Walter Fried

Studies on erythropoiesis. III. Factors controlling erythropoietin production.

Summary and Conclusion 1. We studied the erythropoietic response to anemic plasma of a variety of experimental conditions in the rat. Rats subjected to hypophysectomy, an atmosphere of high O2, starvation, and transfusion-induced polycythemia have a decreased rate of erythropoiesis and an exaggerated response to the administration of anemic plasma. 2. Treatment with dinitrophenol increases the rate of erythropoiesis and decreases the response to anemic plasma. 3. These findings are in agreement with the hypothesis that the rate of erythropoiesis is determined by the amount of erythropoietin, the production of which is regulated by the relationship between O2 supply and demand, not by either factor alone.

Studies on Erythropoiesis. Part IV. Reticulocyte Response of Hypophysectomized and Polycythemic Rodents to Erythropoietin

Summary and Conclusions 1. Following hypophysectomy of rats, reticulocytes in the peripheral blood fall to a minimal level within a period of 3 to 4 weeks. This reduction represents a 5- to 10-fold decrease from the prehypophysectomy control. This reticulocyte decline is more rapid in 8-week-old rats than in younger animals. 2. Administration of anemic plasma to hypophysectomized rats produces a reticulocyte rise that is many fold greater than that produced by normal plasma or saline. 3. Erythropoiesis, as measured by the number of reticulocytes in peripheral blood, falls to zero in 6 days in mice made polycythemic by repeated intraperitoneal injections of homologous red cells. Anemic plasma obtained from rats or rabbits, when injected intravenously or intra-peritoneally into polycythemic mice, produces a reticulocytosis many times greater than that observed from normal plasma. Normal saline produces no increase in reticulocytes. 4. In polycythemic mice and hypophysectomized rats, reticulocyte determinations in the peripheral blood and Fe59 red cell incorporation studies are of about equal sensitivity in measuring the response to anemic plasma.

THE ERYTHROPOIETIC-STIMULATING EFFECTS OF ANDROGENS

Several authors have reported that testosterone is capable of stimulating erythropoiesis in some patients with various types of anemias.13 More knowledge of the mechanism by which testosterone affects erythropoiesis might improve our understanding of the pathogenesis of these anemias and thereby lead to a more rational basis for their therapy with androgens. Blacher first suggested that the sex hormones influence erythropoiesis after he described differences in the red blood counts of male and female fowl.4 Shortly thereafter Juhn and Domm showed that these differences were dependent on the presence of intact gonads in the male.5 Vollmer et al.6,7 showed that testosterone propionate stimulated erythropoiesis in female rats, in castrated male rats and in hypophysectomized rats. In this paper we shall report the results of studies on the mechanism by which androgens and other anabolic steroids stimulate erythropoiesis in mice. We will also discuss the effect of testosterone on mice with anemia resulting from mutation at the W locus. Finally we shall speculate on the relationship between these findings and the way that testosterone stimulates erythropoiesis in anemic patients.

Erythropoietic effect of plasma from mice receiving testosterone.

THE clinical effectiveness of testosterone in some previously refractory anaemias1–3 has aroused interest in its mechanism of action.

Studies of erythropoietin: the hormone regulating red cell production.

