Circulating microRNA
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OBJECTIVES: MicroRNAs (miRNAs) are short single-stranded RNAs that play a role in the post-transcriptional regulation of gene expression. Their deregulation can be associated with various diseases, such as cancer, neurodegenerative, and immune-related diseases. The aim of our study was to compare miRNA levels in plasma that could potentially influence the progression of hyperuricemia to gout, since the mechanism of progression is still unclear. METHODS: Total RNA, including miRNA, was isolated from the plasma of 45 patients with asymptomatic hyperuricemia, 131 patients with primary gout (including 16 patients having a gout attack), and 130 normouricemic controls. The expression of 18 selected miRNAs (cel-miR-39 and cel-miR-54 as spike-in controls, hsa-miR-16-5p and hsa-miR-25-3p as endogenous controls, hsa-miR-17-5p, hsa-miR-18a-5p, hsa-miR-30a-3p, hsa-miR-30c-5p, hsa-miR-126-3p, hsa-miR-133a-3p, hsa-miR-142-3p, hsa-miR-143-3p, hsa-miR-146a-5p, hsa-miR-155-5p, hsa-miR-222-3p, hsa-miR-223-3p, hsa-miR-488-3p and hsa-miR-920) was measured using qPCR. RESULTS: We found that hsa-miR-17-5p, hsa-miR-18a-5p, hsa-miR-30c-5p, hsa-miR-142-3p, and hsa-miR-223-3p were significantly upregulated (p < 0.001) in the plasma of hyperuricemia and gout patients compared to normouricemic individuals. As part of the follow-up of our previous study, we found a negative correlation between hsa-miR-17-5p, hsa-miR-30c-5p, hsa-miR-126-3p, hsa-miR-142-3p, and hsa-miR-223-3p with plasma levels of chemokine MCP-1. Additionally, we found a positive correlation between CRP and plasma levels of hsa-miR-17-5p, hsa-miR-18a-5p, hsa-miR-30c-5p, hsa-miR-126-3p, hsa-miR-142-3p, hsa-miR-146a-5p, hsa-miR-155-5p, hsa-miR-222-3p, and hsa-miR-223-3p. Five of those miRNAs (hsa-miR-126-3p, hsa-miR-142-3p, hsa-miR-146a-5p, hsa-miR-155-5p, and hsa-miR-222-3p) also had a positive correlation with serum creatinine and therefore a negative correlation with eGFR. CONCLUSION: Five miRNAs were significantly upregulated in the plasma of patients with hyperuricemia and gout (and those during a gout attack) compared to normouricemic controls. We also found a correlation between the plasma levels of several miRNA and plasma levels of MCP-1, CRP, serum creatinine, and eGFR.
In this study, the circulating miRNome from diagnostic neuroblastoma serum was assessed for identification of noninvasive biomarkers with potential in monitoring metastatic disease. After determining the circulating neuroblastoma miRNome, 743 miRNAs were screened in 2 independent cohorts of 131 and 54 patients. Evaluation of serum miRNA variance in a model testing for tumor stage, MYCN status, age at diagnosis, and overall survival revealed tumor stage as the most significant factor impacting miRNA abundance in neuroblastoma serum. Differential abundance analysis between patients with metastatic and localized disease revealed 9 miRNAs strongly associated with metastatic stage 4 disease in both patient cohorts. Increasing levels of these miRNAs were also observed in serum from xenografted mice bearing human neuroblastoma tumors. Moreover, murine serum miRNA levels were strongly associated with tumor volume. These findings were validated in longitudinal serum samples from metastatic neuroblastoma patients, where the 9 miRNAs were associated with disease burden and treatment response.
- MeSH
- cirkulující mikroRNA krev MeSH
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- metastázy nádorů diagnóza MeSH
- mikro RNA krev MeSH
- mladiství MeSH
- mladý dospělý MeSH
- myši MeSH
- nádorové biomarkery krev MeSH
- neuroblastom krev diagnóza MeSH
- předškolní dítě MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- staging nádorů MeSH
- transplantace heterologní MeSH
- zvířata MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- myši MeSH
- předškolní dítě MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Methods in molecular biology. Springer protocols (Series), ISSN 1064-3745 1024
xi, 270 stran : grafy a tabulky ; 28 cm
Poor outcome of extramedullary disease in multiple myeloma patients and lack of outcome predictors prompt continued search for new markers of the disease. In this report, we show circulating microRNA distinguishing multiple myeloma patients with extramedullary disease from myeloma patients without such manifestation and from healthy donors. MicroRNA-130a was identified by TaqMan Low Density Arrays and verified by quantitative PCR on 144 serum samples (59 multiple myeloma, 55 myeloma with extramedullary disease, 30 healthy donors) in test and validation cohorts as being down-regulated in myeloma patients with extramedullary disease. Circulating microRNA-130a distinguished myeloma patients with extramedullary disease from healthy donors with specificity of 90.0% and sensitivity of 77.1%, patients with extramedullary disease from newly diagnosed multiple myeloma patients with specificity of 77.1% and sensitivity of 34.3% in the test cohort and with specificity of 91.7% and sensitivity of 30.0% in the validation cohort of patients. Circulating microRNA-130a in patients with extramedullary myeloma was associated with bone marrow plasma cells infiltration. Further, microRNA-130a was decreased in bone marrow plasma cells obtained from patients with extramedullary myeloma in comparison to bone marrow plasma cells of myeloma patients without such manifestation, but it was increased in tumor site plasma cells of patients with extramedullary disease compared to bone marrow plasma cells of such patients (p<0.0001). Together, our data suggest connection between lower level of microRNA-130a and extramedullary disease and prompt further work to evaluate this miRNA as a marker of extramedullary disease in multiple myeloma.
