[Hereditary iron overload]. / Surcharges héréditaires en fer.

The field of hereditary iron overload has known, in the recent period, deep changes mainly related to major advances in molecular biology. It encompasses now a series of genetic entities. The mechanistic understanding of iron overload development and iron toxicity has greatly improved. The diagnostic approach has become essentially noninvasive with a major role for biological tests. From the therapeutic viewpoint, the phlebotomy treatment is now enriched by the possibility of resorting to oral chelation and by innovative perspectives directly linked to our improvement in the molecular understanding of these diseases.

The 16189 variant of mitochondrial DNA occurs more frequently in C282Y homozygotes with haemochromatosis than those without iron loading.

BACKGROUND: Patients with hereditary haemochromatosis (HH) are usually homozygous for the C282Y mutation in the HFE gene. They have variable expression of iron overload and present with a variety of complications, including liver disease, diabetes, arthropathy, fatigue, and cardiomyopathy. The mitochondrial 16189 variant is associated with diabetes, dilated cardiomyopathy, and low body fat at birth, and might contribute to genetic predisposition in further multifactorial disorders. The objective of this study was to determine the frequency of the 16189 variant in a range of patients with haemochromatosis, who had mutations in the HFE gene. METHODS: Blood DNA was analysed for the presence of the 16189 variant in British, French, and Australian C282Y homozygotes and controls, with known iron status, and in birth cohorts. RESULTS: The frequency of the mitochondrial 16189 variant was found to be elevated in individuals with haemochromatosis who were homozygous for the C282Y allele, compared with population controls and with C282Y homozygotes who were asymptomatic (42/292 (14.4%); 102/1186 (8.6%) (p = 0.003); and 2/64 (3.1%) (p = 0.023), respectively). CONCLUSIONS: Iron loading in C282Y homozygotes with HH was exacerbated by the presence of the mitochondrial 16189 variant.

Serum paraoxonase activity and paraoxonase gene polymorphism in type 2 diabetic patients with or without vascular complications.

BACKGROUND: Serum paraoxonase (PON) activity and the relevance of PON gene polymorphism in vascular complications of type 2 diabetic patients were investigated in a case-control study. METHODS: The population included 105 control subjects, 96 diabetic patients without vascular complications and 71 diabetics with vascular complications. RESULTS: Serum PON activity was significantly decreased (p<0.001) in diabetic patients without vascular complications: 207 IU (25-817) compared with the controls: 259 IU (24-950). Although serum PON activity was also decreased: 232 IU (34-797) in the population with vascular complications, the difference was not statistically significant (p=0.11). The Q192 allele frequency is significantly higher (p<0.005) in diabetics without vascular complications (77%), and with vascular complications (73%) than in the controls (63%). No significant association was found between either PON(1)55 L/M and PON(2)311 C/S gene polymorphisms and vascular complications. CONCLUSIONS: The difference in allele frequency for the PON(1) Q/R 192 gene polymorphism may be the cause of the low paraoxonase activity observed in type 2 diabetes mellitus. Further studies need to be conducted to elucidate the role of the enzyme in the development of vascular complications in diabetes.

Low penetrant hemochromatosis phenotype in eight families: no evidence of modifiers in the MHC region.

The gene responsible for hemochromatosis (HFE) has been identified on the short arm of chromosome 6, 4.5 Mb telomeric to HLA-A. A major mutation C282Y is closely correlated with the disease, as it accounts for 68 to 100\% of the cases of hemochromatosis. Nevertheless, some C282Y homozygotes subjects have no clinical or biological expression of the disease. Moreover, in Northern European populations a large discrepancy is observed between the number of C282Y homozygotes and the number of diagnosed hemochromatosis patients, suggesting incomplete penetrance of the mutation. To localize and identify the modifying genes, we investigated eight families including C282Y homozygous relatives showing no clinical signs of the disease, in addition to the hemochromatosis patients. Genomic DNA from 20 C282Y homozygotes (10 patients and 10 siblings presenting no or minor biological abnormalities) were studied. Five polymorphisms from the HFE gene were determined by PCR restriction. Extended haplotypes of the 6p21.3 region were constructed with 10 microsatellite markers. All the C282Y homozygotes shared the same HFE polymorphism. The haplotypes presented no significant difference between the probands and their unaffected relatives. These studies suggest that neither HFE polymorphism nor genes surrounding HFE are able to modulate HFE expression.

