Hemochromatosis: As a conformational disorder

Hereditary hemochroamtosis (HH) refers to a unique clinicopathologic subset of iron overload syndromes that includes the disorder related to C282Y homozygous mutation of the hemochromatosis protein (HFE), the most common form of hereditary hemochromatosis. Recent reports have highlighted analogies with the class of disorders, known as the conformational diseases whereby HFE C282Y mutant protein forms aggregates and is subsequently retained in the endoplasmic reticulum (ER). In conformational disorders, accumulation of unfolded or misfolded proteins in the ER can activate a complex cascade linked to the regulation of diverse physiologic processes, disease onset and progression. To-date, reviews on HFE C282Y HH have largely dealt with the end-stage consequence of this disorder (iron overload). However, our review focuses on upstream molecular events resulting from the mislocalization of the aggregation-prone HFE C282Y protein leading to potential advances in treatment and diagnosis.     

Possible roles of the hereditary hemochromatosis protein, HFE, in regulating cellular iron homeostasis

Hereditary hemochromatosis (HH) is the most common inherited disorder in people of Northern European descent. Over 83% of the cases of HH result from a single mutation of a Cys to Tyr in the HH protein. HFE. This mutation causes a recessive disease resulting in an accumulation of iron in selected tissues. Iron overload damages these organs leading to cirrhosis of the liver, diabetes, cardiomyopathy, and arthritis. The mechanism by which HFE influences iron homeostasis in cells and in the body remains elusive. Lack of functional HFE in humans produces the opposite effects in different cell types in the body. In the early stages of the disease. Kupffer cells in the liver and enterocytes in the intestine cells are iron depleted and have low intracellular ferritin levels, whereas hepatocytes in the liver are iron overloaded and have high intracellular iron levels. This review gives the background and a model as to possible mechanisms of how HFE could exert different effects on iron homeostasis in different cell types.     

Uncommon mutations and polymorphisms in the hemochromatosis gene

Hereditary hemochromatosis (HH) is a common autosomal recessive disorder of iron metabolism. Iron absorption from the gut is inappropriately high, resulting in increasing iron overload. The hemochromatosis gene (HFE) was identified in 1996 by extensive positional cloning by many groups over a period of about 20 years. Two missense mutations were identified. Homozygosity for one of these, a substitution of a tyrosine for a conserved cysteine (C282Y), has now clearly been shown to be associated with HH in 60-100% of patients. The role of the second mutation, the substitution of an aspartic acid for a histidine (H63D), is not so clear but compound heterozygotes for both these mutations have a significant risk of developing HH. Here we review other putative mutations in the HFE gene and document a number of diallelic polymorphisms in HFE introns.     

Nijmegen breakage syndrome (NBS)

Most inborn errors of metabolism (IEM) are recessive, genetically transmitted diseases and are classified into 3 main groups according to their mechanisms: cellular intoxication, energy deficiency, and defects of complex molecules. They can be associated with endocrine manifestations, which may be complications from a previously diagnosed IEM of childhood onset. More rarely, endocrinopathies can signal an IEM in adulthood, which should be suspected when an endocrine disorder is associated with multisystemic involvement (neurological, muscular, hepatic features, etc.). IEM can affect all glands, but diabetes mellitus, thyroid dysfunction and hypogonadism are the most frequent disorders. A single IEM can present with multiple endocrine dysfunctions, especially those involving energy deficiency (respiratory chain defects), and metal (hemochromatosis) and storage disorders (cystinosis). Non-autoimmune diabetes mellitus, thyroid dysfunction and/or goiter and sometimes hypoparathyroidism should steer the diagnosis towards a respiratory chain defect. Hypogonadotropic hypogonadism is frequent in haemochromatosis (often associated with diabetes), whereas primary hypogonadism is reported in Alstr?m disease and cystinosis (both associated with diabetes, the latter also with thyroid dysfunction) and galactosemia. Hypogonadism is also frequent in X-linked adrenoleukodystrophy (with adrenal failure), congenital disorders of glycosylation, and Fabry and glycogen storage diseases (along with thyroid dysfunction in the first 3 and diabetes in the last). This is a new and growing field and is not yet very well recognized in adulthood despite its consequences on growth, bone metabolism and fertility. For this reason, physicians managing adult patients should be aware of these diagnoses.     

