DISORDER/SETTING

Question 1:  What is the specific clinical disorder to be studied? 

The specific clinical disorder is primary iron overload of adult onset sufficient to cause significant morbidity and mortality.

·        Iron overload refers to excess deposition of iron in parenchymal cells in the liver, pancreas and heart, and/or increased total body mobilizable iron.

·        Primary refers to a genetically determined abnormality of iron absorption, metabolism, or both. 

·        Morbidity refers to organ damage that results in physical disability over and above that seen in the absence of iron overload. 

A single inherited disorder, HFE-related hereditary hemochromatosis (HHC) accounts for the vast majority of cases of primary iron overload in Caucasian adults in the United States.  The HFE gene is linked to HLA-A on the short arm of chromosome 6.  HFE-related HHC is a recessive disorder.  A small proportion of primary iron overload cases is explained by inherited disorders other than HFE-related HHC.  Juvenile hemochromatosis (HFE2) is a rare autosomal recessive disorder associated with a gene mapped to the long arm of chromosome 1.  Iron overload occurs earlier, with patients having a more severe clinical presentation in the second and third decades.  A third type of autosomal recessive, non-HFE-related primary iron overload results from mutations in the transferrin receptor 2 gene (TfR2) located on chromosome 7, and has been reported in a few Italian families (Camaschella et al., 2000; Girelli et al., 2002).  A fourth reported type shows autosomal dominant transmission and is associated with mutations in the ferroportin gene (SLC11A3) on the long arm of chromosome 2.  Other rare inherited disorders of iron metabolism include atransferrinemia, hyperferritinemia, and aceruloplasminemia (Fletcher and Halliday, 2001).  Although iron absorption is enhanced in a number of other inherited disorders (e.g., thalassemia major, iron-loading anemias, hereditary spherocytosis), these are considered secondary, or acquired, forms of iron overload and are not considered in this report. 

The original clinical diagnosis of hereditary hemochromatosis was based on the triad of hepatic cirrhosis, diabetes mellitus, and skin pigmentation.  However, by this point in the natural history of the disease, tissue damage due to iron overload has progressed too far for treatment to be more than palliative.  If population screening for iron overload were to be considered, two strategies for initial testing might be employed:

·        direct DNA testing for the homozygous C282Y HFE genotype

·        biochemical measurement(s) of serum transferrin saturation

Both strategies require that those with positive screening results undergo further testing to quantify the extent of iron overload.  For C282Y homozygous individuals, serum transferrin saturation and ferritin would be measured as follow-up tests.  For individuals with elevated transferrin saturation, further testing would also be needed to measure extent of iron overload (ferritin) and to differentiate HFE-related HHC from other causes of primary iron overload.  This report focuses on the first strategy employing DNA testing as the primary screening test.

DISORDER/SETTING 

Question 2:  What are the clinical findings defining this disorder? 

In 1880, Tosier described a group of patients with the triad of hepatic siderosis and cirrhosis, diabetes mellitus, and skin pigmentation; von Recklinghausen postulated that the origin of the iron deposited in the liver was the blood and named this disorder “hemochromatosis”.  Subsequently, it was shown that the iron deposition in HC is due to excessive absorption of iron from the gastrointestinal tract (Powell et al., 1975; Valberg et al., 1980).  Although iron may be deposited in many tissues, the primary site of uptake is the hepatic parenchymal cells (Powell et al., 1975; Worwood, 1997).  The clinical findings of iron overload are all directly or indirectly due to tissue iron deposition and damage by the iron, possibly from lipid peroxidation (Gutteridge et al., 1985; Britton et al., 1987; Sherwood et al., 1998).  The following are the most commonly involved organ systems:  

Liver:  Early in the course of iron overload, hepatic iron deposition may result in hepatomegaly, abdominal pain, and abnormal liver function tests.  If the deposition progresses and is not treated, hepatic fibrosis, cirrhosis, liver failure, and possibly hepatic carcinoma (either hepatocellular carcinoma or cholangiocarcinoma) may occur (Powell et al., 1975; Niederau et al., 1985; Cuthbert, 1997; Racchi et al., 1999; Blanc et al., 2000; Bonkowsky and Lambrecht, 2000).   

