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The nervous system is rich in miRNAs, indicating an important role of these molecules in regulating processes associated with cognition, memory, and others. Therefore, qualitative and quantitative imbalances involving such miRNAs may be involved in dementia contexts, including Late-Onset Alzheimer's Disease (LOAD). To test the viability of circulating miRNAs (c-miRNAs) as biomarkers for LOAD, we proceed accordingly to the following reasoning. The first stage was to discover and identify profile of c-miRNAs by RNA sequencing (RNA-Seq). For this purpose, blood serum samples were used from LOAD patients (n = 5) and cognitively healthy elderly control group (CTRL_CH) (n = 5), all over 70 years old. We identified seven c-miRNAs differentially expressed (p ≤ 0.05) in the serum of LOAD patients compared to CTRL_CH (miR-10a-5p; miR-29b-2-5p; miR-125a-5p; miR-342-3p, miR-708-5p, miR-380-5p and miR-340-3p). Of these, five (p ≤ 0.01) were selected for in silico analysis (miR-10a-5p; miR-29b-2-5p; miR-125a-5p; miR-342-3p, miR-708-5p), for which 44 relevant target genes were found regulated by these c-miRNAs and related to LOAD. Through the analysis of these target genes in databases, it was possible to observe that they have functions related to the development and progress of LOAD, directly or indirectly connecting the different Alzheimer's pathways. Thus, this work found five promising serum c-miRNAs as options for biomarkers contributing to LOAD diagnosis. Our study shows the complex network between these molecules and LOAD, supporting the relevance of studies using c-miRNAs in dementia contexts.
Assuntos
Doença de Alzheimer , Biomarcadores , MicroRNAs , Humanos , Doença de Alzheimer/sangue , Doença de Alzheimer/genética , Doença de Alzheimer/diagnóstico , MicroRNAs/sangue , MicroRNAs/genética , Masculino , Idoso , Biomarcadores/sangue , Feminino , Idoso de 80 Anos ou maisRESUMO
The growing amount of genome information and transcriptomes data available allows for a better understanding of biological processes. However, analysis of complex transcriptomic experimental designs involving different conditions, tissues, or times is relevant. This study proposes a novel approach to analyze complex data sets combining transcriptomes and miRNAs at the chromosome-level genome. Atlantic salmon smolts were transferred to seawater under two strategies: (i) fish group exposed to gradual salinity changes (GSC) and (ii) fish group exposed to a salinity shock (SS). Gills, intestine, and head kidney samples were used for total RNA extraction, followed by mRNA and small RNA illumina sequencing. Different expression patterns among the tissues and treatments were observed through a whole-genome transcriptomic approach. Chromosome regions highly expressed between experimental conditions included a great abundance of transposable elements. In addition, differential expression analysis showed a greater number of transcripts modulated in response to SS in gills and head kidney. miRNA expression analysis suggested a small number of miRNAs involved in the smoltification process. However, target analysis of these miRNAs showed a regulatory role in growth, stress response, and immunity. This study is the first to evidence the interplaying among mRNAs and miRNAs and the structural relationship at the genome level during Atlantic salmon smoltification.
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Axonal protein synthesis has been shown to play a role in developmental and regenerative growth, as well as in the maintenance of the axoplasm in a steady state. Recent studies have begun to identify the mRNAs localized in axons, which could be translated locally under different conditions. Despite that by now hundreds or thousands of mRNAs have been shown to be localized into the axonal compartment of cultured neurons in vitro, knowledge of which mRNAs are localized in mature myelinated axons is quite limited. With the purpose of characterizing the transcriptome of mature myelinated motor axons of peripheral nervous systems, we modified the axon microdissection method devised by Koenig, enabling the isolation of the axoplasm RNA to perform RNA-seq analysis. The transcriptome analysis indicates that the number of RNAs detected in mature axons is lower in comparison with in vitro data, depleted of glial markers, and enriched in neuronal markers. The mature myelinated axons are enriched for mRNAs related to cytoskeleton, translation, and oxidative phosphorylation. Moreover, it was possible to define core genes present in axons when comparing our data with transcriptomic data of axons grown in different conditions. This work provides evidence that axon microdissection is a valuable method to obtain genome-wide data from mature and myelinated axons of the peripheral nervous system, and could be especially useful for the study of axonal involvement in neurodegenerative pathologies of motor neurons such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophies (SMA).
