Lactoferrin effects of phagocytic cell function. II. The presence of iron is required for the lactoferrin molecule to stimulate intracellular killing by macrophages but not to enhance the uptake of particles and microorganisms.

Human lactoferrin (LF)--a neutrophil glycoprotein, the body fluid levels of which increase in inflammatory conditions--stimulates the phagocytic and cytotoxic properties of macrophages. We found in this work that, whereas the presence of iron in the LF molecule was not required to increase the capacity of mouse peritoneal macrophages (MPM) to take up Trypanosoma cruzi amastigotes (AMA), Listeria monocytogenes, or latex particles, it was necessary for LF to enhance intracellular killing of the two microorganisms. Thus, iron-free human lactoferrin (ApoLF), which did not increase MPM cytotoxicity, after restoration of ferric ions prior to its use in MPM treatments or when ferric citrate was added to the culture medium immediately after ApoLF treatment of the MPM, does increase MPM cytotoxicity. In that iron ions cannot be internalized as such, the latter observation suggested that ApoLF had taken up iron while membrane bound and then enhanced killing. Immunofluorescence studies revealed that comparable proportions of MPM-bound ApoLF or LF at either 20 or 100% iron saturation without appreciable differences in fluorescence intensity. Therefore, reduced binding of ApoLF compared with LF was not a likely explanation for the lack of effect of ApoLF on MPM killing. LF did not enhance AMA killing by MPM in the presence of the iron chelator deferoxamine. Diethylaminetriamine-pentaacetic acid, an iron chelator which is not incorporated into cells, had a similar effect. The iron-binding protein transferrin did not alter the capacity of MPM to either take up or kill the AMA, indicating that the noted LF effects were not shared by all iron-binding proteins. However, prior treatment of MPM with transferrin enabled the cells to display a greater parasite killing capacity after ApoLF treatment, suggesting a role for iron in this activity. Whether iron is required for LF to impart the signal that elicits enhanced killing, to satisfy a biochemical requirement for more effective killing, or both, remains to be clarified. We also found that killing of internalized AMA by LF-treated MPM--previously reported to be mediated in part by H2O2, O2-., and 1O2--was inhibitable by scavengers of OH., and therefore, appears to involve this oxygen metabolite as well.