How Often Should You Eat Liver for Iron Intake? | Healthy Eating

By Jody Braverman Updated December 06, 2018

Love it or hate it, liver is a good-for-you food. Packed with vitamins and minerals, including iron, that you need for optimal health, it is also low in calories and fat. Even though liver is a great source of easily metabolized iron, it’s best to eat only a maximum of a couple times a week because of its high levels of vitamin A.

Your Liver Limit

Liver is the most concentrated dietary source of vitamin A, a fat-soluble vitamin important for healthy vision, immune function and reproduction. A 4-ounce portion of beef liver has 19,095 international units, or IU, of vitamin A, and 4 ounces of lamb liver has 27,812 IU. That is about 400 to 550 percent of the daily value for vitamin A.

In addition, the upper limit – the amount of a nutrient you should not exceed in a day – s set at 10,000 IU for adults. As a fat-soluble vitamin, excess vitamin A is stored in the body for a long period of time. Consistently exceeding the upper limit for vitamin A can lead to toxicity, causing nausea, dizziness, headaches, joint and bone pain, and even coma and death. Some research also suggests that high intakes of vitamin A may be associated with a reduction in bone mineral density and increased risk of fractures, according to the National Institutes of Health.

How Much Is Too Much?

Liver is also high in cholesterol, with 311 milligrams in 4 ounces of beef liver and 419mg in the same portion of lamb liver. As it becomes more clear that dietary cholesterol has little effect on cholesterol levels in the body, a maximum intake recommendation of 300mg per day has been removed from the current Dietary Guidelines for Americans. Still, people with heart disease and diabetes should be careful about how much dietary cholesterol they consume.

For everyone else, the cholesterol in an occasional serving of liver isn’t a big deal. And, most likely, neither is the vitamin A content. However, it’s still a good idea to keep your liver intake in check. Berkeley Wellness recommends eating small portions of liver only on occasion, and the Weston A. Price Foundation recommends limiting your intake to a 4-ounce serving no more than once or twice per week.

Mix It Up

Eating a wide variety of foods will help you get all the iron you need, in addition to adequate amounts of other nutrients. Other rich sources of dietary iron include oysters, chickpeas, lentils, sardines, beef and dark chocolate. The iron in plant foods isn’t as highly bioavailable as the iron in animal foods. However, you can improve absorption by eating plant foods with animal foods or by consuming a food rich in vitamin C together with the iron source.

10 Iron-Packed Foods for Combatting Anemia and Low Energy

If you’ve been told you’re not getting enough iron, you’re not alone. Iron deficiency is the most common nutritional deficiency globally — especially among children and pregnant women — and the only nutrient deficiency that is widely prevalent in developed countries, according to the World Health Organization. That’s a problem because the mineral plays a number of critical roles in the body, says Sarah Gold Anzlovar, RDN, the Boston-based owner of Sarah Gold Nutrition. “Most well known is that it’s a key component of red blood cells and helps transport oxygen from your lungs to the rest of the body,” says Anzlovar.

Iron deficiency, a condition called anemia, makes it difficult for your red blood cells to deliver oxygen, according to the Mayo Clinic. Symptoms of anemia may include fatigue, chest pain or shortness of breath, cold hands and feet, dizziness and headache, poor appetite, and unusual cravings for substances like ice, dirt, or starch.

How Much Iron Do You Need Per Day?

According to the National Institutes of Health (NIH), here’s how much iron different groups of people need per day:

Nonpregnant Women ages 19 to 50 18 milligrams (mg)

Pregnant Women 27 mg

Women Age 51 and Older 8 mg

Men Age 19 and Older 8 mg

Infants and Children 7 to 16 mg, depending on age

RELATED: A Detailed Guide to Using MyPlate for Healthy Eating

Avoid Consuming Too Much Iron

The NIH cautions against taking in more than 45 mg of iron per day if you are a teenager or adult and more than 40 mg per day among those age 13 and younger.

Heme vs. Non-Heme Iron: What’s the Difference?

“There are two types of iron: heme iron from animal sources and non-heme iron from plant sources,” says Frances Largeman-Roth, RD, author of Eating in Color: Delicious, Healthy Recipes for You and Your Family and a nutrition counselor in private practice in New York City. The NIH also notes that meat, poultry, and seafood contain both heme and non-heme iron.

Heme iron is more easily absorbed by the body than plant-based non-heme iron according to the Cleveland Clinic, so it can be beneficial to get both types of the nutrient in your diet, Largeman-Roth adds. You’ll need to aim for nearly twice as much iron per day (about 1.8 times as much, per the NIH) if you don’t eat meat.

RELATED: Why Are Healthy Eating Habits Important?

Common Foods Can Help You Get Enough Iron

The good news is that a lot of common foods contain iron — from oysters and pumpkin seeds to fortified cereals and red meat.

Here are 10 foods high in iron that can help you get all of the mineral you need.

Iron in Foods | HealthLinkBC File 68d

There are 2 types of iron found in foods:

Your body absorbs heme iron more easily than non-heme iron. However, foods containing non-heme iron are also very important sources of iron in your diet.

See HealthLinkBC File #68c Iron and Your Health for more information on how much iron you need and how to get the most iron from foods.

What foods have heme iron?

Food	Iron (mg) per 75g (2 ½ oz serving)
Pork liver*	13.4
Chicken liver*	9.2
Oysters**	6.3
Mussels	5.0
Beef liver*	4.8
Liver pate, canned*	4.1
Beef	2. 4
Clams	2.1
Sardines, canned	2.0
Lamb	1.5
Tuna/herring/trout/mackerel	1.2
Chicken	0.9
Pork	0.9
Shrimp	0.9
Salmon	0.5
Turkey	0.5
Flounder/sole/plaice	0.2

g = gram, mg = milligram, oz = ounce

*Liver and liver products (e.g. liverwurst spread and liver sausages) are high in vitamin A. Too much vitamin A may cause birth defects, especially during the first trimester. The safest choice is to limit these foods during pregnancy. If you choose to eat liver or liver products, have no more than 75g (2 ½ ounces) per week.

**Pacific oysters tend to be higher in cadmium. Health Canada recommends that adults eat no more than 12 B.C. oysters per month and that children eat no more than 1.5 B.C. oysters per month.

What foods have non-heme iron?

Food	Serving	Iron (mg)
Infant cereal, dry***	28 g (5 tbsp)	7.0
Dried soybeans, boiled	175 mL (3/4 cup)	6.5
Lentils, cooked	175 mL (3/4 cup)	4.9
Pumpkin seeds/kernels, roasted	60 mL (1/4 cup)	4.7
Enriched cold cereal***	30 g	4.5
Dark red kidney beans, cooked	175 mL (3/4 cup)	3.9
Blackstrap molasses	15 mL (1 tbsp)	3.6
Instant enriched hot cereal***	175 mL (3/4 cup)	3. 4
Spinach, cooked	125 mL (1/2 cup)	3.4
Refried beans, canned	175 mL (3/4 cup)	2.7
Edamame, green soybeans, cooked and shelled	125 mL (1/2 cup)	2.4
Medium firm or firm tofu	150 g (3/4 cup)	2.4
Tahini (sesame seed butter)	30 mL (2 tbsp)	2.3
Chickpeas, canned	175 mL (3/4 cup)	2.2
Lima beans, boiled	125 mL (1/2 cup)	2.2
Swiss chard, cooked	125 mL (1/2 cup)	2.1
Bagel	1/2 bagel	1.9
Potato, baked with skin	1 medium	1.9
Seaweed, agar (dried)	8 g (1/2 cup)	1. 7
Prune puree	60 mL (1/4 cup)	1.7
Beet greens, cooked	125 mL (1/2 cup)	1.5
Quinoa, cooked	125 mL (1/2 cup)	1.5
Eggs	2	1.4
Green peas, boiled	125 mL (1/2 cup)	1.3
Quick or large flake oats, prepared	175 mL (3/4 cup)	1.3
Hummus	60 mL (1/4 cup)	1.2
Sunflower seeds/ kernels, dry roasted	60 mL (1/4 cup)	1.2
Tomato sauce, canned	125 mL (1/2 cup)	1.2
Pearled barley, cooked	125 mL (1/2 cup)	1.1
Sauerkraut	125 mL (1/2 cup)	1. 1
Fortified soy beverage	250 mL (1 cup)	1.1
Fancy molasses	15 mL (1 tbsp)	1.0
Shredded wheat***	30 g	1.0
Spinach, raw	250 mL (1 cup)	0.9
Whole wheat bread	35 g (1 slice)	0.9
Whole wheat pasta, cooked	125 mL (1/2 cup)	0.8
Beets, sliced, boiled	125 mL (1/2 cup)	0.7

g = gram, mg = milligram, mL = milliliter, tbsp = tablespoon

***Iron amounts in enriched and prepared foods vary. Check the nutrition label for more information. By 2022, all labels will list the amount of iron in milligrams. Until then, some labels will only list the iron as a percent daily value (%DV). The daily value used is 14 mg (or 7 mg for infant cereals). For example, if a serving of cereal has 25% of the daily value, it has 3.5 mg of iron (0.25 x 14 mg).

Note: Most of the iron values in the above tables come from the Canadian Nutrient File (CNF). If more than one entry for that food item was available in the CNF, an average of the entries was taken.

For More Information

For more nutrition information, call 8-1-1 to speak with a registered dietitian.

Iron homeostasis in the liver

Compr Physiol. Author manuscript; available in PMC 2014 Feb 27.

Published in final edited form as:

PMCID: PMC3936199

NIHMSID: NIHMS551241

Erik R Anderson

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

Yatrik M Shah

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

The publisher’s final edited version of this article is available at Compr PhysiolSee other articles in PMC that cite the published article.

Abstract

Iron is an essential nutrient that is tightly regulated. A principal function of the liver is the regulation of iron homeostasis. The liver senses changes in systemic iron requirements and can regulate iron concentrations in a robust and rapid manner. The last 10 years have led to the discovery of several regulatory mechanisms in the liver which control the production of iron regulatory genes, storage capacity, and iron mobilization. Dysregulation of these functions leads to an imbalance of iron, which is the primary causes of iron-related disorders. Anemia and iron overload are two of the most prevalent disorders worldwide and affect over a billion people. Several mutations in liver-derived genes have been identified, demonstrating the central role of the liver in iron homeostasis. During conditions of excess iron, the liver increases iron storage and protects other tissues, namely the heart and pancreas from iron-induced cellular damage. However, a chronic increase in liver iron stores results in excess reactive oxygen species production and liver injury. Excess liver iron is one of the major mechanisms leading to increased steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma.

INTRODUCTION

Iron is an essential micronutrient that is a critical component of oxygen transport proteins (hemoglobin and myoglobin) and of numerous metabolic and redox enzymes. The average adult has 2–4 grams of iron, and over 80% is contained in hemoglobin of red blood cells (RBC). Chronic iron deficiency results in decreased hemoglobin production and anemia. Systemic iron levels are tightly controlled through an integrative mechanism that involves iron absorption, storage, and recycling. The past decade has been termed “The Golden Age of Iron Biology”, due to the significant increase in the understanding of the molecular underpinnings of systemic iron homeostasis (7). When iron regulatory pathways are dysregulated, this leads to either excess tissue iron or iron deficiencies, which affect over a billion people worldwide. Four major cell types or tissues have been shown to be critical for systemic iron homeostasis ():

Systemic iron regulation

Dietary iron is absorbed through the small intestine and mainly utilized for RBC production. Hepatic and splenic macrophages recycle iron from senescent RBCs. The iron derived from recycling is used for production of RBCs. During times of iron excess the liver can store iron and during increased systemic needs the liver can mobilize iron stores for utilization.

Enterocyte

Dietary iron absorption is tightly regulated in the small intestine. Dietary iron enters the body through absorptive cells in the duodenum. Dietary ferric iron (Fe⁺³) is reduced by the apical ferric reductase duodenal cytochrome b (DcytB) to ferrous iron (Fe⁺²) and transported into the enterocyte via an apical iron transporter, divalent metal transporter-1 (DMT1, also known as Nramp2, SLC11a2 and DCT1) (49, 69, 100, 107, 109). Once iron enters into the cytoplasm through DMT1, iron is either stored or exported by the iron exporter ferroportin (FPN, also known as SLC40A1)(1, 43, 110) located on the basolateral side of the enterocyte. Disruption of DMT1 or FPN leads to inhibition of iron absorption and dysregulation of systemic iron homeostasis (43, 68). Following export of iron through FPN, iron is oxidized back to the ferric form by the ferroxidase hephaestin and ceruloplasmin (Cp) and loaded onto transferrin (Tf) and circulated through the body (70).

Erythroblast

A significant portion of circulating Tf-bound iron is utilized in early RBC precursors called erythroblasts for the synthesis of hemoglobin. Hemoglobin comprises of about 95% of the total cellular protein of the mature RBC and more than 80% of functional iron in the body is found in hemoglobin (28). Iron is an essential cofactor for the ability of RBCs to transport oxygen, and thus a decrease in body iron levels is the most predominant cause of anemia worldwide.

Macrophage

Only 1–2 mg per day of dietary iron is required to be absorbed through the intestine. This is due to the highly efficient recycling of iron from senescent erythrocytes. Recycling is performed by splenic and hepatic macrophages. Aging erythrocytes at 120 days old undergo specific changes that can be recognized by macrophages, thus initiating erythrophagocytosis. Iron is recovered from the degradation of hemoglobin and heme by hydrolytic enzymes in the phagocytic vesicles and heme-oxygenase-1 (HO-1) (58). Iron is routed back to circulation through the basolateral iron transporter FPN. Recent studies using macrophage-specific conditional deletion of FPN demonstrate the importance of FPN in exporting iron out of macrophages following erythrophagocytosis (189, 190).

Liver

The liver performs three essential functions in maintaining systemic iron homeostasis: 1) The liver is the major site for production of proteins that maintain systemic iron balance, 2) It is a storage site for excess iron and 3) the liver is critical for the mobilization of iron from hepatocytes to the circulation to meet metabolic requirements. Dysregulation of the liver’s ability to maintain balance of these three parameters leads to iron-related disorders. This review will focus on the normal function of the liver in iron homeostasis and the role of liver in iron-related disorders.

PRODUCTION OF PROTEINS FOR IRON HOMEOSTASIS

(Tf)

Tf is an 80 kDa plasma glycoprotein generated predominantly in the liver (138). However, small amounts are also made in the brain and testis (14, 157). Tf is the major serum iron-binding protein and is essential for systemic circulation of iron. Tf consists of two globular lobes of α-helices and β-sheets, which have a significant amount of homology between the N- and C- terminal halves of the molecule. This homology is thought to be due to a gene duplication arising from a 40kDa ancestral protein containing a single lobe (3, 96-98). Each lobe includes metal binding amino acid residues (2 Tyr, 1 His, and 1 Asp). Tf binds to ferric iron reversibly and with high affinity, but does not bind to ferrous iron (3, 4). In addition, Tf can also bind other metals with lower affinity (3). The affinity of iron is also regulated by pH, as iron binding to Tf decreases as the pH drops, with no detectable amounts observed below pH 4.5 (3, 4). The reversible binding of iron to Tf is important and allows it to be a cellular iron donor or iron acceptor depending on the systemic requirements of iron.

(Cp)

Cp is a copper-dependent serum ferroxidase which works in tandem with FPN for cellular iron export into circulation (129, 130). Cellular iron is usually in the ferrous form and is exported out of the cell by FPN, however the affinity of Tf for ferrous iron is relatively weak and Cp is required to oxidize iron to its ferric form, which binds with high affinity to Tf (129, 130). Disruption of Cp in mouse models further supports the role of Cp in systemic iron homeostasis (30, 70).

Ferritin

Ferritin is a multi-subunit protein that consists of a heavy and light chain. Ferritin from different tissues can vary in the ratio of heavy compared to light chains. Liver ferritin contains mainly the light chain and can store up to 4500 atoms of iron. Hepatocytes are the major site for ferritin synthesis, however most cells that have been assessed can synthesize ferritin to a smaller degree (12, 167). Cells with high iron levels are capable of an adaptive increase in ferritin synthesis. Both ferritin subunit synthesis are controlled through a post-transcriptional mechanism by the iron response element (IRE)/iron regulatory protein (IRP) system. The ferritin mRNAs contain a single IRE in the 5’ UTR. Under iron deficient conditions, the IRPs bind to this IRE and repress translation. However, as intracellular iron increases in a cell, the repression is relieved and ferritin synthesis is increased. For more details of the IRE/IRP system, refer to the comprehensive review by Muckenthaler, M.U et al. (113). Although the IRP/IRE regulation of ferritin synthesis is the predominant pathway that allows an adaptive increase, several other mechanisms, such as IL-1, TNFα, hypoxia, and oxidative stress have also been observed to regulate ferritin expression (23, 169, 170, 173, 184). It is likely that a combination of these pathways allow an efficient increase in ferritin when required.

Hepcidin

Hepcidin is a 25 amino acid antimicrobial peptide that is produced in hepatocytes and secreted into the circulation (91). Hepcidin is translated as an 84 amino acid pro-protein, and cleaved by the pro-hormone convertase furin to produce the active peptide (175). Subtractive hybridization experiments between iron loaded and control mouse livers initially identified hepcidin as an iron-regulated protein (136). The role of hepcidin in iron homeostasis was further confirmed when hepcidin was inadvertently disrupted in addition to USF2 and these mice were severely iron overloaded (119). The effect of hepcidin knockout on iron homeostasis was confirmed in a subsequent paper, which showed that animals deficient in hepcidin, but having normal USF2 expression developed severe hemochromatosis (101). Conversely, animals that overexpress hepcidin in the liver demonstrate severe iron-deficiency anemia (120). A clue to the function of hepcidin was discovered after subjecting rats to an iron-deficient diet. Following iron deficiency, rats rapidly repress liver hepcidin and upregulate the intestinal iron transporters DMT1, DcytB, and FPN (54). Hepcidin was believed to regulate iron homeostasis due to an interaction with iron transporters. A breakthrough came when the hepcidin receptor was discovered. Through careful in vitro analysis, it was determined that hepcidin binds to FPN which leads to its internalization and proteasomal degradation (117). Hepcidin is increased in iron loading, which leads to a decrease in duodenal iron absorption to normalize iron levels. In the case of iron deficiency, hepcidin is repressed which allows more iron to be transported from the enterocyte to the serum (). In addition, FPN is increased in macrophages following erythrophagocytosis, and hepcidin represses FPN in these cells (89). In macrophages during iron loading, hepcidin decreases iron transport following iron recycling from senescent RBCs. Whereas during iron deficiency, a significant increase in iron efflux would be expected following erythrophagocytosis (). In addition to iron levels resulting in the regulation of hepcidin levels, hypoxia and erythropoiesis are major repressors of hepcidin expression. This provides a novel link between oxygen homeostasis and iron levels. Moreover, inflammation is a major activator of hepcidin expression. This results in restriction of serum iron levels during an infection, and therefore is less conducive for growth of pathogenic bacteria. The last decade has shown the importance of hepcidin as the master regulator of both duodenal iron absorption and red blood cell (RBC) iron recycling. Studying the regulation of hepcidin expression in the liver has been a priority for understanding regulation of systemic iron homeostasis. The transcriptional regulation of hepcidin has been reviewed elsewhere (56, 114) and is summarized in . Here, we only briefly mention a few major pathways that are critical in hepcidin regulation. In addition to the major pathways, several other accessory proteins have been identified, which are mutated in iron-related disorders. Their importance in hepcidin regulation is covered in more detail below in the liver iron overload and anemia section.

Hepcidin regulation of FPN protein expression during changes in systemic iron levels

High iron levels increase hepcidin expression, which decrease iron export from the small intestine and macrophage due to an internalization and degradation of FPN. Iron deficiency results in a decrease in hepcidin levels and stabilization of FPN protein expression.

Regulation of hepcidin by BMP/SMAD, inflammatory and hypoxia/erythropoietic signaling in the liver

Three major pathways are critical for regulating basal and stimuli-induced hepcidin expression. Binding of iron containing Tf to Tfr1 causes a dissociation of Tfr1-High FE (HFE) complex and an interaction of HFE with Tfr2. Increased stabilization of Tfr2 increases BMP6 mediated phosphorylation of SMAD1/5/ 8 and recruitment of SMAD 1/5/8 and SMAD4 to the hepcidin proximal promoter. BMP/SMAD signaling is the major pathway by which hepcidin expression is coordinated to meet systemic iron requirements. Activation of hepcidin by inflammation is thought to act independently of the BMP/SMAD pathway. The best-studied mechanism is via the pro-inflammatory mediator IL-6. Binding of IL-6 to its receptor IL-6 receptor (IL-6R) initiates activation of the JAK-STAT3 pathway. STAT3 binds directly to the proximal promoter to increase hepcidin expression. Hypoxia and erythropoiesis are inhibitors of hepcidin expression and these are the least understood pathways by which hepcidin expression is regulated. Hypoxia and erythropoiesis have been shown to inhibit hepcidin expression via direct binding of HIF to the proximal promoter, an EPO-EPO receptor (EPOR) mediated decrease in C/EBPα expression, and through increase in an unknown erythroid derived factor which signals through an undefined pathway.

