Avaliação dos níveis da citocina pró-inflamatória fator de necrose tumoral-α em pacientes com diabetes tipo 2 e sua relação com variações genéticas de enzimas antioxidantes e obesidade / Evaluation of proinflammatory cytokine tumor necrosis factor-α levels in obese vs nonobese patients with type 2 diabetes and their relationship to CAT C-262T polymorphism

Introdução: Diabetes tipo 2 (DM2) é um distúrbio multifatorial caracterizado pelo aumento dos níveis de radicais livres. Tanto o estresse oxidativo quanto a obesidade contribuem para um estado inflamatório da doença, principalmente pelo aumento da citocina TNF-α. Sabendo-se que a genética individual pode contribuir para o estresse oxidativo, o estudo avaliou o impacto das variações genéticas de enzimas antioxidantes C262T no gene CAT e polimorfismos nulos dos genes GSTM1 e GSTT1 nos níveis de TNF-α, assim como, avaliou se as variantes genéticas atuariam sinergicamente com a obesidade aumentando os níveis da citocina em diabéticos da Grande Vitória/ES, Brasil.Métodos: O polimorfismo no gene CAT foi avaliado pela técnica PCR/RFLP e nos genes GSTM1 e GSTT1 por PCR multiplex, em 56 pacientes, sendo 28 obesos e 28 não obesos. Níveis de TNF-α foram medidos pela técnica de ELISA sanduíche.Resultados: Frequências das variantes nulas de GSTM1 e GSTT1 foram 44,6% e 17,9%, respectivamente. As frequências genotípicas C262T-CAT foram 73,2%, 25% e 1,8% para homozigoto normal, heterozigoto e homozigoto polimórfico, respectivamente. Não houve associação entre genótipos polimórficos e aumento dos níveis de TNF-α, assim como, não foi demonstrado aumento significante da citocina quando avaliado o sinergismo entre obesidade e genética individual do paciente.Conclusão: Níveis de TNF-α não se elevam em diabéticos tipo 2 na presença dos polimorfismos nos genes CAT, GSTM1 e GSTT1, e a obesidade não atua no aumento dessa citocina na população estudada, separadamente ou em conjunto com a genética individual de variantes nos genes CAT, GSTM1 e GSTT1.

Introduction: Type 2 diabetes is a multifactorial disorder characterized by increased levels of free radicals. Both oxidative stress and obesity contribute to an inflammatory state of the disease, mainly by increasing the levels of the proinflammatory cytokine tumor necrosis factor-α (TNF-α). Considering that personal genetics may contribute to oxidative stress, this study assessed the impact of CAT C-262T polymorphism and GSTM1 and GSTT1 null polymorphisms on TNF-α levels in patients with type 2. diabetes. The study also evaluated whether the genetic variants act synergistically with obesity to increase TNF-α levels in patients with diabetes from Grande Vitória, Brazil.Methods: Fifty-six patients were included, of whom 28 were obese and 28 were nonobese. The CAT gene polymorphism was assessed using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method, whereas GSTM1 and GSTT1 polymorphisms were assessed using multiplex PCR. TNF-α levels were measured using the sandwich ELISA technique.Results: Frequencies of GSTM1 and GSTT1 null polymorphisms were 44.6% and 17.9%, respectively. The genotype frequencies of CATC-262T polymorphism were 73.2%, 25.0%, and 1.8% for normal homozygote, heterozygote, and polymorphic homozygote, respectively. Polymorphic genotypes were not associated with increased TNF-α levels, and there was no significant increase in TNF-α levels when evaluating the synergism between obesity and personal genetics.Conclusion: The presence of CAT, GSTM1, and GSTT1 gene polymorphisms was not associated with increased TNF-α levels in patients with type 2 diabetes. Obesity alone or combined with personal genetics of CAT, GSTM1, and GSTT1gene polymorphisms did not promote increased TNF-α levels in the study population.

