Analysis of the HFE gene (H63D and C282Y) mutations in patients with iron overload, family members and controls from Antioquia, Northwest Colombia

To the Editor: Hereditary hemochromatosis (HH; OMIM 235200) is an autosomal recessive genetic disorder characterized by excessive accumulation of iron in many tissues of the body. Consequently, the disorder manifests clinically as cirrhosis, carcinoma of the liver, diabetes mellitus, bronze pigmentation of the skin, hypogonadism, and chronic fatigue. Classical HH is linked to H63D and C282Y mutations in the HFE gene (1). Published data suggest that those mutations have an unequal worldwide distribution profile. However, no data are available in Colombia (South America) to determine whether clinical cases of iron overload are associated to new mutations in the HFE gene, H63D and C282Y mutations, or mutations in any other gene. The Antioquia region in northwest Colombia has approximately 5,000,000 inhabitants. This population (paisa community) was established in the middle of the sixteenth and/or early seventeenth century and grew in relative isolation until the late nineteenth century. Genetic evidence suggests that this community originated predominantly from southern/northern Spain and Sephardic Jewish population (2). In this study, 13 index cases (2 females and 11 males, average age 52.3 12.8 years; range from 42 to 83 years) of iron overload, 28 family members from three probands (19 females and 9 males), and 190 unrelated individuals (98 females and 92 males) of Antioquian ethnicity were tested for the HFE H63D (nucleotide change c.187 C.G) and C282Y (c.845 G.A) mutations by polymerase chain reaction (PCR). Amplification mixtures were digested with MboI (exon 2) and RsaI (exon 4). Digestions were analyzed by restriction fragment length polymorphism (RFLP) as published elsewhere (3). Patients were selected from University Hospital San Vicente de Paul, University of Antioquia. This study was approved by the University of Antioquia Ethics Committee and financially supported by Colciencias grant 1115-04-16337 to C. V. P. Written informed consent was obtained from all subjects. Nine out of 13 index cases of iron overload were found to carry the H63D (one heterozygote) and C282Y (seven homozygotes and one heterozygote) mutations in theHFE gene. Although the number of patients is small, identification of HFEmutations reveals their common presence in the paisa community, as described in other racially diverse populations (4). Indeed, most patients were C282Y homozygotes, reflecting a 57.7% genotype frequency (one in two approximately). This result is similar to that reported in a group of 35 patients (57.1%) from a Basque population (5). Interestingly, the H63D (case 7184) and C282Y (case 6656) heterozygotes displayed the typical features of classical HH. These observations suggest that other genetic and environmental factors might influence the severity of the disorder. Alternatively, heterozygotes may present additional genetic mutation(s) in the HFE gene not detected by PCR/RFLP methods. Thus, our data suggest that genotyping by this technique might be highly informative and in most cases is confirmatory of the diagnosis of HH, but for definitive genetic implication of other mutations or variants in HH, DNA sequencing of the entire HFE gene should be performed, although this method is not cost-effective. In conjunction, these observations comply with the notion that simple C282Y heterozygous virtually never cause phenotypic HH. Family screening of index cases is necessary to establish the expression variability of these mutations. Of the 28 family members screened for mutations, two homozygous (two females) and nine heterozygous (six female and three male) for the C282Y alteration and four heterozygous (three female and one male) for the H63D