SMAD/ bone morphogenetic protein (BMP) signaling

A key finding in the regulation of hepcidin is the essential role of the BMP-SMAD signaling cascade. BMPs are ligands that belong to the transforming growth factor-β (TGF-β) superfamily. BMPs bind to type I and type II serine threonine kinase receptors, which phosphorylate specific intracellular SMAD proteins (SMAD1/5/8). Phosphorylated SMAD1/5/8 (P-SMAD1/5/8) binds to the common mediator SMAD4, and the SMAD complex translocates to the nucleus to modulate transcription of target genes. BMPs, but not TGF-β signaling induced hepcidin expression in cultured liver cell lines and in vivo (11). Several BMPs signal through SMAD activation, however BMP6 is the endogenous ligand that modulates hepcidin expression. BMP6 knockout mice have decreased hepcidin expression and an increase in tissue iron (8, 10, 11, 111). BMP6 knockout mice still have the ability to increase hepcidin expression following inflammatory stimuli. The importance of SMAD signaling in hepcidin regulation was demonstrated in mice with a hepatocyte-specific disruption of SMAD4. A near complete loss of hepcidin expression was observed in these mice and eventually the mice die of severe iron overload in multiple tissues (183). Interestingly, liver SMAD4 knockout mice are unable to increase hepcidin expression in response to iron loading, suggesting that SMAD4 mediates the response of hepcidin to changes in systemic iron requirements. Signaling through the BMP receptor leads to SMAD1/5/8 phosphorylation, which is required for SMAD4 transcriptional activity (6). The regulation of hepcidin by iron loading and iron deficiency is correlated with phosphorylation of the SMAD1/5/8 proteins. In iron deficiency, pSMAD1/5/8 is decreased dramatically, and is substantially increased in conditions of iron overload (85).

STAT3 and inflammatory pathways

Hepcidin is induced in response to inflammatory stimuli (57, 118). Hepcidin induction by inflammation leads to iron sequestration, which can decrease bacterial growth. However, chronic diseases are associated with anemia (150). The mechanism for the induction of hepcidin by inflammation is mediated by IL-6 in cultured cells, mice, and humans (115). Subsequently, it was shown that IL-6 regulates expression of hepcidin by inducing STAT3 binding to the hepcidin promoter (176, 185). To support this, it was shown that anti-IL-6 receptor antibody improves anemia of inflammation (158).

Hypoxia and erythropoietic pathways

Hypoxia, or low oxygen tension, is a physiological condition that results in numerous adaptive changes in gene expression. Hypoxia represses hepcidin expression both in cultured cells and in mice (121). Many mechanisms have been proposed to play a role in hypoxic hepcidin repression (29, 31, 94, 134, 179). Hypoxia-inducible factor (HIF) is the major transcription factor activated following hypoxia and HIF is thought be critical in the hypoxia-mediated repression of hepcidin (20). HIF was shown to bind directly to the hepcidin promoter resulting in repression (134). However, recent studies refute these findings (108, 179). It has been reported that hypoxia through a HIF2α mediated increase in erythropoiesis is the critical pathway leading to hepcidin repression (108). Interestingly, a mutation that leads to HIF stabilization causes Chuvash polycythemia. In these patients a decrease in hepcidin expression is observed without a significant association with erythropoiesis (65). There is no clear mechanism that mediates hepcidin repression during hypoxia, and this is an active area of study

Erythropoiesis is a well-characterized pathway leading to hepcidin repression. In several mouse models that induce erythropoiesis, a significant decrease in hepcidin is observed (16, 53, 61). The decrease in hepcidin allows the increase of iron required for RBCs during erythropoiesis. In a model of intensive care anemia, erythropoietin injections or phlebotomies were able to repress hepcidin expression despite high levels of IL-6, which is known to strongly increase hepcidin expression (99). Similarly, hypoxia is able to repress hepcidin expression under conditions of high IL-6. One of the mechanisms by which erythropoiesis represses hepcidin is through erythropoietin (EPO) binding to its receptor in hepatocytes, which leads to downregulation of C/EBPα and hepcidin repression (137). C/EBPα is a liver-enriched transcription factor that is important in hepcidin regulation. Mice with a liver specific deletion of C/EBPα have low levels of hepcidin expression (35). However, erythropoietic blockers prevented the suppression of hepcidin (178), suggesting an EPO-independent erythropoietic derived mechanism is responsible for the decrease in hepcidin. Growth differentiation factor 15 (GDF15) and twisted grastrulation 1 (TWGS1) are secreted during erythroblast maturation and can inhibit hepcidin expression (165, 166). However, their role in hepcidin repression during erythropoiesis is still unclear (9, 82).

LIVER IRON IMPORT

Tf-bound iron

The major mechanism for iron uptake in the liver and most other tissues is through the Tf/transferrin receptor (Tfr) system (). Tf as discussed above is a constitutively expressed protein. Tfr1 transcript stability is regulated by the IRP/IRE system. Unlike ferritin, which has a single IRE in its 5’ UTR, Tfr1 transcript contains several IREs in its 3’ UTR. IRPs bind to the IRE in the Tfr1 transcript, which increases mRNA stability (77). Under low iron conditions more Tfr1 is translated allowing for increased iron uptake through Tf. Under conditions of high iron the IRPs are inactivated that leads to decreased Tfr1 mRNA stability and decreased iron uptake. Iron circulates bound to Tf. When both lobes are occupied with iron (Diferric Tf), this complex binds with high affinity to Tfr1 (3, 4). Diferric Tf-TFR1 binding activates cellular iron uptake by receptor-mediated endocytosis, and this pathway is a model system to study the precise mechanisms of receptor-mediated endocytosis (15, 32, 33, 95). Diferric Tf internalization into endocytic vesicles initiates the release of iron from Tf via acidification of endosomes (36, 72). Ferric iron is reduced to ferrous iron by an endosomal ferric reductase (126). Through a positional cloning strategy a transmembrane protein Steap3 was found to be a critical reductase in the endosome. Mutations in the steap3 gene lead to microcytic anemia (126). Steap3 is highly expressed in the hematopoietic cell lineage, but the role of steap3 in other cell types is not clear. However, there are three other family members (Steap1, 2, and 4) that also contain ferric reductase activity (127). Ferrous iron is then transported to the cytosol via DMT1, which in addition to being localized on the brush border cells of the small intestine, is also observed on recycling endosomes (25, 67). Interestingly, mice that lack DMT1 are still capable of accumulating hepatic iron, suggesting that DMT1 is not essential for Tf-bound iron uptake or other transporters have a redundant role (68). ZIP14, a family member of the ZIP metal transporters is also localized to endosomes and is important for the movement of iron from endocytic compartment to the cytosol (191). Following iron transport to the cytosol Tf and Tfr1 are recycled back to circulation and to the cell membrane, respectively (15, 32, 33, 95).

Mechanisms of liver iron uptake

Iron is imported into the liver via Tf/Tfr mediated endocytosis. As the pH of the endocytic vesicle drops, iron is released, reduced to Fe²⁺ by an endocytic reductase, and transported out by DMT1 and/or ZIP14. During iron overload a significant amount of NTBI is present. Iron can be directly transported into the liver through membrane bound DMT1 and/or ZIP14. During conditions of increased hemolysis the liver is capable of transport of hemoglobin and heme. Free hemoglobin binds with high affinity to haptoglobin, whereas free heme binds to hemopexin. These complexes bind to their respective receptors CD163 and Lrp/CD91, which initiate receptor-meditated endocytosis. Hemoglobin is degraded in the endosome and heme is released from the endocytic vesicle. Heme is further degraded by HO-1 releasing iron.

Non-transferrin bound iron (NTBI)

In cases of severe iron overload, the level of iron will exceed the capacity of Tf, and there is a greater ratio of iron in the plasma that occurs as NTBI. NTBI is bound by a number of non-protein ligands including citrate which is likely to be the predominate form of plasma NTBI found in hemochromatosis (21, 66). The liver uptake of ferric citrate involves dissociation of citrate and transport of iron into the hepatocyte (). NTBI is efficiently taken up by hepatocytes and the uptake is not downregulated by excess iron in the liver as observed with Tf-bound iron through the IRE/IRP system. Several mechanisms have been shown to contribute to NTBI uptake including membrane bound DMT1 and ZIP14 facilitating direct uptake of iron into the hepatocytes (105, 154). In addition, several other mechanisms are capable of NTBI transport into the cells. L-type calcium channels facilitate transport of NTBI into cardiac myocytes. Calcium channel blockers inhibit NTBI uptake into the heart (131). Lipocalin 2 is a multi-functional protein, which has iron-sequestering properties and is critical in binding to siderophores and limiting iron to pathogenic bacteria (51). However, some data suggest that lipocalin 2 could meditate NTBI uptake (83). Scara5 is a ferritin receptor that mediates NTBI uptake into the kidney (103). However, the role of these pathways for hepatic NBTI uptake is not clear.

Heme and hemoglobin associated iron

The liver also has the capacity to acquire iron from heme or hemoglobin (). These pathways under normal conditions contribute to a negligible amount of iron uptake in the liver. However, during hemolysis this could lead to a substantial amount of heme or hemoglobin being taken up by the liver. In several diseases such as hemolytic anemia, gram-positive bacterial infection, and malaria, increased hemolysis leads to excess hemoglobin and heme. The liver derived scavenging proteins, haptoglobin and hemopexin rapidly sequesters free hemoglobin and heme. Haptoglobin and hemopexin proteins bind with high affinity to free hemoglobin and heme, respectively (79, 168). Once sequestered, the haptoglobin-hemoglobin complex binds to CD163, which is highly expressed on mature tissue macrophages, including Kupffer cells (92). The hemopexin-heme complex binds to LRP/CD91, which is expressed in several cell types, including macrophages and hepatocytes (78). Following binding to their respective receptor the complex is endocytosed and degraded through a lysosomal pathway. Iron is released from heme by the HO-1 and enters the same intracellular pool as iron from other sources as mentioned above.

LIVER IRON STORAGE

Regardless of the source, iron that enters the hepatocyte enters the same intracellular pool. This pool of iron is stored, mobilized for systemic metabolic demands, used in intracellular enzymes, or used in mitochondrial iron sulfur proteins. Since free intracellular iron is toxic, the majority of iron in the cells is stored in ferritin, which is discussed above in more detail. Within the liver all cell types can store iron, but under normal conditions hepatocytes represent the major storage site. During severe iron overload, as the ferritin storage becomes saturated, storage in hemosiderin is elevated. Hemosiderin is an insoluble complex made up of degraded ferritin and large ferric hydroxide chains. Iron stored in hemosiderin is poorly mobilized (104, 149).

LIVER IRON EXPORT

Iron export from the liver, both in Kupffer cells and hepatocytes, is unclear and far less is known about the molecular mechanisms as compared to iron uptake and storage in the liver. Iron that is stored in ferritin has the ability to be mobilized from the liver during times of high systemic demand of iron. This is the rationale for therapies of patients with hemochromatosis (discussed in more detail below). Patients with high liver iron are periodically bled initiating mobilization of iron from the liver to circulation. Rodent studies using radioactive iron tracers estimate that up to 6% of iron is released from hepatocytes daily (13). Several cues have been shown to regulate iron mobilization from the liver. Erythropoiesis and a systemic change in iron levels in rats increase the mobilization. Iron export is inhibited following inflammation. In addition, the Kupffer cells also contribute significantly to iron release from the liver via erythrophagocytosis and release of iron from RBCs (13). The first step in iron mobilization is the regulated release of iron from ferritin. This is thought to be an autonomous property of ferritin controlled by cytosolic iron levels (38, 142). Expression of FPN in cells increases iron release from ferritin (117). The only known iron exporter is FPN, which has been shown to be critical for iron transport in animals (43). FPN is expressed highly in macrophages and to a lesser degree in hepatocytes (144). As mentioned above, FPN is regulated by hepcidin-mediated binding and degradation. This pathway is well characterized in in vitro cell systems. However, the role hepcidin plays in regulating hepatic FPN protein stability in vivo is not clear. FPN is also regulated by the IRE/IRP system, and has an IRE in its 5’ UTR. Under conditions of cellular iron deficiency, IRP proteins bind to the IRE in the FPN transcript, blocking its translation. This leads to decreased protein expression of FPN on the membrane and allows the cell to retain iron through decreased export (113). Recent conditional disruption studies underscore the importance of FPN in the liver. Macrophage-specific deletion of FPN led to iron sequestration in Kupffer cells. The deletion did not have a profound affect on RBC parameters and only mild anemia was observed (189). This finding is quite surprising since most of the iron for daily requirements is derived from macrophage-mediated recycling of senescent RBCs. These data suggest there must be compensatory mechanisms when macrophage iron export is ablated. A hepatocyte-specific FPN deletion led to mild iron sequestration in hepatocytes. However, RBC parameters were normal. Under low iron conditions these mice developed anemia; RBC and hemoglobin values were significantly lower (190).

LIVER IRON OVERLOAD

The liver is central to iron homeostasis and depends on a complex feedback mechanism between body iron requirements, intestinal absorption, and recycling from senescent RBCs. Dysregulation of these mechanisms can lead to iron overload. This section will discuss common and rare disorders of iron overload.

Hereditary Hemochromatosis (HH)

HH is a genetic disorder and a common cause of iron overload. 1 in 200 will be affected by this disorder (128). It was first described by Armand Trousseau in 1865 and was referred to as bronze diabetes. A change in the hue of the skin, liver, and pancreas was observed, although the cause was not known at this time. Over 30 years later Von Recklinghausen named this condition hemochromatosis following further analysis showing iron accumulation in liver cells. In 1996 it was identified that a mutation in the HFE gene was associated with HH (46). It is now known that HH is an autosomal recessive disorder and 1 in 8 people in the United States have a mutation in a single copy of the gene (128). Further study of patients with HH has led to the identification of several other iron regulatory genes that cause HH. These genes demonstrate that iron sensing and regulation of hepcidin is a concerted effort of several proteins. All HH disorders demonstrate a dysregulation in the hepcidin-FPN homeostasis, and are classified into 5 types.

Type 1

High FE (HFE) encodes an atypical major histocompatibility complex protein, and mutations in this gene are the most common cause of HH (46). The most common mutation observed is a missense mutation of cysteine 282 to tyrosine (Cys282Tyr) (152). However, several other mutations are characterized leading to iron overload (152). HFE mutations that lead to iron overload are associated with a significant decrease in hepcidin expression. Consistent with these data, mouse models which are deleted for HFE or have a knock-in Cys282Tyr mutation also have iron overload and a decrease in hepcidin expression (102). Since HFE is abundant in several tissues including enterocytes and liver, a conditional disruption of HFE in the liver and intestine was generated. In this study, hepatocyte-specific disruption of HFE recapitulated a similar phenotype as the whole body knockout mouse model, characterized by iron overload and decrease in hepcidin expression (180). Mice with HFE disruption in the intestine were similar to normal controls (181). These data demonstrate that HFE in the hepatocytes is critical for iron homeostasis. The molecular function of HFE and its precise role in regulating hepcidin expression has been a subject of great interest. Several lines of evidence suggest that HFE binding to Tfr1 and Tfr2 may be the mechanism by which HFE regulates hepcidin expression (60, 153). Mutations in HFE that increased binding to Trf1 blocked hepcidin expression. Mutations that weakened HFE and Tfr1 interaction increased hepcidin expression (153). In addition, HFE and Tfr2 interact and disruption of Tfr2 leads to decreased hepcidin expression (50, 86, 116). Lastly HFE and Tfr2 interaction is required for regulation of hepcidin by iron containing Tf (60). Together the data suggest a mechanism where Tf binding to Tfr1 releases HFE, which then can bind to Tfr2 and stabilize its protein expression leading to an increase in SMAD signaling ().

Type 2A

Juvenile hemochromatosis (JH) is a rare autosomal recessive disorder of iron overload and symptoms become apparent before the age of 30. JH leads to organ damage, and usually causes cardiomyopathy, hypogonadism, liver injury, and diabetes. JH is a caused by mutations in the gene for HFE2 which encodes the hemojuvelin (HJV) protein (132, 152). HJV is a glycophosphatidylinositol anchored membrane protein. Several HFE2 mutations have been found in patients. However, the glycine 320 to valine is the most frequent mutation that is reported (152). To confirm that HJV is causative in this type of hemochromatosis, an HJV knockout mouse model was generated (125). This mouse model demonstrates severe iron overload associated with very low levels of hepcidin expression, similar to that observed in patients with HJV mutations (132). Hepcidin expression was appropriately increased in response to inflammatory stimuli, suggesting that HJV is involved in iron sensing but does not play a role in hepcidin regulation during inflammation. The early onset of iron overload in JH is due to a robust repression of hepcidin. In HH due to HFE mutations there is only a moderate decrease in hepcidin expression leading to iron overload that is symptomatic at later ages. HJV is expressed in several tissues, and in the liver HJV primarily expressed in hepatocytes. Restoring HJV expression in hepatocytes of HJV knockout mice completely restored hepcidin expression and ablated the iron overload (188). Further mechanistic studies demonstrated that HJV functions as a BMP co-receptor and is important for induction of hepcidin expression in response to BMP signaling (10). HJV binds to BMPs and enhances the activity of the SMAD signaling cascade (10).

Type 2B

Similar to HJV mutations, mutations in the HAMP gene, which encodes for hepcidin, are a very rare cause of JH. Currently 12 known mutations occur on the HAMP gene leading to a decrease in the normal production of hepcidin (74). Since hepcidin function or expression is dramatically diminished, the iron overload symptoms are observed before the age of 30.

Type 3

Tfr2 mutations lead to an autosomal recessive iron overload disease similar to HFE related-HH phenotype. Tfr2 as mentioned above is capable of binding to HFE and this interaction is critical in maintaining hepcidin expression (60). Unlike Tfr1, which is ubiquitously expressed, Tfr2 is expressed only in hepatocytes and erythroid precursors (87, 159). Tfr2 cannot compensate for the loss of Tfr1 (171). The knockout mouse model and the liver-specific disruption of Tfr2 confirm its importance in regulating hepcidin levels (86, 182). Hepcidin levels are decreased significantly in these mouse models compared to littermate controls, and tissue iron is increased. The most common mutation observed is in amino acid 245, which is converted into a stop codon resulting in a protein product that is not expressed (24). As mentioned above, Tfr2 binding to HFE promotes SMAD activation and hepcidin expression. Upon its deletion this signaling pathway is decreased causing a significant drop in hepcidin levels.

Type 4

SLC40a1 is the gene that encodes for the iron exporter FPN. FPN is the target of hepcidin, which causes rapid internalization and degradation of FPN (117). More precise work on the mechanism of FPN degradation by hepcidin demonstrates that following hepcidin binding, FPN is phosphorylated on tyrosine residues, which lead to its endocytic shuttling and degradation by the proteasome pathway (39). JAK2 is the critical kinase phosphorylating FPN (37). However, recent data demonstrate that both phosphorylation of FPN and JAK2 are not essential for FPN degradation (141, 148). Mutations in HFE, HJV, hepcidin, and Tfr2 are all recessive mutations. However, mutations in FPN are dominant. Patients that are heterozygous for the mutation develop the disease. This is due to FPN functioning as a dimer and the mutant protein can act as a dominant negative (40, 41). Several mutations of FPN have been observed. Detailed molecular studies demonstrate that the mutations inhibit proper membrane localization of FPN, inhibit the export function of FPN, disrupt hepcidin binding, or inhibit FPN internalization (84). Therefore, depending on the mutation in FPN the patients can present with very different phenotypes. Mutations that inhibit membrane localization or export function can lead to macrophage iron overload. While those mutations that inhibit hepcidin binding or hepcidin-meditated internalization lead to continuous export of iron into serum and eventually iron overload in the hepatocytes.

