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1.
Arq. gastroenterol ; 58(1): 55-60, Jan.-Mar. 2021. tab, graf
Article in English | LILACS | ID: biblio-1248983

ABSTRACT

ABSTRACT BACKGROUND: Colorectal cancer is the third most common neoplasm in the world. Methylation of tumor related genes in CpG islands can cause gene silencing and been involved in the development of cancer. The potential role of DKK2 as a biomarker for early diagnosis of colorectal cancer remains unclear. OBJECTIVE: The aim of the study was to evaluate the profile of methylation and RNAm expression of DKK2 as potential predictors of colorectal cancer diagnosis and prognosis. METHODS: Expression of mRNAs encoding DKK2 in 35 colorectal cancer tissues was quantified using real-time polymerase chain reaction analysis. The DNA methylation was studied by high resolution melting analysis. The general characteristics of the patients were collected. DKK2 methylation and expression were compared to clinical, pathological aspects and overall survival. RESULTS: Among the 35 patients studied, 18 were male, 10 were on right colon and 25 on left colon. Among the 20 patients with high hypermethylation, 15 of them had mRNA low expression of DKK2. There was no significant association between DKK2 promoter methylation and mRNA DKK2 expression and clinical or pathological features. DKK2 promoter methylation (P=0.154) and DKK2 RNA expression (P=0.345) did not show significant correlation with overall survival. CONCLUSION: DKK2 promoter methylation and DKK2 RNA status appear to be biomarkers of cancer diagnosis but not predictors of prognosis.


RESUMO CONTEXTO: O câncer colorretal é a terceira neoplasia mais comum no mundo. A metilação de alguns genes nas ilhas CpG podem causar silenciamento gênico e estar envolvida no desenvolvimento de câncer. O potencial papel de DKK2 como um biomarcador no diagnóstico precoce de CCR permanece incerto. OBJETIVO: O objetivo do estudo foi avaliar o perfil de metilação e expressão de RNAm do gene DKK2 para identificar preditores potenciais de diagnóstico e prognóstico de CCR. MÉTODOS: A expressão de mRNAs que codificam DKK2 em 35 tecidos de câncer colorretal foi quantificada por reação em cadeia da polimerase em tempo real e a metilação do DNA foi verificada por análise de alta resolução. As características gerais dos pacientes foram coletadas. A metilação e expressão de DKK2 foram comparadas aos aspectos clínicos, patológicos e à sobrevida global. RESULTADOS: Entre os 35 pacientes estudados, 18 eram do sexo masculino, 10 tumores eram do cólon ascendente ou transverso e 25 do descendente ou reto. Entre os 20 pacientes com hipermetilação, 12 deles apresentaram baixa expressão de RNAm do gene DKK2. Não houve associação significativa entre a metilação do promotor de DKK2 e a expressão de RNAm de DKK2 e características clínicas ou patológicas. A metilação do promotor de DKK2 e a expressão do RNA de DKK2 não mostraram correlação com sobrevida global dos pacientes com CCR. CONCLUSÃO: A metilação do gene promotor e a expressão do RNAm do gene DKK2 parecem ser biomarcadores de diagnóstico de câncer, mas não se mostraram úteis na avaliação prognóstica.


Subject(s)
Humans , Male , Female , Colorectal Neoplasms/genetics , DNA Methylation , Prognosis , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Promoter Regions, Genetic , CpG Islands , Intercellular Signaling Peptides and Proteins/genetics
2.
Journal of Experimental Hematology ; (6): 1768-1774, 2021.
Article in Chinese | WPRIM | ID: wpr-922332

