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Master's Dissertation
DOI
https://doi.org/10.11606/D.99.2016.tde-17022016-104522
Document
Author
Full name
Jerenice Esdras Ferreira
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2015
Supervisor
Committee
Sabino, Ester Cerdeira (President)
Almeida Neto, César de
Levi, José Eduardo
Title in Portuguese
Avaliação de polimorfismos do gene SLC40A1 em pacientes com hemocromatose hereditária e adquirida
Keywords in Portuguese
Ferro
Hemocromatose
Metabolismo (Alteração)
Mutação
Polimorfismo
Proteínas
Abstract in Portuguese
A hemocromatose é uma desordem caracterizada por armazenamento alterado de ferro em órgãos vitais. A hemocromatose hereditária pode ser causada por mutações nos genes HFE, SLC40A1, HAMP, HJV e TfR2. A forma adquirida da doença pode ser ocasionada por sobrecarga de ferro, alcoolismo, infecção pelo vírus da hepatite C, anemias hemolíticas e doença hepática crônica. O objetivo deste estudo foi descrever as variantes do gene SLC401 em uma série de casos de pacientes em seguimento no ambulatório de Hematologia do Hospital das Clínicas da FMUSP com hemocromatose hereditária e adquirida e verificar se estas variantes poderiam estar associadas a um pior quadro de hemocromatose. No período de 2008 a 2011 foram coletadas 54 amostras de pacientes com diagnóstico prévio de hemocromatose, acompanhados no ambulatório de anemias do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. Os pacientes foram separados, de acordo com a suspeita clínica, em dois grupos: 38 pacientes com Hemocromatose Hereditária e 16 pacientes com Hemocromatose Adquirida. O DNA foi extraído de sangue periférico pelo kit Qiamp Blood, e as amplificações dos exomas descrito por Wakusawa (2005). As reações de sequenciamento foram realizadas no aparelho AB 3130 e as sequências do gene editadas, alinhadas no programa Sequencer e comparadas com GenBank. Os pacientes com hemocromatose hereditária e adquirida (secundária) apresentaram uma frequência acima de 45% entre a faixa etária de 40 a 60 anos, mostrando uma sobrecarga de ferro mais tardia. O gênero masculino apresentou maior frequência em ambos os grupos (superior a 60%), já em relação à etnia/cor da pele, mais de 80% eram brancos. No diagnóstico laboratorial de ferritina, ferro, CTLF e IST dos pacientes com hemocromatose hereditária e adquirida (secundária) mostra que não houve diferenças significativas. As mutações no gene HFE nos pacientes com hemocromatose hereditária foram C282Y homozigose (21%) e na adquirida (secundária) H63D heterozigose (25%). Entre as mutações do gene HFE, não houve diferença estatística significativa nos grupos. A mutação foi identificada no éxon 3 do gene SLC40A1, no códon 80, descrita p.I80L (c.586C>A). No éxon 4 do gene SLC40A1 foram identificados 3 polimorfismos com a substituição de um nucleotídeo nas posições: c. A630G (190437579), c.648A>G (190437597), c.738Y>C (190437688). Já no éxon 6 foram identificados dois polimorfismos c.1014C>T (190430229) e c.1014Y>T (190430229). As mutações encontradas no gene HFE e SLC40A1 estão associadas com a sobrecarga férrica caracterizando a hemocromatose hereditária. Os polimorfismos encontrados no gene SLC40A1 foram identificados nos pacientes com hemocromatose hereditária e adquirida (secundária) numa frequência de 50%, semelhante aos doadores de sangue, sugerindo que este polimorfismo provavelmente não contribui para o desenvolvimento da doença.
Title in English
Polymorphisms evaluation of SLC40A1 gene in patients with Hereditary Hemochromatosis and Acquired
Keywords in English
Hemochromatosis
Iron
Metabolism (Alteration)
Mutation
Polymorphism
Proteins
Abstract in English
Hemochromatosis is a disorder characterized by altered storage iron in vital organs. Hereditary hemochromatosis can be caused by mutations in the HFE gene, SLC40A1, HAMP, and HVJ TFR2. The acquired form of the disease can be caused by iron overload, alcoholism, infection by the hepatitis C virus, hemolytic anemias and chronic liver disease. The objective of this study was to describe variants of the SLC401 gene in a number of cases of patients being followed at hematology clinic Hospital das Clínicas FMUSP with hereditary hemochromatosis and acquired and verify that these variants could be associated with a worse hemochromatosis frame. In the period 2008 to 2011 were collected 54 samples from patients previously diagnosed with hemochromatosis, anemia accompanied the clinic of the Hospital das Clinicas, Faculdade de Medicina, Universidade de Sao Paulo. The patients were divided, according to clinical suspicion, into two groups: 38 patients with Hereditary Hemochromatosis and 16 patients with Acquired Hemochromatosis. The DNA was extracted from peripheral blood by Qiamp Blood Kit, and amplifications of exomas described by Wakusawa (2005). The sequencing reactions were performed in AB 3130 apparatus and the sequences of the gene edited, aligned in Sequencer program and compared with GenBank. Patients with hereditary hemochromatosis and acquired (secondary) had a frequency above 45% between the age group 40-60 years showing a later iron overload. The males showed higher frequency in both groups (over 60%), as compared to ethnicity / skin color, over 80% were white. In the laboratory diagnosis of ferritin, iron, TIBC and STI patients with hereditary hemochromatosis and acquired (secondary) shows no significant differences. Mutations in the HFE gene in patients with hereditary hemochromatosis C282Y homozygosity were (21%) and acquired (secondary) H63D heterozygosity (25%). Between HFE mutations, there was no statistically significant difference in the groups. The mutation was identified in exon 3 of the SLC40A1 gene at codon 80, described p.I80L (c.586C> A). In exon 4 of the SLC40A1 gene were identified polymorphisms 3 with substitution of a nucleotide in position: c. A630G (190,437,579), c.648A> L (190,437,597), c.738Y> C (190 437 688). Already in exon 6 were identified two polymorphisms c.1014C> T (190 430 229) and c.1014Y> T (190 430 229). The mutations found in the HFE gene and SLC40A1 are associated with iron overload characterizing hereditary hemochromatosis. The polymorphisms found in the SLC40A1 gene have been identified in patients with hereditary hemochromatosis and acquired (secondary) at a frequency of 50%, similar to blood donors, suggesting that this polymorphism will probably not contribute to the development of the disease.
 
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Publishing Date
2016-02-18
 
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