Secondary hemochromatosis

Secondary hemochromatosis is the result of another disease, which causes excess liver iron loading. Most of diseases that lead to secondary hemochromatosis are acquired disorders of erythropoiesis (63). The most common causes of secondary hemochromatosis are listed in . A well-studied disorder that leads to secondary hemochromatosis is β-thalassemia. β-Thalassemia is a congenital blood disorder due to mutations in the β-globin gene leading to a partial or complete loss of β-globin synthesis resulting in β-thalassemia intermedia and Cooley’s anemia, respectively. The decrease in β-globin results in ineffective erythropoiesis and erythropoietic stress. Persons with β-thalassemia intermedia have mild anemia with a slight lowering of hemoglobin levels in the blood. In most cases treatment is not necessary, but severe patients with low hemoglobin levels will need occasional blood transfusions (161). Cooley’s anemia results in a striking deficiency in hemoglobin production. Patients will need frequent blood transfusions (161). The blood transfusions lead to dysregulation of the systemic iron homeostasis since donor blood is a rich source of iron. The body cannot eliminate the excess iron efficiently, leading to increased tissue iron. Regular blood transfusions are the most common cause of secondary hemochromatosis (63). Initially it was thought that the iron overload was primarily due to regular blood transfusions. However, mouse models of β-thalassemia hyperabsorb iron. This is the major mechanism leading to iron overload in β-thalassemia intermedia and significantly contributes to the tissue iron overload in Cooley’s anemia (76, 164, 186). It is less clear whether an increase in iron absorption plays a significant role compared to blood transfusions in other disorders of erythropoiesis listed in . However, recent work has shown that effective and ineffective erythropoiesis can stimulate iron absorption, therefore this mechanism of iron overload may be true for other diseases leading to secondary hemochromatosis (5). Increased iron absorption in secondary hemochromatosis may be due to an increase intestinal hypoxia signaling and a decrease in hepcidin expression (5, 133). Increasing hepcidin levels in mouse models of β-thalassemia improved liver iron loading and anemia (62).

Table 1

Common causes of secondary hemochromatosis

Aceruloplasminemia, hypotransferrinemia, and DMT-1 deficiency

Aceruloplasminemia, hypotransferrinemia, and DMT1 deficiency are causes of secondary hemochromatosis, but are not disorders of erythropoiesis; rather these disorders are due to ineffective transport of iron. Aceruloplasminemia is due to a loss-of-function mutation in Cp and is inherited in an autosomal recessive manner. Iron overload in aceruloplasminemia is mainly observed in the brain and liver (73, 187). A similar phenotype is also observed in Cp knockout mouse models. Hypotransferrinemia is an autosomal recessive disorder leading to loss of Tf production. Hypotransferrinemia is associated with severe microcytic anemia, and an adaptive increase in iron absorption, which leads to severe liver iron loading (64, 75). Consistent with hypotransferrinemic patients, the hpx mouse, which produces no Tf has liver iron overload and anemia (171). DMT1 deficiency is an autosomal recessive disorder leading to increase in liver iron (80). This is thought to be due to the role of DMT1 in iron export from the endocytic compartment.

Dysmetabolic iron overload syndrome (DIOS)

DIOS is a newly characterized secondary hemochromatosis disorder. DIOS is associated with features, such as obesity, type 2 diabetes, alcohol use, and chronic hepatitis C (42, 81, 106, 139). This is now the most common cause of iron overload observed in patients. The iron overload is observed in 15% of patients with metabolic syndrome, 50% in patients with non-alcoholic fatty liver disease, over 40% of patients with chronic hepatitis C infection, and significant number of patients with alcoholic liver disease (18, 81, 139, 174). Currently the mechanisms, which contribute to DIOS are unclear. However, a significant decrease in FPN gene expression has been noted in patients with DIOS (2).

IRON-INDUCED LIVER DAMAGE

High levels of iron deposition lead to tissue damage and dysregulation of function. In the liver increased free iron if untreated, leads to fibrosis and cirrhosis, and can increase morbidity and mortality (123, 124, 145). Hepatic tissue injury is directly correlated to the duration and amount of iron loading (123, 124). Cells normally produce basal levels of reactive oxygen species (ROS) through metabolic function of the mitochondria and other organelles. ROS are kept at low basal levels by several antioxidant enzymes, and low levels of ROS are important in normal cell physiology (146). ROS in conjunction with high cellular iron results in a robust increase of hydroxyl radicals, which leads to cell damage. Free iron generates ROS through the Fenton and Haber-Weiss reactions (). The superoxide radical (O₂^•−) reduces ferric iron to ferrous iron, which reacts with hydrogen peroxide (H₂O₂) to generate highly reactive hydroxyl radicals (OH^•) (90). Hydroxyl radicals lead to an increase in peroxidation of phospholipids within organelle and cellular membranes, oxidation of amino acid side-chains, DNA strand breaks, and protein fragmentation. The exact mechanisms by which high intracellular iron leads to liver fibrosis and cirrhosis are unclear, but iron-induced cellular damage has been shown to directly increase hepatocyte cell death and activate Kupffer and stellate cells (135, 160) (). More recently, clinical evidence suggests that high liver iron could play a role in insulin resistance (45, 143). The increase in iron-induced ROS in Kupffer cells can initiate a pro-inflammatory cascade in the liver. Increased ROS production activates NF-κB signaling leading to an increase in IL-6, TNF-α, and IL-1β in the liver (22, 112). Liver inflammation can lead to hepatic insulin resistance, which is a major pathway leading to hyperglycemia in type II diabetes (55). Lastly, liver iron overload increases the risk for hepatocellular carcinoma (HCC). HCC is the major life threatening complication associated with hereditary hemochromatosis (124). Several studies have looked at the risk factor for HCC in hereditary hemochromatosis patients and some have estimated the risk to be 100-200 fold higher in HH patients (19, 162). In addition, other iron overload disorders such as thalassemia are associated with an increased risk for HCC (17). The increase in iron plays an active role in HCC pathogenesis. Chelation of iron or placing mice on an iron deficient diet decreases tumor growth (71, 151).

Schematic diagram of the Fenton and Haber-Weiss reactions

Iron is a potent catalytic cofactor, which increases highly unstable oxygen radicals that cause cellular damage.

Iron-induced liver damage

Iron accumulation in hepatocytes and Kupffer cells leads to an increase in ROS production and pro-inflammatory mediators. Both ROS and pro-inflammatory mediators initiate a feed forward cycle, which activates stellate cells, initiates cell damage, and leads to loss of function contributing to an increase in steatosis, fibrosis, cirrhosis, and HCC.

TREATMENT OF IRON OVERLOAD

In patients with HH, phlebotomy is used to decrease liver iron. Regular bleeding of the patients leads to an increase in erythropoiesis, which mobilizes liver iron stores to meet the demand for iron that is required to generate mature RBCs (59). Secondary hemochromatosis is also associated with severe anemia, thus phlebotomy is not an option. In these patients iron chelators are used to decrease liver iron (59). Deferoxamine has been used for over three decades for iron chelation. More recently two new iron chelators have also been used, deferiprone and deferasirox (93). Iron chelators have been shown to be effective in decreasing liver iron and also morbidity and mortality associated with iron overload. Several rounds of phlebotomy and/or administration of iron chelators are required; these are slow-acting treatment options and may not successfully decrease the liver injury associated with an increase in iron. Alternatives to existing treatments are needed. Recently, hepcidin has been shown to be a very attractive target and good proof of principles studies have been done. Several studies in mouse models of HH have shown that increasing hepcidin levels can ameliorate the iron overload (122, 177). Recently in β-thalassemia models, increasing hepcidin expression resulted in decreased liver iron (62). Currently several strategies are being assessed to increase hepcidin levels. The specific sites that are required for hepcidin-FPN binding are known. Precise structural mutagenesis studies have demonstrated that nine amino acids of hepcidin are critical for the binding to FPN and initiating its internalization (140). Modifications of these nine amino acids has led to several peptides that have increased activity over full length hepcidin. Moreover, the modified hepcidin derived peptides are functional in vivo and can prevent iron overload that is observed in hepcidin knockout mice (140). These agents are stable orally and may provide a well-tolerated form of treatment for hereditary and secondary hemochromatosis. Other approaches may also be useful in increasing hepcidin expression (59). Treatment with BMP6 increases hepcidin expression in HFE-null mouse model and prevents iron overload (34). Moreover, since hepcidin regulatory pathways are well characterized, several possibilities such as SMAD, C/EBPα, and STAT3 activators could have potential roles in increasing hepcidin expression in vivo.

ANEMIA

On the other end of the spectrum, aberrant upregulation of hepcidin is critical in the pathogenesis of anemia of chronic disease, which encompasses several diseases including kidney disease, inflammatory disease, cancer, and aging (26). In healthy human volunteers and mice, studies demonstrate that inflammatory agents cause a robust and rapid decrease in serum iron levels (27, 88, 115, 147). Within hours following induction of inflammation, hepcidin levels are significantly elevated. The decrease in serum iron is due to hepcidin-mediated internalization of FPN leading to iron sequestration in macrophages. This is thought to be a protective mechanism that limits iron available to infectious pathogens. However, in chronic disorders this leads to anemia, which can have detrimental effects in the primary disease pathogenesis. Similarly, in most cancers there is a decrease in serum iron levels and increased anemia (156). The decrease in iron is suggested to be beneficial in limiting tumor growth. The mechanism may vary depending on the primary disease. However, the IL-6-STAT3 pathway is critical in increasing hepcidin expression during inflammation (163). The best treatment for anemia of chronic disease is resolving the primary chronic disease. In severe cases, blood transfusion, EPO, or intravenous administration of iron is used (163). In addition to an increase in hepcidin expression from chronic disorders, rare genetic mutations in the TMPRSS6 gene cause an increase in hepcidin expression and iron-refractory iron deficiency anemia (IRIDA) (47). This was further confirmed in the TMPRSS6 knockout mice, which had hair loss and microcytic anemia associated with high levels of hepcidin expression (48, 52). A similar finding was noted in the mask mutant mouse strain, characterized by a premature stop codon in the TMPRSS6 gene (44). IRIDA is iron deficiency anemia that is unresponsive to oral iron therapy. TMPRSS6 (also known as matriptase-2) encodes a type II transmembrane serine protease, which is expressed predominantly in the liver. The first substrate that was characterized for TMPRSS6 was HJV. The serine protease of TMPRSS6 cleaves membrane HJV, leading to downregulation of hepcidin expression (155). Mutations in TMPRSS6 decrease its protease activity leading to increased protein expression of HJV and a coordinate increase in hepcidin expression.

CONCLUSION

The liver is the central tissue which regulates systemic iron homeostasis by acting as a sensor and regulator of iron levels. In addition, through its role in iron storage, the liver can protect more sensitive tissues from iron-induced cellular injury. The past decade has led to several novel liver-derived players regulating systemic iron homeostasis and identification of new mutations in iron-related disorders. Several pathways in the liver regulate hepcidin, the master hormone for maintaining systemic iron homeostasis. Recent studies have shown that these pathways can have redundancy or act independently based on the stimuli (172). The challenge will be to understand how these pathways crosstalk and are regulated in a coordinate manner to maintain hepcidin levels. Moreover, how can these pathways be targeted in iron-related disorders that could be of therapeutic benefit? Targeted therapies for iron related disorders are actively being pursued such as the case for hepcidin mimetics and BMP agonists, and the coming decade should yield novel therapies.

ACKNOWLEDGEMENTS

This work was supported by grants to Y.M.S from the National Institutes of Health (CA148828 and DK095201), The University of Michigan Gastrointestinal Peptide Center, and Jeffrey A. Colby Colon Cancer Research and the Tom Liu Memorial Funds of the University of Michigan Comprehensive Cancer Center. E.R.A is supported by the Rackham Predoctoral Fellowship, University of Michigan.

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Iron homeostasis in the liver

Compr Physiol. Author manuscript; available in PMC 2014 Feb 27.

Published in final edited form as:

PMCID: PMC3936199

NIHMSID: NIHMS551241

Erik R Anderson

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

Yatrik M Shah

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

The publisher’s final edited version of this article is available at Compr PhysiolSee other articles in PMC that cite the published article.

Abstract

Iron is an essential nutrient that is tightly regulated. A principal function of the liver is the regulation of iron homeostasis. The liver senses changes in systemic iron requirements and can regulate iron concentrations in a robust and rapid manner. The last 10 years have led to the discovery of several regulatory mechanisms in the liver which control the production of iron regulatory genes, storage capacity, and iron mobilization. Dysregulation of these functions leads to an imbalance of iron, which is the primary causes of iron-related disorders. Anemia and iron overload are two of the most prevalent disorders worldwide and affect over a billion people. Several mutations in liver-derived genes have been identified, demonstrating the central role of the liver in iron homeostasis. During conditions of excess iron, the liver increases iron storage and protects other tissues, namely the heart and pancreas from iron-induced cellular damage. However, a chronic increase in liver iron stores results in excess reactive oxygen species production and liver injury. Excess liver iron is one of the major mechanisms leading to increased steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma.

INTRODUCTION

Iron is an essential micronutrient that is a critical component of oxygen transport proteins (hemoglobin and myoglobin) and of numerous metabolic and redox enzymes. The average adult has 2–4 grams of iron, and over 80% is contained in hemoglobin of red blood cells (RBC). Chronic iron deficiency results in decreased hemoglobin production and anemia. Systemic iron levels are tightly controlled through an integrative mechanism that involves iron absorption, storage, and recycling. The past decade has been termed “The Golden Age of Iron Biology”, due to the significant increase in the understanding of the molecular underpinnings of systemic iron homeostasis (7). When iron regulatory pathways are dysregulated, this leads to either excess tissue iron or iron deficiencies, which affect over a billion people worldwide. Four major cell types or tissues have been shown to be critical for systemic iron homeostasis ():

Systemic iron regulation

Dietary iron is absorbed through the small intestine and mainly utilized for RBC production. Hepatic and splenic macrophages recycle iron from senescent RBCs. The iron derived from recycling is used for production of RBCs. During times of iron excess the liver can store iron and during increased systemic needs the liver can mobilize iron stores for utilization.

Enterocyte

Dietary iron absorption is tightly regulated in the small intestine. Dietary iron enters the body through absorptive cells in the duodenum. Dietary ferric iron (Fe⁺³) is reduced by the apical ferric reductase duodenal cytochrome b (DcytB) to ferrous iron (Fe⁺²) and transported into the enterocyte via an apical iron transporter, divalent metal transporter-1 (DMT1, also known as Nramp2, SLC11a2 and DCT1) (49, 69, 100, 107, 109). Once iron enters into the cytoplasm through DMT1, iron is either stored or exported by the iron exporter ferroportin (FPN, also known as SLC40A1)(1, 43, 110) located on the basolateral side of the enterocyte. Disruption of DMT1 or FPN leads to inhibition of iron absorption and dysregulation of systemic iron homeostasis (43, 68). Following export of iron through FPN, iron is oxidized back to the ferric form by the ferroxidase hephaestin and ceruloplasmin (Cp) and loaded onto transferrin (Tf) and circulated through the body (70).

Erythroblast

A significant portion of circulating Tf-bound iron is utilized in early RBC precursors called erythroblasts for the synthesis of hemoglobin. Hemoglobin comprises of about 95% of the total cellular protein of the mature RBC and more than 80% of functional iron in the body is found in hemoglobin (28). Iron is an essential cofactor for the ability of RBCs to transport oxygen, and thus a decrease in body iron levels is the most predominant cause of anemia worldwide.

Macrophage

Only 1–2 mg per day of dietary iron is required to be absorbed through the intestine. This is due to the highly efficient recycling of iron from senescent erythrocytes. Recycling is performed by splenic and hepatic macrophages. Aging erythrocytes at 120 days old undergo specific changes that can be recognized by macrophages, thus initiating erythrophagocytosis. Iron is recovered from the degradation of hemoglobin and heme by hydrolytic enzymes in the phagocytic vesicles and heme-oxygenase-1 (HO-1) (58). Iron is routed back to circulation through the basolateral iron transporter FPN. Recent studies using macrophage-specific conditional deletion of FPN demonstrate the importance of FPN in exporting iron out of macrophages following erythrophagocytosis (189, 190).

Liver

The liver performs three essential functions in maintaining systemic iron homeostasis: 1) The liver is the major site for production of proteins that maintain systemic iron balance, 2) It is a storage site for excess iron and 3) the liver is critical for the mobilization of iron from hepatocytes to the circulation to meet metabolic requirements. Dysregulation of the liver’s ability to maintain balance of these three parameters leads to iron-related disorders. This review will focus on the normal function of the liver in iron homeostasis and the role of liver in iron-related disorders.

PRODUCTION OF PROTEINS FOR IRON HOMEOSTASIS

(Tf)

Tf is an 80 kDa plasma glycoprotein generated predominantly in the liver (138). However, small amounts are also made in the brain and testis (14, 157). Tf is the major serum iron-binding protein and is essential for systemic circulation of iron. Tf consists of two globular lobes of α-helices and β-sheets, which have a significant amount of homology between the N- and C- terminal halves of the molecule. This homology is thought to be due to a gene duplication arising from a 40kDa ancestral protein containing a single lobe (3, 96-98). Each lobe includes metal binding amino acid residues (2 Tyr, 1 His, and 1 Asp). Tf binds to ferric iron reversibly and with high affinity, but does not bind to ferrous iron (3, 4). In addition, Tf can also bind other metals with lower affinity (3). The affinity of iron is also regulated by pH, as iron binding to Tf decreases as the pH drops, with no detectable amounts observed below pH 4.5 (3, 4). The reversible binding of iron to Tf is important and allows it to be a cellular iron donor or iron acceptor depending on the systemic requirements of iron.

(Cp)

Cp is a copper-dependent serum ferroxidase which works in tandem with FPN for cellular iron export into circulation (129, 130). Cellular iron is usually in the ferrous form and is exported out of the cell by FPN, however the affinity of Tf for ferrous iron is relatively weak and Cp is required to oxidize iron to its ferric form, which binds with high affinity to Tf (129, 130). Disruption of Cp in mouse models further supports the role of Cp in systemic iron homeostasis (30, 70).

Ferritin

Ferritin is a multi-subunit protein that consists of a heavy and light chain. Ferritin from different tissues can vary in the ratio of heavy compared to light chains. Liver ferritin contains mainly the light chain and can store up to 4500 atoms of iron. Hepatocytes are the major site for ferritin synthesis, however most cells that have been assessed can synthesize ferritin to a smaller degree (12, 167). Cells with high iron levels are capable of an adaptive increase in ferritin synthesis. Both ferritin subunit synthesis are controlled through a post-transcriptional mechanism by the iron response element (IRE)/iron regulatory protein (IRP) system. The ferritin mRNAs contain a single IRE in the 5’ UTR. Under iron deficient conditions, the IRPs bind to this IRE and repress translation. However, as intracellular iron increases in a cell, the repression is relieved and ferritin synthesis is increased. For more details of the IRE/IRP system, refer to the comprehensive review by Muckenthaler, M.U et al. (113). Although the IRP/IRE regulation of ferritin synthesis is the predominant pathway that allows an adaptive increase, several other mechanisms, such as IL-1, TNFα, hypoxia, and oxidative stress have also been observed to regulate ferritin expression (23, 169, 170, 173, 184). It is likely that a combination of these pathways allow an efficient increase in ferritin when required.

Hepcidin

Hepcidin is a 25 amino acid antimicrobial peptide that is produced in hepatocytes and secreted into the circulation (91). Hepcidin is translated as an 84 amino acid pro-protein, and cleaved by the pro-hormone convertase furin to produce the active peptide (175). Subtractive hybridization experiments between iron loaded and control mouse livers initially identified hepcidin as an iron-regulated protein (136). The role of hepcidin in iron homeostasis was further confirmed when hepcidin was inadvertently disrupted in addition to USF2 and these mice were severely iron overloaded (119). The effect of hepcidin knockout on iron homeostasis was confirmed in a subsequent paper, which showed that animals deficient in hepcidin, but having normal USF2 expression developed severe hemochromatosis (101). Conversely, animals that overexpress hepcidin in the liver demonstrate severe iron-deficiency anemia (120). A clue to the function of hepcidin was discovered after subjecting rats to an iron-deficient diet. Following iron deficiency, rats rapidly repress liver hepcidin and upregulate the intestinal iron transporters DMT1, DcytB, and FPN (54). Hepcidin was believed to regulate iron homeostasis due to an interaction with iron transporters. A breakthrough came when the hepcidin receptor was discovered. Through careful in vitro analysis, it was determined that hepcidin binds to FPN which leads to its internalization and proteasomal degradation (117). Hepcidin is increased in iron loading, which leads to a decrease in duodenal iron absorption to normalize iron levels. In the case of iron deficiency, hepcidin is repressed which allows more iron to be transported from the enterocyte to the serum (). In addition, FPN is increased in macrophages following erythrophagocytosis, and hepcidin represses FPN in these cells (89). In macrophages during iron loading, hepcidin decreases iron transport following iron recycling from senescent RBCs. Whereas during iron deficiency, a significant increase in iron efflux would be expected following erythrophagocytosis (). In addition to iron levels resulting in the regulation of hepcidin levels, hypoxia and erythropoiesis are major repressors of hepcidin expression. This provides a novel link between oxygen homeostasis and iron levels. Moreover, inflammation is a major activator of hepcidin expression. This results in restriction of serum iron levels during an infection, and therefore is less conducive for growth of pathogenic bacteria. The last decade has shown the importance of hepcidin as the master regulator of both duodenal iron absorption and red blood cell (RBC) iron recycling. Studying the regulation of hepcidin expression in the liver has been a priority for understanding regulation of systemic iron homeostasis. The transcriptional regulation of hepcidin has been reviewed elsewhere (56, 114) and is summarized in . Here, we only briefly mention a few major pathways that are critical in hepcidin regulation. In addition to the major pathways, several other accessory proteins have been identified, which are mutated in iron-related disorders. Their importance in hepcidin regulation is covered in more detail below in the liver iron overload and anemia section.