Heart:  Iron deposition in cardiac muscle can produce dilated or restrictive cardiomyopathy, either of which may result in heart failure (Niederau et al., 1985; Niederau et al., 1996).  This is the second most common cause of death, after liver disease, in untreated patients.  Adult males presenting with clinical disease before age 40 have a high prevalence of cardiomyopathy and arrhythmias; 60% under age 40 and nearly all under age 30 died of congestive failure (Finch and Finch., 1955).  Arrhythmias are most often atrial but may be ventricular (Milder et al., 1980; Niederau et al., 1985; Dabestani et al., 1988).  Although the etiology is uncertain in most cases, iron deposits in the myocardium appear to be involved.  Other factors, such as alcohol, probably exacerbate the cardiomyopathy and arrhythmias (Schellhammer et al., 1967).  If the patient can be kept alive by cardiotherapy during phlebotomy therapy, the cardiac disorders are completely reversible (Short et al., 1981; Dabestani et al., 1988). 

Endocrine glands:  Damage to the beta cells of the pancreatic islets, either directly from iron or from autoimmune reactions (presumably secondary to alteration of antigens by oxidation or other means), may result in diabetes mellitus (Niederau et al., 1985; Adams et al., 1991; Moirand et al., 1997),  Insulin resistance secondary to hepatic damage may also contribute to the metabolic dysfunction (Smith, 1990; Mendler et al., 1999).  Deposition of iron in the anterior pituitary gland may result in sexual dysfunction, including loss of libido, impotence, testicular atrophy, and amenorrhea, secondary to reduced production of the gonadotrophic hormones (Bezwoda et al., 1977; Walton et al., 1982). 

Joints:  Arthralgia or frank degenerative or inflammatory arthropathy is the single largest contributor to patient-perceived morbidity (Adams and Speechley, 1996).  Although other joints may be affected, the 2^nd and 3^rd metacarpal joints are most commonly involved (Axford, 1991; McCurdie and Perry, 1999).  Specific radiological findings include bone erosion, hooking osteophytes, and chondrocalcinosis (Axford, 1991; Hamilton et al.,1981; Huaux et al.,1986), similar to changes seen in severe hyperparathyroidism (Huaux et al., 1986). 

Skin:  Bronze skin pigmentation is believed to be due primarily to excessive melanin secretion (Smith, 1990; Adams et al., 1997; Pounder, 1997), although iron deposition itself may also be involved.   

Other:  Non-specific symptoms include lethargy, weakness, chronic fatigue, emotional distress (including frank depression), and abdominal pain.  These are frequently the presenting, and occasionally the only, symptoms of iron overload (Adams and Valberg, 1996; Adams et al., 1997; Moirand et al., 1997). 

DISORDER/SETTING 

Question 3:  What is the setting in which the test is to be performed? 

The setting for this report is population screening of adults.  Several studies have proposed biochemical and/or DNA screening for all adults.  The published recommendations, however, nearly always focus on the Caucasian population because of the higher prevalence of HHC and the high proportion attributable to the HFE gene.  The recommended minimum age for screening ranges from 20 to 40 years (Question 5).  Iron overload is usually not present in males until the second or third decade of life, and clinical signs and symptoms are uncommon before the fifth decade.  Women generally develop iron overload and associated clinical findings 8 to 30 years later than men (Meyer et al., 1990; Adams et al., 1991; Edwards and Kushner, 1993; Bulaj et al., 2000).  Although HFE mutations can be reliably detected at any time during life, the low penetrance in the early decades raises both ethical and medical questions about the appropriateness of genotyping prior to adulthood (Brittenham et al., 1998; Burke et al., 1998; McDonnell et al., 1998; Cogswell et al., 1999; Bhavnani et al., 2000; Hickman et al., 2000; Byrnes et al., 2001; Evans et al., 2001). 

In order for testing to be effective from a public health perspective, a screening program would need to be widely available.  One possibility would be for primary care providers to offer screening as part of routine care (McDonnell et al., 1998; Niederau et al., 1998).  However, a substantial proportion of the adult population does not avail itself of such care.  For that reason, other screening strategies need to be considered, in order to reach a broader segment of the target population.  For example, testing might be offered in a variety of public settings, similar to the model used for cholesterol testing. 

It has been proposed that rheumatology (Olynyk et al., 1994) and diabetic (O’Brien et al., 1994) clinic patients be “screened” for iron overload.  However, this type of routine testing cannot be considered screening.  The more appropriate terminology would be “case finding,” since a pre-selected, symptomatic population is being tested.  Case finding will not be discussed in this report. 