- MeSH
- dospělí MeSH
- kostní dřeň patologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- mikro RNA analýza krev genetika MeSH
- mnohočetný myelom krev diagnóza genetika patologie MeSH
- nádorové biomarkery analýza krev genetika MeSH
- plazmatické buňky patologie MeSH
- prognóza MeSH
- regulace genové exprese u nádorů * MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Circulating cell-free microRNAs are promising candidates for minimally invasive clinical biomarkers for the diagnosis, prognosis and monitoring of many human diseases. Despite substantial efforts invested in the field, the research so far has failed to deliver expected results. One of the contributing factors is general lack of agreement between various studies, partly due to the considerable technical challenges accompanying the workflow. Pre-analytical variables including sample collection, RNA isolation, and quantification are sources of bias that may hamper biological interpretation of the results. Here, we present a Two-tailed RT-qPCR panel for quality control, monitoring of technical performance, and optimization of microRNA profiling experiments from biofluid samples. The Two-tailed QC (quality control) panel is based on two sets of synthetic spike-in molecules and three endogenous microRNAs that are quantified with the highly specific Two-tailed RT-qPCR technology. The QC panel is a cost-effective way to assess quality of isolated microRNA, degree of inhibition, and erythrocyte contamination to ensure technical soundness of the obtained results. We provide assay sequences, detailed experimental protocol and guide to data interpretation. The application of the QC panel is demonstrated on the optimization of RNA isolation from biofluids with the miRNeasy Serum/Plasma Advanced Kit (Qiagen).
- MeSH
- analýza nákladů a výnosů MeSH
- biologické markery krev MeSH
- cirkulující mikroRNA krev izolace a purifikace MeSH
- krysa rodu rattus MeSH
- kvantitativní polymerázová řetězová reakce ekonomika přístrojové vybavení metody normy MeSH
- lidé MeSH
- reagenční diagnostické soupravy normy MeSH
- řízení kvality * MeSH
- studie proveditelnosti MeSH
- zdraví dobrovolníci pro lékařské studie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVE: Hand osteoarthritis (OA) is a frequent musculoskeletal disorder with an increasing prevalence during ageing. This study aimed to evaluate circulating microRNAs (miRNAs) in the plasma of patients with hand OA compared with age- and sex-matched healthy control subjects. METHODS: In total, 238 participants (96 with erosive and 73 with non-erosive hand OA patients and 69 healthy control subjects) were included in this study. All patients underwent clinical examinations, including self-reported measures (AUSCAN and Algofunctional index). Radiographs of both hands were scored with the Kallman scale. The profile of miRNAs in plasma was screened using TaqManTM Low-Density Array, and candidate miRNAs were validated on two quantitative real-time PCR (qRT-PCR) systems (QuantStudio and SmartChip). RESULTS: Of all the 754 miRNAs, 40 miRNAs were different between hand OA patients and healthy control subjects in the screening cohort. Following the two-phase validation process, three miRNAs (miR-23a-3p, miR-146a-5p, and miR-652-3p) were increased in patients with hand OA compared with healthy control subjects and were associated with the AUSCAN sum score and AUSCAN pain. Furthermore, an inverse correlation of miR-222-3p with the Kallman radiographic score was found. The expression of miRNAs did not differ between erosive and non-erosive hand OA. CONCLUSION: The profile of circulating miRNAs could unveil candidate biomarkers associated with hand OA symptoms. Longitudinal studies are required to determine the role of miRNAs in hand OA.