ABSTRACT

OBJECTIVE@#To investigate the effect of silencing DNA methyltransferase 1(DNMT1) to the methylation of the promoter of the tumor suppressor gene wnt-1 (WIF-1) in human chronic myeloid leukemia (CML) cells.@*METHODS@#DNMT1 siRNAi plasmid was constructed and DNMT1 siRNAi was transfected into CML K562 cells. RT-PCR and Western blot were used to detect the expression of DNMT1 gene and related protein, and methylation PCR was used to detect WIF-1 gene promoter methylation level. The trypan blue exclusion and MTT assay were used to detect the cell proliferation, flow cytometry were used to detect the cell apoptosis rate, colony formation assay was used to detect cell colony formation ability. Expression of Wnt/β- catenin and its downstream signaling pathway proteins were detected by Western blot after DNMT1 gene was silenced.@*RESULTS@#The expression level of DNMT1 mRNA and its related protein in the experimental group were significantly lower than those in the control group and negative control group (P<0.05). After 72 hours of successful transfection, the WIF-1 gene in the control group and negative control group were completely methylated, while in the experimental group, the methylation level significantly decreased. The results of MSP showed that the PCR product amplified by the unmethylated WIF-1 primer in the experimental group increased significantly,while by the methylated WIF-1 primer decreased significantly. After 48 h of transfection, the OD value, viable cell number and colony formation of the cells in experimental group were significantly lower than those in the negative control group and the control group (P<0.05). The apoptosis rate of the cells in experimental group was significantly higher than those in the negative control group and control group (P<0.05). The expression levels of β- actin, myc, cyclin D1 and TCF-1 in K562 cells in the experimental group were significantly lower than those in the negative control group and control group (P<0.05).@*CONCLUSION@#Silencing DNMT1 gene can inhibit the proliferation and promote the apoptosis of K562 cells. The mechanism may be related to reverse the hypermethylation level of the WIF-1 gene promoter, thereby inhibit the activity of the Wnt/β- catenin signaling pathway.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , DNA Methylation , Humans , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Repressor Proteins/metabolism
3.
Article in English | WPRIM | ID: wpr-922251

ABSTRACT

In neuronal system, epigenetic modifications are essential for neuronal development, the fate determination of neural stem cells and neuronal function. The dysfunction of epigenetic regulation is closely related to occurrence and development of neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease. Abnormally elevated DNA methylation inhibits the expression of some DNA repair-related genes and affects the progression of Huntington's disease. In the brain of Alzheimer's disease patients, the levels of H3K27ac and H3K9ac histone modifications increased. In addition, the alteration of RNA methylation in animal models of Alzheimer's disease and Parkinson's disease showed discrepancy trends. Therefore, epigenetic modifications may serve as potential therapeutic targets for neurodegenerative diseases. Here, we summarize the recent progress of the roles of epigenetic modifications in neurodegenerative diseases.


Subject(s)
Animals , DNA Methylation , Epigenesis, Genetic , Humans , Neurodegenerative Diseases/genetics , Parkinson Disease/genetics , Protein Processing, Post-Translational
4.
Chinese Journal of Lung Cancer ; (12): 705-713, 2021.
Article in Chinese | WPRIM | ID: wpr-922131

ABSTRACT

Patients with oncogenic driver alterations of non-small cell lung cancer (NSCLC) can benefit from targeted therapy, but acquired resistance is inevitable ultimately. Epigenetic modifications, including DNA methylation, histone modifications, non-coding RNA-mediated regulate and chromatin remodeling, are important mechanisms of acquired resistance in targeted therapy of NSCLC. In recent years, studies have found that epigenetic modifications can effectively reverse drug resistance. Targeted therapy combined with epigenetic modifications may become a promising therapeutic strategy. Here, we review the progress of epigenetic mechanism in acquired resistance of targeted therapy in NSCLC, hoping to provide ideas for screening dominant population and overcoming resistance.
.


Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , DNA Methylation , Epigenesis, Genetic , Humans , Lung Neoplasms/genetics
5.
Acta Physiologica Sinica ; (6): 980-990, 2021.
Article in Chinese | WPRIM | ID: wpr-921303

ABSTRACT

The normal development of follicles involves a series of complex life processes such as ordered transcriptional activation and inhibition, which is crucial for female reproductive ability. Histone methylation can change the chromatin state in cells and affect the transcription activity of genes. Current studies indicate that epigenetic modifications such as histone methylation play an important regulatory role in follicular development in female mammals. This paper summarized the relationship between H3K4, H3K9 methylation and germ cell development, their regulatory effects, including their dynamical changes during follicular development, and the progress of H3K4me3 and other histone methylation binding to promoter regions of different genes to regulate gene expression and thus affect germ cell epigenetic reprogramming, oocyte transcription, meiosis and other processes. This review will provide a reference for the study of mechanisms related to histone methylation modification and the development and maturation of gonadal parenchymal cells.