DISORDER/SETTING 

Question 4:  What DNA tests are associated with this disorder? 

In 1996, Feder et al. reported a 250 kb region on the short arm of chromosome 6, encoding a major histocompatibility complex (MHC) class I-like protein that was mutated in a large proportion of individuals with clinically diagnosed HHC (Feder et al., 1996).  This gene was initially called HLA-H and subsequently renamed HFE.  Two HFE missense mutations, C282Y and H63D, were initially described, and at least 17 other allelic variants of the HFE gene have now been reported (LeGac et al., 2001; Beutler et al., 2002).  By altering HFE protein structure and disrupting b₂-microglobulin binding and cell surface expression, the C282Y mutation results in significant loss of protein function (Feder et al., 1996; Feder et al., 1997).  Homozygosity for this mutation is most strongly correlated with clinically diagnosed primary iron overload due to HHC.  The effects of the other common mutations, H63D and S65C, on protein function are less severe, although both are associated with milder forms of the disorder in a small proportion of individuals who also carry the C282Y mutation (compound heterozygotes).  It is not yet completely clear whether H63D and S65C allelic variants are minor mutations with low penetrance or polymorphisms in linkage disequilibrium with one or more as yet unidentified mutations (Feder et al., 1996; Douabin et al., 1999).  Homozygosity for the C282Y mutation is the dominant genotype in HFE-related HHC and for that reason, it will serve in this report as the only DNA test evaluated. 

DNA-based tests for the two common mutations (C282Y, H63D) have been developed using a wide range of technologies that include the standard polymerase chain reaction (PCR)/restriction enzyme method (Feder et al., 1996), multiplex ARMS PCR (Baty et al., 1998; Bradley et al., 1998), LightCycler PCR (Bollhalder et al., 1999), multiplex PCR and capillary electrophoresis (Lubin et al., 1999), heteroduplex analysis (Jackson et al., 1997), high performance liquid chromatography (HPLC) (Liang et al., 2001), and real-time PCR fluorescent resonance energy transfer (FRET) hybridization (Parks et al., 2001).  Though a wide variety of testing methodologies has been described, most laboratories reporting to the American College of Medical Genetics/College of American Pathologists Molecular Genetics Laboratory external proficiency testing program are currently using the PCR/restriction enzyme and ARMS PCR methods.  In the United States, no kits have been approved by the Food and Drug Administration (FDA) for HFE testing, but some of these have been approved by the FDA as Analyte Specific Reagents (ASRs).  Laboratories offering HFE mutation analysis will come under ‘home brew’ regulations. 

Testing has been successfully performed using anticoagulated blood, buccal samples, and dried blood spots.  Blood samples (obtained by venipuncture) serve as a highly reliable source of DNA and can be readily obtained in many health care settings.  The method of collecting buccal cells by brush, swab or mouthwash is inexpensive and is well suited to collecting samples in primary care offices, at home, and in other non-health care settings.  Blood and buccal samples are stable when transported at ambient temperature, and testing has been successfully performed on buccal lysates stored frozen for 3-4 years (Haddow et al., 1999).  Buccal sample failure rates are generally 1% or less, and results can nearly always be obtained from blood samples.

DISORDER/SETTING 

Question 5:  Are “pre-screening” tests employed (Asking a question about race/ethnicity)? 

An inquiry about racial/ethnic heritage may be appropriate prior to offering HFE mutation analysis as a screening test.  Both the population prevalence of hemochromatosis and the frequencies of the common alleles vary, depending upon race and ethnicity.  On average, heterozygosity for C282Y is found in about 9 percent of Caucasians in Europe and North America, but it is almost never observed in populations from Africa, the Middle East, Asia, the Indian subcontinent, and Australasia (Merryweather-Clarke, 1997; Merryweather-Clarke, 1999; Hanson et al., 2000).

DISORDER/SETTING 

Question 6:  Is it a stand-alone screening test or is it one of a series of screening tests? 