Influência de polimorfismos nos genes antioxidantes GSTM1, GSTT1 e GSTP1 no desenvolvimento de complicações oftalmológicas em diabéticos tipo 2 / Influence of polymorphisms in the GSTM1, GSTT1, and GSTP1 antioxidant genes on the development of ophthalmic complications in persons with type 2 diabetes

Introdução: A diabetes tipo 2 (DM2) é uma desordem metabólica ocasionada pela disfunção das células beta pancreáticas que interferem na produção de insulina e/ou pela resistência dos órgãos alvos a esse hormônio. Níveis elevados de radicais livres em conjunto com o declínio das defesas antioxidantes presente na DM2 podem ocasionar danos a organelas celulares, promovendo complicações da doença. As glutationas S- transferases (GST) são as principais enzimas antioxidantes que participam da defesa celular contra o estresse oxidativo. Os polimorfismos nos genes que codificam essas enzimas podem acarretar o surgimento de complicações oftalmológicas em diabéticos. Este trabalho avaliou a influência dos polimorfismos nos genes GST no desenvolvimento de doenças como a catarata e o glaucoma em pacientes com DM2 na Grande Vitória (ES). Metodologia: Os polimorfismos dos genes GSTM1 e GSTT1 foram investigados através da técnica de PCR multiplex. Para o gene GSTP1 utilizou-se a técnica PCR- RFLP. A análise estatística foi realizada através do teste exato de Fisher ou do teste do qui-quadrado com P-valor < 0.05. Resultados: Não foi encontrada relação entre os polimorfismos nos genes GSTM1, GSTT1 e GSTP1 e o surgimento de doenças como glaucoma e catarata em pacientes com DM2. Conclusão: Nossos dados sugerem que os polimorfismos nulos nos genes GSTM1 e GSTT1 e o polimorfismo Ile105Val no gene GSTP1 não estão associados com a suscetibilidade individual para o desenvolvimento de complicações oftalmológicas em pacientes com DM2. (AU)

Introduction: Type 2 diabetes mellitus (T2DM) is a metabolic disorder caused by beta cell dysfunction that interferes with insulin production and/or by the resistance of target organs to this hormone. An increase in free radicals together with a decline in antioxidant defenses, present in T2DM, can damage cellular organelles and promote the occurrence of disease complications. Glutathione S-transferases (GSTs) are the main antioxidant enzymes involved in cellular defense against oxidative stress, and polymorphisms in genes encoding GSTs can lead to ophthalmic complications in persons with diabetes. In this study, we evaluated the influence of GST polymorphisms on the development of diseases such as cataract and glaucoma in patients with T2DM in Grande Vitória, Espírito Santo, Brazil. Methods: GSTM1 and GSTT1 polymorphisms were investigated using a multiplex PCR technique. PCR-RFLP was used for the GSTP1 gene. Statistical analysis was performed with Fisher's exact test or the chi-square test, with P-value <0.05. Results: There was no relationship between GSTM1, GSTT1, or GSTP1 polymorphisms and the occurrence of diseases such as glaucoma and cataract in patients with T2DM. Conclusion: Our data suggest that the GSTM1 and GSTT1 null polymorphisms and the ile105Val polymorphism in the GSTP1 gene are not associated with individual susceptibility to the development of ophthalmic complications in persons with T2DM. (AU)

Hypermethylation status of DAPK, MGMT and RUNX3 in HPV negative oral and oropharyngeal squamous cell carcinoma.

Squamous cell carcinoma of the oral cavity and oropharynx is the sixth most common type of cancer in the world. During tumorigenesis, gene promoter hypermethylation is considered an important mechanism of transcription silencing of tumor suppressor genes, such as DAPK, MGMT and RUNX3. These genes participate in signaling pathways related to apoptosis, DNA repair and proliferation whose loss of expression is possibly associated with cancer development and progression. In order to investigate associations between hypermethylation and clinicopathological and prognostic parameters, promoter methylation was evaluated in 72 HPV negative oral and oropharyngeal tumors using methylation-specific PCR. Hypermethylation frequencies found for DAPK, MGMT and RUNX3 were 38.88%, 19.44% and 1.38% respectively. Patients with MGMT hypermethylation had a better 2-year overall survival compared to patients without methylation. Being MGMT a repair gene for alkylating agents, it could be a biomarker of treatment response for patients who are candidates for cisplatin chemotherapy, predicting drug resistance. In view of the considerable levels of hypermethylation in cancer cells and, for MGMT, its prognostic relevance, DAPK and MGMT show potential as epigenetic markers, in a way that additional studies may test its viability and efficacy in clinical management.