Detection of the Hereditary Hemochromatosis Gene Mutation by Real-Time Fluorescence Polymerase Chain Reaction and Peptide Nucleic Acid Clamping

Hereditary hemochromatosis (HH), an iron overload disease, is the most common known inheritable disease. The most prevalent form of HH is believed to be the result of a single base-pair mutation. We describe a rapid homogeneous mutation analysis method that does not require post-polymerase chain reaction (PCR) manipulations. This method is a marriage of three emerging technologies: rapid cycling PCR thermal cyclers, peptide nucleic acid (PNA) probes, and a new double-stranded DNA-selective fluorescent dye, Sybr Green I. The LightCycler is a rapid thermal cycler that fluorometrically monitors real-time formation of amplicon with Sybr Green I. PNAs are DNA mimics that are more sensitive to mismatches than DNA probes, and will not serve as primers for DNA polymerases. PNA probes were designed to compete with PCR primers hybridizing to the HH mutation site. Fully complemented PNA probes at an 18:1 ratio over DNA primers with a mismatch result in suppression of amplicon formation. Conversely, PNA probes with a mismatch will not impair the binding of a complementary primer, culminating in amplicon formation. A LightCycler-based rapid genetic assay has been developed to distinguish HH patients from HH carriers and normal individuals using PNA clamping technology.     

Hfe acts in hepatocytes to prevent hemochromatosis

Hereditary hemochromatosis (HH) is a prevalent, potentially fatal disorder of iron metabolism hallmarked by intestinal hyperabsorption of iron, hyperferremia, and hepatic iron overload. In both humans and mice, type I HH is associated with mutations in the broadly expressed HFE/Hfe gene. To identify where Hfe acts to prevent HH, we generated mice with tissue-specific Hfe ablations. This work demonstrates that local Hfe expression in hepatocytes serves to maintain physiological iron homeostasis, answering a long-standing question in medicine and explaining earlier clinical observations.     

Frequency of HFE H63D, S65C, and C282Y mutations in patients with iron overload and controls from Toledo, Spain

Hereditary hemochromatosis (HH) is an autosomal recessive disease caused by a defective iron absorption. C282Y is the most frequent HFE gene mutation causing HH in Northern European populations and their descendants. However, two other mutations, H63D and S65C, have been described as pathogenic changes. In this study, we have tried to evaluate the frequency of these three mutations in our community. Eighty-three patients with clinical and/or biochemical features of hemochromatosis and 150 controls were screened for H63D, S65C, and C282Y mutations using a PCR-restriction fragment length polymorphism (RFLP)-based strategy. In contrast to previous studies, 7% of the patients were homozygous for C282Y mutation. The remaining patients were 20% H63D homozygous, 10% H63D/C282Y compound heterozygous, 1% H63D/S65C compound heterozygous, 22% H63D heterozygous, 2% C282Y heterozygous, 2% S65C heterozygous, and 36% of patients lacked any of the three mutations studied, despite the fact that they showed clinical/biochemical features of hemochromatosis. We observed a high frequency of the H63D mutation in both the control group and patients, whereas the main genotypes implicated in HH in our series were H63D homozygous and H63D/C282Y compound heterozygous. We propose that the H63D mutation be analyzed in HH patients from our geographic area. Moreover, further studies are needed to elucidate the role of this mutation in the development of HH and the genetic, environmental or other factors that affect the genotype-phenotype correlation between H63D and hemochromatosis.     