Subject(s)
Animals , DNA Methylation , Epigenesis, Genetic , Female , Histones , Mammals , Ovarian Follicle/growth & development , Protein Processing, Post-Translational
6.
Chinese Medical Journal ; (24): 2901-2910, 2021.
Article in English | WPRIM | ID: wpr-921119

ABSTRACT

Recent research efforts have provided compelling evidence of genome-wide DNA methylation alterations in pediatrics. It is currently well established that epigenetic clocks, composed of DNA methylation sites, can estimate the gestational and chronological age of cells and tissues from different ages. Also, extensive research is aimed at their correlation with early life exposure and pediatric diseases. This review aimed to systematically summarize the epigenetic clocks in the pediatric population. Publications were collected from PubMed and Web of Science databases up to Apr 2021. Epigenetic clocks, DNA methylation clocks, epigenetic age acceleration or deceleration, pediatric and the pediatric population were used as search criteria. Here, we first review the currently applicative pediatric epigenetic clocks. We then highlight the interpretation for epigenetic age deviations in the pediatric population and their association with external factors, developmental trajectories, and pediatric diseases. Considering the remaining unknown of pediatric clocks, research strategies into them are also discussed. In all, pediatric epigenetic clocks may act as potent tools to understand development, growth and diseases in early life.


Subject(s)
Aging , Child , DNA Methylation/genetics , Epigenesis, Genetic/genetics , Epigenomics , Humans
7.
Article in Chinese | WPRIM | ID: wpr-888404

ABSTRACT

The progress of epigenetic research has led to the discovery and confirmation of age-related markers based on DNA methylation. These DNA methylation indices are called "epigenetic clock/age". The concept of "epigenetic clock/age" and the establishment of its evaluation system are helpful to solve some of the long-standing problems in the field of life and medicine. When facing the current global aging, it is of great significance to refer to the comprehensive health parameters to determine the biological age and life span of an individual, and thus to design a plan to slow down the process of life cycle. This paper has summarized the concept and development of "epigenetic clock/age" in recent years.


Subject(s)
Aging/genetics , Biomarkers , DNA Methylation , Epigenesis, Genetic , Humans
8.
Article in Chinese | WPRIM | ID: wpr-888070

ABSTRACT

This study aims to explore the relationship of DNA methylation with the contents of the index components as well as the growth and development of Pogostemon cablin. The demethylation reagent 5-azacytidine(5-azaC) was used to treat the tissue culture seedlings of patchouliol-type P. cablin. High performance liquid chromatography was employed to evaluate the changes of DNA methy-lation in P. cablin, and GC-MS to detect the contents of index components in P.cablin. The agronomic characters of P.cablin were measured using the common methods. The results showcased that DNA methylation of P.cablin was significantly reduced by 5-azaC in a concentration-dependent manner. Thirty days after treatment with 5-azaC at different concentrations, the content of patchouli alcohol changed slightly; compared with that in the control group, the content of pogostone in 50 μmol·L~(-1) and 100 μmol·L~(-1) 5-azaC groups was significantly up-regulated. The 100 μmol·L~(-1) 5-azaC group had the largest differences in contents of pogostone and patchouli alcohol compared with the control group, followed by the 50 μmol·L~(-1) 5-azaC group. Ninety days after disinhibition, the content of pogostone in the treatment group was significantly increased and the content of patchouli alcohol was significantly decreased. In addition, 5-azaC significantly inhibited the growth and development of P.cablin in a dose-dependent manner. These results indicate that DNA methylation regulates the biosynthesis of the index components in patchouliol-type P.cablin and proper demethylation can directly promote the synthesis of pogostone and indirectly affect the accumulation of patchouli alcohol.


Subject(s)
Azacitidine , DNA Methylation , Gas Chromatography-Mass Spectrometry , Oils, Volatile , Pogostemon/genetics
9.
Chinese Medical Journal ; (24): 1310-1316, 2021.
Article in English | WPRIM | ID: wpr-878104

ABSTRACT

BACKGROUND@#Epigenetics, especially DNA methylation, plays an important role in the pathogenesis of primary Sjogren syndrome (pSS). Our study aimed to reveal the role of DNA methylation in peripheral monocytes of pSS patients.@*METHODS@#A total of 11 pSS patients and five age-matched healthy controls (HCs) were included in this study. Monocytes were isolated from peripheral blood mononuclear cells using magnetic microbeads. DNA methylation profiles were generated using Human Methylation 850K BeadChips.@*RESULTS@#In monocytes from pSS patients, we identified 2819 differentially methylated positions (DMPs), comprising 1977 hypomethylated- and 842 hypermethylated-DMPs, corresponding to 1313 unique genes when compared with HCs. IFI44L, MX1, PAARP9, and IFITM1, which influence the interferon (IFN) signaling pathway, were among the genes hypomethylated in pSS. Functional analysis of genes with a minimum of two DMPs showed involvement in antigen binding, transcriptional regulation, cell adhesion, IFN-γ pathway, type I IFN pathway, antigen presentation, Epstein-Barr virus infection, human T-lymphotropic virus type 1 virus infection, and metabolic disease-related pathways. In addition, patients with higher serum IgG levels exhibited enrichment in Notch signaling and metabolic-related pathways. Upon comparing monocytes with salivary gland epithelial cells, an important overlap was observed in the cell cycle, cell senescence, and interleukin-17 signaling pathways. The differentially methylated genes were more enriched in the ribosome- and AMP-activated protein kinase signaling pathway in anti-Ro/SSA and anti-La/SSB autoantibodies double-positive patients.@*CONCLUSION@#Genome-wide DNA methylation profiling revealed significant differences in DNA methylation in monocytes isolated from patients with pSS.