Hepcidin regulation of FPN protein expression during changes in systemic iron levels

High iron levels increase hepcidin expression, which decrease iron export from the small intestine and macrophage due to an internalization and degradation of FPN. Iron deficiency results in a decrease in hepcidin levels and stabilization of FPN protein expression.

Regulation of hepcidin by BMP/SMAD, inflammatory and hypoxia/erythropoietic signaling in the liver

Three major pathways are critical for regulating basal and stimuli-induced hepcidin expression. Binding of iron containing Tf to Tfr1 causes a dissociation of Tfr1-High FE (HFE) complex and an interaction of HFE with Tfr2. Increased stabilization of Tfr2 increases BMP6 mediated phosphorylation of SMAD1/5/ 8 and recruitment of SMAD 1/5/8 and SMAD4 to the hepcidin proximal promoter. BMP/SMAD signaling is the major pathway by which hepcidin expression is coordinated to meet systemic iron requirements. Activation of hepcidin by inflammation is thought to act independently of the BMP/SMAD pathway. The best-studied mechanism is via the pro-inflammatory mediator IL-6. Binding of IL-6 to its receptor IL-6 receptor (IL-6R) initiates activation of the JAK-STAT3 pathway. STAT3 binds directly to the proximal promoter to increase hepcidin expression. Hypoxia and erythropoiesis are inhibitors of hepcidin expression and these are the least understood pathways by which hepcidin expression is regulated. Hypoxia and erythropoiesis have been shown to inhibit hepcidin expression via direct binding of HIF to the proximal promoter, an EPO-EPO receptor (EPOR) mediated decrease in C/EBPα expression, and through increase in an unknown erythroid derived factor which signals through an undefined pathway.

SMAD/ bone morphogenetic protein (BMP) signaling

A key finding in the regulation of hepcidin is the essential role of the BMP-SMAD signaling cascade. BMPs are ligands that belong to the transforming growth factor-β (TGF-β) superfamily. BMPs bind to type I and type II serine threonine kinase receptors, which phosphorylate specific intracellular SMAD proteins (SMAD1/5/8). Phosphorylated SMAD1/5/8 (P-SMAD1/5/8) binds to the common mediator SMAD4, and the SMAD complex translocates to the nucleus to modulate transcription of target genes. BMPs, but not TGF-β signaling induced hepcidin expression in cultured liver cell lines and in vivo (11). Several BMPs signal through SMAD activation, however BMP6 is the endogenous ligand that modulates hepcidin expression. BMP6 knockout mice have decreased hepcidin expression and an increase in tissue iron (8, 10, 11, 111). BMP6 knockout mice still have the ability to increase hepcidin expression following inflammatory stimuli. The importance of SMAD signaling in hepcidin regulation was demonstrated in mice with a hepatocyte-specific disruption of SMAD4. A near complete loss of hepcidin expression was observed in these mice and eventually the mice die of severe iron overload in multiple tissues (183). Interestingly, liver SMAD4 knockout mice are unable to increase hepcidin expression in response to iron loading, suggesting that SMAD4 mediates the response of hepcidin to changes in systemic iron requirements. Signaling through the BMP receptor leads to SMAD1/5/8 phosphorylation, which is required for SMAD4 transcriptional activity (6). The regulation of hepcidin by iron loading and iron deficiency is correlated with phosphorylation of the SMAD1/5/8 proteins. In iron deficiency, pSMAD1/5/8 is decreased dramatically, and is substantially increased in conditions of iron overload (85).

STAT3 and inflammatory pathways

Hepcidin is induced in response to inflammatory stimuli (57, 118). Hepcidin induction by inflammation leads to iron sequestration, which can decrease bacterial growth. However, chronic diseases are associated with anemia (150). The mechanism for the induction of hepcidin by inflammation is mediated by IL-6 in cultured cells, mice, and humans (115). Subsequently, it was shown that IL-6 regulates expression of hepcidin by inducing STAT3 binding to the hepcidin promoter (176, 185). To support this, it was shown that anti-IL-6 receptor antibody improves anemia of inflammation (158).

Hypoxia and erythropoietic pathways

Hypoxia, or low oxygen tension, is a physiological condition that results in numerous adaptive changes in gene expression. Hypoxia represses hepcidin expression both in cultured cells and in mice (121). Many mechanisms have been proposed to play a role in hypoxic hepcidin repression (29, 31, 94, 134, 179). Hypoxia-inducible factor (HIF) is the major transcription factor activated following hypoxia and HIF is thought be critical in the hypoxia-mediated repression of hepcidin (20). HIF was shown to bind directly to the hepcidin promoter resulting in repression (134). However, recent studies refute these findings (108, 179). It has been reported that hypoxia through a HIF2α mediated increase in erythropoiesis is the critical pathway leading to hepcidin repression (108). Interestingly, a mutation that leads to HIF stabilization causes Chuvash polycythemia. In these patients a decrease in hepcidin expression is observed without a significant association with erythropoiesis (65). There is no clear mechanism that mediates hepcidin repression during hypoxia, and this is an active area of study

Erythropoiesis is a well-characterized pathway leading to hepcidin repression. In several mouse models that induce erythropoiesis, a significant decrease in hepcidin is observed (16, 53, 61). The decrease in hepcidin allows the increase of iron required for RBCs during erythropoiesis. In a model of intensive care anemia, erythropoietin injections or phlebotomies were able to repress hepcidin expression despite high levels of IL-6, which is known to strongly increase hepcidin expression (99). Similarly, hypoxia is able to repress hepcidin expression under conditions of high IL-6. One of the mechanisms by which erythropoiesis represses hepcidin is through erythropoietin (EPO) binding to its receptor in hepatocytes, which leads to downregulation of C/EBPα and hepcidin repression (137). C/EBPα is a liver-enriched transcription factor that is important in hepcidin regulation. Mice with a liver specific deletion of C/EBPα have low levels of hepcidin expression (35). However, erythropoietic blockers prevented the suppression of hepcidin (178), suggesting an EPO-independent erythropoietic derived mechanism is responsible for the decrease in hepcidin. Growth differentiation factor 15 (GDF15) and twisted grastrulation 1 (TWGS1) are secreted during erythroblast maturation and can inhibit hepcidin expression (165, 166). However, their role in hepcidin repression during erythropoiesis is still unclear (9, 82).

LIVER IRON IMPORT

Tf-bound iron

The major mechanism for iron uptake in the liver and most other tissues is through the Tf/transferrin receptor (Tfr) system (). Tf as discussed above is a constitutively expressed protein. Tfr1 transcript stability is regulated by the IRP/IRE system. Unlike ferritin, which has a single IRE in its 5’ UTR, Tfr1 transcript contains several IREs in its 3’ UTR. IRPs bind to the IRE in the Tfr1 transcript, which increases mRNA stability (77). Under low iron conditions more Tfr1 is translated allowing for increased iron uptake through Tf. Under conditions of high iron the IRPs are inactivated that leads to decreased Tfr1 mRNA stability and decreased iron uptake. Iron circulates bound to Tf. When both lobes are occupied with iron (Diferric Tf), this complex binds with high affinity to Tfr1 (3, 4). Diferric Tf-TFR1 binding activates cellular iron uptake by receptor-mediated endocytosis, and this pathway is a model system to study the precise mechanisms of receptor-mediated endocytosis (15, 32, 33, 95). Diferric Tf internalization into endocytic vesicles initiates the release of iron from Tf via acidification of endosomes (36, 72). Ferric iron is reduced to ferrous iron by an endosomal ferric reductase (126). Through a positional cloning strategy a transmembrane protein Steap3 was found to be a critical reductase in the endosome. Mutations in the steap3 gene lead to microcytic anemia (126). Steap3 is highly expressed in the hematopoietic cell lineage, but the role of steap3 in other cell types is not clear. However, there are three other family members (Steap1, 2, and 4) that also contain ferric reductase activity (127). Ferrous iron is then transported to the cytosol via DMT1, which in addition to being localized on the brush border cells of the small intestine, is also observed on recycling endosomes (25, 67). Interestingly, mice that lack DMT1 are still capable of accumulating hepatic iron, suggesting that DMT1 is not essential for Tf-bound iron uptake or other transporters have a redundant role (68). ZIP14, a family member of the ZIP metal transporters is also localized to endosomes and is important for the movement of iron from endocytic compartment to the cytosol (191). Following iron transport to the cytosol Tf and Tfr1 are recycled back to circulation and to the cell membrane, respectively (15, 32, 33, 95).

Mechanisms of liver iron uptake

Iron is imported into the liver via Tf/Tfr mediated endocytosis. As the pH of the endocytic vesicle drops, iron is released, reduced to Fe²⁺ by an endocytic reductase, and transported out by DMT1 and/or ZIP14. During iron overload a significant amount of NTBI is present. Iron can be directly transported into the liver through membrane bound DMT1 and/or ZIP14. During conditions of increased hemolysis the liver is capable of transport of hemoglobin and heme. Free hemoglobin binds with high affinity to haptoglobin, whereas free heme binds to hemopexin. These complexes bind to their respective receptors CD163 and Lrp/CD91, which initiate receptor-meditated endocytosis. Hemoglobin is degraded in the endosome and heme is released from the endocytic vesicle. Heme is further degraded by HO-1 releasing iron.

Non-transferrin bound iron (NTBI)

In cases of severe iron overload, the level of iron will exceed the capacity of Tf, and there is a greater ratio of iron in the plasma that occurs as NTBI. NTBI is bound by a number of non-protein ligands including citrate which is likely to be the predominate form of plasma NTBI found in hemochromatosis (21, 66). The liver uptake of ferric citrate involves dissociation of citrate and transport of iron into the hepatocyte (). NTBI is efficiently taken up by hepatocytes and the uptake is not downregulated by excess iron in the liver as observed with Tf-bound iron through the IRE/IRP system. Several mechanisms have been shown to contribute to NTBI uptake including membrane bound DMT1 and ZIP14 facilitating direct uptake of iron into the hepatocytes (105, 154). In addition, several other mechanisms are capable of NTBI transport into the cells. L-type calcium channels facilitate transport of NTBI into cardiac myocytes. Calcium channel blockers inhibit NTBI uptake into the heart (131). Lipocalin 2 is a multi-functional protein, which has iron-sequestering properties and is critical in binding to siderophores and limiting iron to pathogenic bacteria (51). However, some data suggest that lipocalin 2 could meditate NTBI uptake (83). Scara5 is a ferritin receptor that mediates NTBI uptake into the kidney (103). However, the role of these pathways for hepatic NBTI uptake is not clear.

Heme and hemoglobin associated iron

The liver also has the capacity to acquire iron from heme or hemoglobin (). These pathways under normal conditions contribute to a negligible amount of iron uptake in the liver. However, during hemolysis this could lead to a substantial amount of heme or hemoglobin being taken up by the liver. In several diseases such as hemolytic anemia, gram-positive bacterial infection, and malaria, increased hemolysis leads to excess hemoglobin and heme. The liver derived scavenging proteins, haptoglobin and hemopexin rapidly sequesters free hemoglobin and heme. Haptoglobin and hemopexin proteins bind with high affinity to free hemoglobin and heme, respectively (79, 168). Once sequestered, the haptoglobin-hemoglobin complex binds to CD163, which is highly expressed on mature tissue macrophages, including Kupffer cells (92). The hemopexin-heme complex binds to LRP/CD91, which is expressed in several cell types, including macrophages and hepatocytes (78). Following binding to their respective receptor the complex is endocytosed and degraded through a lysosomal pathway. Iron is released from heme by the HO-1 and enters the same intracellular pool as iron from other sources as mentioned above.

LIVER IRON STORAGE

Regardless of the source, iron that enters the hepatocyte enters the same intracellular pool. This pool of iron is stored, mobilized for systemic metabolic demands, used in intracellular enzymes, or used in mitochondrial iron sulfur proteins. Since free intracellular iron is toxic, the majority of iron in the cells is stored in ferritin, which is discussed above in more detail. Within the liver all cell types can store iron, but under normal conditions hepatocytes represent the major storage site. During severe iron overload, as the ferritin storage becomes saturated, storage in hemosiderin is elevated. Hemosiderin is an insoluble complex made up of degraded ferritin and large ferric hydroxide chains. Iron stored in hemosiderin is poorly mobilized (104, 149).

LIVER IRON EXPORT

Iron export from the liver, both in Kupffer cells and hepatocytes, is unclear and far less is known about the molecular mechanisms as compared to iron uptake and storage in the liver. Iron that is stored in ferritin has the ability to be mobilized from the liver during times of high systemic demand of iron. This is the rationale for therapies of patients with hemochromatosis (discussed in more detail below). Patients with high liver iron are periodically bled initiating mobilization of iron from the liver to circulation. Rodent studies using radioactive iron tracers estimate that up to 6% of iron is released from hepatocytes daily (13). Several cues have been shown to regulate iron mobilization from the liver. Erythropoiesis and a systemic change in iron levels in rats increase the mobilization. Iron export is inhibited following inflammation. In addition, the Kupffer cells also contribute significantly to iron release from the liver via erythrophagocytosis and release of iron from RBCs (13). The first step in iron mobilization is the regulated release of iron from ferritin. This is thought to be an autonomous property of ferritin controlled by cytosolic iron levels (38, 142). Expression of FPN in cells increases iron release from ferritin (117). The only known iron exporter is FPN, which has been shown to be critical for iron transport in animals (43). FPN is expressed highly in macrophages and to a lesser degree in hepatocytes (144). As mentioned above, FPN is regulated by hepcidin-mediated binding and degradation. This pathway is well characterized in in vitro cell systems. However, the role hepcidin plays in regulating hepatic FPN protein stability in vivo is not clear. FPN is also regulated by the IRE/IRP system, and has an IRE in its 5’ UTR. Under conditions of cellular iron deficiency, IRP proteins bind to the IRE in the FPN transcript, blocking its translation. This leads to decreased protein expression of FPN on the membrane and allows the cell to retain iron through decreased export (113). Recent conditional disruption studies underscore the importance of FPN in the liver. Macrophage-specific deletion of FPN led to iron sequestration in Kupffer cells. The deletion did not have a profound affect on RBC parameters and only mild anemia was observed (189). This finding is quite surprising since most of the iron for daily requirements is derived from macrophage-mediated recycling of senescent RBCs. These data suggest there must be compensatory mechanisms when macrophage iron export is ablated. A hepatocyte-specific FPN deletion led to mild iron sequestration in hepatocytes. However, RBC parameters were normal. Under low iron conditions these mice developed anemia; RBC and hemoglobin values were significantly lower (190).

LIVER IRON OVERLOAD

The liver is central to iron homeostasis and depends on a complex feedback mechanism between body iron requirements, intestinal absorption, and recycling from senescent RBCs. Dysregulation of these mechanisms can lead to iron overload. This section will discuss common and rare disorders of iron overload.

Hereditary Hemochromatosis (HH)

HH is a genetic disorder and a common cause of iron overload. 1 in 200 will be affected by this disorder (128). It was first described by Armand Trousseau in 1865 and was referred to as bronze diabetes. A change in the hue of the skin, liver, and pancreas was observed, although the cause was not known at this time. Over 30 years later Von Recklinghausen named this condition hemochromatosis following further analysis showing iron accumulation in liver cells. In 1996 it was identified that a mutation in the HFE gene was associated with HH (46). It is now known that HH is an autosomal recessive disorder and 1 in 8 people in the United States have a mutation in a single copy of the gene (128). Further study of patients with HH has led to the identification of several other iron regulatory genes that cause HH. These genes demonstrate that iron sensing and regulation of hepcidin is a concerted effort of several proteins. All HH disorders demonstrate a dysregulation in the hepcidin-FPN homeostasis, and are classified into 5 types.

Type 1

High FE (HFE) encodes an atypical major histocompatibility complex protein, and mutations in this gene are the most common cause of HH (46). The most common mutation observed is a missense mutation of cysteine 282 to tyrosine (Cys282Tyr) (152). However, several other mutations are characterized leading to iron overload (152). HFE mutations that lead to iron overload are associated with a significant decrease in hepcidin expression. Consistent with these data, mouse models which are deleted for HFE or have a knock-in Cys282Tyr mutation also have iron overload and a decrease in hepcidin expression (102). Since HFE is abundant in several tissues including enterocytes and liver, a conditional disruption of HFE in the liver and intestine was generated. In this study, hepatocyte-specific disruption of HFE recapitulated a similar phenotype as the whole body knockout mouse model, characterized by iron overload and decrease in hepcidin expression (180). Mice with HFE disruption in the intestine were similar to normal controls (181). These data demonstrate that HFE in the hepatocytes is critical for iron homeostasis. The molecular function of HFE and its precise role in regulating hepcidin expression has been a subject of great interest. Several lines of evidence suggest that HFE binding to Tfr1 and Tfr2 may be the mechanism by which HFE regulates hepcidin expression (60, 153). Mutations in HFE that increased binding to Trf1 blocked hepcidin expression. Mutations that weakened HFE and Tfr1 interaction increased hepcidin expression (153). In addition, HFE and Tfr2 interact and disruption of Tfr2 leads to decreased hepcidin expression (50, 86, 116). Lastly HFE and Tfr2 interaction is required for regulation of hepcidin by iron containing Tf (60). Together the data suggest a mechanism where Tf binding to Tfr1 releases HFE, which then can bind to Tfr2 and stabilize its protein expression leading to an increase in SMAD signaling ().

Type 2A

Juvenile hemochromatosis (JH) is a rare autosomal recessive disorder of iron overload and symptoms become apparent before the age of 30. JH leads to organ damage, and usually causes cardiomyopathy, hypogonadism, liver injury, and diabetes. JH is a caused by mutations in the gene for HFE2 which encodes the hemojuvelin (HJV) protein (132, 152). HJV is a glycophosphatidylinositol anchored membrane protein. Several HFE2 mutations have been found in patients. However, the glycine 320 to valine is the most frequent mutation that is reported (152). To confirm that HJV is causative in this type of hemochromatosis, an HJV knockout mouse model was generated (125). This mouse model demonstrates severe iron overload associated with very low levels of hepcidin expression, similar to that observed in patients with HJV mutations (132). Hepcidin expression was appropriately increased in response to inflammatory stimuli, suggesting that HJV is involved in iron sensing but does not play a role in hepcidin regulation during inflammation. The early onset of iron overload in JH is due to a robust repression of hepcidin. In HH due to HFE mutations there is only a moderate decrease in hepcidin expression leading to iron overload that is symptomatic at later ages. HJV is expressed in several tissues, and in the liver HJV primarily expressed in hepatocytes. Restoring HJV expression in hepatocytes of HJV knockout mice completely restored hepcidin expression and ablated the iron overload (188). Further mechanistic studies demonstrated that HJV functions as a BMP co-receptor and is important for induction of hepcidin expression in response to BMP signaling (10). HJV binds to BMPs and enhances the activity of the SMAD signaling cascade (10).

Type 2B

Similar to HJV mutations, mutations in the HAMP gene, which encodes for hepcidin, are a very rare cause of JH. Currently 12 known mutations occur on the HAMP gene leading to a decrease in the normal production of hepcidin (74). Since hepcidin function or expression is dramatically diminished, the iron overload symptoms are observed before the age of 30.

Type 3

Tfr2 mutations lead to an autosomal recessive iron overload disease similar to HFE related-HH phenotype. Tfr2 as mentioned above is capable of binding to HFE and this interaction is critical in maintaining hepcidin expression (60). Unlike Tfr1, which is ubiquitously expressed, Tfr2 is expressed only in hepatocytes and erythroid precursors (87, 159). Tfr2 cannot compensate for the loss of Tfr1 (171). The knockout mouse model and the liver-specific disruption of Tfr2 confirm its importance in regulating hepcidin levels (86, 182). Hepcidin levels are decreased significantly in these mouse models compared to littermate controls, and tissue iron is increased. The most common mutation observed is in amino acid 245, which is converted into a stop codon resulting in a protein product that is not expressed (24). As mentioned above, Tfr2 binding to HFE promotes SMAD activation and hepcidin expression. Upon its deletion this signaling pathway is decreased causing a significant drop in hepcidin levels.