The DNA-based testing that is used for screening individuals for predisposition to primary iron overload due to HFE-related HHC is a “stand-alone” test.  It may be preceded in some programs by a screening question about race/ethnicity, intended to determine what individuals should be offered screening, or to provide specific information about the efficacy of testing.  HFE mutation analysis identifies individuals who are at risk for iron overload because they are homozygous for the C282Y mutation.  Follow-up testing to determine the extent of iron overload could identify individuals who may benefit treatment.

DISORDER/SETTING 

Question 7:  If it is part of a series of screening tests, are all tests performed in all instances (parallel) or are some tests only performed on the basis of other results (series)? 

HFE testing may be preceded in some programs by a screening question about race/ethnicity, intended to determine what individuals should be offered screening or to provide specific information about the efficacy of testing.

References 

Adams PC, Chakrabarti S.  1998.  Genotypic-phenotypic correlations in genetic hemochromatosis: evolution of diagnostic criteria.  Gastroenterology 114:319-23.

Adams PC, Deugnier Y, Moirand R, Brissot P.  1997.  The relationship between iron overload, clinical symptoms, and age in 410 patients with genetic hemochromatosis.  Hepatology  25:162-166.

Adams PC, Gregor JC, Kertesz, Valberg LS.  1995.  Screening blood donors for hereditary hemochromatosis: decision analysis model based on a 30-year database.  Gastroenterology  109:177-88.

Adams PC, Kertesz AE, McLaren CE, Barr R, Bamford A, Chakrabarti S.  2000.  Population screening for hemochromatosis: a comparison of unbound iron-binding capacity, transferrin saturation, and C282Y genotyping in 5,211 voluntary blood donors.  Hepatology 31:1160-4. 

Adams PC, Kertesz, Valberg LS.  1991.  Clinical presentation of hemochromatosis: a changing scene.  Am J Med  90:445-459.

Adams PC, Speechley M.  1996.  The effect of arthritis on the quality of life in hereditary hemochromatosis.  J Rheumatol  23:707-710.

Adams PC, Valberg LS.  1996.  Evolving expression of hereditary hemochromatosis.  Semin Liver Dis  16:47-54.

Axford JS.  1991.  Rheumatic manifestations of haemochromatosis.  Baillière’s Clin Rheumatol  5:351-365. 

Baer DM, Simons JL, Staples RL, Rumore GH, Morton CJ.  1995.  Hemochromatosis screening in asymptomatic ambulatory men 30 years of age and older.  Am J Med 98:464-8.

Bakker AJ.  1991.  Influence of monoclonal immunoglobulins in direct determinations of iron in serum.  Clin Chem 37:690-4. 

Balabaud C, Bioulac-Sage P.  2000.  Increased incidence of HFE C282Y mutations in patients with iron overload and hepatocellular carcinoma developed in non-cirrhotic liver.  J Hepatol  32:805-811. 

Barton JC, Rothenberg BE, Bertoli LF, Acton RT.  1999.  Diagnosis of hemochromatosis in family members of probands: a comparison of phenotyping and HFE genotyping.  Genetics in Medicine 1:89-93.

Bassett ML, Halliday JW, Bryant S, Dent O, Powell LW.  1988.  Screening for hemochromatosis.  Ann NY Acad Sci 526:274-89.

Baty D, Terron Kwiatkowski A, Mechan D, Harris A, Pippard MJ, Goudie D.  Development of a multiplex ARMS test for mutations in the HFE gene associated with hereditary hemochromatosis.  J Clin Pathol, 51: 73-4.

Beutler E, Griffin MJ, Gelbart T, West C.  A previously undescribed nonsense mutation of the HFE gene.  Clin Genet 2002;61:40-42.

Bezwoda WR, Bothwell TH, Van der Walt LA, Kronheim S, Pimstone BL.  1977.  An investigation into gonadal dysfunction in patients with idiopathic haemochromatosis.  Clin Endocrin  6:377-385.

Bhavnani M, Lloyd D, Bhattacharyya A, Marples J, Elton P, Worwood M.  2000.  Screening for genetic haemochromatosis in blood samples with raised alanine aminotransferase.  Gut 46:707-10. 

Blanc JF, de Ledinghen V, Bernard P-H, de Verneuil H, Winnock M, Le Bail B, Carles J, Saric J, Balabaud C, Bioulac-Sage P.  2000.  Increased incidence of HFE C282Y mutations in patients with iron overload and hepatocellular carcinoma developed in non-cirrhotic liver.  J Hepatol 32:805-11. 