Geography of HFE C282Y and H63D mutations.

Hereditary hemochromatosis (HH) is a common autosomal recessive disorder causing inappropriate dietary iron absorption that affects North Europeans. HH is associated with the C282Y mutation of the HFE gene, and the H63D mutation to a lesser degree. Both mutations are abundant in Europe, with H63D also appearing in North Africa, the Middle East, and Asia. Emigration from Europe over the past 500 years has introduced C282Y and H63D to America, Australia, New Zealand, and South Africa in an essentially predictable fashion. The distinctive characteristics of the population genetics of HH are the confined racial distribution and high frequency in North European peoples. C282Y frequencies in North Europeans are typically between 5% and 10%, with homozygotes accounting for between 1/100 and 1/400 of these populations. The scarcity of the C282Y mutation in other populations accounts for the lack of HH in non-Europeans.

[Molecular genetics of hemochromatosis]. / Génétique moléculaire de l'hémochromatose.

Hemochromatosis is a recessive disorder of iron metabolism characterized by progressive iron loading of parenchymal organs, which accounts for clinical complications such as cirrhosis, diabetes mellitus, cardiopathy, endocrine dysfunctions and arthropathy. Clinical complications, which usually develop after the third or fourth decade of life, can be fatal but may be prevented by phlebotomy if iron excess is detected at a very early stage. The hemochromatosis gene (HFE), located 4.5 megabases telomeric to the HLA-A locus, encodes an HLA class I like protein and two missense mutations, C282Y and H63D in complete disequilibrium have been identified within this gene. Due to its high frequency in the general population, the involvement of H63D in the pathogenesis of the disease remains controversial, and it might correspond to a minor mutation. Conversely, the C282Y mutation is tightly linked to the disease, as it accounts for 80 to 100% of the hemochromatosis cases in Northern Europe. The lower frequency observed, in the patients, in Italy and South of France led to imagine either the implication of other mutations or of other genes. The C282Y mutation is absent in Asia and Africa and is present in the general population with a decreasing gradient of frequency from Northern to Southern Europe. The prevalence of the disease was usually estimated to be 3% but the observed frequency of the C282Y homozygotes is 5% in our breton population raising the question of the penetrance of the disease, and consequently the use of the genotypic test for its systematic screening. As HFE encodes a membrane protein similar to HLA class I protein, its contribution to iron overload is not obvious. The normal protein is predicted to to be expressed at the cell surface in association with beta 2-microglobulin, a localization for which C282Y is critical as it disrupts this association. This protein has also been shown to form a stable complex with the transferrin receptor leading to a decreased affinity for transferrin. A better knowledge of its function will help to decipher iron and different metal-ions metabolism. Although the exact role of the HFE protein is unknown, the genotypic test allows the clinicians to ascertain their diagnosis and genetic counselling.

[The diagnosis of hemochromatosis in the era of the gene]. / Le diagnostic de l'hémochromatose à l'heure du gène.

The discovery of the hemochromatosis gene has deeply changed and simplified the diagnosis of the disease. In a given individual, establishing the diagnosis relies, from now on, on a simple blood sample showing the couple: elevated transferrin saturation and homozygous C282Y mutation (= C282Y +/+). Liver biopsy should only be performed when iron overload is massive in order to detect cirrhosis (or bridging fibrosis), i.e. in a prognostic view. Practically, liver biopsy is confined to the following two situations: when the C282Y +/+ patient exhibits hepatomegaly and/or an increase in serum transaminases and/or a serum ferritin level above 1,000 micrograms/L; whenever, despite a strong bio-clinical suspicion of iron overload, genetic testing does not show the expected homozygosity for C282Y. At the family level, evaluating the risk for hemochromatosis is now "instantaneous" thanks to genetic testing. One must, however, keep in mind in interpreting the data of the family members that: clinical expression of the homozygous status is not constant; heterozygosity for C282Y does not per se lead to significant iron overload, but may constitute a co-factor exacerbating (or increasing the risk of) other hepatic or non hepatic diseases. Heterozygosity exposes also to the risk of homozygosity among the offspring; this knowledge of C282Y status must be balanced by the negative impact from the standpoint of possible societal genetic discrimination.

Insulin resistance-associated hepatic iron overload.