There is mounting evidence that erythropoiesis is controlled by a humoral factor or factors. Since Carnot and DCflandrel first published this thesis in 1906, many investigators have attempted to substantiate their work, but it has been largely in the last twenty years that such investigators as Reissmann,Z Erslevj3 Gordon and his co-~orkers ,~ Borsook et u Z . , ~ and Stohlman et aL6 have succeeded in bringing the problem to the point of serious study and acceptance. In 1952 Grant and Root7 wrote a comprehensive review of the literature on the subject. The first convincing support of Carnot’s hypothesis is found in the work of Reissmann,2 and Stohlman et uL6 Reissmann demonstrated that increased erythropoiesis occurred in both parabionts even though only one of the pair was maintained in low O2 tension. Stohlman and Rath found that erythroblastic hyperplasia was not confined to the areas of regional hypoxia in a human being with regional hypoxia secondary to a patent ductus arteriosus. Hyperplasia was also observed in the bone marrow that was supplied by blood with a normal 0 2 saturation. Of necessity, the relationship of the humoral factor that is present in anemic plasma to the maintenance of a normal amount of red blood cells and hemoglobin has been one of conjecture until recently, when Gurney and his associates demonstrated its presence in normal plasma.* The exact relationship of arterial oxygen saturation to the production of circulatory erythropoietint and, thus, to erythropoiesis has likewise not been established. During the past several years we have been concerned with: (1) applying the technique of Fe69 incorporation as a measure of red cell production; (2) finding more sensitive assay preparations for erythropoietin; (3) elucidating the role of erythropoietin in maintaining the dynamic equilibrium of erythropoiesis; (4) determining the basic conditions that control erythropoietin blood levels and, thus, the rate of erythropoiesis; (5) studying the effect of cobaltous ion on erythropoietin production; (6) searching for the site of erythropoietin production; and, of course, (7) using the information obtained to solve certain clinical problems. A brief discussion of our research and some interpretations of the data are the subjects of this paper.

Erythropoiesis. II. Assay of erythropoietin in hypophysectomized rats.

Summary and Conclusions Using incorporation of Fe59 into newly-formed red cells as index of red cell production, it has been shown that erythropoiesis is gradually reduced, reaching a minimum in rats at 8 to 13 days after hypophysectomy. A factor of 10 exists between incorporation of Fe59 into red cells of normal control (37 ± 4) and the hypophysectomized rat (4 ± 2) at this interval. We have also found that the hypophysectomized rat is an extremely sensitive preparation for assay of factor(s) in anemic plasma that stimulates or mediates erythropoiesis. Our observations may be summarized as follows: 1. Administration of anemic plasma to hypophysectomized assay animal increases incorporation of Fe59 3- to 7-fold. 2. Plasma from hypophysectomized animals and un-operated controls made anemic by repeated phlebotomy increases Fe59 red cell incorporation to the same extent when administered to the hypophysectomized assay animal. 3. A single injection of 2 ml of anemic plasma to the hypophysectomized assay animal elicits a 2- to 3-fold increase in Fe59 incorporation. The mechanism of rapid reduction of erythropoiesis that follows hypophysectomy in rats and its relationship to increased sensitivity of the hypophysectomized animal to anemic plasma are discussed briefly.

Erythropoietic Effect of Testosterone in the Polycythemic Mouse

Summary Testosterone has been demonstrated to stimulate erythropoiesis in the polycythemic mouse. A response can be observed following a single injection of 0.5 mg of testosterone. One mg is as effective as larger single doses. A greater erythropoietic response was observed with 10 daily injections. Possible mechanisms of action of testosterone on erythropoiesis are discussed.

Use of Mild Plethora to Demonstrate an Erythropoietic Effect from Small Amounts of Androgens

Summary Two daily injections of as little as 0.1 mg testosterone are capable of stimulating erythropoiesis in mildly plethoric mice. Twenty-five hundredths of a milligram of testosterone is sufficient to elevate the erythropoietin titers of hypoxic mice.

Combined Effect of Cobalt and Testosterone on Erythropoiesis

Summary It has been demonstrated that a combination of cobalt and testosterone serves to potentiate erythropoiesis in the polycythemic mouse, and acts synergistically to increase erythropoietin levels in the plasma of the normal mouse. When these 2 substances are used together, the erythropoietic response in plethoric animals and the amount of erythropoietin elaborated in normal animals are substantially greater than the sum of the responses produced by each agent separately.

Hematologic complications of chronic renal failure.

Uremia interferes with erythropoiesis, granulocyte, platelet, and immune functions. As a result, uremic patients are almost invariably anemic, and have a high incidence of infections and hemorrhagic complications. The anemia of renal failure, which is caused primarily by damage to the site of erythropoietin production is often complex, and complicated by hemolysis from a variety of mechanisms, iron deficiency, and so forth. Although hemodialysis ameliorates some of the hematologic complications to a variable degree, they remain a serious hinderance to the well being of this group of patients. Progress in understanding the mechanism of these problems and their therapy has been reviewed here.