Frequency of hemochromatosis C282Y and H63D mutations in Sardinia

Hereditary hemochromatosis (HH) is one of the most common autosomal recessive disorders of iron metabolism among Caucasians, and it is associated with C282Y mutation of the HFE gene in populations of Celtic origins. A second mutation, H63D, shows a very high widespread frequency, although its role in iron metabolism is still inconclusive. There are no data on the frequencies of these two mutations in Sardinia, an island in the Mediterranean sea that has not been invaded by Celtic peoples. We examined 836 chromosomes from Sardinian subjects and tested for the mutation by restriction enzyme digestion of PCR products. Among the 836 analyzed chromosomes, we found a C282Y allele frequency of 0.0036 and an H63D allele frequency of 0.173. These data could explain the observed rarity of HH in Sardinia. The high allele frequency of H63D and the rarity of HH in Sardinia is suggestive that this mutation is not a major contributor to this disease.     

Population genetics of hereditary hemochromatosis in Saguenay Lac-Saint-Jean (Quebec, Canada).

Hereditary hemochromatosis (HH) is an autosomal recessive disorder that has a high prevalence in Caucasian populations. Based on HLA typing in 18 families, the gene frequency was estimated 0.12. The homozygote frequency was 0.014 and the heterozygote frequency was 0.21 in Saguenay Lac-Saint-Jean (SLSJ), a geographically isolated region of northeastern Quebec. The genealogical reconstruction showed that 15 of the 57 obligate carriers of the HH gene could be traced back to a unique ancestor in the 18th century. The mean coefficients of inbreeding and kinship were 17 and 15 times, respectively, higher in the HH group than in three control groups. The values of both coefficients were much higher than those found in other HH populations and in most of the other recessive disorders prevalent in SLSJ.     

A reverse-hybridization assay for the rapid and simultaneous detection of nine HFE gene mutations

Hereditary hemochromatosis (HH) is a very common autosomal recessive disorder of iron metabolism and frequently associated with mutations in the HFE gene. Molecular genetic testing for HFE mutations is considered valuable for carrier identification, as well as for early diagnosis of the disease, allowing simple treatment by phlebotomy and normal survival of patients. We have developed a reverse-hybridization assay for the routine diagnosis of eight previously described and one novel (E168Q) HFE point mutations. The test is based on multiplex DNA amplification and ready-to-use membrane teststrips, which contain oligonucleotide probes for each wild-type and mutated allele immobilized as an array of parallel lines. The procedure is rapid and accessible to automation on commercially available equipment, and by adding new probes the teststrip can easily be adapted to cover an increasing number of mutations.     

Allele frequencies of hereditary hemochromatosis gene mutations in a local population of west Brittany

Hereditary hemochromatosis (HH) gene mutations, C282Y and H63D, have been screened in a cohort of 254presumably healthy persons originating from a western region of France.The carrier frequencies of these mutations and the incidence of HH have been estimated and compared with those of other studies. This cohort contains two C282Y/C282Y genotypes and has the highest C282Y heterozygosity frequency (17.46%) ever reported. Received: 25 May 1997 / Accepted: 19 November 1997     

血色病的临床与基础研究进展

遗传性血色病（Hereditary Hemochromatosis,HH）是一种西方常见的遗传性铁过载性疾病。目前已知的血色病基因主要包括剧硒、YfR2、HJV、FPN及HAMP。这些基因突变导致大量铁离子逐渐沉积在肝、心、胰腺等脏器的实质细胞,造成组织纤维化和结构改变,最终引起器官功能障碍和衰竭,常见症状有肝硬化、肝癌、糖尿病、心力衰竭、垂体及性腺功能减退、关节疾病和皮肤色素沉着等。当前,机体铁代谢分子机制研究的飞速发展,为深入了解血色病带来了契机。综合铁代谢研究领域最新进展,着重对血色病发展历程、发病机制、临床表现、诊断、治疗及中国血色病现状等方面展开综述。     

Olfactory function in patients with hypogonadotropic hypogonadism: an all-or-none phenomenon?