Polymorphisms in methylenetetrahydrofolate reductase and cystathionine beta-synthase in oral cancer - a case-control study in southeastern Brazilians / Polimorfismos em metilenotetrahidrofolato redutase, cistationina beta-sintase no câncer de boca - um estudo de caso-controle no Sudeste brasileiro

ABSTRACT INTRODUCTION: Oral squamous cell carcinoma (OSCC) is a serious public health problem, due to its high mortality rate and worldwide rising incidence. OSCC susceptibility is mediated by interactions between genetic and environmental factors. Studies suggest that genetic variants encoding enzymes involved in folate metabolism may modulate OSCC risk by altering DNA synthesis/repair and methylation process. OBJECTIVE: The goals of this study were to evaluate the association of three genotypic polymorphism (MTHFR C677T, MTHFR A1298C and CBS 844ins68) and oral cancer risk in southeastern Brazilians and evaluate the interactions between polymorphisms and clinical histopathological parameters. METHODS: This case-control study included 101 cases and 102 controls in the state of Espírito Santo, Brazil. MTHFR genotyping was done by PCR-RFLP (polymerase chain reaction - restriction fragment length polymorphism) and CBS genotyping by PCR (polymerase chain reaction) analysis. RESULTS: MTHFR C677T polymorphism was associated with lymph node involvement. Genotype CT + TT acted as a protective factor. MTHFR A1298C AC + CC genotype was associated with tumor differentiation, and possibly with a better prognosis. In risk analysis, no correlation was observed between genotypes and OSCC. CONCLUSION: We concluded that MTHFR C677T, MTHFR A1298C and CBS 844ins68 polymorphisms were not associated with OSCC risk in southeastern Brazilians; however, we suggest a prognosis effect associated with MTHFR C677T and A1298C polymorphisms in OSCC.

Resumo Introdução: O carcinoma espinocelular oral (CECO) trata-se de um importante problema de saúde pública, devido à elevada taxa de mortalidade e incidência crescente em todo o mundo. A susceptibilidade ao CECO é mediada por interações entre fatores genéticos e ambientais. Estudos sugerem que as variantes genéticas que codificam as enzimas envolvidas no metabolismo do folato podem modular o risco de CECO, alterando a síntese/reparação do DNA e o processo de metilação. Objetivo: Os objetivos deste estudo foram avaliar a associação de três polimorfismos genotípicos (MTHFR C677T, MTHFR A1298C e CBS 844ins68) e o risco de câncer oral em brasileiros da região Sudeste, e avaliar as interações entre polimorfismos e parâmetros clínico-histopatológicos. Método: Este estudo de caso-controle incluiu 101 casos e 102 controles no estado do Espírito Santo, Brasil. A genotipagem do polimorfismo MTHFR foi realizada por PCR-RFLP (Reação de Polimerase em Cadeia - Polimorfismo no Comprimento de Fragmento de Restrição) e a do CBS por análise da PCR (Reação de Polimerase em Cadeia). Resultados: O polimorfismo MTHFR C677T foi associado ao envolvimento de gânglios linfáticos. O genótipo CT + TT atuou como um fator protetor. O genótipo MTHFR A1298C AC + CC foi associado à diferenciação do tumor e, possivelmente, a um prognóstico melhor. Na análise de risco, a correlação entre os genótipos e o CECO não foi observada. Conclusão: Concluímos que os polimorfismos MTHFR C677T, MTHFR A1298C e CBS 844ins68 não estão associados ao risco de CECO nos brasileiros da região Sudeste; no entanto, sugerimos um efeito prognóstico associado aos polimorfismos MTHFR C677T e A1298C em CECO.