Type 4

SLC40a1 is the gene that encodes for the iron exporter FPN. FPN is the target of hepcidin, which causes rapid internalization and degradation of FPN (117). More precise work on the mechanism of FPN degradation by hepcidin demonstrates that following hepcidin binding, FPN is phosphorylated on tyrosine residues, which lead to its endocytic shuttling and degradation by the proteasome pathway (39). JAK2 is the critical kinase phosphorylating FPN (37). However, recent data demonstrate that both phosphorylation of FPN and JAK2 are not essential for FPN degradation (141, 148). Mutations in HFE, HJV, hepcidin, and Tfr2 are all recessive mutations. However, mutations in FPN are dominant. Patients that are heterozygous for the mutation develop the disease. This is due to FPN functioning as a dimer and the mutant protein can act as a dominant negative (40, 41). Several mutations of FPN have been observed. Detailed molecular studies demonstrate that the mutations inhibit proper membrane localization of FPN, inhibit the export function of FPN, disrupt hepcidin binding, or inhibit FPN internalization (84). Therefore, depending on the mutation in FPN the patients can present with very different phenotypes. Mutations that inhibit membrane localization or export function can lead to macrophage iron overload. While those mutations that inhibit hepcidin binding or hepcidin-meditated internalization lead to continuous export of iron into serum and eventually iron overload in the hepatocytes.

Secondary hemochromatosis

Secondary hemochromatosis is the result of another disease, which causes excess liver iron loading. Most of diseases that lead to secondary hemochromatosis are acquired disorders of erythropoiesis (63). The most common causes of secondary hemochromatosis are listed in . A well-studied disorder that leads to secondary hemochromatosis is β-thalassemia. β-Thalassemia is a congenital blood disorder due to mutations in the β-globin gene leading to a partial or complete loss of β-globin synthesis resulting in β-thalassemia intermedia and Cooley’s anemia, respectively. The decrease in β-globin results in ineffective erythropoiesis and erythropoietic stress. Persons with β-thalassemia intermedia have mild anemia with a slight lowering of hemoglobin levels in the blood. In most cases treatment is not necessary, but severe patients with low hemoglobin levels will need occasional blood transfusions (161). Cooley’s anemia results in a striking deficiency in hemoglobin production. Patients will need frequent blood transfusions (161). The blood transfusions lead to dysregulation of the systemic iron homeostasis since donor blood is a rich source of iron. The body cannot eliminate the excess iron efficiently, leading to increased tissue iron. Regular blood transfusions are the most common cause of secondary hemochromatosis (63). Initially it was thought that the iron overload was primarily due to regular blood transfusions. However, mouse models of β-thalassemia hyperabsorb iron. This is the major mechanism leading to iron overload in β-thalassemia intermedia and significantly contributes to the tissue iron overload in Cooley’s anemia (76, 164, 186). It is less clear whether an increase in iron absorption plays a significant role compared to blood transfusions in other disorders of erythropoiesis listed in . However, recent work has shown that effective and ineffective erythropoiesis can stimulate iron absorption, therefore this mechanism of iron overload may be true for other diseases leading to secondary hemochromatosis (5). Increased iron absorption in secondary hemochromatosis may be due to an increase intestinal hypoxia signaling and a decrease in hepcidin expression (5, 133). Increasing hepcidin levels in mouse models of β-thalassemia improved liver iron loading and anemia (62).

Table 1

Common causes of secondary hemochromatosis

Aceruloplasminemia, hypotransferrinemia, and DMT-1 deficiency

Aceruloplasminemia, hypotransferrinemia, and DMT1 deficiency are causes of secondary hemochromatosis, but are not disorders of erythropoiesis; rather these disorders are due to ineffective transport of iron. Aceruloplasminemia is due to a loss-of-function mutation in Cp and is inherited in an autosomal recessive manner. Iron overload in aceruloplasminemia is mainly observed in the brain and liver (73, 187). A similar phenotype is also observed in Cp knockout mouse models. Hypotransferrinemia is an autosomal recessive disorder leading to loss of Tf production. Hypotransferrinemia is associated with severe microcytic anemia, and an adaptive increase in iron absorption, which leads to severe liver iron loading (64, 75). Consistent with hypotransferrinemic patients, the hpx mouse, which produces no Tf has liver iron overload and anemia (171). DMT1 deficiency is an autosomal recessive disorder leading to increase in liver iron (80). This is thought to be due to the role of DMT1 in iron export from the endocytic compartment.

Dysmetabolic iron overload syndrome (DIOS)

DIOS is a newly characterized secondary hemochromatosis disorder. DIOS is associated with features, such as obesity, type 2 diabetes, alcohol use, and chronic hepatitis C (42, 81, 106, 139). This is now the most common cause of iron overload observed in patients. The iron overload is observed in 15% of patients with metabolic syndrome, 50% in patients with non-alcoholic fatty liver disease, over 40% of patients with chronic hepatitis C infection, and significant number of patients with alcoholic liver disease (18, 81, 139, 174). Currently the mechanisms, which contribute to DIOS are unclear. However, a significant decrease in FPN gene expression has been noted in patients with DIOS (2).

IRON-INDUCED LIVER DAMAGE

High levels of iron deposition lead to tissue damage and dysregulation of function. In the liver increased free iron if untreated, leads to fibrosis and cirrhosis, and can increase morbidity and mortality (123, 124, 145). Hepatic tissue injury is directly correlated to the duration and amount of iron loading (123, 124). Cells normally produce basal levels of reactive oxygen species (ROS) through metabolic function of the mitochondria and other organelles. ROS are kept at low basal levels by several antioxidant enzymes, and low levels of ROS are important in normal cell physiology (146). ROS in conjunction with high cellular iron results in a robust increase of hydroxyl radicals, which leads to cell damage. Free iron generates ROS through the Fenton and Haber-Weiss reactions (). The superoxide radical (O₂^•−) reduces ferric iron to ferrous iron, which reacts with hydrogen peroxide (H₂O₂) to generate highly reactive hydroxyl radicals (OH^•) (90). Hydroxyl radicals lead to an increase in peroxidation of phospholipids within organelle and cellular membranes, oxidation of amino acid side-chains, DNA strand breaks, and protein fragmentation. The exact mechanisms by which high intracellular iron leads to liver fibrosis and cirrhosis are unclear, but iron-induced cellular damage has been shown to directly increase hepatocyte cell death and activate Kupffer and stellate cells (135, 160) (). More recently, clinical evidence suggests that high liver iron could play a role in insulin resistance (45, 143). The increase in iron-induced ROS in Kupffer cells can initiate a pro-inflammatory cascade in the liver. Increased ROS production activates NF-κB signaling leading to an increase in IL-6, TNF-α, and IL-1β in the liver (22, 112). Liver inflammation can lead to hepatic insulin resistance, which is a major pathway leading to hyperglycemia in type II diabetes (55). Lastly, liver iron overload increases the risk for hepatocellular carcinoma (HCC). HCC is the major life threatening complication associated with hereditary hemochromatosis (124). Several studies have looked at the risk factor for HCC in hereditary hemochromatosis patients and some have estimated the risk to be 100-200 fold higher in HH patients (19, 162). In addition, other iron overload disorders such as thalassemia are associated with an increased risk for HCC (17). The increase in iron plays an active role in HCC pathogenesis. Chelation of iron or placing mice on an iron deficient diet decreases tumor growth (71, 151).

Schematic diagram of the Fenton and Haber-Weiss reactions

Iron is a potent catalytic cofactor, which increases highly unstable oxygen radicals that cause cellular damage.

Iron-induced liver damage

Iron accumulation in hepatocytes and Kupffer cells leads to an increase in ROS production and pro-inflammatory mediators. Both ROS and pro-inflammatory mediators initiate a feed forward cycle, which activates stellate cells, initiates cell damage, and leads to loss of function contributing to an increase in steatosis, fibrosis, cirrhosis, and HCC.

TREATMENT OF IRON OVERLOAD

In patients with HH, phlebotomy is used to decrease liver iron. Regular bleeding of the patients leads to an increase in erythropoiesis, which mobilizes liver iron stores to meet the demand for iron that is required to generate mature RBCs (59). Secondary hemochromatosis is also associated with severe anemia, thus phlebotomy is not an option. In these patients iron chelators are used to decrease liver iron (59). Deferoxamine has been used for over three decades for iron chelation. More recently two new iron chelators have also been used, deferiprone and deferasirox (93). Iron chelators have been shown to be effective in decreasing liver iron and also morbidity and mortality associated with iron overload. Several rounds of phlebotomy and/or administration of iron chelators are required; these are slow-acting treatment options and may not successfully decrease the liver injury associated with an increase in iron. Alternatives to existing treatments are needed. Recently, hepcidin has been shown to be a very attractive target and good proof of principles studies have been done. Several studies in mouse models of HH have shown that increasing hepcidin levels can ameliorate the iron overload (122, 177). Recently in β-thalassemia models, increasing hepcidin expression resulted in decreased liver iron (62). Currently several strategies are being assessed to increase hepcidin levels. The specific sites that are required for hepcidin-FPN binding are known. Precise structural mutagenesis studies have demonstrated that nine amino acids of hepcidin are critical for the binding to FPN and initiating its internalization (140). Modifications of these nine amino acids has led to several peptides that have increased activity over full length hepcidin. Moreover, the modified hepcidin derived peptides are functional in vivo and can prevent iron overload that is observed in hepcidin knockout mice (140). These agents are stable orally and may provide a well-tolerated form of treatment for hereditary and secondary hemochromatosis. Other approaches may also be useful in increasing hepcidin expression (59). Treatment with BMP6 increases hepcidin expression in HFE-null mouse model and prevents iron overload (34). Moreover, since hepcidin regulatory pathways are well characterized, several possibilities such as SMAD, C/EBPα, and STAT3 activators could have potential roles in increasing hepcidin expression in vivo.

ANEMIA

On the other end of the spectrum, aberrant upregulation of hepcidin is critical in the pathogenesis of anemia of chronic disease, which encompasses several diseases including kidney disease, inflammatory disease, cancer, and aging (26). In healthy human volunteers and mice, studies demonstrate that inflammatory agents cause a robust and rapid decrease in serum iron levels (27, 88, 115, 147). Within hours following induction of inflammation, hepcidin levels are significantly elevated. The decrease in serum iron is due to hepcidin-mediated internalization of FPN leading to iron sequestration in macrophages. This is thought to be a protective mechanism that limits iron available to infectious pathogens. However, in chronic disorders this leads to anemia, which can have detrimental effects in the primary disease pathogenesis. Similarly, in most cancers there is a decrease in serum iron levels and increased anemia (156). The decrease in iron is suggested to be beneficial in limiting tumor growth. The mechanism may vary depending on the primary disease. However, the IL-6-STAT3 pathway is critical in increasing hepcidin expression during inflammation (163). The best treatment for anemia of chronic disease is resolving the primary chronic disease. In severe cases, blood transfusion, EPO, or intravenous administration of iron is used (163). In addition to an increase in hepcidin expression from chronic disorders, rare genetic mutations in the TMPRSS6 gene cause an increase in hepcidin expression and iron-refractory iron deficiency anemia (IRIDA) (47). This was further confirmed in the TMPRSS6 knockout mice, which had hair loss and microcytic anemia associated with high levels of hepcidin expression (48, 52). A similar finding was noted in the mask mutant mouse strain, characterized by a premature stop codon in the TMPRSS6 gene (44). IRIDA is iron deficiency anemia that is unresponsive to oral iron therapy. TMPRSS6 (also known as matriptase-2) encodes a type II transmembrane serine protease, which is expressed predominantly in the liver. The first substrate that was characterized for TMPRSS6 was HJV. The serine protease of TMPRSS6 cleaves membrane HJV, leading to downregulation of hepcidin expression (155). Mutations in TMPRSS6 decrease its protease activity leading to increased protein expression of HJV and a coordinate increase in hepcidin expression.

CONCLUSION

The liver is the central tissue which regulates systemic iron homeostasis by acting as a sensor and regulator of iron levels. In addition, through its role in iron storage, the liver can protect more sensitive tissues from iron-induced cellular injury. The past decade has led to several novel liver-derived players regulating systemic iron homeostasis and identification of new mutations in iron-related disorders. Several pathways in the liver regulate hepcidin, the master hormone for maintaining systemic iron homeostasis. Recent studies have shown that these pathways can have redundancy or act independently based on the stimuli (172). The challenge will be to understand how these pathways crosstalk and are regulated in a coordinate manner to maintain hepcidin levels. Moreover, how can these pathways be targeted in iron-related disorders that could be of therapeutic benefit? Targeted therapies for iron related disorders are actively being pursued such as the case for hepcidin mimetics and BMP agonists, and the coming decade should yield novel therapies.

ACKNOWLEDGEMENTS

This work was supported by grants to Y.M.S from the National Institutes of Health (CA148828 and DK095201), The University of Michigan Gastrointestinal Peptide Center, and Jeffrey A. Colby Colon Cancer Research and the Tom Liu Memorial Funds of the University of Michigan Comprehensive Cancer Center. E.R.A is supported by the Rackham Predoctoral Fellowship, University of Michigan.

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Iron homeostasis in the liver

Compr Physiol. Author manuscript; available in PMC 2014 Feb 27.

Published in final edited form as:

PMCID: PMC3936199

NIHMSID: NIHMS551241

Erik R Anderson

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

Yatrik M Shah

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

¹Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor MI. 481109

²Department of Internal Medicine Division of Gastroenterology, University of Michigan, Ann Arbor MI. 481109

The publisher’s final edited version of this article is available at Compr PhysiolSee other articles in PMC that cite the published article.

Abstract

Iron is an essential nutrient that is tightly regulated. A principal function of the liver is the regulation of iron homeostasis. The liver senses changes in systemic iron requirements and can regulate iron concentrations in a robust and rapid manner. The last 10 years have led to the discovery of several regulatory mechanisms in the liver which control the production of iron regulatory genes, storage capacity, and iron mobilization. Dysregulation of these functions leads to an imbalance of iron, which is the primary causes of iron-related disorders. Anemia and iron overload are two of the most prevalent disorders worldwide and affect over a billion people. Several mutations in liver-derived genes have been identified, demonstrating the central role of the liver in iron homeostasis. During conditions of excess iron, the liver increases iron storage and protects other tissues, namely the heart and pancreas from iron-induced cellular damage. However, a chronic increase in liver iron stores results in excess reactive oxygen species production and liver injury. Excess liver iron is one of the major mechanisms leading to increased steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma.

INTRODUCTION

Iron is an essential micronutrient that is a critical component of oxygen transport proteins (hemoglobin and myoglobin) and of numerous metabolic and redox enzymes. The average adult has 2–4 grams of iron, and over 80% is contained in hemoglobin of red blood cells (RBC). Chronic iron deficiency results in decreased hemoglobin production and anemia. Systemic iron levels are tightly controlled through an integrative mechanism that involves iron absorption, storage, and recycling. The past decade has been termed “The Golden Age of Iron Biology”, due to the significant increase in the understanding of the molecular underpinnings of systemic iron homeostasis (7). When iron regulatory pathways are dysregulated, this leads to either excess tissue iron or iron deficiencies, which affect over a billion people worldwide. Four major cell types or tissues have been shown to be critical for systemic iron homeostasis ():

Systemic iron regulation

Dietary iron is absorbed through the small intestine and mainly utilized for RBC production. Hepatic and splenic macrophages recycle iron from senescent RBCs. The iron derived from recycling is used for production of RBCs. During times of iron excess the liver can store iron and during increased systemic needs the liver can mobilize iron stores for utilization.

Enterocyte

Dietary iron absorption is tightly regulated in the small intestine. Dietary iron enters the body through absorptive cells in the duodenum. Dietary ferric iron (Fe⁺³) is reduced by the apical ferric reductase duodenal cytochrome b (DcytB) to ferrous iron (Fe⁺²) and transported into the enterocyte via an apical iron transporter, divalent metal transporter-1 (DMT1, also known as Nramp2, SLC11a2 and DCT1) (49, 69, 100, 107, 109). Once iron enters into the cytoplasm through DMT1, iron is either stored or exported by the iron exporter ferroportin (FPN, also known as SLC40A1)(1, 43, 110) located on the basolateral side of the enterocyte. Disruption of DMT1 or FPN leads to inhibition of iron absorption and dysregulation of systemic iron homeostasis (43, 68). Following export of iron through FPN, iron is oxidized back to the ferric form by the ferroxidase hephaestin and ceruloplasmin (Cp) and loaded onto transferrin (Tf) and circulated through the body (70).

Erythroblast

A significant portion of circulating Tf-bound iron is utilized in early RBC precursors called erythroblasts for the synthesis of hemoglobin. Hemoglobin comprises of about 95% of the total cellular protein of the mature RBC and more than 80% of functional iron in the body is found in hemoglobin (28). Iron is an essential cofactor for the ability of RBCs to transport oxygen, and thus a decrease in body iron levels is the most predominant cause of anemia worldwide.

Macrophage

Only 1–2 mg per day of dietary iron is required to be absorbed through the intestine. This is due to the highly efficient recycling of iron from senescent erythrocytes. Recycling is performed by splenic and hepatic macrophages. Aging erythrocytes at 120 days old undergo specific changes that can be recognized by macrophages, thus initiating erythrophagocytosis. Iron is recovered from the degradation of hemoglobin and heme by hydrolytic enzymes in the phagocytic vesicles and heme-oxygenase-1 (HO-1) (58). Iron is routed back to circulation through the basolateral iron transporter FPN. Recent studies using macrophage-specific conditional deletion of FPN demonstrate the importance of FPN in exporting iron out of macrophages following erythrophagocytosis (189, 190).

Liver

The liver performs three essential functions in maintaining systemic iron homeostasis: 1) The liver is the major site for production of proteins that maintain systemic iron balance, 2) It is a storage site for excess iron and 3) the liver is critical for the mobilization of iron from hepatocytes to the circulation to meet metabolic requirements. Dysregulation of the liver’s ability to maintain balance of these three parameters leads to iron-related disorders. This review will focus on the normal function of the liver in iron homeostasis and the role of liver in iron-related disorders.

PRODUCTION OF PROTEINS FOR IRON HOMEOSTASIS

(Tf)

Tf is an 80 kDa plasma glycoprotein generated predominantly in the liver (138). However, small amounts are also made in the brain and testis (14, 157). Tf is the major serum iron-binding protein and is essential for systemic circulation of iron. Tf consists of two globular lobes of α-helices and β-sheets, which have a significant amount of homology between the N- and C- terminal halves of the molecule. This homology is thought to be due to a gene duplication arising from a 40kDa ancestral protein containing a single lobe (3, 96-98). Each lobe includes metal binding amino acid residues (2 Tyr, 1 His, and 1 Asp). Tf binds to ferric iron reversibly and with high affinity, but does not bind to ferrous iron (3, 4). In addition, Tf can also bind other metals with lower affinity (3). The affinity of iron is also regulated by pH, as iron binding to Tf decreases as the pH drops, with no detectable amounts observed below pH 4.5 (3, 4). The reversible binding of iron to Tf is important and allows it to be a cellular iron donor or iron acceptor depending on the systemic requirements of iron.

(Cp)

Cp is a copper-dependent serum ferroxidase which works in tandem with FPN for cellular iron export into circulation (129, 130). Cellular iron is usually in the ferrous form and is exported out of the cell by FPN, however the affinity of Tf for ferrous iron is relatively weak and Cp is required to oxidize iron to its ferric form, which binds with high affinity to Tf (129, 130). Disruption of Cp in mouse models further supports the role of Cp in systemic iron homeostasis (30, 70).

Ferritin

Ferritin is a multi-subunit protein that consists of a heavy and light chain. Ferritin from different tissues can vary in the ratio of heavy compared to light chains. Liver ferritin contains mainly the light chain and can store up to 4500 atoms of iron. Hepatocytes are the major site for ferritin synthesis, however most cells that have been assessed can synthesize ferritin to a smaller degree (12, 167). Cells with high iron levels are capable of an adaptive increase in ferritin synthesis. Both ferritin subunit synthesis are controlled through a post-transcriptional mechanism by the iron response element (IRE)/iron regulatory protein (IRP) system. The ferritin mRNAs contain a single IRE in the 5’ UTR. Under iron deficient conditions, the IRPs bind to this IRE and repress translation. However, as intracellular iron increases in a cell, the repression is relieved and ferritin synthesis is increased. For more details of the IRE/IRP system, refer to the comprehensive review by Muckenthaler, M.U et al. (113). Although the IRP/IRE regulation of ferritin synthesis is the predominant pathway that allows an adaptive increase, several other mechanisms, such as IL-1, TNFα, hypoxia, and oxidative stress have also been observed to regulate ferritin expression (23, 169, 170, 173, 184). It is likely that a combination of these pathways allow an efficient increase in ferritin when required.