BACKGROUND & AIMS: Hepatic iron overload has been reported in various metabolic conditions, including the insulin-resistance syndrome (IRS) and nonalcoholic steatohepatitis (NASH). The aim of this study was to show that such hepatic iron overload is part of a unique and unrecognized entity. METHODS: A total of 161 non-C282Y-homozygous patients with unexplained hepatic iron overload were included. We determined the age; sex; presence of IRS (1 or more of the following: body mass index of >25, diabetes, or hyperlipidemia); serum iron tests and liver iron concentration (LIC; reference value, <36 micromol/g); liver function test results; C282Y and H63D HFE mutations; and liver histological status. RESULTS: Patients were predominantly male and middle-aged. Most (94%) had IRS. Transferrin saturation was increased in 35% (median, 42%; range, 13%-94%). LIC ranged from 38 to 332 micromol/g (median, 90 micromol/g), and LIC/age ratio ranged from 0.5 to 4.8 (median, 1.8). Allelic frequencies of both HFE mutations were significantly increased compared with values in normal controls (C282Y, 20% vs. 9%; H63D, 30% vs. 17%), only because of a higher prevalence of compound heterozygotes. Patients with no HFE mutations had similar degrees of iron overload as those with other genotypes, except for compound heterozygotes, who had slightly more iron burden. Steatosis was present in 25% of patients and NASH in 27%. Portal fibrosis (grades 0-3) was present in 62% of patients (grade 2 or 3 in 12%) in association with steatosis, inflammation, and increased age. Sex ratio, IRS, transferrin saturation, and LIC did not vary with liver damage. Serum ferritin concentration, liver function test results, and fibrosis grade were more elevated in patients with steatosis and NASH than in others, but LIC and allelic frequencies of HFE mutations were similar. CONCLUSIONS: This study shows that patients with unexplained hepatic iron overload are characterized by a mild to moderate iron burden and the nearly constant association of an IRS irrespective of liver damage.

Elevated parathyroid hormone 44-68 and osteoarticular changes in patients with genetic hemochromatosis.

OBJECTIVE: To determine whether the osteoarticular changes associated with genetic hemochromatosis could be explained by metabolic parathyroid hormone (PTH) disorders. METHODS: The study involved 210 patients with liver iron overload syndromes. Osteoarticular changes were numerically scored as the number of damaged joints. PTH 1-84 and 44-68 were assayed. RESULTS: An increase in serum PTH 44-68 levels was found in one-third of untreated patients who had no calcium or PTH 1-84 abnormalities. Serum PTH 44-68 levels correlated positively with serum ferritin levels. In multivariate analyses, the number of affected joints correlated positively with age, serum PTH 44-68 levels, and serum ferritin levels. CONCLUSION: Liver iron overload syndromes, especially genetic hemochromatosis, are associated with elevated circulating levels of PTH fragments containing the 44-68 region, which appears to play a role in osteoarticular changes. This increase seems to be a consequence of iron overload.

A genotypic study of 217 unrelated probands diagnosed as "genetic hemochromatosis" on "classical" phenotypic criteria.

BACKGROUND/AIMS: The HFE gene is a crucial candidate gene for hemochromatosis. The aims of this study were to assess the HFE genotypic profile in a large series of unrelated probands diagnosed as having phenotypic hemochromatosis, to characterize the sub-group of patients who were not homozygous for the major C282Y mutation, and to report the iron status of the detected HFE-identical siblings. METHODS: In 217 patients, the phenotypic diagnosis of hemochromatosis was based on strict bioclinical and/or histological criteria, and their genotypic profile (C282Y and H63D mutations) was determined. RESULTS: 1) 209 of the 217 probands were C282Y +/+. In 33 cases, an HFE-identical sibling was identified. Two of them had neither a clinical nor a biochemical phenotypic profile of hemochromatosis in the absence of any external factor which might have attenuated this expression. 2) Eight patients (seven males) were not C282Y +/+. Their genotypic profiles were: (C282Y +/-): six cases (four were H63D +/- and two H63D -/-); (C282Y -/-): two cases (one was H63D +/+, one H63D +/-). Phenotypic expression consisted of six cases of mild liver siderosis (among whom were the four compound heterozygotes and one case of alcoholic cirrhosis) and two severe cases of hepatic iron overload (one with alcoholic cirrhosis). Three HFE-identical siblings were identified, none of them presenting with iron excess. CONCLUSIONS: In our population: 1) The classical phenotypic criteria fitted, in 96.3% of cases, with a homogeneous genotypic entity defined by homozygosity for the C282Y mutation. Incomplete penetrance of the homozygous status was shown by the absence of the hemochromatosis phenotypic profile in 6% of the HFE-identical siblings. 2) A minority (3.7%) were not homozygous for C282Y. These were essentially men with mild iron overload, and might present with distinct iron overload entity(ies) as suggested by the presence in three of an HFE-identical sibling with absence of iron overload.