Polymorphisms in methylenetetrahydrofolate reductase and cystathionine beta-synthase in oral cancer - a case-control study in southeastern Brazilians.

INTRODUCTION: Oral squamous cell carcinoma (OSCC) is a serious public health problem, due to its high mortality rate and worldwide rising incidence. OSCC susceptibility is mediated by interactions between genetic and environmental factors. Studies suggest that genetic variants encoding enzymes involved in folate metabolism may modulate OSCC risk by altering DNA synthesis/repair and methylation process. OBJECTIVE: The goals of this study were to evaluate the association of three genotypic polymorphism (MTHFR C677T, MTHFR A1298C and CBS 844ins68) and oral cancer risk in southeastern Brazilians and evaluate the interactions between polymorphisms and clinical histopathological parameters. METHODS: This case-control study included 101 cases and 102 controls in the state of Espírito Santo, Brazil. MTHFR genotyping was done by PCR-RFLP (polymerase chain reaction - restriction fragment length polymorphism) and CBS genotyping by PCR (polymerase chain reaction) analysis. RESULTS: MTHFR C677T polymorphism was associated with lymph node involvement. Genotype CT+TT acted as a protective factor. MTHFR A1298C AC+CC genotype was associated with tumor differentiation, and possibly with a better prognosis. In risk analysis, no correlation was observed between genotypes and OSCC. CONCLUSION: We concluded that MTHFR C677T, MTHFR A1298C and CBS 844ins68 polymorphisms were not associated with OSCC risk in southeastern Brazilians; however, we suggest a prognosis effect associated with MTHFR C677T and A1298C polymorphisms in OSCC.

Mutation analysis of GJB2 and GJB6 genes in Southeastern Brazilians with hereditary nonsyndromic deafness.

In developed countries deafness has a genetic cause in over 60% of the cases. Contrastingly, in Brazil, it is estimated that only 16% of all deafnesses are caused by genetic factors. Among hereditary hearing deficiencies, approximately half is caused by mutations in the Gap Junction Protein Beta-2 (GJB2) gene, which encodes the protein Connexin 26 (Cx26). There are four mutations in this gene that present high prevalence in specific ethnical groups, namely, 35delG, 167delT, 235delC, and W24X. The 35delG mutation is the most frequent one, occurring in homozygosity or in compound heterozygosity with mutations in the GJB2 and GJB6 genes. This study aims to determine the prevalence of GJB2-35delG, GJB2-167delT, GJB2-235delC, GJB2-W24X, del (GJB6-D13S1830), and del (GJB6-D13S1854) mutations in patients with nonsyndromic deafness in the Espirito Santo State, Brazil. A total of 77 individuals were evaluated, from which 88.3% presented normal genotypes for all analyzed mutations, 1.3% were compound heterozygotes for 35delG-GJB2/D13S1830-GJB6, 1.3% were compound heterozygotes for 35delG/D13S1854-GJB6, 3.9% were homozygotes for the 35delG mutation and 5.2% were heterozygotes for 35delG/GJB2. The frequency of mutant alleles 35delG/GJB2, del (D13S1830/GJB6), and del (D13S1854/GJB6) was 7.8, 0.65, and 0.65%, respectively. Mutations 167delT, 235delC, and W24X were not detected. Determining the prevalence of specific mutations related to inherited deafness in a population can contribute to the development of more efficient and affordable molecular diagnostic protocols, and help in the genetic counseling of patients and their families.

Prevalence of 35delG/GJB2 and del (GJB6-D13S1830) mutations in patients with non-syndromic deafness from a population of Espírito Santo-Brazil.

