A compensated hemolytic state was induced in rats by injection of phenylhydrazine (PHZ) over a 6-week period. The liver and kidney were perfused to determine the levels and time of appearance of a hepatic erythropoietic factor (HEF) which induces the production of hepatic erythropoietin (Ep) and its antagonist, a renal inhibitory factor (RIF). Erythropoietin assays on the perfusates have been previously reported. The amount of HEF in perfusates recovered from the livers of the PHZ-treated rats was significantly higher during the 4th to 5th weeks of treatment, coinciding with the time of increased liver production of Ep. During the 6th week of PHZ treatment, the titer of RIF in perfusates recovered from the kidney was markedly increased and the HEF titer was decreased to near the control level, suggesting inhibitory action of RIF on HEF synthesis and/or effect. These findings indicate that an HEF/RIF control mechanism regulates hepatic Ep production in this compensated hemolytic condition.

Rats were injected with phenylhydrazine (PHZ) for periods of up to 6 months, during which time a marked leukocytosis was induced. The highest leukocyte counts occurred within 4-5 days following injection. An initial injection of 4 mg/100 gm body weight evoked a mean total leukocyte count of 129 X 10(3) cells/microliter. Successive weekly injections of 2 mg/100 gm resulted in a mean total leukocyte count of 70 X 10(3) cells/microliter compared to a mean total leukocyte count of 12.5 X 10(3) cells/microliter in saline-injected rats. Lymphocytes and monocytes accounted for approximately 75% of the total cell counts in both the PHZ-treated and control rats. The presence of increased numbers of mononuclear cells was confirmed by Percoll gradient separation and by phase-contrast microscopy. Although a leukocytosis was evident when using the automated Coulter electronic cell counter, it was not discernible when blood samples were counted manually in a hemocytometer by light microscopy. Histological examination of the thymus, lymph nodes, and spleen of the PHZ-treated rats indicated that lymphocytes and monocytes were mobilized from these sites. Lymphocyte depletion was evident, and germinal centers were found in all these lymphoid organs, indicating that PHZ induced a lymphopoietic response. A possible autoimmune etiology for PHZ-induced red blood cell destruction is discussed.

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The immunoactive steroid dexamethasone (DXM) was administered to rats injected with a dose of phenylhydrazine (PHZ) known to induce anemia. PHZ treatment alone resulted in a hemolytic anemia that was most pronounced on Days 1-7 after injection. This anemia was accompanied by a leukocytosis that was greatest on Days 2-7 following PHZ treatment. Lymphocytes accounted for greater than 75% of this incremental increase. In contrast, rats treated with PHZ as well as with DXM displayed erythrocyte counts and hematocrits within the normal range. Although the reticulocyte counts of these rats were higher than those of controls, they were significantly lower than those of animals receiving PHZ alone. In addition, DXM suppressed the leukocytosis and splenomegaly resulting from PHZ administration and inhibited the rise in plasma IgG titers induced by PHZ exposure. DXM also altered the ratio of peripheral blood T and B lymphocytes of PHZ-treated rats. DXM suppression of PHZ-induced anemia is further confirmation that this anemia is associated with immune activation.

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