Hemochromatosis is an autosomal recessive disorder of iron metabolism that is characterized by increased iron absorption and iron accumulation in the liver, pancreas, heart, synovia (A clear, viscid lubricating fluid secreted by membranes in joint cavities), and pituitary gland. Iron accumulation in the liver leads to hepatic scarring, cirrhosis and in some cases, cancer. Iron accumulation in other tissues causes diabetes, caridomyopathy, arrhythmias, arthritis, hypogonadism, impotence, hyperpigmentation and some non-specific symptoms such as fatigue and weakness. Approximately one of every 200 to 400 people is affected, while 1 in 10 is a carrier. (1)
Hemochromatosis (HH) is classified as either primary or secondary. Primary or hemochromatosis is genetically influenced. In 1996, the gene for HHC was mapped on the short arm of chromosome 6 (6p21.3). This location is near the class 1 major histocompatibilty (MHC) gene. The linkage of HH to HLA-A has been utilized clinically to track affected pedigrees. A serologic test for HLA-A3 (Human Leucocitary Antigen- H) has often been used to assess whether relatives of a known hemochromatosis patient will have the disease. Ten mutations have been identified within the HFE (Hemochromatosis Gene) gene as of April 2000. Two of those mutations, C282Y and H63D, have been linked with the phenotypic expression of the disease. Phenotypic expression of HHC is due to a mixture of genetic susceptibility, age, gender and some environmental influences. (2) Hereditary Hemochromatosis is further broken down into four types. Type 1, which affects the HFE gene and is thought to interfere with the regulation of intestinal iron absorption. Patients develop chronic iron deposition in liver, hair, skin, and pancreas. Type 2 HH is similar to type 1 but there is accelerated iron loading that results in rapid death, which is why type 2 is known as the juvenile form of hereditary hemochromatosis. Type 3 HH is due to a defect in the Transferrin Receptor 2 (TRF 2) gene. TRF 2 affects regulation of intestinal iron absorption and this type has the same clinical manifestations as type 1. Finally, type 4 hemochromatosis is caused by a defect in the ferroportin gene (SLC11A3) that affects transmembrane export of iron from intestinal epithelial cells, especially macrophages. Ferroportin is highly expressed by reticuloendothelial macrophages. Which operate to upload iron form hemoglobin scavenged from senescent red blood cells. The flux of iron through reticuloendothelial macrophages is much greater than the flux through the intestinal epithelial cells. The loss of one functional copy of ferroportin in macrophages makes iron recycling less efficient, leading to a misinterpretation of body iron stores. As a consequence, regulatory mechanisms are thought to convey a signal to intestinal epithelial cells that iron absorption should increase to meet demand and iron overload results. (3)
Generally patients with HH are homozygous for a mutation of the HFE gene; usually a cysteine substituted for a tyrosine at position 282 (C282Y). Another prevalent mutation found in the HFE gene results in the substitution of histamine for aspartic acid at position 63 (H63D). The HFE protein is a transmembrane glycoprotein that is homologous to MHC. Researchers who found that the HFE gene's normal protein product had a resemblance to an HLA molecule first noted this. Initially it was called HLA-H, but using linkage-disequilibrium and full haplotype analysis scientist where able to find 2 distinct proteins. Like an HLA protein the HFE spans the cell membrane. Its external portion includes a nonfunctional peptide-binding domain, the site where a true HLA protein would hold an antigen for inspection by immune cells. Both proteins also have an alpha 3 loop, a region where they associate with an accessory protein called beta 2- microglobulin. (See figure 1). (4)
Three pathways exist in the small intestine for uptake of food iron (see figure 2). There are separate uptake pathways for heme, ferric (Fe 3+) and ferrous (Fe 2+) iron. Most dietary iron is in the ferric state. It is mobilized from food in the stomach acid for chelation with mucins ascorbace (this is a dietary factors that influences iron absorption). Ascorbate and citrate increase iron uptake in part by acting as weak chelators to help to solubilize the metal in the duodenum. Iron is readily transferred from these compounds into the mucosal lining cells. Ferric iron enters the absorptive cell via ß 3 integrin in combination with mobilferrin. A large complex named paraferritin (P), which contains intergrin; mobliferrin flavin, monooxygenase, and ß 2 microglobulin, is found in the cell cytosol; it permits ferrireduction so ferrous iron is available for incorporation in to compounds such as ferritin and heme. Heme iron is digested free of globin in the intestinal lumen and enters the absorptive cell as an intact metalloporphyrin. This process is believed to be endosomal. (5) Within the cell, heme is degraded by heme oxygenase and this releases inorganic iron. On the basolateral membranes there are transferrin receptors, which permit entry of the transferrin-iron complex into the endosome vesicles. (4)
Iron metabolism starts in the intestinal lumen where it is reduced from the ferric to the ferrous state and is then transported into the enterocytes by the divalent mental transporter 1 (DMT-1). There is also another protein that encourages iron transport called stimulator of iron transport (SFT) that has finally been cloned in recent years. SFT is a transmembrane protein whose transcript levels in liver were upregulated in hemochromatosis patients. Little is known about relationship and role in iron metabolism of SFT extensive research is currently underway. Once iron leaves the lumen, through DMT-1 and SFT, it enters the portal circulation for delivery by transferrin to target cells such as hepatocytes and erythrocytes.
