Iron is one of the essential elements required for proper cell growth and proliferation1. Iron is distributed throughout the body in two forms: (i)hemic-iron, mostly found in the hemoglobin, myoglobin and cytochromes and (ii) non-hemic-iron, as a cofactor of numerous enzymes1. Dietary iron is absorbed in the small intestine1. Most of the iron in the body (2-3g of total iron) is found in hemoglobin, a major oxygen-transport metalloprotein found in red blood cells2. Furthermore, once an erythrocyte is degraded at the end of its life span, its iron content is immediately bound by iron-binding plasma proteins -transferrins, as an iron source for erythropoiesis1,2. Correct iron concentration in the body is crucial, as its deficiency or excess causes tissue and cell dysfunction. Iron deficiency restricts the process of iron uptake by erythrocyte precursors, thus limiting hemoglobin synthesis and causing anemia2. Furthermore, synthesis of ferroproteins in other cells may be endangered, which can affect muscle performance and maintenance of epithelial surfaces2. Excessive iron accumulation leads to tissue and cell damage, through a presumable generation of reactive oxygen species1,2.


Iron homeostasis and proteins involved

There are multiple proteins and processes that allow a controlled storage and transport of iron throughout the body. Iron is imported to human cells through endocytosis in a transferrin-bound form1. Consequently, transferrin-bound iron is released from the endosome into the cytoplasm by a transporter protein - divalent metal transporter 1 (DMT1)1. Further iron retention in the cell is possible by presence of a major iron storage protein - ferritin, that sequesters and stores iron ions in ferric form (Fe3+)1. Release of iron into the cytoplasm occurs through reduction of ferric (Fe3+) to ferrous (Fe2+) ions 1. Finally, an iron exporting protein -ferroportin, together with other ferroxidases, delivers iron ions to the plasma1. During that proces iron is converted from ferrous to ferric form1. These reduction and oxidation steps are crucial, for cell storage of iron ions and further iron binding to transferrins in the blood.

Ferroportin and hepcidin, a cationic peptide hormone synthesized mainly by hepatocytes, are one of the important players involved in iron homeostasis. It has been shown that ferroportin is downregulated by a pro-inflammatory cytokine interleukin-6 (IL-6)3. Ferroportin can also by regulated by hepcidin, another essential player, which controls iron homeostasis by binding, and degradation of ferroportin4. Hepcidin, like ferroportin is also controlled by other factors. Specifically, hepcidin is regulated transcriptionally by iron stores5, a mechanism involving several pathways that allow direct sensing of iron levels by hepatocytes. Moreover, inflammation, and pro-inflammatory cytokines such as IL-6, IL-1 alpha and IL-1 beta upregulate hepcidin synthesis6. Together, these data present multiple factors that can lead to ferroportin degradation and consequent decrease in iron export from cells into plasma.

As a consequence of decreased iron export form the cells, iron concentration increases within the cells that leads to overload. Simultaneously at the systemic level iron deficiency (ID), iron deficiency anemia (IDA) and anemia of inflammation (AI) becomes established1. ID, IDA and AI are the most common iron disorders affecting over 30% of the world´s population1. Programs that intend to prevent the development of ID and IDA rely on oral iron administration. However, studies show that ferrous iron supplementation fails to restore iron homeostasis in patients with ID or IDA, while causing side effects such as nausea, vomiting, diarrhea and gastrointestinal discomfort7-9.Furthermore, a previous in vivostudy revealed that rats fed with an iron-enriched diet produced a higher amount of reactive oxygen species, than rats fed with normal diet10. Together these data show that iron deficiency is a challenging condition to treat.


Lactoferrin, and its function in iron homeostasis

Lactoferrin is an iron-binding glycoprotein, with multifunctional activities, identified in 1939 in bovine milk and isolated in 1960 from human milk11. Interestingly, lactoferrin exists in two conformational states: (i) metal-free, open state (apo-lactoferrin), and (ii) metal-bound, closed state (holo-lactoferrin) 12,13. Lactoferrin can reversibly chelate two Fe3+ions per molecule with high affinity, Kd ~ 10-20M1.In addition, lactoferrin can also bound other ions such as copper (Cu2+)14and manganese (Mn3+)15, however with lower affinity than to Fe3+without change in structure1.

Majority of in vivo and in vitro studies have used bovine lactoferrin, and recognized it as a safe substance (GRAS) by the Food and Drug Administration1. Recently lactoferrin was found to bind to heme-iron16. This is of importance since human absorption of iron low, when compared to other nutrients, specifically absorption of heme-iron (contained in animal products) is 15-25% while the nonheme iron (contained in vegetables and cereals) is 2-5%. Thus lactoferrin could improve delivery of iron in iron-depleted individuals. Interestingly, a preventive study from 2008 by Koikawa et al.17, suggests that intake of lactoferrin increases the utilization and absorption of iron and can be useful in prevention of anemia among female long distance runners. More recently, lactoferrin was found to counteract inflammation-caused changes in iron balance in macrophages3, which could open up new venues for therapeutic use. Together, these data suggest that lactoferrin could be a candidate for use in individuals suffering from iron-deficiency or anemia.


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