Diverse membrane-active agents modify the hemolytic response to ferriprotoporphyrin IX.

Because ferriprotoporphyrin IX (ferriheme, FP) is a lytic agent that can be released by degradation or oxidative denaturation of hemoglobin, we measured the hemolytic responses of human erythrocytes to FP alone or to FP in combination with various membrane-active agents. Suspensions of erythrocytes (0.5%) incubated at pH 7.4 and 37 degrees C were hemolyzed by FP alone in concentrations of 10 microM or greater. Preincubation of the erythrocytes with nonhemolytic concentrations of chloroquine, mefloquine, quinine, calcium, lanthanum or manganese potentiated the hemolytic response to FP. For example, hemolysis in the presence of 5 microM FP was 5%; in the presence of 5 microM FP and 20 microM chloroquine, hemolysis exceeded 80%. For 5 microM FP, maximal potentiation was obtained with 20 microM chloroquine, 200 microM quinine or 1 mM calcium. Paradoxically, with 5 microM FP and a high concentration of chloroquine (1 mM), hemolysis did not exceed the base-line value of 5%. In addition, all of the agents that individually potentiated the hemolytic response to 5 microM FP also inhibited hemolysis when used in combination with 5 microM chloroquine and 5 microM FP. Detailed studies of calcium showed that it inhibited the increase in osmotic fragility, which was induced by FP and chloroquine, without inhibiting the increase in binding of [14C]chloroquine to erythrocytes treated with FP. These results can be rationalized by proposing that the structure of the erythrocyte membrane at the time of exposure to FP and membrane-active agents determines whether the hemolytic response will be potentiated or inhibited.