Subject(s)
DNA Methylation/genetics , Epstein-Barr Virus Infections , Herpesvirus 4, Human , Humans , Leukocytes, Mononuclear , Monocytes , Sjogren's Syndrome/genetics
10.
Protein & Cell ; (12): 7-28, 2021.
Article in English | WPRIM | ID: wpr-880895

ABSTRACT

Mammalian fertilization begins with the fusion of two specialized gametes, followed by major epigenetic remodeling leading to the formation of a totipotent embryo. During the development of the pre-implantation embryo, precise reprogramming progress is a prerequisite for avoiding developmental defects or embryonic lethality, but the underlying molecular mechanisms remain elusive. For the past few years, unprecedented breakthroughs have been made in mapping the regulatory network of dynamic epigenomes during mammalian early embryo development, taking advantage of multiple advances and innovations in low-input genome-wide chromatin analysis technologies. The aim of this review is to highlight the most recent progress in understanding the mechanisms of epigenetic remodeling during early embryogenesis in mammals, including DNA methylation, histone modifications, chromatin accessibility and 3D chromatin organization.


Subject(s)
Animals , Chromatin Assembly and Disassembly , DNA Methylation , DNA Transposable Elements , Embryo, Mammalian , Embryonic Development/genetics , Epigenesis, Genetic , Epigenome , Female , Fertilization/physiology , Gene Expression Regulation, Developmental , Histone Code , Histones/metabolism , Male , Mice , Oocytes/metabolism , Spermatozoa/metabolism
11.
Article in Chinese | WPRIM | ID: wpr-880834

ABSTRACT

OBJECTIVE@#To explore the mechanism by which ginsenoside 20(S)-Rg3 upregulates the expression of tumor suppressor von Hippel-Lindau (VHL) gene in ovarian cancer cells.@*METHODS@#Ovarian cancer cell line SKOV3 treated with 20(S)-Rg3 were examined for mRNA and protein levels of VHL, DNMT1, DNMT3A and DNMT3B by real-time PCR and Western blotting, respectively. The changes in VHL mRNA expression in SKOV3 cells in response to treatment with 5-Aza-CdR, a DNA methyltransferase inhibitor, were detected using real-time PCR. VHL gene promoter methylation was examined with methylation-specific PCR and VHL expression levels were determined with real-time PCR and Western blotting in non-treated or 20(S)-Rg3-treated SKOV3 cells and in 20(S)-Rg3-treated DNMT3A-overexpressing SKOV3 cells. VHL and DNMT3A protein levels were detected by immunohistochemistry in subcutaneous SKOV3 cell xenografts in nude mice.@*RESULTS@#Treatment of SKOV3 cells with 20(S)-Rg3 significantly upregulated VHL and downregulated DNMT3A expressions at both the mRNA and protein levels (@*CONCLUSIONS@#Ginsenoside 20(S)-Rg3 upregulates VHL expression in ovarian cancer cells by suppressing DNMT3A-mediated DNA methylation.


Subject(s)
Animals , Cell Line, Tumor , DNA Methylation , Female , Gene Expression , Ginsenosides/pharmacology , Humans , Mice , Mice, Nude , Ovarian Neoplasms/genetics , Promoter Regions, Genetic , Von Hippel-Lindau Tumor Suppressor Protein/genetics
12.
Article in English | WPRIM | ID: wpr-880633

ABSTRACT

OBJECTIVES@#To explore the correlation between cytosine-phosphoric-guanylic (CpG) site of Septin 9 gene and colorectal cancer, and to develop a real-time PCR detection system in plasma in patients with colorectal cancer.@*METHODS@#The methylation of training samples was detected by high-throughput sequencing technology, and the sites highly consistent with the clinical information of colorectal cancer were identified. Then the detection system of real-time PCR was designed to analyze the consistency of plasma and tissue based on methylationa sensitive enzyme digestion. Finally, 100 clinical trials were conducted to evaluate the performance of the detection system with the methylation sensitive enzyme digestion-real-time PCR.@*RESULTS@#The highly consistent sites, which were selected by high-throughput sequencing from 71 training set samples, was the 38th CpG. Based on the detection region, the screened methylation sensitive enzymes were @*CONCLUSIONS@#The 38th CpG site of Septin 9 detected by the detection system of methylation sensitive enzyme digestion-real-time PCR can highly predict the occurrence of colorectal cancer with great clinical application value.


