Primaquine-induced hemolytic anemia: Role of splenic macrophages in the fate of 5-hydroxyprimaquine-treated rat erythrocytes

Primaquine- induced hemolytic anemia is known to result from premature sequestration of damaged ( but intact) erythrocytes by the spleen. We have shown previously that a phenolic metabolite, 5- hydroxyprimaquine ( 5- HPQ), is a direct- acting hemolytic agent in rats, suggesting that 5- HPQ is a mediator of the hemolytic response to primaquine. To investigate the fate of erythrocytes in vivo after in vitro exposure to 5- HPQ, rat Cr-51-labeled erythrocytes were incubated with hemolytic concentrations of 5- HPQ and then readministered intravenously to rats. The time course of loss of radioactivity from blood and uptake into the spleen and liver was measured. In rats given 5- HPQ-treated erythrocytes, an increased rate of removal of radioactivity from the circulation was observed as compared with the vehicle control. The loss of blood radioactivity was accompanied by a corresponding increase in radioactivity appearing in the spleen but not in the liver. When rats were pretreated with clodronate- loaded liposomes to deplete splenic macrophages, there was a decreased rate of removal of radioactivity from the circulation and a markedly diminished uptake into the spleen. A role for phagocytic removal of 5- HPQ- treated red cells was confirmed in vitro using the J774A. 1 macrophage cell line. Furthermore, depletion of red cell GSH with diethyl maleate significantly enhanced in vitro phagocytosis of 5- HPQ- treated red cells. The data indicate that splenic macrophages are responsible for removing 5- HPQ- treated red cells and support the postulate that this metabolite is a contributor to the hemolytic anemia induced after administration of the parent compound.