Regulation of iron metabolism by the HFE protein comes from it's binding to the transferrin receptor. This binding increases the receptor's affinity for iron-carrying transferrin. Transferrin and its receptor are recycled into the circulation and free to bind and transport additional iron atoms. The HFE protein associates with the transferrin receptor and prevents internalization of iron-transferrin complex into cells. The HFE protein, in effect, acts as a brake on cellular iron uptake and increases the receptor's affinity for iron-carrying transferrin. (6). Absence or dysfunction of the HFE gene is therefore the cause of HH. The major mutation C282Y abolishes the B2-microglobulin binding site of the HFE gene, permitting HFE degradation. In the absence of HFE iron transport into the cytoplasm proceeds without negative regulation. The minor mutation H63D decreases the capacity of HFE to inhibit endosomal iron release in this manner increasing iron flux into the cytoplasm. (7)
Studies on iron metabolism at the molecular level have focused on the function of transferrin and the transferrin receptor. This bilobal protein is the only way our body has of transporting iron to the endosomes. The two lobes that make up transferrin have been designated the C lobe and the N lobe due to their sequences. The N lobe has a pair of lysines that when protonated induce the release of iron. The protonation of lysine is facilitated by low pH in the endosomal cells. The release of iron is also assisted by the transferrin receptor, which also lowers local pH by, having partially negative amino acids on its active site, thus facilitating protonation of the lysines on transferrin. The mechanisms whereby HFE influences iron absorption through the duodenal mucus in relation with the maintenance of iron homeostasis is still unknown.(8)
The discovery of DMT-1 has given great insight into the mechanism of Fe absorption. On important step was the isolation of Fe+2-regulated transporter 1 (IREG1). This molecule was also named ferroportin 1 and metal transport protein 1 (MTP 1). Three different methods where used to isolate this protein. In one study the gene responsible for hypochromic anemia in zebra fish was cloned. Subtractive cloning from the cDNA (DNA of strong, cloned copies of otherwise fragile mRNA) of hypotrans-ferrinemic mice was applied. This technique is used to identify genes expressed differentially between two tissue samples. A large excess of mRNA from one sample is hybridized to cDNA from the other, and the double stranded hybrids removed by physical means. Remaining cDNAs are those not represented as RNA in the first sample, and thus presumably expressed uniquely in the second. To improve specificity, the process is often repeated several times. Another study was based on systemic evolution of ligands by exponential enrichment (SELEX) in order to find the ferroportin 1 gene The SELEX method is c straightforward: in a standard DNA-oligonucleotide synthesizer a starting pool is generated. The machine synthesizes an oligonucleotide with a completely random base-sequence which is flanked by defined primer binding sites. In this way, up to 10exp15 different DNA molecules can be synthesized at once. The immense complexity of the generated pool lets the scientist assume that it contains a few molecules with the correct receptor structure; these are selected, for example by affinity chromatography or filter binding. Because a pool of such high complexity can be expected to contain only a very small fraction of functional molecules, several purification steps are usually required. Therefore, the very rare active molecules are amplified by the polymerase chain reaction (PCR) or in a transcription-based step. In this way cycles of selection can be carried out. Successive selection and amplification cycles result in an exponential increase in the abundance of functional sequences, until they dominate the population, and this is how the ferroportin was establish.