Subject(s)
Colorectal Neoplasms/genetics , CpG Islands/genetics , DNA , DNA Methylation , Humans , Plasma/metabolism , Septins/metabolism
13.
Article in English | WPRIM | ID: wpr-880627

ABSTRACT

The causes for male sexual orientation are complicated, which have not yet been clarified. Recent years have witnessed fruitful progress in the field of biology, while the impact of environment has received little attention. Adverse childhood experiences (ACEs), identified as a special environment in the early stage of development, can affect the individual phenotype by DNA methylation. Given the relationships among male sexual orientation, ACEs, and DNA methylation, as well as based on the existing theory, this article proposes the model "ACEs-DNA methylation-male sexual orientation"from the perspective of environment and epigenetics, aiming to provide a theoretical basis for future research.


Subject(s)
Adverse Childhood Experiences , Child , DNA Methylation , Female , Humans , Male , Sexual Behavior
14.
Article in English | WPRIM | ID: wpr-879956

ABSTRACT

Epigenetics concerns gene regulatory mechanisms beyond DNA sequence,such as DNA methylation,histone modification,chromatin remodeling,and non-coding RNA. Epigenetic mechanisms play a key role in development,cell fate decision and tumorigenesis. Chromatin modifications and its high order structure across our genome are major forms of epigenetic information,and its establishment and maintenance are closely related to cell metabolism. Metabolic changes in cancer cells include aerobic glycolysis,increased glucose uptake,abnormally active glutamine metabolism,and the use of non-conventional energy supply. These changes meet the vigorous energy and matter needs for the development and spread of cancer,and help tumor cells adapt to hypoxia microenvironment for their survival,proliferation,invasion and migration. There is a complex relationship between epigenetic modifications and cell metabolism in tumor. On the one hand,metabolites in tumor cells may act as cofactors,modification donors or antagonists of epigenetic enzymes,thus modulating the epigenetic landscape. On the other hand,epigenetic modifications can directly regulate the expression of metabolic enzymes,transporters,signaling pathway and transcription factors to affect cell metabolism. This article reviews the crosstalk between epigenetics and cancer metabolism,to explore their potential future applications in the treatment of tumors.


Subject(s)
Carcinogenesis , DNA Methylation , Epigenesis, Genetic , Gene Expression Regulation , Humans , Neoplasms/genetics , Tumor Microenvironment
15.
Article in Chinese | WPRIM | ID: wpr-879619

ABSTRACT

OBJECTIVE@#To study the correlation between DNA methylation patterns and gene expression in Down syndrome (DS).@*METHODS@#Induced pluripotent stem cells (iPSCs) derived from normal controls and DS patients were subjected to whole genome bisulfite sequencing and differentially methylated region (DMR) screening. Statistical analysis for chromosomal and gene element distribution were carried out for DMR. Gene ontology (GO) and enrichment-based cluster analysis were used to explore the molecular function of differentially expressed genes.@*RESULTS@#A total of 1569 DMR were identified in iPSCs derived from DS patients, for which the proportion of hypermethylation in promoter regions was significantly greater than that of the genebody. No DMR enrichment was noted on chromosome 21. Hypermethylation of the promoter and genebody was predicted to be inhibitory for gene expression. Functional clustering revealed the pathways related to neurodevelopmental, stem cell pluripotency and organ size regulation to be significantly correlated with differentially methylated genes.@*CONCLUSION@#Extensive and stochastic anomalies of genome-wide DNA methylation has been discovered in iPSCs derived from DS patients, for which the pattern and molecular regulation of methylation were significantly different from those of normal controls. Above findings suggested that DNA methylation pattern may play a vital role in both the pathogenesis of neurodevelopmental disorders and other phenotypic abnormalities during early embryonic development.