Hepcidin

Hepcidin is a 25 amino acid antimicrobial peptide that is produced in hepatocytes and secreted into the circulation (91). Hepcidin is translated as an 84 amino acid pro-protein, and cleaved by the pro-hormone convertase furin to produce the active peptide (175). Subtractive hybridization experiments between iron loaded and control mouse livers initially identified hepcidin as an iron-regulated protein (136). The role of hepcidin in iron homeostasis was further confirmed when hepcidin was inadvertently disrupted in addition to USF2 and these mice were severely iron overloaded (119). The effect of hepcidin knockout on iron homeostasis was confirmed in a subsequent paper, which showed that animals deficient in hepcidin, but having normal USF2 expression developed severe hemochromatosis (101). Conversely, animals that overexpress hepcidin in the liver demonstrate severe iron-deficiency anemia (120). A clue to the function of hepcidin was discovered after subjecting rats to an iron-deficient diet. Following iron deficiency, rats rapidly repress liver hepcidin and upregulate the intestinal iron transporters DMT1, DcytB, and FPN (54). Hepcidin was believed to regulate iron homeostasis due to an interaction with iron transporters. A breakthrough came when the hepcidin receptor was discovered. Through careful in vitro analysis, it was determined that hepcidin binds to FPN which leads to its internalization and proteasomal degradation (117). Hepcidin is increased in iron loading, which leads to a decrease in duodenal iron absorption to normalize iron levels. In the case of iron deficiency, hepcidin is repressed which allows more iron to be transported from the enterocyte to the serum (). In addition, FPN is increased in macrophages following erythrophagocytosis, and hepcidin represses FPN in these cells (89). In macrophages during iron loading, hepcidin decreases iron transport following iron recycling from senescent RBCs. Whereas during iron deficiency, a significant increase in iron efflux would be expected following erythrophagocytosis (). In addition to iron levels resulting in the regulation of hepcidin levels, hypoxia and erythropoiesis are major repressors of hepcidin expression. This provides a novel link between oxygen homeostasis and iron levels. Moreover, inflammation is a major activator of hepcidin expression. This results in restriction of serum iron levels during an infection, and therefore is less conducive for growth of pathogenic bacteria. The last decade has shown the importance of hepcidin as the master regulator of both duodenal iron absorption and red blood cell (RBC) iron recycling. Studying the regulation of hepcidin expression in the liver has been a priority for understanding regulation of systemic iron homeostasis. The transcriptional regulation of hepcidin has been reviewed elsewhere (56, 114) and is summarized in . Here, we only briefly mention a few major pathways that are critical in hepcidin regulation. In addition to the major pathways, several other accessory proteins have been identified, which are mutated in iron-related disorders. Their importance in hepcidin regulation is covered in more detail below in the liver iron overload and anemia section.

Hepcidin regulation of FPN protein expression during changes in systemic iron levels

High iron levels increase hepcidin expression, which decrease iron export from the small intestine and macrophage due to an internalization and degradation of FPN. Iron deficiency results in a decrease in hepcidin levels and stabilization of FPN protein expression.

Regulation of hepcidin by BMP/SMAD, inflammatory and hypoxia/erythropoietic signaling in the liver

Three major pathways are critical for regulating basal and stimuli-induced hepcidin expression. Binding of iron containing Tf to Tfr1 causes a dissociation of Tfr1-High FE (HFE) complex and an interaction of HFE with Tfr2. Increased stabilization of Tfr2 increases BMP6 mediated phosphorylation of SMAD1/5/ 8 and recruitment of SMAD 1/5/8 and SMAD4 to the hepcidin proximal promoter. BMP/SMAD signaling is the major pathway by which hepcidin expression is coordinated to meet systemic iron requirements. Activation of hepcidin by inflammation is thought to act independently of the BMP/SMAD pathway. The best-studied mechanism is via the pro-inflammatory mediator IL-6. Binding of IL-6 to its receptor IL-6 receptor (IL-6R) initiates activation of the JAK-STAT3 pathway. STAT3 binds directly to the proximal promoter to increase hepcidin expression. Hypoxia and erythropoiesis are inhibitors of hepcidin expression and these are the least understood pathways by which hepcidin expression is regulated. Hypoxia and erythropoiesis have been shown to inhibit hepcidin expression via direct binding of HIF to the proximal promoter, an EPO-EPO receptor (EPOR) mediated decrease in C/EBPα expression, and through increase in an unknown erythroid derived factor which signals through an undefined pathway.

SMAD/ bone morphogenetic protein (BMP) signaling

A key finding in the regulation of hepcidin is the essential role of the BMP-SMAD signaling cascade. BMPs are ligands that belong to the transforming growth factor-β (TGF-β) superfamily. BMPs bind to type I and type II serine threonine kinase receptors, which phosphorylate specific intracellular SMAD proteins (SMAD1/5/8). Phosphorylated SMAD1/5/8 (P-SMAD1/5/8) binds to the common mediator SMAD4, and the SMAD complex translocates to the nucleus to modulate transcription of target genes. BMPs, but not TGF-β signaling induced hepcidin expression in cultured liver cell lines and in vivo (11). Several BMPs signal through SMAD activation, however BMP6 is the endogenous ligand that modulates hepcidin expression. BMP6 knockout mice have decreased hepcidin expression and an increase in tissue iron (8, 10, 11, 111). BMP6 knockout mice still have the ability to increase hepcidin expression following inflammatory stimuli. The importance of SMAD signaling in hepcidin regulation was demonstrated in mice with a hepatocyte-specific disruption of SMAD4. A near complete loss of hepcidin expression was observed in these mice and eventually the mice die of severe iron overload in multiple tissues (183). Interestingly, liver SMAD4 knockout mice are unable to increase hepcidin expression in response to iron loading, suggesting that SMAD4 mediates the response of hepcidin to changes in systemic iron requirements. Signaling through the BMP receptor leads to SMAD1/5/8 phosphorylation, which is required for SMAD4 transcriptional activity (6). The regulation of hepcidin by iron loading and iron deficiency is correlated with phosphorylation of the SMAD1/5/8 proteins. In iron deficiency, pSMAD1/5/8 is decreased dramatically, and is substantially increased in conditions of iron overload (85).

STAT3 and inflammatory pathways

Hepcidin is induced in response to inflammatory stimuli (57, 118). Hepcidin induction by inflammation leads to iron sequestration, which can decrease bacterial growth. However, chronic diseases are associated with anemia (150). The mechanism for the induction of hepcidin by inflammation is mediated by IL-6 in cultured cells, mice, and humans (115). Subsequently, it was shown that IL-6 regulates expression of hepcidin by inducing STAT3 binding to the hepcidin promoter (176, 185). To support this, it was shown that anti-IL-6 receptor antibody improves anemia of inflammation (158).

Hypoxia and erythropoietic pathways

Hypoxia, or low oxygen tension, is a physiological condition that results in numerous adaptive changes in gene expression. Hypoxia represses hepcidin expression both in cultured cells and in mice (121). Many mechanisms have been proposed to play a role in hypoxic hepcidin repression (29, 31, 94, 134, 179). Hypoxia-inducible factor (HIF) is the major transcription factor activated following hypoxia and HIF is thought be critical in the hypoxia-mediated repression of hepcidin (20). HIF was shown to bind directly to the hepcidin promoter resulting in repression (134). However, recent studies refute these findings (108, 179). It has been reported that hypoxia through a HIF2α mediated increase in erythropoiesis is the critical pathway leading to hepcidin repression (108). Interestingly, a mutation that leads to HIF stabilization causes Chuvash polycythemia. In these patients a decrease in hepcidin expression is observed without a significant association with erythropoiesis (65). There is no clear mechanism that mediates hepcidin repression during hypoxia, and this is an active area of study

Erythropoiesis is a well-characterized pathway leading to hepcidin repression. In several mouse models that induce erythropoiesis, a significant decrease in hepcidin is observed (16, 53, 61). The decrease in hepcidin allows the increase of iron required for RBCs during erythropoiesis. In a model of intensive care anemia, erythropoietin injections or phlebotomies were able to repress hepcidin expression despite high levels of IL-6, which is known to strongly increase hepcidin expression (99). Similarly, hypoxia is able to repress hepcidin expression under conditions of high IL-6. One of the mechanisms by which erythropoiesis represses hepcidin is through erythropoietin (EPO) binding to its receptor in hepatocytes, which leads to downregulation of C/EBPα and hepcidin repression (137). C/EBPα is a liver-enriched transcription factor that is important in hepcidin regulation. Mice with a liver specific deletion of C/EBPα have low levels of hepcidin expression (35). However, erythropoietic blockers prevented the suppression of hepcidin (178), suggesting an EPO-independent erythropoietic derived mechanism is responsible for the decrease in hepcidin. Growth differentiation factor 15 (GDF15) and twisted grastrulation 1 (TWGS1) are secreted during erythroblast maturation and can inhibit hepcidin expression (165, 166). However, their role in hepcidin repression during erythropoiesis is still unclear (9, 82).

LIVER IRON IMPORT

Tf-bound iron

The major mechanism for iron uptake in the liver and most other tissues is through the Tf/transferrin receptor (Tfr) system (). Tf as discussed above is a constitutively expressed protein. Tfr1 transcript stability is regulated by the IRP/IRE system. Unlike ferritin, which has a single IRE in its 5’ UTR, Tfr1 transcript contains several IREs in its 3’ UTR. IRPs bind to the IRE in the Tfr1 transcript, which increases mRNA stability (77). Under low iron conditions more Tfr1 is translated allowing for increased iron uptake through Tf. Under conditions of high iron the IRPs are inactivated that leads to decreased Tfr1 mRNA stability and decreased iron uptake. Iron circulates bound to Tf. When both lobes are occupied with iron (Diferric Tf), this complex binds with high affinity to Tfr1 (3, 4). Diferric Tf-TFR1 binding activates cellular iron uptake by receptor-mediated endocytosis, and this pathway is a model system to study the precise mechanisms of receptor-mediated endocytosis (15, 32, 33, 95). Diferric Tf internalization into endocytic vesicles initiates the release of iron from Tf via acidification of endosomes (36, 72). Ferric iron is reduced to ferrous iron by an endosomal ferric reductase (126). Through a positional cloning strategy a transmembrane protein Steap3 was found to be a critical reductase in the endosome. Mutations in the steap3 gene lead to microcytic anemia (126). Steap3 is highly expressed in the hematopoietic cell lineage, but the role of steap3 in other cell types is not clear. However, there are three other family members (Steap1, 2, and 4) that also contain ferric reductase activity (127). Ferrous iron is then transported to the cytosol via DMT1, which in addition to being localized on the brush border cells of the small intestine, is also observed on recycling endosomes (25, 67). Interestingly, mice that lack DMT1 are still capable of accumulating hepatic iron, suggesting that DMT1 is not essential for Tf-bound iron uptake or other transporters have a redundant role (68). ZIP14, a family member of the ZIP metal transporters is also localized to endosomes and is important for the movement of iron from endocytic compartment to the cytosol (191). Following iron transport to the cytosol Tf and Tfr1 are recycled back to circulation and to the cell membrane, respectively (15, 32, 33, 95).

Mechanisms of liver iron uptake

Iron is imported into the liver via Tf/Tfr mediated endocytosis. As the pH of the endocytic vesicle drops, iron is released, reduced to Fe²⁺ by an endocytic reductase, and transported out by DMT1 and/or ZIP14. During iron overload a significant amount of NTBI is present. Iron can be directly transported into the liver through membrane bound DMT1 and/or ZIP14. During conditions of increased hemolysis the liver is capable of transport of hemoglobin and heme. Free hemoglobin binds with high affinity to haptoglobin, whereas free heme binds to hemopexin. These complexes bind to their respective receptors CD163 and Lrp/CD91, which initiate receptor-meditated endocytosis. Hemoglobin is degraded in the endosome and heme is released from the endocytic vesicle. Heme is further degraded by HO-1 releasing iron.

Non-transferrin bound iron (NTBI)

In cases of severe iron overload, the level of iron will exceed the capacity of Tf, and there is a greater ratio of iron in the plasma that occurs as NTBI. NTBI is bound by a number of non-protein ligands including citrate which is likely to be the predominate form of plasma NTBI found in hemochromatosis (21, 66). The liver uptake of ferric citrate involves dissociation of citrate and transport of iron into the hepatocyte (). NTBI is efficiently taken up by hepatocytes and the uptake is not downregulated by excess iron in the liver as observed with Tf-bound iron through the IRE/IRP system. Several mechanisms have been shown to contribute to NTBI uptake including membrane bound DMT1 and ZIP14 facilitating direct uptake of iron into the hepatocytes (105, 154). In addition, several other mechanisms are capable of NTBI transport into the cells. L-type calcium channels facilitate transport of NTBI into cardiac myocytes. Calcium channel blockers inhibit NTBI uptake into the heart (131). Lipocalin 2 is a multi-functional protein, which has iron-sequestering properties and is critical in binding to siderophores and limiting iron to pathogenic bacteria (51). However, some data suggest that lipocalin 2 could meditate NTBI uptake (83). Scara5 is a ferritin receptor that mediates NTBI uptake into the kidney (103). However, the role of these pathways for hepatic NBTI uptake is not clear.

Heme and hemoglobin associated iron

The liver also has the capacity to acquire iron from heme or hemoglobin (). These pathways under normal conditions contribute to a negligible amount of iron uptake in the liver. However, during hemolysis this could lead to a substantial amount of heme or hemoglobin being taken up by the liver. In several diseases such as hemolytic anemia, gram-positive bacterial infection, and malaria, increased hemolysis leads to excess hemoglobin and heme. The liver derived scavenging proteins, haptoglobin and hemopexin rapidly sequesters free hemoglobin and heme. Haptoglobin and hemopexin proteins bind with high affinity to free hemoglobin and heme, respectively (79, 168). Once sequestered, the haptoglobin-hemoglobin complex binds to CD163, which is highly expressed on mature tissue macrophages, including Kupffer cells (92). The hemopexin-heme complex binds to LRP/CD91, which is expressed in several cell types, including macrophages and hepatocytes (78). Following binding to their respective receptor the complex is endocytosed and degraded through a lysosomal pathway. Iron is released from heme by the HO-1 and enters the same intracellular pool as iron from other sources as mentioned above.

LIVER IRON STORAGE

Regardless of the source, iron that enters the hepatocyte enters the same intracellular pool. This pool of iron is stored, mobilized for systemic metabolic demands, used in intracellular enzymes, or used in mitochondrial iron sulfur proteins. Since free intracellular iron is toxic, the majority of iron in the cells is stored in ferritin, which is discussed above in more detail. Within the liver all cell types can store iron, but under normal conditions hepatocytes represent the major storage site. During severe iron overload, as the ferritin storage becomes saturated, storage in hemosiderin is elevated. Hemosiderin is an insoluble complex made up of degraded ferritin and large ferric hydroxide chains. Iron stored in hemosiderin is poorly mobilized (104, 149).

LIVER IRON EXPORT

Iron export from the liver, both in Kupffer cells and hepatocytes, is unclear and far less is known about the molecular mechanisms as compared to iron uptake and storage in the liver. Iron that is stored in ferritin has the ability to be mobilized from the liver during times of high systemic demand of iron. This is the rationale for therapies of patients with hemochromatosis (discussed in more detail below). Patients with high liver iron are periodically bled initiating mobilization of iron from the liver to circulation. Rodent studies using radioactive iron tracers estimate that up to 6% of iron is released from hepatocytes daily (13). Several cues have been shown to regulate iron mobilization from the liver. Erythropoiesis and a systemic change in iron levels in rats increase the mobilization. Iron export is inhibited following inflammation. In addition, the Kupffer cells also contribute significantly to iron release from the liver via erythrophagocytosis and release of iron from RBCs (13). The first step in iron mobilization is the regulated release of iron from ferritin. This is thought to be an autonomous property of ferritin controlled by cytosolic iron levels (38, 142). Expression of FPN in cells increases iron release from ferritin (117). The only known iron exporter is FPN, which has been shown to be critical for iron transport in animals (43). FPN is expressed highly in macrophages and to a lesser degree in hepatocytes (144). As mentioned above, FPN is regulated by hepcidin-mediated binding and degradation. This pathway is well characterized in in vitro cell systems. However, the role hepcidin plays in regulating hepatic FPN protein stability in vivo is not clear. FPN is also regulated by the IRE/IRP system, and has an IRE in its 5’ UTR. Under conditions of cellular iron deficiency, IRP proteins bind to the IRE in the FPN transcript, blocking its translation. This leads to decreased protein expression of FPN on the membrane and allows the cell to retain iron through decreased export (113). Recent conditional disruption studies underscore the importance of FPN in the liver. Macrophage-specific deletion of FPN led to iron sequestration in Kupffer cells. The deletion did not have a profound affect on RBC parameters and only mild anemia was observed (189). This finding is quite surprising since most of the iron for daily requirements is derived from macrophage-mediated recycling of senescent RBCs. These data suggest there must be compensatory mechanisms when macrophage iron export is ablated. A hepatocyte-specific FPN deletion led to mild iron sequestration in hepatocytes. However, RBC parameters were normal. Under low iron conditions these mice developed anemia; RBC and hemoglobin values were significantly lower (190).

LIVER IRON OVERLOAD

The liver is central to iron homeostasis and depends on a complex feedback mechanism between body iron requirements, intestinal absorption, and recycling from senescent RBCs. Dysregulation of these mechanisms can lead to iron overload. This section will discuss common and rare disorders of iron overload.

Hereditary Hemochromatosis (HH)

HH is a genetic disorder and a common cause of iron overload. 1 in 200 will be affected by this disorder (128). It was first described by Armand Trousseau in 1865 and was referred to as bronze diabetes. A change in the hue of the skin, liver, and pancreas was observed, although the cause was not known at this time. Over 30 years later Von Recklinghausen named this condition hemochromatosis following further analysis showing iron accumulation in liver cells. In 1996 it was identified that a mutation in the HFE gene was associated with HH (46). It is now known that HH is an autosomal recessive disorder and 1 in 8 people in the United States have a mutation in a single copy of the gene (128). Further study of patients with HH has led to the identification of several other iron regulatory genes that cause HH. These genes demonstrate that iron sensing and regulation of hepcidin is a concerted effort of several proteins. All HH disorders demonstrate a dysregulation in the hepcidin-FPN homeostasis, and are classified into 5 types.

Type 1

High FE (HFE) encodes an atypical major histocompatibility complex protein, and mutations in this gene are the most common cause of HH (46). The most common mutation observed is a missense mutation of cysteine 282 to tyrosine (Cys282Tyr) (152). However, several other mutations are characterized leading to iron overload (152). HFE mutations that lead to iron overload are associated with a significant decrease in hepcidin expression. Consistent with these data, mouse models which are deleted for HFE or have a knock-in Cys282Tyr mutation also have iron overload and a decrease in hepcidin expression (102). Since HFE is abundant in several tissues including enterocytes and liver, a conditional disruption of HFE in the liver and intestine was generated. In this study, hepatocyte-specific disruption of HFE recapitulated a similar phenotype as the whole body knockout mouse model, characterized by iron overload and decrease in hepcidin expression (180). Mice with HFE disruption in the intestine were similar to normal controls (181). These data demonstrate that HFE in the hepatocytes is critical for iron homeostasis. The molecular function of HFE and its precise role in regulating hepcidin expression has been a subject of great interest. Several lines of evidence suggest that HFE binding to Tfr1 and Tfr2 may be the mechanism by which HFE regulates hepcidin expression (60, 153). Mutations in HFE that increased binding to Trf1 blocked hepcidin expression. Mutations that weakened HFE and Tfr1 interaction increased hepcidin expression (153). In addition, HFE and Tfr2 interact and disruption of Tfr2 leads to decreased hepcidin expression (50, 86, 116). Lastly HFE and Tfr2 interaction is required for regulation of hepcidin by iron containing Tf (60). Together the data suggest a mechanism where Tf binding to Tfr1 releases HFE, which then can bind to Tfr2 and stabilize its protein expression leading to an increase in SMAD signaling ().

Type 2A

Juvenile hemochromatosis (JH) is a rare autosomal recessive disorder of iron overload and symptoms become apparent before the age of 30. JH leads to organ damage, and usually causes cardiomyopathy, hypogonadism, liver injury, and diabetes. JH is a caused by mutations in the gene for HFE2 which encodes the hemojuvelin (HJV) protein (132, 152). HJV is a glycophosphatidylinositol anchored membrane protein. Several HFE2 mutations have been found in patients. However, the glycine 320 to valine is the most frequent mutation that is reported (152). To confirm that HJV is causative in this type of hemochromatosis, an HJV knockout mouse model was generated (125). This mouse model demonstrates severe iron overload associated with very low levels of hepcidin expression, similar to that observed in patients with HJV mutations (132). Hepcidin expression was appropriately increased in response to inflammatory stimuli, suggesting that HJV is involved in iron sensing but does not play a role in hepcidin regulation during inflammation. The early onset of iron overload in JH is due to a robust repression of hepcidin. In HH due to HFE mutations there is only a moderate decrease in hepcidin expression leading to iron overload that is symptomatic at later ages. HJV is expressed in several tissues, and in the liver HJV primarily expressed in hepatocytes. Restoring HJV expression in hepatocytes of HJV knockout mice completely restored hepcidin expression and ablated the iron overload (188). Further mechanistic studies demonstrated that HJV functions as a BMP co-receptor and is important for induction of hepcidin expression in response to BMP signaling (10). HJV binds to BMPs and enhances the activity of the SMAD signaling cascade (10).

