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©The Author(s) 2019. Published by Baishideng Publishing Group Inc. All rights reserved.
Comprehensive analysis of HFE gene in hereditary hemochromatosis and in diseases associated with acquired iron overload
Wagner Narciso de Campos, Juliana Doblas Massaro, Eduardo Luiz Rachid Cançado, Cláudia Emília Vieira Wiezel, Aguinaldo Luiz Simões, Andreza Correa Teixeira, Fernanda Fernandes de Souza, Celso Teixeira Mendes-Junior, Ana de Lourdes Candolo Martinelli, Eduardo Antônio Donadi
Wagner Narciso de Campos, Juliana Doblas Massaro, Eduardo Antônio Donadi, Division of Clinical Immunology, Department of Medicine, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto 14048-900, Brazil
Eduardo Luiz Rachid Cançado, Department of Gastroenterology, Clinical Gastroenterology and Clinical Hepatology of Clinical Hospital, University of São Paulo School of Medicine, São Paulo 01329-000, Brazil
Cláudia Emília Vieira Wiezel, Aguinaldo Luiz Simões, Department of Genetics, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto 14048-900, Brazil
Andreza Correa Teixeira, Fernanda Fernandes de Souza, Ana de Lourdes Candolo Martinelli, Division of Gastroenterology, Department of Medicine, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto 14048-900, Brazil
Celso Teixeira Mendes-Junior, Departamento de Química, Laboratório de Pesquisas Forenses e Genômicas, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto 14040-901, Brazil
Author contributions: de Campos WN and Massaro JD contributed equally in the production of the paper; Campos WN performed all experiments and statistical analysis, wrote the final manuscript; Massaro JD wrote the initial project, standardized HLA-HFE procedures, performed the statistical analysis, helped on the writing of the final manuscript; Cançado ELR selected and provided patients; Wiezel CEV performed cloning experiments for defining HFE alleles; Simões AL provided facilities to perform the HFE cloning experiments; Teixeira AC selected and provided patients; Souza FF selected and provided patients; Mendes-Junior CT helped on statistical analyses; Martinelli ALC responsible for the outpatients followed-up at gastroenterology clinics; Donadi EA mentor of the research, provided financial support, technical facilities and performed final review.
Supported by: “Conselho Nacional de Desenvolvimento Cientifico e Tecnologico”(CNPq, Brazil), No. 304931/2014-1 and No. 148638/2010-4.
Institutional review board statement: This study was approved by the local Ethics Research Committee, process number HCRP 4822/2011. The samples are deposited in the Bank of Samples of the Nucleus in Research in Immunogenetics (BAMPI), process number HCFMRP 3530/2007, under the coordination of Eduardo Antônio Donadi, and in the Bank of Samples HCFMRP 3416/2003, under the responsibility of Ana de Lourdes Candolo Martinelli.
Informed consent statement: All individuals gave their informed consent to participate in the study.
Conflict-of-interest statement: The authors declare that there are no competing interests associated with the manuscript.
STROBE statement: The STROBE Statement has been adopted.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Corresponding author: Eduardo Antônio Donadi, MD, PhD, Associate Professor, Division of Clinical Immunology, Department of Medicine, Ribeirão Preto Medical School, University of São Paulo, Avenida Bandeirantes 3900, Monte Alegre, Ribeirão Preto 14048-900, Brazil. eadonadi@fmrp.usp.br
Telephone: +55-16-36022566 Fax: +55-16-36020229
Received: December 8, 2018
Peer-review started: December 10, 2018
First decision: January 5, 2019
Revised: January 17, 2019
Accepted: January 26, 2019
Article in press: January 26, 2019
Published online: February 27, 2019
ARTICLE HIGHLIGHTS
Research background
HFE gene controls the iron uptake from gut, and defects of the encoded molecule have been associated with iron overload (IO), particularly in hemochromatosis hereditary (HH), which can cause serious damage to the liver. Besides HH, patients with hepatitis C virus (HCV) and hepatocellular carcinoma (HCC) may or not develop IO.
Research motivation
The search for markers associated with IO may be very useful for the early diagnosis of these patients, which is essential for their survival.
Research objectives
The main objectives of this work is to identify associations between HFE coding region variable sites in patients exhibiting HH and in diseases associated with acquired IO.
Research methods
We sequenced exons 2 to 5 and boundary introns of the HFE gene to evaluate all polymorphic sites in patients presenting HH or acquired IO (HCV and HCC), and in healthy controls, using Sanger sequencing. We also determined the extended haplotype in healthy controls, including other major histocompatibility genes (HLA-A/-B/-C/-DRB1/-DQB1 alleles, and HLA-G 14bp INDEL and TNFa-d microsatellites). Haplotype reconstruction was performed using the Arlequin and Phase softwares, and linkage disequilibrium (LD) between histocompatibility loci and HFE gene was performed using the Haploview software.
Research results
The HFE*003 allele was overrepresented (f = 71%) and HFE*001 allele was underrepresented (f = 14%) in HH patients compared to all groups. A strong LD was observed among the previously reported H63D-G, IVS2(+4)-C and C282Y-G gene variants, particularly in HH; however, the mutation IVS2(+4)T>C was not associated with HH susceptibility. The HFE*001/HFE*002 genotype conferred susceptibility to HCC in HCV patients exhibiting IO (P = 0.02, OR = 14.14). Although HFE is telomeric to other histocompatibility genes, the H63D-G/IVS2(+4)-C (P ≤ 0.00001/P ≤ 0.0057) combination was in LD with HLA-B*44 allele group in healthy controls.
Research conclusions
This study systematically evaluated variation sites along the HFE gene using the HLA official nomenclature, previously described by our group. The HFE*003 allele that was overrepresented in HH patients encompasses major variation sites previously described in association with HH in several worldwide populations, in contrast with the HFE*001 allele which does not present HH-associated variation sites and predominates among healthy controls. On the other hand, the HFE*001/HFE*002 genotype was identified as a risk factor for HCC and HCV patients exhibiting IO. Although the HFE gene is distant from other histocompatibility genes, the HFE H63D-G/IVS2(+4)-C alleles were in weak LD with the HLA-B*44 allele. Thus, a differential HFE association was observed for HH and for diseases associated with acquired IO (HCV, HCC).
Research perspectives
Besides the identification of markers associated with IO, which may permit an early detection of patients prone to develop iron deposits, the knowledgement of the major gene associated with iron uptake may help on the understanding of the IO pathogenesis.
