RESUMO
Our previous study identified 8 risk and 9 protective plasma miRNAs associated with progression to end-stage kidney disease (ESKD) in diabetes. This study aimed to elucidate preanalytical factors that influence the quantification of circulating miRNAs. Using the EdgeSeq platform, which quantifies 2,002 miRNAs in plasma, including ESKD-associated miRNAs, we compared miRNA profiles in whole plasma versus miRNA profiles in RNA extracted from the same plasma specimens. Less than half of the miRNAs were detected in standard RNA extraction from plasma. Detection of individual and concentrations of miRNAs were much lower when RNA extracted from plasma was quantified by RNA sequencing (RNA-Seq) or quantitative reverse transcription PCR (qRT-PCR) platforms compared with EdgeSeq. Plasma profiles of miRNAs determined by the EdgeSeq platform had excellent reproducibility in assessment and had no variation with age, sex, hemoglobin A1c, BMI, and cryostorage time. The risk ESKD-associated miRNAs were detected and measured accurately only in whole plasma and using the EdgeSeq platform. Protective ESKD-associated miRNAs were detected by all platforms except qRT-PCR; however, correlations among concentrations obtained with different platforms were weak or nonexistent. In conclusion, preanalytical factors have a profound effect on detection and quantification of circulating miRNAs in ESKD in diabetes. Quantification of miRNAs in whole plasma and using the EdgeSeq platform may be the preferable method to study profiles of circulating cell-free miRNAs associated with ESKD and possibly other diseases.
Assuntos
MicroRNA Circulante , Falência Renal Crônica , Humanos , MicroRNA Circulante/sangue , MicroRNA Circulante/genética , Falência Renal Crônica/sangue , Falência Renal Crônica/genética , Masculino , Feminino , Pessoa de Meia-Idade , Nefropatias Diabéticas/sangue , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/diagnóstico , Biomarcadores/sangue , Idoso , Reprodutibilidade dos Testes , Adulto , MicroRNAs/sangue , MicroRNAs/genética , Progressão da Doença , Diabetes Mellitus/sangue , Diabetes Mellitus/genética , Diabetes Mellitus/diagnósticoRESUMO
Aim: To investigate the integrated epigenetic regulation of acquired drug resistance in cancer. Materials & methods: Our gene expression data of five induced drug-tolerant cell models, one resistant cell line and one publicly available drug-resistant dataset were integrated to identify common differentially expressed genes and pathways. ChIP-seq and DNA methylation by HM450K beadchip were used to study the epigenetic profile of differential expressed genes. Results & conclusion: Integrated transcriptomic analysis identified a common 'viral mimicry' related gene signature in induced drug-tolerant cells and the resistant state. Analysis of the epigenetic regulation revealed a common set of down-regulated genes, which are marked and regulated by a concomitant loss of H3K4me3, gain of H3K9me3 and increment of regional DNA methylation levels associated with tumor suppressor genes and apoptotic signaling.
Assuntos
Neoplasias/patologia , Linhagem Celular Tumoral , Metilação de DNA , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Tolerância a Medicamentos/genética , Perfilação da Expressão Gênica/métodos , Histonas/genética , Histonas/metabolismo , Humanos , Proteína 5 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteína 5 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Neoplasias/genética , Neoplasias/mortalidade , Regiões Promotoras Genéticas , Análise de SobrevidaRESUMO
DNA methylation is an epigenetic mechanism that plays a key role in regulating gene expression and other functions. Although this modification is seen in different sequence contexts, the most frequently detected DNA methylation in mammals involves cytosine-guanine dinucleotides. Pathological alterations in DNA methylation patterns are described in a variety of human diseases, including cancer. Unlike genetic changes, DNA methylation is heavily influenced by subtle modifications in the cellular microenvironment. In all cancers, aberrant DNA methylation is involved in the alteration of a large number of oncological pathways with relevant theranostic utility. Several technologies for DNA methylation mapping have been developed recently and successfully applied in cancer studies. The scope of these technologies varies from assessing a single cytosine-guanine locus to genome-wide distribution of DNA methylation. Here, we review the strengths and weaknesses of these approaches in the context of clinical utility for the molecular diagnosis of human cancers.