Subject(s)
DNA Methylation , Down Syndrome/genetics , Female , Humans , Induced Pluripotent Stem Cells , Pregnancy , Promoter Regions, Genetic , Whole Genome Sequencing
16.
Article in Chinese | WPRIM | ID: wpr-879553

ABSTRACT

DNA methylation as an important aspect of epigenetics plays an important role in spermatogenesis and embryonic development. In recent years, researchers have found that male infertility, in particular abnormal semen quality, is related to abnormal DNA methylation. To further delineate the pathogenesis of male infertility and inspire new ideas for the treatment of male infertility, a comprehensive review over the correlation between abnormal methylation of imprinted genes, repetitive DNA elements and non-imprinted genes, semen quality (including sperm count, morphology, and vitality) and male infertility is provided.


Subject(s)
DNA Methylation , Humans , Infertility, Male/genetics , Male , Semen Analysis , Sperm Count , Spermatogenesis , Spermatozoa/pathology
17.
Pesqui. vet. bras ; 40(12): 1063-1072, Dec. 2020. tab, graf, ilus
Article in English | LILACS, VETINDEX | ID: biblio-1155041

ABSTRACT

Somatic Cell Nuclear Transfer (SCNT-Cloning) is a promising technique in many areas and is based on genetically identical individuals. However, its efficiency is low. Studies suggest that the leading cause is inadequate epigenetic reprogramming. This study aimed to characterize the methylation pattern of the exon 10 regions of the IGF2 gene and the Imprinting Control Region (ICR) of the H19 gene in the placenta of cloned calves. For this study, female and male cloned calves presenting different phenotypes were used. Genomic DNA from these animals' placenta was isolated, then treated with sodium bisulfite and amplified to the ICR/H19 and IGF2 loci. PCR products were cloned into competent bacteria and finally sequenced. A significant difference was found between controls and clones with healthy phenotypes for the ICR/H19 region. In this region, controls showed a hemimethylated pattern, as predicted in the literature due to this region has an imprinted control, while clones were showed less methylated. For the IGF2, no significant differences were found between controls and clones. These results suggest that different genomic regions in the genome may be independently reprogrammed and that failures in reprogramming the DNA methylation patterns of imprinted genes may be one of the causes of the low efficiency of SCNT.(AU)


A Transferência Nuclear de Células Somáticas (TNCS-Clonagem) é uma técnica promissora em várias áreas, e se baseia na produção de indivíduos geneticamente idênticos. No entanto, sua eficiência é baixa. Estudos sugerem que a principal causa seja uma reprogramação epigenética inadequada. O objetivo desse trabalho é caracterizar o padrão de metilação da região éxon 10 do gene IGF2 e da Região Controladora de Imprinting (ICR) do gene H19 na placenta de bezerros clonados. Para a execução do trabalho foram selecionados clones bovinos fêmeas e machos, apresentando diferentes fenótipos. O DNA da placenta desses animais foi extraído, e em seguida foi tratado com bissulfito de sódio e amplificado para os loci ICR/H19 e IGF2. Os produtos da PCR foram clonados em bactérias competentes e, por fim, sequenciados. Foi encontrada uma diferença significativa entre os controles e os clones com fenótipos saudáveis para a região da ICR/H19. Nesta região, os controles tiveram um padrão hemimetilado, como previsto pela literatura, devido essa região ser imprinted. Enquanto os clones encontravam-se menos metilados. Para a região do éxon 10 do IGF2, não foi encontrada diferença significativa entre controles e clones. Estes resultados sugerem que as diferentes regiões do genoma podem se reprogramar independente umas das outras e que falhas na reprogramação do padrão de metilação do DNA de genes imprinted podem ser uma das causas da baixa eficiência da TNCS.(AU)


Subject(s)
Animals , Cattle , Placenta , Cattle/genetics , Clone Cells , Epigenomics , Insulin-Like Growth Factor II/analysis , DNA Methylation
18.
Braz. oral res. (Online) ; 34: e087, 2020. tab, graf
Article in English | LILACS, BBO | ID: biblio-1132726

ABSTRACT

Abstract Inflammatory external root resorption (IERR) is a pathological process defined by the progressive loss of dental hard tissue, dentin, and cementum, resulting from the combination of the loss of external root protective apparatus and root canal infection. It has been suggested that healing patterns after tooth replantation may be influenced by the genetic and immunological profiles of the patients. The purpose of the present investigation was to evaluate the DNA methylation patterns of 22 immune response-related genes in extracted human teeth presenting with IERR. Methylation assays were performed on samples of root fragments showing IERR and compared with healthy bone tissue collected during the surgical extraction of impacted teeth. The methylation patterns were quantified using EpiTect Methyl II Signature Human Cytokine Production PCR Array. The results revealed significantly higher hypermethylation of the FOXP3 gene promoter in IERR (65.95%) than in the bone group (23.43%) (p < 0.001). The ELANE gene was also highly methylated in the pooled IERR sample, although the difference was not statistically significant (p= 0.054). Our study suggests that the differential methylation patterns of immune response-related genes, such as FOXP3 and ELANE, may be involved in IERR modulation, and this could be related to the presence of root canal infection. However, further studies are needed to corroborate these findings to determine the functional relevance of these alterations and their role in the pathogenesis of IERR.