Bollhalder M, Mura C, Landt O, Maly FE.  1999.  LightCycler PCR assay for simultaneous detection of the H63D and S65C mutations in the HEF hemochromatosis gene based on opposite melting temperature shifts.  45: 2275-8.

Bonkovsky HL, Lambrecht RW.  2000.  Iron-induced liver injury.  Clin Liver Dis  4:409-429.

Borwein S, Ghent CN, Valberg LS.  1984.  Diagnostic efficacy of screening tests for hereditary hemochromatosis.  Can Med  Assoc J 131:895-901.

Bradley LA, Johnson DD, Palomaki GE, Haddow JE, Robertson NH, Ferrie RM.  1998.  Hereditary haemochromatosis mutation frequencies in the general population.  J Med Screen  5: 34-6.

Brittenham GM, Franks AL, Rickles FR.  1998.  Research priorities in hereditary hemochromatosis.  Ann Intern Med 129:993-6.

Britton RS, Bacon BR, Recknagel RO.  1987.  Lipid peroxidation and associated hepatic organelle dysfunction in iron overload.  Chem Phys Lipids  45:207-239.

Bulaj ZJ, Ajioka RS, Phillips JD, LaSalle BA, Jorde LB, Griffen LM, Edwards CQ, Kushner JP.  2000.  Disease-related conditions in relatives of patients with hemochromatosis.  N Engl J Med 343:1529-35.

Burke W, Thomson E, Khoury MJ, McDonnell SM, Press N, Adams PC, Barton JC, Beutler E, Brittenham G, Buchanan A, Clayton EW, Cogswell ME, Meslin EM, Motulsky AG, Powell LW, Sigal E, Wilfond BS, Collins FS.  1998.  Hereditary hemochromatosis.  Gene discovery and its implications for population-based screening.  JAMA 280:172-8.

Byrnes V, Ryan E, Barrett S, Kenny P, Mayne P, Crowe J.  2001.  Genetic hemochromatosis, a Celtic disease:  Is it now time for population screening?  Genet Test 5:127-30.

Camaschella C, Roetto A, Cali A, De Gobbi M, Garozzo G, Carella M, Majorano N, Totaro A, Gasparini P.  The gene TFR2 is mutated in a new type of haemochromatosis mapping to 7q22.  Nat Genet 2000;25:14-15.

Chakraborty R, Kamboh MI, Nwankwo M, Ferrell RE.  1992.  Caucasian genes in American Blacks: new data.  Am J Hum Genet 50:145-55

Cogswell ME, Burke W, McDonnell SM, Franks AL.  1999.  Screening for hemochromatosis.  A public health perspective.  Am J Prev Med 16:134-40.

Conte D, Manachino D, Colli A, Guala A, Aimo G, Andreoletti M, Corsetti M, Fraquelli M.  1998.  Prevalence of genetic hemochromatosis in a cohort of Italian patients with diabetes mellitus.  Ann Intern Med 128:370-3.

Cuthbert JA.  1997.  Iron, HFE, and hemochromatosis update.  J Invest Med  45:518-529.

Dabestani A. Child JS, Perloff JK, Figueroa WG, Schelbert HR, Engel TR.  1988.  Cardiac abnormalities in primary hemochromatosis.  Ann NY Acad Sci  526:234-244.

Douabin V, Moirand R, Jouanolle A, Brissot P, LeGall J, Deugnier Y, David V.  Polymorphisms in the HFE gene.  Hum Hered  49: 21-6.

Edwards C, Kushner JP.  1993.  Screening for hemochromatosis.  N Eng J Med 328:1616-20.

Edwards CQ, Carroll M, Bray P, Cartwright GE.  1977.  Hereditary hemochromatosis. Diagnosis in siblings and children.  N Engl J Med 197:7-13. 

Emond MJ, Bronner MP, Carlson TH, Lin M, Labbe RF, Kowdley KV.  1999.  Quantitative study of the variability of hepatic iron concentrations.  Clin Chem 45:340-6.

Evans JP, Skrzynia C, Burke W.   2001.  The complexities of predictive genetic testing. BMJ 322:1052-6. 

Feder JN, Gnirke A, Thomas W, Tsuchihashi Z, Ruddy DA, Basava A, et al.  1996.  A novel MHC class 1-like gene is mutated in patients with hereditary haemochromatosis.  Nat Genet  13: 399-408.