Further evidence for the role of ferroportin iron transport is its in tissues that are critical in iron absorption is. Ferroportin 1 is expressed at the basolateral membrane of villus in enterocytes and in the liver and spleen, large intestine heart, kidney, and skeletal muscle. Researchers hope that ferroportin 1 can be used to diagnose hemochromatosis patients. This is because studies have found that a mutation in ferroportin 1 has been associated with the development of type 4 autosomal dominant hemochromatosis. (9) Another area of research on hemochromatosis has been on the hormone hepcidin. It is thought to be the "missing link" involved in communication between sites of iron storage and the uptake. Scientist working with lab mice found that their animals had hepatic overload resembling that of human HH. They soon discovered that mice with the USF2 (Upstream Stimulatory Factor 2), which increases iron responsiveness, failed to express copies of hepcidin genes downstream of USF2. A connection was made between iron loading and the absence of hepcidin. Hepcidin now joins a list of gene deficiencies that have been demonstrated to cause HH-like phenotypes in previous studies: HFE, transferrin receptor, b2- microglobulin (b2M) and hepcidin. The products of these genes are interrelated; transferrin receptor 1 (TfR 1) binds to HFE in the duodenal crypt cells (which line the pits in the small intestine that lead to the tubular intestinal glands). An additional gene, Transferrin receptor 2 (TfR 2), mediates iron uptake by hepatocytes, which, in turn regulates the expression of hepcidin. Hepcidin interacts with HFE, b2M and TfR in the duodenal crypt cell to regulate dietary absorption of iron. There is stipulation that this is the model for regulation of iron homeostasis. Further studies must be done to confirm that the loss of Hepcidin itself produces the hereditary hemochromatosis phenotype. However there is hope that it will provide a potential drug target to modulate iron absorption. (10) Post-translation iron regulatory proteins (IRPs) 1 and 2 play an important role in regulating iron homeostasis. IREs were first identified in the ferritin 5' untranslated region (UTR). Binding of IRPs to the ferritin 5' iron responsive element (IRE) represses translation by preventing recruitment of the 40s small ribosome subunit to the RNA. The binding of IRPs to the transferrin receptor 3' IRE protects this mRNA from endonucleolytic cleavage. Iron regulation of IRPs results in increased ferritin translation, leading to iron seizures and decline of TfR mRNA. (7)
Early diagnosis is the best treatment for Hemochromatosis. Physicians use the "rule of three A's " in which each "A" correspond major symptoms. For example, asthenia refers to unexplained chronic fatigue, with a sexual component in males. There is also arthralgia, which is sign of pain in the joints especially the wrists and knees. The last symptom would be aminotransferase increase (Iron depletion leads to a reduction in aminotransferase levels) which is three times the normal absorption of iron in patients.(2) The focal point of diagnosis is the serum transferrin saturation (TS). This establishes biochemical defects of iron metabolism. Increased TS reflects the basic metabolic abnormality of Hemochromatosis and is acknowledged as the most sensitive single test for phenotypic identification of the disease. (8)
Treatment for HH patients does not necessarily involve a low iron diet. The benefits of keeping a strict, low iron, diet for a year are equal to having 2 or 3 blood phlebotomies in a year. Instead of a special diet supplemental iron and vitamin c, which contribute to iron absorption, are removed from a person's diet. Another change involves increased consumption of foods or drinks that decrease iron absorption such as tea. Periodic phlebotomy (periodic removal of blood) is a safe, effective and inexpensive treatment for HH patients. A one-unit phlebotomy (500ml of whole blood) will remove approximately 250 mg of iron. (11) Liver transplantation has become an accepted form of treatment for chronic, severe liver damage. However, it must be initiated before organ damage has occurred. Advances in surgical techniques and the use of new drugs to suppress rejection have improved the success rate of transplantation. The outcome is excellent for hemochromatosis patients, and with the slow progression of this disease, it is possible to plan elective transplant surgery. For those people who cannot be bled because of extreme anemia, there is chelation. This approach lacks the complete efficiency of bloodletting and is used only when absolutely necessary. For some it is infused over night with a portable pump at home during sleep over a 12-hour period. In most cases, the infusion pump is installed in the body of the patient.(12)
Hemochromatosis is a very common disease but the outlook is optimistic. Research is making leaps and bounds. The most immediate public-health effort is to make doctors more aware of the symptoms in order to increase early detection. As HFE screening is moving toward clinical realization, problems arise. Consider this, if HFE screening were widespread throughout the United States 1 to 1.5 million people would be placed on a phlebotomy program. For the general population it would form a costly swell of now-limited medical resources. The benefits of genetic testing by far compensate for the drawbacks. Genetic testing can give an opportunity for prevention and can tell you if your family members are at risk.
Copyright © 2002 Claudia Cortes and Koni Stone
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