Type 2B

Similar to HJV mutations, mutations in the HAMP gene, which encodes for hepcidin, are a very rare cause of JH. Currently 12 known mutations occur on the HAMP gene leading to a decrease in the normal production of hepcidin (74). Since hepcidin function or expression is dramatically diminished, the iron overload symptoms are observed before the age of 30.

Type 3

Tfr2 mutations lead to an autosomal recessive iron overload disease similar to HFE related-HH phenotype. Tfr2 as mentioned above is capable of binding to HFE and this interaction is critical in maintaining hepcidin expression (60). Unlike Tfr1, which is ubiquitously expressed, Tfr2 is expressed only in hepatocytes and erythroid precursors (87, 159). Tfr2 cannot compensate for the loss of Tfr1 (171). The knockout mouse model and the liver-specific disruption of Tfr2 confirm its importance in regulating hepcidin levels (86, 182). Hepcidin levels are decreased significantly in these mouse models compared to littermate controls, and tissue iron is increased. The most common mutation observed is in amino acid 245, which is converted into a stop codon resulting in a protein product that is not expressed (24). As mentioned above, Tfr2 binding to HFE promotes SMAD activation and hepcidin expression. Upon its deletion this signaling pathway is decreased causing a significant drop in hepcidin levels.

Type 4

SLC40a1 is the gene that encodes for the iron exporter FPN. FPN is the target of hepcidin, which causes rapid internalization and degradation of FPN (117). More precise work on the mechanism of FPN degradation by hepcidin demonstrates that following hepcidin binding, FPN is phosphorylated on tyrosine residues, which lead to its endocytic shuttling and degradation by the proteasome pathway (39). JAK2 is the critical kinase phosphorylating FPN (37). However, recent data demonstrate that both phosphorylation of FPN and JAK2 are not essential for FPN degradation (141, 148). Mutations in HFE, HJV, hepcidin, and Tfr2 are all recessive mutations. However, mutations in FPN are dominant. Patients that are heterozygous for the mutation develop the disease. This is due to FPN functioning as a dimer and the mutant protein can act as a dominant negative (40, 41). Several mutations of FPN have been observed. Detailed molecular studies demonstrate that the mutations inhibit proper membrane localization of FPN, inhibit the export function of FPN, disrupt hepcidin binding, or inhibit FPN internalization (84). Therefore, depending on the mutation in FPN the patients can present with very different phenotypes. Mutations that inhibit membrane localization or export function can lead to macrophage iron overload. While those mutations that inhibit hepcidin binding or hepcidin-meditated internalization lead to continuous export of iron into serum and eventually iron overload in the hepatocytes.

Secondary hemochromatosis

Secondary hemochromatosis is the result of another disease, which causes excess liver iron loading. Most of diseases that lead to secondary hemochromatosis are acquired disorders of erythropoiesis (63). The most common causes of secondary hemochromatosis are listed in . A well-studied disorder that leads to secondary hemochromatosis is β-thalassemia. β-Thalassemia is a congenital blood disorder due to mutations in the β-globin gene leading to a partial or complete loss of β-globin synthesis resulting in β-thalassemia intermedia and Cooley’s anemia, respectively. The decrease in β-globin results in ineffective erythropoiesis and erythropoietic stress. Persons with β-thalassemia intermedia have mild anemia with a slight lowering of hemoglobin levels in the blood. In most cases treatment is not necessary, but severe patients with low hemoglobin levels will need occasional blood transfusions (161). Cooley’s anemia results in a striking deficiency in hemoglobin production. Patients will need frequent blood transfusions (161). The blood transfusions lead to dysregulation of the systemic iron homeostasis since donor blood is a rich source of iron. The body cannot eliminate the excess iron efficiently, leading to increased tissue iron. Regular blood transfusions are the most common cause of secondary hemochromatosis (63). Initially it was thought that the iron overload was primarily due to regular blood transfusions. However, mouse models of β-thalassemia hyperabsorb iron. This is the major mechanism leading to iron overload in β-thalassemia intermedia and significantly contributes to the tissue iron overload in Cooley’s anemia (76, 164, 186). It is less clear whether an increase in iron absorption plays a significant role compared to blood transfusions in other disorders of erythropoiesis listed in . However, recent work has shown that effective and ineffective erythropoiesis can stimulate iron absorption, therefore this mechanism of iron overload may be true for other diseases leading to secondary hemochromatosis (5). Increased iron absorption in secondary hemochromatosis may be due to an increase intestinal hypoxia signaling and a decrease in hepcidin expression (5, 133). Increasing hepcidin levels in mouse models of β-thalassemia improved liver iron loading and anemia (62).

Table 1

Common causes of secondary hemochromatosis

Aceruloplasminemia, hypotransferrinemia, and DMT-1 deficiency

Aceruloplasminemia, hypotransferrinemia, and DMT1 deficiency are causes of secondary hemochromatosis, but are not disorders of erythropoiesis; rather these disorders are due to ineffective transport of iron. Aceruloplasminemia is due to a loss-of-function mutation in Cp and is inherited in an autosomal recessive manner. Iron overload in aceruloplasminemia is mainly observed in the brain and liver (73, 187). A similar phenotype is also observed in Cp knockout mouse models. Hypotransferrinemia is an autosomal recessive disorder leading to loss of Tf production. Hypotransferrinemia is associated with severe microcytic anemia, and an adaptive increase in iron absorption, which leads to severe liver iron loading (64, 75). Consistent with hypotransferrinemic patients, the hpx mouse, which produces no Tf has liver iron overload and anemia (171). DMT1 deficiency is an autosomal recessive disorder leading to increase in liver iron (80). This is thought to be due to the role of DMT1 in iron export from the endocytic compartment.

Dysmetabolic iron overload syndrome (DIOS)

DIOS is a newly characterized secondary hemochromatosis disorder. DIOS is associated with features, such as obesity, type 2 diabetes, alcohol use, and chronic hepatitis C (42, 81, 106, 139). This is now the most common cause of iron overload observed in patients. The iron overload is observed in 15% of patients with metabolic syndrome, 50% in patients with non-alcoholic fatty liver disease, over 40% of patients with chronic hepatitis C infection, and significant number of patients with alcoholic liver disease (18, 81, 139, 174). Currently the mechanisms, which contribute to DIOS are unclear. However, a significant decrease in FPN gene expression has been noted in patients with DIOS (2).

IRON-INDUCED LIVER DAMAGE

High levels of iron deposition lead to tissue damage and dysregulation of function. In the liver increased free iron if untreated, leads to fibrosis and cirrhosis, and can increase morbidity and mortality (123, 124, 145). Hepatic tissue injury is directly correlated to the duration and amount of iron loading (123, 124). Cells normally produce basal levels of reactive oxygen species (ROS) through metabolic function of the mitochondria and other organelles. ROS are kept at low basal levels by several antioxidant enzymes, and low levels of ROS are important in normal cell physiology (146). ROS in conjunction with high cellular iron results in a robust increase of hydroxyl radicals, which leads to cell damage. Free iron generates ROS through the Fenton and Haber-Weiss reactions (). The superoxide radical (O₂^•−) reduces ferric iron to ferrous iron, which reacts with hydrogen peroxide (H₂O₂) to generate highly reactive hydroxyl radicals (OH^•) (90). Hydroxyl radicals lead to an increase in peroxidation of phospholipids within organelle and cellular membranes, oxidation of amino acid side-chains, DNA strand breaks, and protein fragmentation. The exact mechanisms by which high intracellular iron leads to liver fibrosis and cirrhosis are unclear, but iron-induced cellular damage has been shown to directly increase hepatocyte cell death and activate Kupffer and stellate cells (135, 160) (). More recently, clinical evidence suggests that high liver iron could play a role in insulin resistance (45, 143). The increase in iron-induced ROS in Kupffer cells can initiate a pro-inflammatory cascade in the liver. Increased ROS production activates NF-κB signaling leading to an increase in IL-6, TNF-α, and IL-1β in the liver (22, 112). Liver inflammation can lead to hepatic insulin resistance, which is a major pathway leading to hyperglycemia in type II diabetes (55). Lastly, liver iron overload increases the risk for hepatocellular carcinoma (HCC). HCC is the major life threatening complication associated with hereditary hemochromatosis (124). Several studies have looked at the risk factor for HCC in hereditary hemochromatosis patients and some have estimated the risk to be 100-200 fold higher in HH patients (19, 162). In addition, other iron overload disorders such as thalassemia are associated with an increased risk for HCC (17). The increase in iron plays an active role in HCC pathogenesis. Chelation of iron or placing mice on an iron deficient diet decreases tumor growth (71, 151).

Schematic diagram of the Fenton and Haber-Weiss reactions

Iron is a potent catalytic cofactor, which increases highly unstable oxygen radicals that cause cellular damage.

Iron-induced liver damage

Iron accumulation in hepatocytes and Kupffer cells leads to an increase in ROS production and pro-inflammatory mediators. Both ROS and pro-inflammatory mediators initiate a feed forward cycle, which activates stellate cells, initiates cell damage, and leads to loss of function contributing to an increase in steatosis, fibrosis, cirrhosis, and HCC.

TREATMENT OF IRON OVERLOAD

In patients with HH, phlebotomy is used to decrease liver iron. Regular bleeding of the patients leads to an increase in erythropoiesis, which mobilizes liver iron stores to meet the demand for iron that is required to generate mature RBCs (59). Secondary hemochromatosis is also associated with severe anemia, thus phlebotomy is not an option. In these patients iron chelators are used to decrease liver iron (59). Deferoxamine has been used for over three decades for iron chelation. More recently two new iron chelators have also been used, deferiprone and deferasirox (93). Iron chelators have been shown to be effective in decreasing liver iron and also morbidity and mortality associated with iron overload. Several rounds of phlebotomy and/or administration of iron chelators are required; these are slow-acting treatment options and may not successfully decrease the liver injury associated with an increase in iron. Alternatives to existing treatments are needed. Recently, hepcidin has been shown to be a very attractive target and good proof of principles studies have been done. Several studies in mouse models of HH have shown that increasing hepcidin levels can ameliorate the iron overload (122, 177). Recently in β-thalassemia models, increasing hepcidin expression resulted in decreased liver iron (62). Currently several strategies are being assessed to increase hepcidin levels. The specific sites that are required for hepcidin-FPN binding are known. Precise structural mutagenesis studies have demonstrated that nine amino acids of hepcidin are critical for the binding to FPN and initiating its internalization (140). Modifications of these nine amino acids has led to several peptides that have increased activity over full length hepcidin. Moreover, the modified hepcidin derived peptides are functional in vivo and can prevent iron overload that is observed in hepcidin knockout mice (140). These agents are stable orally and may provide a well-tolerated form of treatment for hereditary and secondary hemochromatosis. Other approaches may also be useful in increasing hepcidin expression (59). Treatment with BMP6 increases hepcidin expression in HFE-null mouse model and prevents iron overload (34). Moreover, since hepcidin regulatory pathways are well characterized, several possibilities such as SMAD, C/EBPα, and STAT3 activators could have potential roles in increasing hepcidin expression in vivo.

ANEMIA

On the other end of the spectrum, aberrant upregulation of hepcidin is critical in the pathogenesis of anemia of chronic disease, which encompasses several diseases including kidney disease, inflammatory disease, cancer, and aging (26). In healthy human volunteers and mice, studies demonstrate that inflammatory agents cause a robust and rapid decrease in serum iron levels (27, 88, 115, 147). Within hours following induction of inflammation, hepcidin levels are significantly elevated. The decrease in serum iron is due to hepcidin-mediated internalization of FPN leading to iron sequestration in macrophages. This is thought to be a protective mechanism that limits iron available to infectious pathogens. However, in chronic disorders this leads to anemia, which can have detrimental effects in the primary disease pathogenesis. Similarly, in most cancers there is a decrease in serum iron levels and increased anemia (156). The decrease in iron is suggested to be beneficial in limiting tumor growth. The mechanism may vary depending on the primary disease. However, the IL-6-STAT3 pathway is critical in increasing hepcidin expression during inflammation (163). The best treatment for anemia of chronic disease is resolving the primary chronic disease. In severe cases, blood transfusion, EPO, or intravenous administration of iron is used (163). In addition to an increase in hepcidin expression from chronic disorders, rare genetic mutations in the TMPRSS6 gene cause an increase in hepcidin expression and iron-refractory iron deficiency anemia (IRIDA) (47). This was further confirmed in the TMPRSS6 knockout mice, which had hair loss and microcytic anemia associated with high levels of hepcidin expression (48, 52). A similar finding was noted in the mask mutant mouse strain, characterized by a premature stop codon in the TMPRSS6 gene (44). IRIDA is iron deficiency anemia that is unresponsive to oral iron therapy. TMPRSS6 (also known as matriptase-2) encodes a type II transmembrane serine protease, which is expressed predominantly in the liver. The first substrate that was characterized for TMPRSS6 was HJV. The serine protease of TMPRSS6 cleaves membrane HJV, leading to downregulation of hepcidin expression (155). Mutations in TMPRSS6 decrease its protease activity leading to increased protein expression of HJV and a coordinate increase in hepcidin expression.

CONCLUSION

The liver is the central tissue which regulates systemic iron homeostasis by acting as a sensor and regulator of iron levels. In addition, through its role in iron storage, the liver can protect more sensitive tissues from iron-induced cellular injury. The past decade has led to several novel liver-derived players regulating systemic iron homeostasis and identification of new mutations in iron-related disorders. Several pathways in the liver regulate hepcidin, the master hormone for maintaining systemic iron homeostasis. Recent studies have shown that these pathways can have redundancy or act independently based on the stimuli (172). The challenge will be to understand how these pathways crosstalk and are regulated in a coordinate manner to maintain hepcidin levels. Moreover, how can these pathways be targeted in iron-related disorders that could be of therapeutic benefit? Targeted therapies for iron related disorders are actively being pursued such as the case for hepcidin mimetics and BMP agonists, and the coming decade should yield novel therapies.

ACKNOWLEDGEMENTS

This work was supported by grants to Y.M.S from the National Institutes of Health (CA148828 and DK095201), The University of Michigan Gastrointestinal Peptide Center, and Jeffrey A. Colby Colon Cancer Research and the Tom Liu Memorial Funds of the University of Michigan Comprehensive Cancer Center. E.R.A is supported by the Rackham Predoctoral Fellowship, University of Michigan.

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Iron With Chicken Livers Vs. Beef Liver

Chicken and beef liver contain the more easily absorbed heme iron.

Image Credit: Ivan MajtA!n/iStock/Getty Images

Iron deficiency is considered the most common nutritional deficiency in the world, according to the World Health Organization. Iron is an essential mineral found in all cells of the body and integral to a variety of processes. Iron deficiency is a condition in which too little iron is in the body, resulting in weakness and fatigue, among other symptoms. Chicken and beef liver contain high amounts of iron. They’re good sources to meet daily iron intake recommendations.

Iron Content

Chicken liver is one of the richest sources of iron; beef liver provides a good amount of iron. A 100g slice of pan-fried chicken liver contains 13mg iron, or approximately 72 percent daily value. A 100g slice of simmered chicken liver contains 11.6mg or 65 percent daily value, based upon a 2,000-calorie diet, according to “Calorie Lab.” Raw chicken liver contains 9mg iron or 50 percent daily value per 100g. Raw beef, on the other hand, contains 4.9mg or 27 percent daily value of iron. A 68g slice of braised beef provides 4.4mg or 25 percent daily value of iron; an 81g slice of pan-fried beef provides 5mg or 28 percent daily value of iron, according to “Calorie Lab.”

Iron

Iron is a dietary mineral that plays a key role in a variety of bodily processes. A major function of iron is the role it plays in supporting the transport of oxygen in the body; nearly two-thirds of iron in the body is found in hemoglobin. Hemoglobin is a protein found in red blood cells that transports oxygen throughout the body. Iron is also required for certain chemical reactions in the body and the production of amino acids, neurotransmitters and hormones, according to Harvard Health Publications. The two forms of dietary iron include heme, which comes from animal sources, and nonheme, which comes from plant sources. Heme iron, the form found in chicken and beef liver, is more easily absorbed by the body as compared to nonheme iron.

Dietary Reference Intakes

The Food and Nutrition Board of the Institute of Medicine recommends 8mg iron per day for adult males and 18mg per day for females ages 19 to 50. Pregnant women need the highest amount of iron per day, with a recommendation of 27mg per day; adolescent women between ages 14 and 18 need 15mg per day. Lactating women need approximately 9mg to 10mg per day. Children ages 1 to 3 need 7mg per day, and those 4 to 8 need 10mg per day. The tolerable upper limit intake levels for iron are 40mg to 45mg iron per day. Toxicity can occur with high iron intake because it’s not easily eliminated from the body. Keep iron supplements locked and away from children to prevent accidental iron toxicity, which can be fatal.

Deficiency

Iron deficiency, a common health problem around the world, most often leads to iron deficiency anemia or low levels of iron in the blood. It can be caused by low dietary intake, blood loss and iron absorption problems and cause symptoms such as fatigue, decreased performance and a weakened immune system. Pregnant and menstruating women as well as children are at the highest risk of developing iron deficiency, according to the Office of Dietary Supplements. Iron supplements are often prescribed for the treatment of iron deficiency.

90,000 10 iron-rich foods (list) :: Health :: RBK Style

Iron is an important trace element necessary for all living organisms. It helps to synthesize collagen and serotonin, supports the immune system and participates in metabolic processes [1]. But the main function of iron is cellular respiration. This trace element is part of hemoglobin – the protein that makes up red blood cells.It is iron that helps blood cells bind oxygen and deliver it to tissues, and then remove waste carbon dioxide from the body. By the way, it also stains the blood red.

Our body is not able to produce iron on its own. He gets it from food, so it is important that the food is varied. There are two types of iron: heme and non-heme. The first is assimilated more efficiently [2]. It can be found in meat, fish, and seafood. The source of the second is plant food.Here is a list of the foods with the highest iron content of both types. Including them in the diet will help replenish micronutrient reserves.

Daily iron intake

Women 19-50 years of age need iron most of all. They need to receive at least 18 mg of a trace element per day. During pregnancy, the need for it increases to 27 mg. Adolescents 14-18 years old also require an increased iron content: girls – 15 mg, boys – 11 mg.The average daily intake of iron for adult men and older adults of both sexes is 8 mg [3]. It increases significantly with intense exercise, regular heavy physical exertion and heavy menstruation.

Foods high in iron

Shellfish

Almost all types of shellfish are rich in iron. Thus, one hundred-gram serving of oysters contains about 3 mg of iron, which is 17% of the daily requirement [4]. In addition, this amount also provides 24% of the daily value for vitamin C and 4% of the daily value for vitamin B12.Shellfish are also low in calories, high in protein, and raise the level of “good” cholesterol, which prevents heart disease.

Offal

Liver, kidneys, brain, heart, stomachs and other by-products contain large amounts of iron. Although not everyone likes their taste, by-products are often superior to meat in terms of nutrient content. For example, to get 36% of the daily value of iron and meet the daily need for vitamin A, it is enough to eat only 100 g of beef liver [5].Plus, offal is a good source of protein, copper, selenium and choline, which is important for the liver.

Red meat

This is the main source of easily digestible heme iron. Moreover, the darker the meat, the more of this microelement it contains. One 100 g of ground beef patty, steamed, contains 2.7 mg of iron. This meets the daily requirement by 15% [6]. Meat also serves as a source of protein, zinc, selenium and B vitamins. But poultry is not so rich in iron: its content in 100 g of turkey does not exceed 0.7 mg [7].

© Andrijana Bozic / Unsplash

Spinach

Such a rich set of nutrients as in spinach is rare. It contains folate, lutein, beta-carotene, calcium, vitamins A and E. In addition, 100 g of the product replenish 15% of the daily value of iron. It is non-heme, but at the same time it is quite well absorbed due to the high concentration of vitamin C in spinach.Doctors advise boiling the leaves a little – this will help reduce the amount of oxalic acid, which interferes with iron absorption [8].

But keep in mind: 100g fresh spinach is a big bag. It is designed for several people, and it is hardly possible to eat it at a time. In addition, spinach tends to accumulate nitrates, which are often used in its cultivation. Buy the product in proven farm shops or in special organic packaging. Or try growing it yourself – on a windowsill.In winter, instead of fresh spinach, you can take frozen spinach: all its beneficial properties and taste are preserved.

Legumes

This is a must-have for vegetarians and vegans. Legumes are one of the best plant sources of iron. Chickpeas, peas, lentils, beans, soy – choose what you love. One cup of cooked lentils contains 6.6 mg of iron. This is 37% of the daily value [9]. And half a glass of cooked beans is enough to fill 10% of the daily requirement for an element [10]. In addition, legumes can help you feel fuller for a long time and reduce your calorie intake [11].

Pumpkin seeds

Pumpkin seeds can be a snack option. 100 g of the product contains 9 mg of iron, or half the daily recommended value [12]. But you can’t get carried away with them. First, it can cause gastrointestinal problems. Secondly, pumpkin seeds are very nutritious. A 100-gram serving provides the body with 559 kcal. To increase iron levels without harming your health, add a small handful of seeds to salad, porridge, or soup.