Feder JN, Tsuchihashi Z, Irrinki A, Lee VK, Mapa FA, Morikang E, et al.  1997.  The hemochromatosis founder mutation in HLA-H disrupts beta-2-microglobulin interaction and cell surface expression.  J Biol Chem 272: 14025-8.

Finch SC, Finch CA.  1955.  Idiopathic hemochromatosis, an iron storage disease. A. Iron metabolism in hemochromatosis.  Medicine  34:381-430.

George DK, Powell LW.  1997.  Review article: the screening, diagnosis and optimal management of haemochromatosis.  Aliment Pharacol Ther 11:631-9.

Girelli D, Bozzini C, Roetto A, Alberti F, Daraio F, Colombari R, et al.  Clinical and pathologic findings in hemochromatosis type 3 due to a novel mutation in Transferrin Receptor 2 gene.  Gastroenterology 2002;122:1295-1302.

Gordeuk V, Mukiibi J, Hasstedt SJ, Samowitz W, Edwards CQ, West G, Ndambire S, Emmanual J, Nkanza N, Chapanduka Z, et al. 1992.  Iron overload in Africa. Interaction between a gene and dietary iron content. N Engl J Med 326:95-100.

Guyader D, Jacquelinet C, Moirand R, Turlin B, Mendler MH, Chaperon J, et al.  Noninvasive prediction of fibrosis in C282Y homozygous haemochromatosis.  Gastroenterology 1998;115:929-36.

Haddow JE, Bradley LA.  1999.  Hereditary hemochromatosis: to screen or not.  Conditions for screening are not yet fulfilled.  BMJ 7209: 531-2.

Hamilton EBD, Bomford AB, Laws JW, Williams R.  1981.  The natural history of arthritis in idiopathic haemochromatosis: progression (sic) of the clinical and radiological features over ten years.  Q J Med  L199:321-329.

Huaux JP, Geubel A, Koch MC, Malghem J, Devogelaer JP, de Deuxchaisnes CN.  The arthritis of hemochromatosis.  1986.  A review of 25 cases with special reference to chondrocalcinosis, and a comparison with patients with primary hyperparathyroidism and controls.  Clin Rhematol  3:317-324.

Le Gac G, Mura C, Ferec C.  Complete scanning of the hereditary hemochromastosis gene (HFE) by use of denaturing HPLC.  Clin Chem 2001;47:1633-40.

Liang Q, Davis PA, Thompson BH, Simpson JT.  2001.  High-performance liquid chromatography multiplex detection of two single nucleotide mutations associated with hereditary hemochromatosis.  J Chromatogr B Biomed Sci Appl  754: 265-70.

Lubin IM, Yamada NA, Stansel RM, Pace RG, Rohlfs EM, Silverman LM.  1999.  HFE genotyping using Multiplex allele-specific polymerase chain reaction and capillary electrophoresis.  Arch Pathol Lab Med  123: 1277-81.

Ludwig J, Batts KP, Moyer TP, Baldus WP, Fairbanks VF.  1993.  Liver biopsy diagnosis of homozygous hemochromatosis: a diagnostic algorithm.  Mayo Clin Proc 68:263-7.

Lyon E, Frank EL.  2001.  Hereditary hemochromatosis since discovery of the HFE gene. Clin Chem 47:1147-56. 

McCurdie I, Perry JD.  1999.  Haemochromatosis and exercise related joint pains.  BMJ  318:449-451.

McDonnell SM, Phatak PD, Felitti V, Hover A, McLaren GD.  1998.  Screening for hemochromatosis in primary care settings.  Ann Intern Med 129:962-70.

Mendler M-H, Turlin B, Moirand R, Jouanolle A-M, Sapey T, Guyader D, le Gall J-Y, Brissot P, David V, Deugnier Y.  1999.  Insulin resistance-associated hepatic iron overload.  Gastroenterology  117:1155-1163.

Meyer T, Baynes R, Bothwell T, Jenkins T, Jooste P, du Toit E, Martell R, Jacobs P.  1990.  Phenotypic expression of the HLA linked iron-binding gene in males over the age of 40 years: a population study using serial serum ferritin estimations.  J Intern Med 227:397-406.

