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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
BACKGROUND
Patients with hepatitis C virus (HCV) and hepatocellular carcinoma (HCC) may or not develop iron overload (IO), which is associated with worst prognosis, because can cause serious damage to organs. HFE gene controls the iron uptake from gut, particularly in patients with hereditary hemochromatosis (HH).
AIM
To identify associations between HFE coding region in patients exhibiting hereditary hemochromatosis and in diseases associated with acquired IO.
METHODS
We sequenced exons 2 to 5 and boundary introns of HFE gene, evaluating all polymorphic sites in patients presenting hereditary (hemochromatosis) or acquired iron overload HCV and HCC) and in healthy controls, using Sanger sequencing. We also determined the ensemble of extended haplotype in healthy control individuals, including several major histocompatibility complex loci, using sequence specific probes. 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.
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 linkage disequilibrium was observed among the H63D-G, IVS2(+4)-C and C282Y-G gene variants, particularly in HH; however, the mutation IVS2(+4)T>C was not directly 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. No LD was observed between HFE alleles and other major histocompatibility loci.
CONCLUSION
A differential HFE association was observed for HH and for diseases associated with acquired IO (HCV, HCC). Since HFE is very distant from other histocompatibility loci, only weak associations were observed with these alleles.
Core tip: Patients with hepatitis C virus (HCV) and hepatocellular carcinoma (HCC) may or not develop iron overload (IO), which is associated with worset prognosis. The sequencing of the HFE gene permitted to assemble the previously described variation sites (H63DC>G-, S65CA>T and C282YG>A) associated with hereditary hemochromatosis into HFE haplotypes, under the standardized HLA nomenclature. A differential association of HFE alleles was observed for hereditary and acquired IO (HCV, HCC). In addition to the HFE gene, we also typed other major histocompatibility loci (HLA-A/-B/-C/ DRB1/-DQB1, and HLA-G 14bp INDEL and TNFa-d microsatellites) in the healthy population to understand how the HFE gene variability is associated with these loci.