Assuntos
Esclerose Lateral Amiotrófica/genética , Neurônios Motores/metabolismo , Atrofia Muscular Espinal/genética , RNA/genética , Esclerose Lateral Amiotrófica/metabolismo , Animais , Axônios/metabolismo , Axônios/patologia , Diferenciação Celular/genética , Perfilação da Expressão Gênica , Humanos , Microdissecção , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/patologia , Sistema Nervoso Periférico/metabolismo , Sistema Nervoso Periférico/patologia , RNA Mensageiro/genética , RNA-Seq , Transcriptoma/genéticaRESUMO
Cancer is a complex disease, and its study requires deep understanding of several biological processes and their regulation. It is an accepted fact that non-coding RNAs are vital components of the regulation and cross-talk among cancer-related signaling pathways that favor tumor aggressiveness and metastasis, such as neovascularization, angiogenesis, and vasculogenic mimicry. Both long non-coding RNAs (lncRNAs) and micro-RNAs (miRNAs) have been described as master regulators of cancer on their own; yet there is accumulating evidence that, besides regulating mRNA expression through independent mechanisms, these classes of non-coding RNAs interact with each other directly, fine-tuning the effects of their regulation. While still relatively scant, research on the lncRNA-miRNA-mRNA axis regulation is growing at a fast rate, it is only in the last 5 years, that lncRNA-miRNA interactions have been identified in tumor-related vascular processes. In this review, we summarize the current progress of research on the cross-talk between lncRNAs and miRNAs in the regulation of neovascularization, angiogenesis and vasculogenic mimicry.
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Background: Human biological material has become an important resource for biomedical research. Tumor Biobanks are facilities that collect, store and distribute samples of tumor and normal tissue for further use in basic and translational cancer research. mRNA-translation has been demonstrated to modulate protein levels and is considered a fundamental post-transcriptional mechanism of gene expression regulation. Thus, determining translation efficiencies of individual mRNAs in human tumors may add another layer of information that contributes to the understanding of tumorigenic pathways. To analyze the RNAs actively engaged in translation, RNAs associated with ribosomes (polysomes) are isolated, identified and compared to total RNA. However, the application of this technique in human tumors depends on the stability of the polysomal structure under Biobank storage conditions that usually consists of ultra-low temperature. Since the effect of freezing on the stability of the polysomal structure in stored tumor samples is not known, it is essential to evaluate this factor in the frozen samples, validating the use of biobank samples in studies of translational efficiency. Methods: Xenograft tumors were divided in two parts, half was subject to immediate processing, and half was frozen for posterior analysis. Both parts were subject to polysomal separation, RNA extraction and identification through RNAseq. Results: It was possible to successfully extract and identify total and polysomal RNA from both fresh and frozen tumoral tissue. The quantification of the polysome profile indicated no difference in the translational efficiency estimated in fresh versus frozen tissue. Gene expression data from the fresh versus frozen tissues were compared and the correlation between the polysome associated fresh x frozen (R = 0,89) and total fresh x frozen (0,90) mRNAs was calculated. No difference was identified between the two conditions. Conclusions: We demonstrated that tissue freezing does not affect the polysomal structure, consequently validating the viability of the use of biobank stored tissue for polysome associated RNA analysis (AU)
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Humanos , Polirribossomos , RNA , Expressão Gênica , Regulação da Expressão Gênica , NeoplasiasRESUMO
Investigation on functional genome research may contribute to the knowledge of functional roles of different mRNAs and miRNAs in bone cells of osteoporotic animals. Currently, few studies indicate the changes in gene modulation that osteoporosis causes in osteoblastic cells from different sites. Thus, the purpose of this investigation was to evaluate cell viability, alkaline phosphatase activity and modulation of mRNAs/miRNAs in osteoblastic cells from calvaria and bone marrow by means of microarray technology. Wistar female rats were divided in sham operated and ovariectomized groups. After 150 days of ovariectomy, cells were isolated from both sites to perform cell culture. Results showed that calvaria cells from ovariectomized rats had a decrease in viability when compared to control groups and to bone marrow cells from osteoporotic rats after 3 days. Alkaline phosphatase activity decreased in calvaria cells from ovariectomized rats whereas it was increased in bone marrow osteoblastic cells in the same group. Microarray data analysis showed 5447 differentially expressed mRNAs and 82 differentially expressed miRNAs in calvaria cells. The same way, 4399 mRNAs and 54 miRNAs were expressed in bone marrow cells. mRNAs associated with bone metabolism such as Anxa5, Sp7, Spp1, Notch1 were distinctively modulated in both sites, as well as miRNAs such as miR-350, miR-542-3p, miR-204-5p, and miR-30e-3p. The RNA species identified in this study could be further used as targets for treatment or prevention of osteoporosis.
