Iron chelation suppresses mononuclear cell activation, modifies lymphocyte migration patterns, and prolongs rat cardiac allograft survival in rats.

Iron influences host immunity, in part by affecting the function and migration patterns of T cell subpopulations. Removal of iron stores from the body by chelation decreases proliferation and differentiation of T cells, as shown in models of autoimmunity and pancreatic islet transplantation. We have examined the influence of iron chelation on rejection of vascularized heart allografts in rats. Two protocols were investigated: "treated" recipients received desferrithiocin (30 mg/kg/day) orally for 10 days beginning the day of transplantation; "pretreated" hosts received a similar dose of the drug for 10 days before engraftment. Graft survival increased from about 7 days in untreated animals to 14-16 days in both treatment groups (P < 0.001). Histological and immunoperoxidase studies of allografts at day 7 showed that iron chelation resulted in only a mild reduction in cell infiltration, but in a marked decrease in graft edema and interstitial hemorrhage and essentially complete suppression of mononuclear cell activation and cytokine production. Chelation therapy was also found to inhibit profoundly cytokine (TNF-alpha) production in rats treated with LPS, consistent with the effects observed in situ in the allografts. In vitro studies showed that pretreatment significantly inhibited the mixed lymphocyte reaction. Chelation also influenced migration of T lymphocyte subsets: treatment stimulated migration of CD4+ lymphocytes to peripheral lymph nodes; pretreatment strikingly and selectively increased CD8+ cell migration into parathymic lymph nodes draining the graft, with the opposite effect on nondraining node groups. We conclude that treatment with iron-chelating agents has several effects on host alloresponsiveness in a rat heart graft model secondary to inhibition of immune activation; these include prolongation of graft survival, inhibition of the mixed lymphocyte reaction (pretreatment), marked depression of cytokine production, and alteration in recirculation patterns of lymphocyte subpopulations.

Augmentation of human monocyte chemiluminescence by iron-saturated lactoferrin.

The effect of Fe3+ bound lactoferrin (LF) on chemiluminescence (CL) generation in human monocytes in the presence and absence of opsonized zymosan was examined. Fe3+ bound LF (10(-6) or 10(-7) M) augmented the CL response in the presence but not the absence of zymosan. In contrast, Apo LF, Fe3+ bound transferrin and Fe3+ itself had no significant effect. It appears that the effect of LF on zymosan-induced chemiluminescence is dependent on iron and is receptor specific. The augmentation of CL by LF was inhibited by superoxide dismutase (SOD) and ETYA (fatty acid cyclooxygenase and lipoxygenase inhibitor), but not indomethacin (fatty acid cyclooxygenase inhibitor). Thus the enhancement of CL by LF depends on superoxide anion production and is related to the lipoxygenase pathway. The significance of this observation in terms of microbicidal or tumoricidal mechanisms remains to be determined.

A subpopulation of human polymorphonuclear neutrophils contains an active form of lactoferrin capable of binding to human monocytes and inhibiting production of granulocyte-macrophage colony stimulatory activities.

Subpopulations of human peripheral blood polymorphonuclear neutrophils (PMN) differing in functional capacity were separated by a rosetting procedure by using rabbit IgG antibody-coated sheep erythrocytes (EA). EA+ and EA- populations of PMN both contained lactoferrin (LF) as determined by radioimmunoassay. However, only LF (10(-16) to 10(-6) M) obtained from EA+ (= FC receptor (FcR)+) PMN was able to suppress the production of granulocyte-macrophage colony stimulatory activity derived from human monocytes. LF from EA- PMN was inactive at concentrations as high as 10(-5) M. Extracts of EA- PMN contained proteolytic enzymes, not apparent in FcR+ PMN, that inactivated the inhibitory activity of LF obtained from FcR+ PMN. These results were substantiated by immunofluorescence studies. LF from FcR+ PMN bound to monocytes whereas LF from EA- PMN demonstrated negligible binding to the monocytes and extracts from EA- PMN drastically decreased the capability of LF obtained from FcR+ PMN to bind to monocytes. These studies demonstrate functional heterogeneity of peripheral blood PMN and suggest a role for PMN subpopulations in the regulation of myelopoiesis.

The transfer of retinol from serum retinol-binding protein to cellular retinol-binding protein is mediated by a membrane receptor.

The hypothesis that the cellular uptake of retinol involves the specific interaction of a plasma membrane receptor with serum retinol-binding protein (RBP) at the extracellular surface followed by ligand transfer to cytoplasmic cellular retinol-binding protein (CRBP) has been investigated. The experimental system consisted of the [3H]retinol-RBP complex, Escherichia coli-expressed recombinant apo-CRBP containing the 10 amino acid long streptavidin-binding peptide sequence at its C terminus (designated as CRBP-Strep) and permeabilized human placental membranes. [3H]Retinol transfer from RBP to CRBP-Strep was monitored by measuring the radioactivity associated with CRBP-Strep retained by an immobilized streptavidin resin. Using this assay system, we have demonstrated that optimal retinol uptake is achieved with holo-RBP, the membrane receptor and apo-CRBP. The effects are specific: other binding proteins, including beta-lactoglobulin and serum albumin, despite their ability to bind retinol, failed to substitute for either RBP or apo-CRBP. The process is facilitated by membranes containing the native receptor suggesting that this protein is an important component in the transfer mechanism. Taken together, the data suggest that the RBP receptor, through specific interactions with the binding proteins, participates (either directly or via associated proteins) in the mechanism which mediates the transfer of retinol from extracellular RBP to intracellular CRBP.