Hypogonadotropic hypogonadism (HH) refers to an endocrine defectof hypothalamic origin resulting in gonadal hypoplasia and frequentlyassociated with anosmia or severely impaired olfactory function(Kallmann's syndrome). This apparently results from a disruptionin the migration of neurons from the olfactory placode to thebulb and hypothalamus early in development, and so providesa unique opportunity to investigate olfactory function in humansubjects with congenitally incomplete peripheral systems. Olfactoryperformance in 37 HH patients and 37 age-matched controls wascompared using a modified version of the Munich Olfaction Test.This test is based on the sniff-bottle method and includes testsof (i) odor quality discrimination, (ii) intensity discrimination,(iii) detection thresholds, and (iv) recognition, hedonic evaluationand identification ability. The patients could be divided intotwo distinct groups differing significantly on all four subtestsand showing no overlap in performance: 20 anosmics, conformingto Kallmann's syndrome, and 17 apparent normosmics whose performancewas slightly poorer, but not significantly different to thatof the controls. The unexpected failure to find a continuumof olfactory dysfunction now raises the question whether HHwith or without anosmia represents two syndromes with distinctetiologies, or rather reflects the ability of the olfactorysystem to function well despite morphological impairment.     

Hereditary hemochromatosis: the clinical significance of the S65C mutation

Hereditary hemochromatosis (HH) is a common genetic disease with iron overload in certain organs, especially the liver. Most cases are homozygous for the C282Y mutation in the HFE gene; a few are C282Y heterozygous, compound C282Y/H63D heterozygous, or have no known mutation. A third mutation, S65C, has been associated with HH, but this finding is disputed. We have studied the clinical significance of various genotypes with the S65C mutation. In a population-based screening for HH in 65,238 persons, 613 had high serum transferrin saturation in two blood samples and were invited for HFE genotyping. In 556 persons with complete data sets, we studied the serum ferritin concentration and the risk of being diagnosed with phenotypic HH in the various genotypic groups. The phenotypic diagnosis was given without knowing the genotypic result. Except for the C282Y homozygotes, no differences in median serum ferritin concentrations were found between the various genotypic groups. However, the C282Y/S65C compound heterozygous group had a higher risk of being diagnosed with phenotypic HH than the wild-type group, as did the C282Y homozygous and the C282Y/H63D compound heterozygous groups. When combined with the C282Y mutation, the S65C mutation is associated with an increased risk of being diagnosed with phenotypic HH.     

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.     

Hereditary hemochromatosis in a Brazilian university hospital in São Paulo State (1990-2000)

Hereditary hemochromatosis (HH) is the most common genetic disease among individuals of European descent. Two mutations (845G-->A, C282Y and 187C-->G, H63D) in the hemochromatosis gene (HFE gene) are associated with HH. About 85-90% of patients of northern European descent with HH are C282Y homozygous. The prevalence of HH in the Brazilian population, which has a very high level of racial admixture, is unknown. The aims of the present study were to identify individuals with diagnostic criteria for HH among patients with a body iron overload attended at the university hospital of the Faculty of Medicine of Ribeirao Preto from 1990 to 2000, and to evaluate the prevalence of HFE mutations. We screened first-degree relatives for HFE mutations. Four of 72 patients (three men and one woman, mean age 47 years) fulfilled the criteria for HH. HFE mutations were studied in three patients [two C282Y homozygotes (patients 1 and 2) and one H63D heterozygote]. Patient 1 had four children (all C282Y heterozygotes with no iron overload) and seven brothers and sisters: two sisters (66 and 76 years old) were C282Y homozygotes and both had an iron overload (a liver biopsy in one showed severe iron deposits), one sister (79 years old) was a compound heterozygote with no iron overload, one brother (78 years old) was a C282Y heterozygote with no iron overload, two individuals were H63D heterozygotes (one brother, 49 years old, obese, with a body iron overload and abnormal liver enzymes - a biopsy showed non-alcoholic steatohepatitis, and one 70-year-old sister with no iron overload). Patient 2 had two children (22 and 24 years old who were C282Y heterozygotes with no iron overload) but no brothers or sisters. These results showed that HH was uncommon among individuals attended at our hospital, although HFE mutations were found in all patients. Familial screening is valuable for the early diagnosis of individuals at risk since it allows treatment to be initiated before the onset of the clinical manifestations of organ damage associated with HH.     