Subject(s)
Humans , Root Resorption , Tooth Replantation , Tooth Root , DNA Methylation , Dental Cementum
19.
Belo Horizonte; s.n; 2020. 124 p. ilus, graf, tab.
Thesis in English, Portuguese | LILACS, BBO | ID: biblio-1099625

ABSTRACT

A reabsorção radicular externa inflamatória (RREI) é um processo patológico definido como a perda progressiva de tecido mineralizado radicular, dentina e cemento, resultante da combinação entre a lesão às camadas protetoras da superfície externa da raiz e a presença de microrganismos no interior do sistema de canais radiculares. Estudos clínicos demonstraram o papel da idade e de fatores relacionados ao manejo e tratamento do dente avulsionado na etiopatogenia e evolução das RREI após reimplantes. Entretanto, não existem informações sobre a interação destes fatores, bem como poucos estudos avaliaram a influência do perfil genético e imunológico do paciente no padrão de cicatrização após reimplantes dentários. O presente estudo objetivou (1) avaliar a interação de fatores prognósticos para o desenvolvimento da RREI após o reimplante de dentes permanentes, bem como (2) investigar o papel da epigenética nos processos imunomediados das RREI pós-traumáticas. Para estudo dos determinantes clínicos e suas interações, o universo da pesquisa envolveu 427 pacientes (idade média de 12,6 anos) portadores de 581 dentes permanentes reimplantados, com rizogênese completa no momento do trauma, tratados na Clínica de Traumatismos dentários da Faculdade de Odontologia da Universidade Federal de Minas Gerais entre 1994 e 2018. Dados relativos à idade do paciente no momento do trauma, grau de rizogênese, condições de armazenamento e período extra alveolar do dente avulsionado, uso de antibioticoterapia sistêmica, tempo decorrido entre o reimplante e o início da terapia endodôntica radical (TER) e a duração do período de imobilização foram coletados dos prontuários dos pacientes. Tomadas radiográficas realizadas na consulta de início do TER foram utilizadas para diagnóstico da atividade de reabsorção. Sinais radiográficos de RREI foram encontrados em 80,7% da amostra (469 dentes). Os resultados demonstraram que a idade do paciente no momento do trauma e o tempo decorrido até o início do TER representaram importantes fatores prognósticos para a ocorrência de RREI. Além disso foi observada uma interação quantitativa entre estas duas variáveis uma vez que o aumento na idade do paciente atenuou significativamente o efeito do tempo até o início da terapia endodôntica. Este resultado inédito evidencia a maior vulnerabilidade do paciente mais jovem e enfatiza a importância de se considerar estas duas covariáveis conjuntamente durante a tomada de decisão clínica. Para o estudo epigenético, o perfil de metilação do DNA de 22 genes envolvidos na resposta imune foi avaliado em um pool de 08 amostras de fragmentos radiculares de dentes reimplantados portadores de RREI, indicados para exodontia. O grupo controle consistiu em um pool de 06 amostras de tecido ósseo saudável coletado durante a extração cirúrgica de dentes impactados. Os padrões de metilação do DNA dos 22 genes foram quantificados utilizando EpiTect Methyl II Signature Human Cytokine Production PCR Array. Os resultados do estudo da epigenética revelou que o pool de amostras com RREI apresentou nível mais alto de metilação do DNA na região promotora da FOXP3, em comparação com o pool de osso normal (65,95% e 23,43%, respectivamente). Esta é a primeira evidência de uma possível participação de eventos epigenéticos na modulação da RREI e especula-se se o padrão hipermetilado da FOXP3 poderia estar relacionado à presença da infecção endodôntica.