Phenotypic expression of HFE mutations: a French study of 1110 unrelated iron-overloaded patients and relatives.

BACKGROUND & AIMS: Two mutations have been described in the HFE gene: C282Y and H63D. The aim of this study was to determine the phenotype of the different HFE genotypes. METHODS: Clinical symptoms and iron data were examined according to HFE genotypes in 531 unrelated patients with unexplained liver iron overload and 579 relatives of hemochromatotic patients. RESULTS: Non-C282Y +/+ patients did not markedly differ in terms of iron overload or clinical expression according to genotype, except for compound heterozygotes, who had slightly increased transferrin saturation. This contrasted with the strikingly increased expression in C282Y homozygotes. Similar phenotype/genotype correlations were observed in relatives based on serum iron test results. Family transmission of iron overload linked to HFE was exceptional in non-C282Y +/+ siblings and frequent in C282Y homozygotes. CONCLUSIONS: Iron overload in patients with the non-C282Y +/+ genotype is mild to moderate, strikingly lower than in C282Y homozygotes, and is not influenced by HFE genotype, except, to a small extent, for compound heterozygotes. The role of H63D mutation therefore seems to be marginal.

Non-C282Y familial iron overload: evidence for locus heterogeneity in haemochromatosis.

Haemochromatosis (HC) is an autosomal recessive disease with progressive iron overload leading to midlife onset of clinical complications. The causal gene (HFE) maps to 6p, in close linkage with the HLA class I genes. An HFE candidate gene recently identified has two missense mutations (C282Y and H63D) associated with the disease. Here we document the phenotypic and genetic analysis of a nuclear family comprising two sibs with symptomatic and massive iron overload before the age of 25. The disease seemed to be recessively transmitted and fitted the agreed criteria for haemochromatosis, but was neither associated with the C282Y and H63D mutations nor linked with HLA markers. Our data strongly support locus heterogeneity in haemochromatosis by showing evidence that the gene responsible for juvenile haemochromatosis (JH) does not map to 6p. In the absence of clear cut phenotypic distinction from typical haemochromatosis, patients below 30 years of age and C282Y negative should be considered as putative juvenile cases. This has practical implications in genetic counselling and family management.

Prevalence of the C282Y mutation in Brittany: penetrance of genetic hemochromatosis?

Hemochromatosis (GH) is an inborn error of iron metabolism, characterized by progressive iron loading that, if untreated, causes high morbidity and death. The gene responsible for the disease (HFE), located 4.5 megabases telomeric to the HLA-A locus, encodes a protein homologous to class I MHC molecules. A main mutation, C282Y, has been identified within the gene. Although hemochromatosis is considered as the most frequent inherited disease in the populations of Northern European origin, its prevalence in Brittany had not been evaluated yet. In this issue we report the C282Y mutation frequency in a cohort of 1000 newborns from maternity hospitals of the four breton départements. The homozygote frequency was 5/1000 and heterozygote frequency was 12%; such high frequencies raise the question of the penetrance of the disease and the relevance of systematic genotypic screening for hemochromatosis.

A candidate gene for hemochromatosis: frequency of the C282Y and H63D mutations.

The gene whose alteration causes hereditary hemochromatosis (HFE according to the international nomenclature) was, more than 20 years ago, shown to map to 6p21.3. It has since escaped all efforts to identify it by positional cloning strategies. Quite recently, a gene named HLA-H was reported as being responsible for the disease. Two missense mutations, Cys282Tyr (C282Y) and His63Asp (H63D), were observed, but no proof was produced that the gene described is the hemochromatosis gene. To validate this gene as the actual site of the alteration causing hemochromatosis, we decided to look for the two mutations in 132 unrelated patients from Brittany. Our results indicate that more than 92% of these patients are homozygous for the C282Y mutation, and that all 264 chromosomes but 5 carry either mutation. These findings confirm the direct implication of HLA-H in hemochromatosis.