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Células da Medula Óssea/metabolismo , Menopausa/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Medula Óssea/metabolismo , Diferenciação Celular/genética , Feminino , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Osteoporose/metabolismo , Ovariectomia/métodos , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Crânio/metabolismo , Transcriptoma/genéticaRESUMO
BACKGROUND: Acute respiratory distress syndrome (ARDS) is a potentially devastating form of acute inflammatory lung injury as well as a major cause of acute respiratory failure. Although researchers have made significant progresses in elucidating the pathophysiology of this complex syndrome over the years, the absence of a universal detail disease mechanism up until now has led to a series of practical problems for a definitive treatment. This study aimed to predict some genes or pathways associated with sepsis-related ARDS based on a public microarray dataset and to further explore the molecular mechanism of ARDS. RESULTS: A total of 122 up-regulated DEGs and 91 down-regulated differentially expressed genes (DEGs) were obtained. The up- and down-regulated DEGs were mainly involved in functions like mitotic cell cycle and pathway like cell cycle. Protein-protein interaction network of ARDS analysis revealed 20 hub genes including cyclin B1 (CCNB1), cyclin B2 (CCNB2) and topoisomerase II alpha (TOP2A). A total of seven transcription factors including forkhead box protein M1 (FOXM1) and 30 target genes were revealed in the transcription factor-target gene regulation network. Furthermore, co-cited genes including CCNB2-CCNB1 were revealed in literature mining for the relations ARDS related genes. CONCLUSIONS: Pathways like mitotic cell cycle were closed related with the development of ARDS. Genes including CCNB1, CCNB2 and TOP2A, as well as transcription factors like FOXM1 might be used as the novel gene therapy targets for sepsis related ARDS.
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Estudos de Associação Genética , Síndrome do Desconforto Respiratório/genética , Sepse/complicações , Sepse/genética , Transcriptoma , Ciclo Celular/genética , Bases de Dados Genéticas , Regulação para Baixo , Perfilação da Expressão Gênica , Marcação de Genes , Humanos , Mapas de Interação de Proteínas , Fatores de Transcrição , Regulação para CimaRESUMO
BACKGROUND: Acute respiratory distress syndrome (ARDS) is a potentially devastating form of acute inflammatory lung injury as well as a major cause of acute respiratory failure. Although researchers have made significant progresses in elucidating the pathophysiology of this complex syndrome over the years, the absence of a universal detail disease mechanism up until now has led to a series of practical problems for a definitive treatment. This study aimed to predict some genes or pathways associated with sepsis-related ARDS based on a public microarray dataset and to further explore the molecular mechanism of ARDS. RESULTS: A total of 122 up-regulated DEGs and 91 down-regulated differentially expressed genes (DEGs) were obtained. The up- and down-regulated DEGs were mainly involved in functions like mitotic cell cycle and pathway like cell cycle. Protein-protein interaction network of ARDS analysis revealed 20 hub genes including cyclin B1 (CCNB1), cyclin B2 (CCNB2) and topoisomerase II alpha (TOP2A). A total of seven transcription factors including forkhead box protein M1 (FOXM1) and 30 target genes were revealed in the transcription factor-target gene regulation network. Furthermore, co-cited genes including CCNB2-CCNB1 were revealed in literature mining for the relations ARDS related genes. CONCLUSIONS: Pathways like mitotic cell cycle were closed related with the development of ARDS. Genes including CCNB1, CCNB2 and TOP2A, as well as transcription factors like FOXM1 might be used as the novel gene therapy targets for sepsis related ARDS
Assuntos
Humanos , Transtornos Respiratórios/genética , Sepse/complicações , Sepse/genética , Estudos de Associação Genética , Transcriptoma , Fatores de Transcrição , Regulação para Baixo , Ciclo Celular/genética , Regulação para Cima , Marcação de Genes , Perfilação da Expressão Gênica , Bases de Dados Genéticas , Mapas de Interação de ProteínasRESUMO