Milder MS, Cook JD, Stray S, Finch CA.  1980.  Idiopathic hemochromatosis, an interim report.  Medicine  59:34-49.

Moirand R, Adams PC, Bicheler V, Brissot P, Deugnier Y.  1997.  Clinical features of genetic hemochromatosis in women compared with men.  Ann Intern Med  127:105-110.

Mura C, Raguenes O.  1999.  HFE mutation analysis in 711 hemochromatosis probands: evidence for S65C implication in the mild form of hemochromatosis.  Blood  932: 2502-5.

Niederau C, Fischer R, Pürschel  A, Stremmel W, Häussinger D, Strohmeyer G.  1996.  Long-term survival in patients with hereditary hemochromatosis.  Gastroenterology  110:1107-1119.

Niederau C, Fischer R, Sonnenberg A, Stremmel W, Trampisch HJ, Strohmeyer G.  1985.  Survival and causes of death in cirrhotic and in noncirrhotic patients with primary hemochromatosis.  N Eng J Med  313:1256-1262.

Niederau C, Niederau CM, Lange S, Littauer A, Abdel-Jalil N, Maurer M, Häussinger H, Strohmeyer G.  1998.  Screening for hemochromatosis and iron deficiency in employees and primary care patients in Western Germany.  Ann Intern Med 128:337-45.

Olsson KS, Ritter B, Rosèn U, Heedman PA, Staugård F.  1983.  Prevalence of iron overload in central Sweden.  Acta Med Scand 213:145-50.

Olynyk JK, Cullen DJ, Aquilia S, Rossi E, Summerville L, Powell LW.  1999.  A population-based study of the clinical expression of the hemochromatosis gene.  N Engl J Med 341:718-24. 

Parks SB, Popovich BW, Press RD.  2001.  Real-time polymerase chain reaction with fluorescent hybridization probes for the detection of prevalent mutations causing common thrombophilic and iron overload phenotypes.  Am J Clin Pathol  115: 439-47.

Piperno A.  Classification and diagnosis of iron overload.  1998.  Haematologica 83:447-55.

Pointon JJ, Wallace D, Merryweather-Clarke AT., et al.  2000.  Uncommon mutations and polymorphisms in the hemochromatosis gene.  Genet Test  4: 151-61.

Pounder R.  1997.  Genetic haemochromatosis.  Lancet  349:1688-1693.

Powell LW, Kerr JF.  1975.  The pathology of the liver in hemochromatosis.  Pathobiol Annu  5:317-337.

Racchi O, Mangerini R, Rapezzi D, Gaetani GF, Nobile MT, Picciotto A, Ferraris AM.  1999.  Mutations of the HFE gene and the risk of hepatocellular carcinoma.  Blood Cells Mol Dis  25:350-353. 

Reed TE.  1992.  Issues in estimating Caucasian admixture in American Blacks.  Am J Hum Genet 51:678-9.

Schellhammer PE, Engle MA, Hagstrom JW.  1967.  Histochemical studies of the myocardium and conduction system in acquired iron-storage disease.  Circulation 35:631-7.

Sham RL, Raubertas RF, Braggins C, Cappuccio J, Gallagher M, Phatak PD.  2000.  Asymptomatic hemochromatosis subjects: genotypic and phenotypic profiles.  Blood 96:3707-11. 

Short EM, Winkle RA, Billingham ME.  1981.  Myocardial involvement in idiopathic hemochromatosis.  Morphologic and clinical improvement following venesection.  Am J Med 70:1275-9.

Smith  LH.  1990.  Overview of hemochromatosis.  West J Med 153:296-308.

Tavill AS.  2001.  Diagnosis and management of hemochromatosis.  Hepatology 33:1321-8. 

Valberg LS, Lloyd DA, Ghent CN, Flanagan PR, Sinclair NR, Stiller CR, Chamberlain MJ.  1980.  Clinical and biochemical expression of the genetic abnormality in idiopathic hemochromatosis.  Gastroenterology 79:884-92.

Villeneuve J-P, Bilodeau M, Lepage R, Côté J, Lefebvre M.  1996.  Variability in hepatic iron concentration measurement from needle-biopsy specimens.  J Hepatol 25:172-7.

Walton C, Kelly WF, Laing I, Bullock DE.  1983.  Endocrine abnormalities in idiopathic haemochromatosis.  Q J Med  L11:99-110.