Inflammatory external root resorption (IERR) is a pathological process defined as the progressive loss of root mineralized tissue, dentin and cement, resulting from both: damage to the protective layers in the root external surface and the presence of endodontic infection inside the root canal. Clinical studies have demonstrated the role of age and factors related to the management and treatment of avulsed teeth in the etiopathogenesis and progression of RREI. However, there is no information on the interaction of these factors and few studies have evaluated the influence of the patient's genetic and immunological profile on the healing pattern after dental replantation. The present study aimed to (1) evaluate the interaction of prognostic factors for the development of RREI after replantation of permanent teeth, as well as (2) to investigate the role of epigenetics in the immunomediated processes of posttraumatic RREI. To study the clinical determinants and their interactions, the sample comprised 427 patients (mean age 12.6 years) with 581 replanted mature permanent teeth treated at the Dental Trauma Clinic of the Faculty of Dentistry from the Federal University of Minas Gerais between 1994 and 2018. Patients' records were evaluated to collect data such as patient's age at the time of the trauma, storage conditions and extra alveolar period of the avulsed tooth, systemic antibiotic therapy prescription, time elapsed between reimplantation and onset of endodontic therapy (TER) and splinting timing. The presence and index of IERR was assessed radiographically at the visit of pulpectomy. Radiographic signs of IEER were found in 80.7% of the sample (469 teeth) and were absent in 19.7% of cases (112 teeth). The results showed that the patient's age at the time of the trauma and the time that elapsed until the beginning of TER represented important prognostic factors for the occurrence of RREI. In addition, a quantitative interaction was observed between these two variables since the increase in the patient's age significantly attenuated the effect of time until the beginning of endodontic therapy. This is an original result that highlights the greater vulnerability of the younger patients and emphasizes the importance of considering these two covariates together during clinical decision-making. For the epigenetic study, the DNA methylation profile of 22 genes involved in the immune response was evaluated in a pool of 08 samples of root fragments of replanted teeth with RREI, referred to extraction. The control group consisted of a pool of 06 samples of healthy bone tissue collected during surgical extraction of impacted teeth. The DNA methylation pattern was quantified using EpiTect Methyl II Signature Human Cytokine Production PCR Array. The results of the epigenetics study revealed that the sample pool with RREI showed a higher level of DNA methylation in the FOXP3 promoter region, compared to the normal bone pool (65.95% and 23.43%, respectively). This is the first evidence of a possible participation of epigenetic events in the modulation of RREI and it is speculated whether the hypermethylated pattern of FOXP3 could be related to the presence of endodontic infection.


Subject(s)
Periodontal Ligament , Root Resorption , Tooth Avulsion , Tooth Replantation , Dentition, Permanent , DNA Methylation , Epigenomics , Cross-Sectional Studies , Anti-Bacterial Agents
20.
J. appl. oral sci ; 28: e20190583, 2020. tab, graf
Article in English | LILACS, BBO | ID: biblio-1090773

ABSTRACT

Abstract Genetic and epigenetic changes have been associated with periodontitis in various genes; however, little is known about genes involved in epigenetic mechanisms and in oxidative stress. Objective: This study aims to investigate the association of polymorphisms C677T in MTHFR (rs1801133) and −149C→T in DNMT3B (rs2424913), as well as the methylation profiles of MTHFR, miR-9-1, miR-9-3, SOD1, and CAT with periodontitis. The association between polymorphisms and DNA methylation profiles was also analyzed. Methodology: The population studied was composed of 100 nonsmokers of both sexes, divided into healthy and periodontitis groups. Genomic DNA was extracted from the epithelial buccal cells, which were collected through a mouthwash. Polymorphism analysis was performed through polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), while methylation-specific PCR (MSP) or combined bisulfite restriction analysis techniques were applied for methylation analysis. Results: For DNMT3B, the T allele and the TT genotype were detected more frequently in the periodontitis group, as well as the methylated profile on the miR-9-1 promoter region. There was also a tendency towards promoter region methylation on the CAT sequence of individuals with periodontal disease. Conclusion: The polymorphism −149C→T in DNMT3B (rs2424913) and the methylated profile of the miR-9-1 promoter region are associated with periodontitis.


Subject(s)
Humans , Male , Female , Adult , Middle Aged , Aged , Periodontitis/genetics , Polymorphism, Genetic , DNA Methylation/genetics , MicroRNAs/genetics , DNA (Cytosine-5-)-Methyltransferases/genetics , Polymorphism, Restriction Fragment Length , Catalase/genetics , Case-Control Studies , Polymerase Chain Reaction , Methylenetetrahydrofolate Reductase (NADPH2)/genetics , Genetic Association Studies , Superoxide Dismutase-1/genetics , Genotype
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