transfusion. Three days later the animals were given the standard inoculum of approximately 30,000 parasitized red cells. Daily smears were prepared to follow the per cent parasitemia for 10 days and at 12, 15, and 24 days. Two groups of polycythemic animals receiving 750 R were given 106 and 10s parasitized red cells, respectively. Two animals from each group were killed and the blood from each animal was subinoculated into three normal mice on days 6 and 12. Thin smears from the subinoculated animals were prepared on day 6 for per cent parasitemia. Anemic mice were prepared by being bled once daily for 3 days from the medial aspect of the eye with a fine glass pipette. Hematocrit and reticulocyte counts were prepared on day 4 and the standard inoculum of parasitized red cells was given on day 5. Blood smears stained with Wright's stain were prepared daily from the tail veins of each mouse

THE SPECIFIC SUPPRESSION OF THE DIFFERENTIATION OF ERYTHROID CLONES IN POLYCYTHEMIC ANIMALS.

A study was made of the effect of transfusion-induced polycythemia on the formation of intrasplenic clones in X-irradiated mice injected with bone marrow cells. Polycythemic animals showed a 3-fold decrease in the number of macroscopic clones obtained, as compared to non-polycythemic animals. Microscopic examination of the cell types of the spleen colonies revealed that the suppression effect of transfusion-induced polycythemia was cell specific: no erythroid clones were formed in the spleens of the polycythemic animals. Both cell replication and differentiation of the erythroid lines were completely inhibited. No such inhibition was observed among the granuloid colonies; in fact, an increased incidence of granuloid colonies was found in the polycythemic animals.

Systemic oxygen transport in induced normovolemic anemia and polycythemia.

The hemodynamic effects of normovolemic anemia and polycythemia were studied in 14 dogs. Anemia (5 dogs) and polycythemia (5 dogs) were induced by bleeding and simultaneously infusing dextran or packed erythrocytes. Measurements included cardiac output, arterial oxygen saturation, peripheral vascular resistance, and systemic oxygen transport (cardiac output X arterial oxygen content). Cardiac output had a significant negative linear relationship to hematocrit ( r = –0.74, P &lt; 0.01) over the range studied (13–74%). Peripheral resistance fell 46% in anemic animals and increased 152% in four of five polycythemic animals. Arterial saturation was significantly correlated to changes in hematocrit ( r = 0.62, P &lt; 0.01) and cardiac output ( r = –0.55, P &lt; 0.01); these values were due primarily to the linearity encountered in the anemia experiments and a reversal in these relationships tended to occur at high hematocrits. Systemic oxygen transport was maximum at normal hematocrits and decreased in anemia and polycythemia. The data indicate that hemodynamic adjustments in normovolemic anemia and polycythemia are insufficient to maintain normal oxygen delivery.

Response to Erythropoietin.

Alpen and Cranmore (1) and Erslev (2) have concluded that erythropoietin increases red cell output by accelerating the rate of stem cell differentiation into erythroid precursors. Observations on erythropoiesis in polycythemic mice by Gurney, et al.(3) have supported this conclusion. However, observations by several investigators (4-7) seem to indicate that the action of the erythropoietic humoral factor (s) is not limited to regulation of stem cell differentiation. Experiments reported here tested the effect of erythropoietin on later phases of erythroid formation and the results suggest an action here as well as in the early stem cell phase. Methods. Female Sprague-Dawley rats, weighing 180 to 200 g, were made polycythemic by one or several transfusions of packed homologous red blood cells. Each transfusion equaled 50% of the animal's original red cell mass. At various intervals following the initial transfusion the polycythemic rats were given a single injection of human urine erythropoietin (8) or purified sheep erythropoietin (9). Subsequent changes in erythropoiesis were assessed by reticulocyte counts (10) and measurement of the 18-hour erythrocyte radio-iron incorporation(8). Erythroid elements per 1000 nucleated cells were enumerated in smears of marrow from control polycythemic animals. In 3 experiments, polycythemic rats were given single intraperitoneal injections of 2 ml of urine concentrate on either the 5th, 15th or 31st day after their first transfusion. Control polycythemic rats were injected with saline. At the end of each successive 24-hour period following erythropoietin or saline injection, representative animals from each group were killed and the reticulocyte counts, hematocrits and erythrocyte radio-iron incorporation were determined. The radio-iron was injected 18 hours before the rats were killed. All groups given urine concentrate included at least 5 rats.

Hemoglobin content and blood volume of normal, anemic and polycythemic rats, determined by carbon monoxide and radiochromium

AbstractIn normal, anemic and polycythemic rats the blood volume and the total amount of hemoglobin have been determined by means of a modified alveolar CO method and Cr51 tagged erythrocytes. The Cr51 method gives on an average 10 per cent lower values than the CO method, both in normal, anemic and polycythemic animals. Reasons for this difference are discussed.

Blood proteins and their distribution in rats with parabiosis intoxication; determined by paper electrophoresis.

SummaryRats which have developed parabiosis intoxication show, in addition to the other hematologic aberrations which characterize the condition, changes in the blood proteins. Anemic partners have a low plasma protein level and polycythemic animals an elevated plasma protein titre. The partitions which albumin and the respective globulins contribute toward the total serum proteins average the same in both anemic and polycythemic twins, although as compared to control levels there is a diminished serum albumin and an elevated α2-globulin component in each.

Observations on Cobalt Polycythemia. I. Studies on the Peripheral Blood of Rats.

Conclusions1. Injections of cobaltous chloride continued over a period of 8 months without apparent serious toxicity has resulted in persistent polycythemia during this period of a degree comparable to that occurring in polycythemia vera in man. 2. The average increases in blood volume and total erythrocyte mass as compared with controls was 80% and 192% respectively, due entirely to increase in erythrocytes since plasma volume decreased 16% on the average. 3. The erythrocyte of polycythemic animals as compared with the normal increased in volume 41% due almost entirely to an increase in the thickness of the cell. The hemoglobin content remains essentially unchanged.