Duplicação genica é uma das principais forças levando a evolução dos genomas eucarioto. O impacto de duplicações gênicas/genômicas vem sendo investigado a muito tempo em humanos e outros primatas. Um segundo mecanismo de duplicação gênica, a retrotransposição baseada em RNA maduros, vem sendo menos estudada devido ao seu potencial menor de gerar cópias funcionais. No entanto, recentemente, publicações descreveram retrocópias funcionais em humanos, roedores e mosca de fruta. Nesta tese, para investigar sobre retrocópias causando variabilidade genética no genoma de primatas, nós desenvolvemos a implementamos os métodos para detectar estas inserções. Utilizando nove genomas e transcriptomas publicamente disponíveis (sete primatas e dois roedores) nós confirmamos um número similar, porém, com origem independente, de retrocópias em primatas e roedores. Nós também encontramos um enriquecimento de retrocópias no genoma de Platyrrhini, possivelmente explicado pela expansão de L1PA7 e L1P3 nestes genomas. Posteriormente, nós analisamos a ortologia de retrocópias no genoma de primatas e encontramos 127 eventos específicos à linhagem humana. Nós também exploramos dados do projeto 1000 Genomes para detectar retrocópias polimórficas (retroCNVs germinativos) e encontramos 17 eventos, presentes no genoma referência humano, mas ausentes em mais de um indivíduo. Similarmente, nós investigamos novas retroduplicações de mRNAs no genoma humano, detectando 21 eventos ausentes do genoma referência. Finalmente, investigamos a existência de retroCNVs somáticos e descrevemos sete possíveis retrocópias somáticas. Apesar de sua possível insignificância, nós encontramos que algumas retrocópias compartilhadas entre todos os primatas, espécie específicas, e polimórficas podem ser expressas per se ou como transcritos quiméricos com genes hospedeiros. Sobretudo, nós encontramos que retrocópias são um fator importante da variabilidade genética inter-espécie, intra-espécie e intra-indivíduo e podem estar influenciando a evolução de mamíferos ao criar reservatórios de duplicações potencialmente funcionais
Gene duplication is a major driving force of evolution in eukaryotic genome. The impact of gene/genomic duplication has long been investigated in human and other primates. A second mechanism of gene duplication, retrotransposition, which is based on mature RNA, has been traditionally less studied due to their lower potential to generate functional copies. Recently, however, publications described functional retrocopies in humans, murines and drosophila. Here, to gain insights of the genetic variability arising from retrocopies on primate genomes, we developed and implemented the methods to detect these insertions. Using nine publicly available reference genomes and transcriptomes (seven primates and two rodents) we described a similar number independently arisen retrocopies in primates and rodents. We also found an enrichment of retrocopies in Platyrhinni genomes, putatively explained by the expansion of L1PA7 and L1P3 in these genomes. Next, we evaluated the orthology of retrocopies in primate genomes and found 127 events specific to human lineage. We also explored 1000 Genomes Project data to detect polymorphic events (germinative retroCNVs) on human populations and found 17 events, present on the reference genome, absent in more than one individual. Conversely, we also investigated new insertions of mRNA retroduplications in the human genome, detecting 21 events absent to the human reference genome. Finally, we evaluated the existence of somatic retroCNVs and described seven putative somatic retrocopies. Despite their putative insignificance, we found that some of these shared, specie-specific and polymorphic events may be expressed per se and as chimeric transcripts within host genes. Taken together, we found that retrocopies are a great factor of genetic variation interspecie, intraspecie e intraindividual and may be affecting mammal evolution by creating reservoirs of potentially functional duplications