Quinoa

South American cereals are often used as a substitute for gluten-containing cereals.Add 100 g of boiled seeds to your favorite salad to replenish 8% of your daily iron requirement [13]. Unlike traditional grains, quinoa contains a lot of protein, which contains essential amino acids [14]. Interestingly, our body perceives quinoa as a protein from cow’s milk.

Broccoli

A diet rich in broccoli helps improve vision, reduces inflammation and slows down aging. Broccoli cleanses the body, removes cholesterol and excess sugar.Use it as a side dish — a glass of cooked broccoli provides 6% of your daily iron requirement [15]. For maximum benefits, steam broccoli for no longer than 5 minutes. This will help preserve vitamin C.

Tofu

The production of tofu is similar to the process of making cheese from milk – which is why many call it soy cheese. In terms of its nutritional properties, it is almost equal to dairy products – for this it is loved by vegans and people with lactose intolerance. 100 g of tofu cheese contains 17 g of protein, which is easily and quickly absorbed by the body.In addition, the same amount of the product helps to cover 15% of the daily value of iron [16].

Dark chocolate

Chocolate not only brings pleasure and stimulates the production of the “happiness hormone”, but also allows you to normalize the level of iron. Choose chocolate that contains at least 70% cocoa [17]. Nutritionists advise eating no more than a quarter of a bar of chocolate a day. This will be enough to offset 17% of your daily iron requirement, improve your gut microflora, and lift your mood.

© Dovile Ramoskaite / Unsplash

Why iron deficiency is dangerous

Iron deficiency is usually asymptomatic at first. But if you do not replenish its reserves in time, you can provoke the development of iron deficiency anemia [18]. Its main symptoms are weakness, fatigue, shortness of breath, pallor, drowsiness, loss of appetite, heart palpitations and headaches [19].You may be tempted to eat something inedible – chalk, clay, paper or ice. With a lack of iron, the cells begin to “suffocate”, which is why many vital metabolic processes are disrupted in the body.

Iron deficiency also contributes to decreased immunity and a high risk of infections [20]. It is also one of the causes of hair loss. The trace element is responsible for delivering oxygen to the follicles, thereby strengthening and nourishing the roots. With its deficiency, hair becomes dry and weak and may begin to fall out [21].Other external signs include sores in the corners of the mouth, dry skin, brittle peeling nails. According to a study by Japanese scientists, in some cases, iron deficiency causes depression [22].

If you notice signs of iron deficiency, seek medical attention. He will order blood tests, determine the source of the problem, and be able to draw up a treatment plan based on your individual characteristics.

Expert commentary

Evgeniya Maevskaya, MD, PhD, gastroenterologist and nutritionist, GMS Clinic

How often should a blood test be taken in order to find out about the lack of iron in time?

The frequency depends on many factors: general health status, clinical signs of an overt or latent deficiency, getting into the risk group for iron deficiency or the presence of chronic diseases, including the gastrointestinal tract.

For a potentially healthy person, it is enough to monitor blood counts every six months. At the same time, a general analysis is not enough. At a minimum, it should be supplemented with a test for serum iron and ferritin, otherwise the signs of latent deficiency can be missed. In some cases, a more rare test is needed – for soluble transferrin receptors. This is determined only by the doctor.

Is it possible to fill the iron deficiency only with plant foods? What are the best tips for vegetarians and vegans?

Treatment of anemia only with iron in food is impossible due to its low content and low bioavailability.Anemia can only be treated with iron supplements.

Vegetarians and vegans should eat as varied as possible, including plant-based sources of iron, such as seaweed, in the diet. If applicable, shrimp, mussels and sea fish can serve as a good source of iron. For vegetarians, it is better to get tested and make sure that there is no atrophy in the stomach and problems in the intestines. With atrophy and insufficient acidity of the stomach, the transition of non-heme iron from plant food to the assimilable heme form is significantly difficult, which means that it will not be absorbed.

What if a person notices symptoms of iron deficiency?

At the first symptoms of iron deficiency, consult a doctor. It is important not only to correct the deficit, but also, most importantly, to identify its cause. It is impossible to do this on your own.

What is the danger of excess iron in the body?

The so-called iron overload is certainly dangerous. It can lead to damage to internal organs, fibrosis in organs and tissues. There is also evidence of direct damage to the genetic apparatus of cells.Most often, the liver, pancreas and myocardium are affected – this manifests itself in the form of toxic cardiomyopathy and arrhythmias. This situation is more likely with parenteral or enteral uncontrolled iron administration. Food cannot be the cause of excess iron in any way.

90,000 WHERE it is contained and HOW MUCH is needed per day

Iron is the most important element of hematopoiesis. In the form of an iron-containing protein (hemoglobin), it is contained in erythrocytes and respiratory enzymes involved in tissue respiration.It is used to treat anemias of various etiologies.

Content:

What foods are rich in iron

Sources of iron – all kinds of plant and animal food. When choosing, it is advisable to focus on the norm. An excess of a trace element is as dangerous as a deficiency. 12 – 15 mm with an average weight of 70 kg – this is the daily rate for adults.

The best foods containing large amounts of iron are meat products. By-products are the leaders. 100 g of pork, chicken, beef liver more than covers the body’s daily requirement for a mineral.

There is a lot of iron in rabbit meat, beef brains, pork heart, beef tongue (from 4 to 6 mg).

Dried mushrooms are the leading plant products. 100 g contains a double daily norm of a trace element – 35 mg. They are followed by seaweed – 16 mg, prunes – 13 mg. There is a lot of it in dried apricots, beans, cocoa powder, lentils, rose hips, dried apples – from 11 to 12 mg.

The use of a microelement by the body, its absorption through the intestinal wall largely depends on the composition of the food consumed at one time.

Ascorbic acid promotes absorption. Reduce the digestibility of calcium carbonate, phosphates, magnesia, baking soda.

This fact should be taken into account when drawing up the menu. Combining meat food with vegetables, rosehip broth, juices and other dishes rich in vitamin “C”, you can replenish the deficient element.

Table of iron content in 100 g of product

In the table, foods are listed in descending order of micronutrient.

Iron in products for pregnant women and children

The amount of iron in products for pregnant women is 30 mg per day. For a six-month-old child, the same indicator is 12 – 16 mg. Insufficient intake of a trace element with food is dangerous to the health of the fetus and mother.

The main supplier of iron is lean meat and offal – liver, beef tongue, heart.

If there are no contraindications, the menu includes nuts, eggs, blueberries, lingonberries, cranberries, seaweed salads.From drinks – rosehip infusion, apple, peach, apricot juices. Compotes with dried fruits are useful – prunes, dried apricots, apples, pears. Oatmeal, buckwheat porridge, dishes with lentils, peas are required. For children, products are selected according to age and on the recommendation of a doctor.

Foods with high iron for anemia

There are several types of anemias, all of them are associated with a decrease in the number of red blood cells in the blood and a decrease in hemoglobin. Since animal food is better absorbed, with anemia, preference is given to it.

A large amount of a trace element is contained in:

• pork, chicken, veal liver;
• beef tongue;
• beef heart;
• beef brains;
• oysters;
• egg yolk.

Along with meat, plant foods that increase hemoglobin are an indispensable component of the daily menu. Blood is normalized with the use of buckwheat and oatmeal, prunes, dried apricots, dishes from lentils, peas, seaweed.

Hemoglobin will be normal if nutrition is properly organized and the body is provided with nutrients, vitamins, macro and microelements on a daily basis. Food that contains enough iron is an excellent prevention of new and old (in the acute stage) iron deficiency pathologies.

90,000 Eight foods with the highest iron content were named

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Eight foods with the highest iron content were named

Eight foods with the highest iron content were named – RIA Sports News, 01.10.2021

Eight foods with the highest iron content were named

Doctor and weight loss specialist at the clinic Pavel Isanbaev, in a conversation with RIA Novosti, named leading products in iron content, and explained why this … RIA Novosti Sport, 01.10.2021

2021-08-24T03: 35

2021-08-24T03: 35

2021-10-01T18: 30

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MOSCOW, 24 Aug – RIA Novosti. Doctor and weight loss specialist at the clinic Pavel Isanbayev, in a conversation with RIA Novosti, named the leading products in iron content, and explained why this mineral may not be absorbed. “Most iron is in animal products: meat, eggs, offal, especially liver and In meat, iron is in the most bioavailable form – this is the so-called heme iron, which is easier for the body to assimilate, “says Pavel Isanbayev.He stressed that the iron contained in the product will not be fully absorbed. “From plant foods, for example, legumes, spinach, cereals – buckwheat, barley, oatmeal – the body” takes “about 2-12% of the iron content. Thus, it is more difficult for vegans to get the trace element only from food”, – explains the weight loss specialist . According to Isanbayev, some carbohydrate foods block the absorption of iron: first of all, it is fiber, especially if it is eaten in large quantities; Foods rich in polyphenols – such as nuts and coffee – and phytates, an anti-nutrient found in legumes and cereals.Thus, if you want to eat meat with buckwheat, it is better to soak buckwheat before cooking: you will simplify the process of iron absorption. Therefore, its reserves are regularly depleted. And if a person has an iron deficiency with adequate nutrition (or even medication), then it makes sense to consult a doctor and comprehensively check the body “, – recommends Pavel Isanbayev.

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03:35 24.08.2021 (updated: 18:30 01.10.2021)

Named eight products with the highest iron content

MOSCOW, Aug 24 – RIA Novosti. Doctor and weight loss specialist at the clinic Pavel Isanbaev, in a conversation with RIA Novosti, named the leading products in iron content, and explained why this mineral may not be absorbed.

“Most iron is in animal products: meat, eggs, offal, especially liver and blood sausages. In meat, iron is in the most bioavailable form – the so-called heme iron, which is easier for the body to assimilate,” says Pavel Isanbayev.

August 23, 04:15 AM The doctor told who should not eat rice

He stressed that the iron contained in the product will not be completely absorbed.”From plant foods, for example, legumes, spinach, cereals – buckwheat, barley, oatmeal – the body” takes “about 2-12% of the iron content. Thus, it is more difficult for vegans to get the trace element only from food”, – explains the weight loss specialist …

According to Isanbayev, some carbohydrate products block the absorption of iron: first of all, it is fiber, especially if it is eaten in large quantities; Foods rich in polyphenols – such as nuts and coffee – and phytates, an anti-nutrient found in legumes and cereals.Thus, if you want to eat meat with buckwheat, it is better to soak the buckwheat before cooking: you will simplify the process of iron absorption.

October 1, 18:27

Buckwheat with kefir: doctors talked about the pros and cons of the popular diet

“The health of the digestive tract and the state of the body as a whole are equally important: any foci of inflammation in the body, from caries to neoplasms, increase iron consumption Therefore, its reserves are regularly depleted. And if a person has an iron deficiency with adequate nutrition (or even medication), then it makes sense to consult a doctor and comprehensively check the body “, – recommends Pavel Isanbayev.

Iron in foods – what is there to increase hemoglobin

Iron, which is necessary for a growing child’s body, is found mainly in meat, but not only. It is real to choose and form the child’s diet correctly, knowing what nutrients are contained in the products.

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Lack of iron in a child’s body, as well as in adults, causes a serious and intractable condition called iron deficiency anemia.This is due to the fact that a sufficient amount of hemoglobin is not formed in the blood – a special iron-containing protein that transports oxygen to all organs, transferring it with the bloodstream. Iron is best absorbed from animal products, but it is also abundant in plant foods.

Pork and beef liver

This product is considered the main and most accessible source of iron. 100 g of animal liver contains up to 20 mg or more. mineral, and it is heme iron, which is absorbed most efficiently – by 20-35%.Liver of poultry (turkey and chicken) is also good, it contains about 15 mg of iron per 100 g.

Red meat

Beef, veal and pork are also excellent sources of heme iron. Moreover, the redder the meat, the more iron it contains – beef is more useful than pork and lamb.

Source: Fotolia

Fish and seafood

Suitable shellfish, oysters, mussels, sardines, shrimps, tuna, red and black caviar.

9014 9014 black caviar

Legumes

Excellent sources of non-heme iron (which is less digestible) than heme), all possible legumes – beans, lentils, chickpeas, peanuts, soybeans.

Seeds

Seeds that can add iron to the body include seeds (both sunflower and pumpkin seeds), sesame seeds. And, accordingly, sunflower, pumpkin and sesame oil. And also hummus, which includes chickpeas and sesame paste.

Fruit

Of the fruits, persimmons, black currants, dogwoods, apples and especially fresh rose hips are the richest in iron.

Source: Fotolia

Vegetables

Almost all vegetables contain iron in varying amounts, but it is important to remember that this is only an additional source of the mineral that cannot replace liver and meat.Most of all iron is found in cauliflower and Brussels sprouts, celery, dill, beets, and garlic. Sometimes it is recommended to drink freshly squeezed juice from these products to increase hemoglobin.

Dried fruits

Dried fruits will also support the production of hemoglobin – rose hips, dried apricots, raisins, dates, prunes.

Nuts

Among nuts, the most iron is found in pine nuts, almonds and pistachios.

9014 9014 walnut

Source: Shutterstock

Cereals

Wheat bran is the best source of iron among cereals.Eat more bread and other bran foods. In second place is our wonderful buckwheat. And there is also a small amount of iron in oatmeal and corn.

9014

9014 corn

What helps and hinders the absorption of iron

In addition to the correct selection of products – sources of iron, it is important to correctly combine them with others.Knowing what will help you absorb iron as much as possible, and what prevents its absorption, you definitely won’t go wrong. So for the assimilation of iron, ascorbic acid, vitamin B12 and vitamin A are needed. Interfere with the absorption of iron – excess of calcium, vitamin E, zinc, tannin (in tea, coffee, quince, blueberries), oxalic acid , phosphates (in dairy products and eggs).

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Top 7 foods with a high iron content

It is necessary to monitor your diet all year round. The diet of a healthy person should include a full range of vitamins and minerals in order to function without problems.Therefore, do not forget about iron-containing products. Next – why they are important.

What does our body need iron for? It participates in many processes, the most important of which is the transfer of oxygen to tissues. And this mineral is also involved in the formation of DNA. According to the World Health Organization, more than 20% of women of childbearing age suffer from anemia – iron deficiency. The fact is that the body itself does not know how to produce this mineral and gets it from food.Therefore, it is important to monitor your diet and include foods high in iron.

1. Seafood

They are all high in iron, especially shrimp, oysters and mussels. Thus, a 100-gram portion of seafood can contain 17% of the daily requirement of the mineral. It is important to remember that the iron in shellfish (heme iron) is more easily absorbed by the body than what we get from vegetables. Seafood also increases the level of “good cholesterol”, which is beneficial for the health of the heart and blood vessels.

2. Liver and meat

Beef liver, kidney, heart and meat itself contain the highest amount of iron. 100 grams of the product accounts for 36% of the daily value of the mineral. It is also heme iron, which has been shown to be absorbed five times better than non-heme iron. Do not forget that for the good of the body, all of the above should be cooked as correctly as possible: steamed, cooked or stewed in the oven. Vitamin C helps to absorb iron, so combine meat with vegetables containing it (cabbage, bell peppers, tomatoes, etc.)etc.).

3. Legumes

Beans, lentils, chickpeas, peas and soy should be present in the diet, especially if you do not consume meat. In addition to all the other advantages, the presence of folic acid, magnesium, legumes contain about 18% of the daily value of iron. But there is one subtlety. Beans also contain substances (phytic acid) that prevent iron from being absorbed by the body. Dealing with this is quite simple. If you’re planning on eating legumes, soak them in warm water overnight.

4. Spinach

Spinach is considered to be one of the leaders in iron content among plants. And this is absolutely true. 100 g of greens contains 15% of the daily iron requirement. But the problem is that oxalic acid, which is also found in spinach, is an inhibitor. This means that the benefits of spinach will only appear when properly cooked. Boil, simmer, steam with olive oil is the best option.

5.Tofu

Everything is logical: tofu is made from soy, which belongs to legumes, and we wrote above that they contain a lot of iron. Tofu is a great option for vegetarians. 100 grams of this product will supply about 14% of your daily iron requirement. Tofu is also a good source of thiamine and certain minerals such as calcium, magnesium and selenium. Plus, it provides 22 grams of protein per serving. Tofu contains isoflavones that reduce the risk of heart disease.

6. Currant

Fruits are not the best source of iron, but there are exceptions. For example, black currant. With 100 grams of berries, our body will receive 12% of the daily intake of a useful mineral. Currants also contain vitamin C, as we already know, which promotes the absorption of iron in the body. You should also pay attention to mulberries, persimmons, blueberries, elderberries.

7. Dark chocolate

30 grams of dark chocolate – a source of various nutrients.For example, antioxidants, magnesium, copper and, of course, iron, as well as fibers that improve bowel function. Research by American scientists has shown that chocolate lowers cholesterol levels and improves heart health. But the benefits of this product depend on the content of cocoa beans in it. Only dark chocolate containing 70% or more cocoa beans should be included in the diet.

See also: About the benefits of celery for anemia and not only

See also: Loss of strength: reasons and what to do to recover

See also: Iodine-containing products

Iron is essential for both vegetarians and meat-eaters.What foods contain it?

Foods containing iron should be eaten every day to replenish a vital element for the well-coordinated work of the body.

If there is not enough iron

Iron is one of the most important elements involved in many biochemical processes, in particular, it is necessary for the synthesis of hemoglobin. It contains myoglobin, which is responsible for oxygenation of muscles.Its deficiency leads to decreased performance, increased fatigue, dry skin, brittle nails.

Offal and beef

These products are suitable for those who cannot imagine their menu without a piece of meat. Beef liver, heart, kidneys are not only excellent sources of iron, but also low in calories. For those who are afraid of extra pounds and adhere to a balanced diet, this is a very good option.

There is a sufficient amount of iron in beef.But nutritionists advise including it in the diet no more than twice a week, since it has a high cholesterol content.

Plant sources of iron

If you are on a vegetarian diet, plant sources are essential for better iron absorption. Be sure to include foods containing vitamin C in your diet: bell peppers, citrus fruits, berries.

• Spinach will be equally beneficial for vegetarians and meat eaters, as it contains the optimal amount of iron for the daily requirement.Even stewed spinach retains its properties. Therefore, cook omelets, smoothies, salads with it and stew with vegetables.

• Legumes contain a record amount of light and healthy protein for plant foods, and are also rich in iron. Beans, lentils, chickpeas will provide the body with a large amount of fiber. They also work as a natural sorbent, removing all harmful from the body.

• Don’t skip a piece of chocolate .And even at night. If you are counting calories and keeping fit, then it is better to consume dark chocolate. 30 g of dark chocolate contains half the daily value of iron.

• Nuts and seeds are not only a rich source of iron, they are also high in protein and healthy vegetable fats. It is better to eat them during the day. They give a feeling of fullness for a long time.

Earlier, “Kubanskie Novosti” wrote about products that contain more calcium than a glass of milk.

Hemochromatosis, symptomatology, diagnostic examination, treatment

The reason for the development of this hereditary disease is a mutation in the HFE gene, as a result of which metabolic dysfunction occurs, and the body, receiving a false signal about a lack of iron, begins to accumulate it. The consumption of this elementary substance can be several times higher than the norm (3-4). The body is not able to assimilate such an amount, which leads to intoxication.Iron, absorbed in the gastrointestinal tract, accumulates in the internal organs. As a depository organ, the liver is the first to be hit, as a result of which the performance of its barrier, detoxification functions is disrupted, and iron begins to accumulate in other vital parts of the body: the heart, spleen, pancreas and others.

Symptoms

The clinical picture of the initial stage of the disease is not pronounced. Symptoms of the onset of hemochromatosis can be: general weakness, rapid fatigue, decreased libido, pain in the right hypochondrium, weight loss, and others.When the indicator of the level of iron in the body is close to the critical mark (20-40 grams, depending on the physiological characteristics of the body) or reaches it, symptoms specific to the disease appear. The most obvious symptom of progressive hemochromatosis is a change in the pigmentation of the skin and mucous membranes. Hair loss, nail deformity, arthropathy, liver enlargement may occur. With a severe course of the disease, pathologies such as hyperpigmentation (bronze skin, yellowness of the eye sclera), cirrhosis of the liver, renal, heart failure appear.

Diagnostic examination

The examination begins with the collection of anamnesis not only of the patient, but also of his immediate family, in order to identify or exclude the factor of heredity. The most commonly used laboratory diagnostics, which allows you to determine the level of iron (iron-containing proteins) in the blood serum, the rate of its excretion in the urine. Molecular genetic research is aimed at identifying the hereditary nature of the disease. One of the ways to confirm the diagnosis is a puncture biopsy, which is performed to detect the hemosiderin pigment, which is formed during intensive absorption of iron.

Treatment

Treatment can only be determined by a doctor on the basis of a comprehensive examination.