GnRH deficiency: new insights from genetics

The acquisition of a sexually dimorphic phenotype is a critical event in mammalian development. Hypogonadotropic hypogonadism (HH) results from impaired secretion of GnRH. The patients display with delayed puberty, micropenis and cryptorchidism in the male reflecting gonadotropin insufficiency, and amenorrhea in the female. Kallmann's syndrome (KS) is defined by the association of HH and anosmia or hyposmia (absent smelling sense). Segregation analysis in familial cases has demonstrated diverse inheritance patterns, suggesting the existence of several genes regulating GnRH secretion. The X-linked form of the disease was associated with a genetic defect in the KALI gene located on the Xp22.3 region. KAL1 gene encodes an extracellular matrix glycoprotein anosmin-1, which facilitates neuronal growth and migration. Abnormalities in the migratory processes of the GnRH neurons with the olfactory neurons explain the association of HH with anosmia. Recently, mutations in the FGF recepteur 1 (FGFR1) gene were found in KS with autosomal dominant mode of inheritance. The role of FGFR1 in the function of reproduction requires further investigation. Besides HH with anosmia, there are isolated HH (IHH). No human GnRH mutations have been reported although hypogonadal mice due to a GnRH gene deletion exist. In patients with idiopathic HH and without anosmia an increasing number of GnRH receptor (GnRHR) mutations have been described which represent about 50% of familial cases. The clinical features are highly variable and there is a good relationship between genotype and phenotype. A complete loss of function is associated with the most severe phenotype with resistance to pulsatile GnRH treatment, absence of puberty and cryptorchidism in the male. In contrast, milder loss of function mutations causes incomplete failure of pubertal development. The preponderant role of GnRH in the secretion of LH by the gonadotrophs explains the difference of the phenotype between male and female with partial GnRH resistance. Affected females can have spontaneous telarche and normal breast development while affected males exhibit no pubertal development but normal testis volume, a feature described as "fertile-eunuch". High-dose pulsatile GnRH has been used to induce ovulation. Another gene, called GPR54, responsible for idiopathic HH has been recently described by segregation analysis in two different consanguineous families. The GPR54 gene is an orphan receptor, and its putative ligand is the product of the KISS-1 gene, called metastine. Their roles in the function of reproduction are still unknown.     

Hypogonadotropic hypogonadism due to GnRH receptor mutation in a sibling

Hypogonadotropic hypogonadism (HH) is characterised by delayed puberty and infertility. Congenital HH comprises Kallmann syndrome with hypo-/anosmia and idiopathic HH (IHH). The genetic origin remains unknown in most cases, but the defective GnRH receptor gene (GNRHR) accounts for a considerable proportion of IHH. Here we describe a pair of siblings diagnosed with IHH. Aged 17 years, the boy was referred because of short stature (162 cm) and overweight (62.5 kg). He presented no signs of puberty, bone age of 14.5 years and insulin resistance. His sister, aged 16 years, also displayed delayed puberty. She was 166 cm tall and weighed 52 kg; her bone age was 12.5 years. Pelvic ultrasonography showed an infantile uterus and fibrous ovaries. In both siblings, serum gonadotropins were extremely low, and non-responsive to GnRH. Testosterone (1.38 nmol/l) and IGF1 (273 ng/ml) were decreased in the boy, although the girl did not present IFG1 deficiency. Her serum oestradiol was 10 pg/ml. MRIs of the hypothalamo-pituitary region and olfactory bulbs revealed them to be normal. The patients' sense of smell was unaltered. Their parents appeared to be first degree cousins. Considering the clinical data and potentially autosomal recessive HH transmission, the GNRHR gene was screened. The siblings turned out to be homozygous for the G416A transition, which had previously been identified in other HH individuals. The parents were heterozygous mutation carriers. The proband, moderately responding to LH, was started on low dose testosterone replacement, and his sister on transdermal oestradiol. Molecular data indicative of GnRH resistance could guide their future therapy should they desire fertility restoration. Further observations of the male patient may provide insights into androgen's influence on body mass, growth and insulin sensitivity.