UNLABELLED: Mutations in GJB2 gene are the leading cause of deafness in autosomal recessive inheritance, and the 35delG mutation is the most common in many ethnic groups. Besides the 35delG mutation in homozygosis, the mutation is also found in compound heterozygosis, coupled with other mutations in genes GJB2 and GJB6. AIM: To determine the prevalence of 35delG/GJB2 and del (GJB6-D13S1830) mutations in patients with sensorineural hearing impairment in residents from the Espirito Santo state, Brazil. MATERIALS AND METHODS: 77 unrelated individuals with moderate to profound sensorineural hearing loss were evaluated. The 35delG mutation was studied by PCR / RFLP; and the del (GJB6-D13S1830) mutation was screened by the technique of multiplex PCR. RESULTS: 88.3% had normal genotype for the studied mutations, 1.3% were compound heterozygotes, 3.9% homozygotic for the 35delG mutation, 6.5% heterozygotic for 35delG/GJB2. The frequency of 35delG/GJB2 and del (D13S1830/GJB6) alleles in the sample was 7.8% and 0.65%, respectively. CONCLUSION: The data confirmed the existence of the mutations studied in cases of sensorineural hearing loss in a population from Espírito Santo / Brazil. These findings reinforce the importance of genetic diagnosis, which can provide early treatment for children and genetic counseling for the affected families.

Prevalências das mutações 35delG/GJB2 e del (GJB6-D13S1830) em portadores de surdez não-sindrômica na população do Espírito Santo - Brasil / Prevalence of 35delG/GJB2 and del (GJB6-D13S1830) mutations in patients with non-syndromic deafness from a population of Espírito Santo - Brazil

Mutações no gene GJB2 constituem a principal causa de surdez genética de herança autossômica recessiva, sendo a mutação 35delG a mais comum em muitos grupos étnicos. Além da mutação 35delG em homozigose, a mutação também é encontrada em heterozigose composta, associada com outras mutações nos genes GJB2 ou GJB6. OBJETIVOS: Determinar a prevalência das mutações 35delG/GJB2 e del (GJB6-D13S1830) em portadores de deficiência auditiva neurossensorial, residentes no estado do Espírito Santo (Brasil). MATERIAL E MÉTODOS: Foram avaliados 77 indivíduos não relacionados, com perda auditiva neurossensorial de moderada a profunda. A mutação 35delG foi estudada através da técnica de PCR/RFLP e a mutação del (GJB6-D13S1830) foi rastreada por meio da técnica de PCR multiplex. RESULTADOS: 88,3 por cento apresentaram genótipo normal para as mutações estudadas, 1,3 por cento foram heterozigotos compostos, 3,9 por cento homozigotos para a mutação 35delG, 6,5 por cento heterozigotos para 35delG/GJB2. A frequência do alelo 35delG/GJB2 e do alelo del (D13S1830/GJB6) na amostra foi de 7,8 por cento e 0,65 por cento, respectivamente. CONCLUSÃO: Os dados obtidos confirmaram a existência das mutações estudadas em casos de perda auditiva neurossensorial na população do Espírito Santo/Brasil. Esses achados reforçam a importância do diagnóstico genético, que pode propiciar um tratamento precoce para crianças e aconselhamento genético para as famílias dos afetados.

Mutations in GJB2 gene are the leading cause of deafness in autosomal recessive inheritance, and the 35delG mutation is the most common in many ethnic groups. Besides the 35delG mutation in homozygosis, the mutation is also found in compound heterozygosis, coupled with other mutations in genes GJB2 and GJB6. AIM: To determine the prevalence of 35delG/GJB2 and del (GJB6-D13S1830) mutations in patients with sensorineural hearing impairment in residents from the Espirito Santo state, Brazil. MATERIALS AND METHODS: 77 unrelated individuals with moderate to profound sensorineural hearing loss were evaluated. The 35delG mutation was studied by PCR / RFLP; and the del (GJB6-D13S1830) mutation was screened by the technique of multiplex PCR. RESULTS: 88.3 percent had normal genotype for the studied mutations, 1.3 percent were compound heterozygotes, 3.9 percent homozygotic for the 35delG mutation, 6.5 percent heterozygotic for 35delG/GJB2. The frequency of 35delG/GJB2 and del (D13S1830/GJB6) alleles in the sample was 7.8 percent and 0.65 percent, respectively. CONCLUSION: The data confirmed the existence of the mutations studied in cases of sensorineural hearing loss in a population from Espírito Santo / Brazil. These findings reinforce the importance of genetic diagnosis, which can provide early treatment for children and genetic counseling for the affected families.