- MeSH
- biologické markery MeSH
- bolest MeSH
- cirkulující mikroRNA * MeSH
- lidé MeSH
- mikro RNA * MeSH
- osteoartróza * diagnostické zobrazování genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
New non-invasive approaches that can complement and improve on current strategies for colorectal cancer (CRC) screening and management are urgently needed. A growing number of publications have documented that components of tumors, which are shed into the circulation, can be detected in the form of liquid biopsies and can be used to detect CRC at early stages, to predict response to certain therapies and to detect CRC recurrence in a minimally invasive way. The analysis of circulating tumor DNA (ctDNA), tumor-derived cells (CTC, circulating tumor cells) or circulating microRNA (miRNA) in blood and other body fluids, have a great potential to improve different aspects of CRC management. The challenge now is to find which types of components, biofluids and detection methods would be the most suitable to be applied in the different steps of CRC detection and treatment. This chapter will provide an up to date review on ctDNA, CTCs and circulating miRNAs as new biomarkers for CRC, either for clinical management or early detection, highlighting their advantages and limitations.
- MeSH
- časná detekce nádoru MeSH
- cirkulující mikroRNA MeSH
- cirkulující nádorová DNA MeSH
- kolorektální nádory krev diagnóza etiologie terapie MeSH
- lidé MeSH
- management nemoci MeSH
- nádorové biomarkery krev MeSH
- nádorové cirkulující buňky MeSH
- prognóza MeSH
- tekutá biopsie metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
BACKGROUND: The aim of the study was to compare selected extracellular miRNA levels (miR-16, miR-21, miR-93 and miR-222 with the response to 8-week-long explosive strength training (EXPL), hypertrophic strength training (HYP) and high-intensity interval training (HIIT). METHODS: 30 young male athletes of white European origin (mean age: 22.5 ± 4.06 years) recruited at the Faculty of Sports Studies of Masaryk University were enrolled in this study. The study participants were randomly assigned to three possible training scenarios: EXPL, HYP or HITT and participated in 8-week-long program in given arm. Blood plasma samples were collected at the baseline and at week 5 and 8 and anthropometric and physical activity parameters were measured. Pre- and post-intervention characteristics were compared and participants were further evaluated as responders (RES) or non-responders (NRES). RES/NRES status was established for the following characteristics: 300°/s right leg extension (t300), 60°/s right leg extension (t60), isometric extension (IE), vertical jump, isometric extension of the right leg and body fat percentage (BFP). RESULTS: No differences in miRNA levels were apparent between the intervention groups at baseline. No statistically significant prediction role was observed using crude univariate stepwise regression model analysis where RES/NRES status for t300, t60, IE, vertical jump and pFM was used as a dependent variable and miR-21, miR-222, miR-16 and miR-93 levels at baseline were used as independent variables. The baseline levels of miR-93 expressed an independent prediction role for responder status based on isometric extension of the right leg (beta estimate 0.76, 95% CI: -0.01; 1.53, p = 0.052). DISCUSSION: The results of the study indicate that 8-week-long explosive strength training, hypertrophic strength training and high-intensity interval training regimens are associated with significant changes in miR-16, mir-21, miR-222 and miR-93 levels compared to a baseline in athletic young men.
- MeSH
- cvičení * MeSH
- dospělí MeSH
- lidé MeSH
- mikro RNA genetika MeSH
- mladý dospělý MeSH
- sporty * MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
MicroRNAs (miRNAs) are evolutionarily conserved small non-coding RNAs that regulate expression of protein-coding genes involved in important biological processes and (patho)physiological states. Circulating miRNAs are protected against degradation, indicating their relevant biological functions. Many studies have demonstrated an association of the specific profile of circulating miRNAs with a wide range of cancers as well as non-malignant diseases. These findings demonstrate the implication of circulating miRNAs in the pathogenesis of diseases and their potential as non-invasive disease biomarkers. However, methods for measurement of circulating miRNAs have critical technical hotspots, resulting in a discrepancy of the reported results and difficult definition of consensus disease biomarkers that may be implicated in clinical use. Here, we review functions of circulating miRNAs and their aberrant expression in particular diseases. Further, we discuss methodological aspects of their detection and quantification as well as our experience with the methods.
Small RNA-sequencing (RNA-Seq) is being increasingly used for profiling of circulating microRNAs (miRNAs), a new group of promising biomarkers. Unfortunately, small RNA-Seq protocols are prone to biases limiting quantification accuracy, which motivated development of several novel methods. Here, we present comparison of all small RNA-Seq library preparation approaches that are commercially available for quantification of miRNAs in biofluids. Using synthetic and human plasma samples, we compared performance of traditional two-adaptor ligation protocols (Lexogen, Norgen), as well as methods using randomized adaptors (NEXTflex), polyadenylation (SMARTer), circularization (RealSeq), capture probes (EdgeSeq), or unique molecular identifiers (QIAseq). There was no single protocol outperforming others across all metrics. Limited overlap of measured miRNA profiles was documented between methods largely owing to protocol-specific biases. Methods designed to minimize bias largely differ in their performance, and contributing factors were identified. Usage of unique molecular identifiers has rather negligible effect and, if designed incorrectly, can even introduce spurious results. Together, these results identify strengths and weaknesses of all current methods and provide guidelines for applications of small RNA-Seq in biomarker research.
