MicroRNAs (miRNAs) are small, non-coding RNAs that regulate various biological processes, primarily through interaction with messenger RNAs. The levels of specific, circulating miRNAs in blood have ...been shown to associate with various pathological conditions including cancers. These miRNAs have great potential as biomarkers for various pathophysiological conditions. In this study we focused on different sample types' effects on the spectrum of circulating miRNA in blood. Using serum and corresponding plasma samples from the same individuals, we observed higher miRNA concentrations in serum samples compared to the corresponding plasma samples. The difference between serum and plasma miRNA concentration showed some associations with miRNA from platelets, which may indicate that the coagulation process may affect the spectrum of extracellular miRNA in blood. Several miRNAs also showed platform dependent variations in measurements. Our results suggest that there are a number of factors that might affect the measurement of circulating miRNA concentration. Caution must be taken when comparing miRNA data generated from different sample types or measurement platforms.
The discovery of microRNAs (miRNAs) as a new class of regulators of gene expression has triggered an explosion of research activities, but has left many unanswered questions about how this regulation ...functions and how it is integrated with other regulatory mechanisms. A number of miRNAs have been found to be present in plasma and other body fluids of humans and mice in surprisingly high concentrations. This observation was unexpected in two respects: first, the fact that these molecules are present at all outside the cell at significant concentrations and second, that these molecules appear to be stable outside of the cell. In light of this it has been suggested that the biological function of miRNAs may also extend outside of the cell and mediate cell-cell communication. We report here that after serum deprivation several human cell lines tested promptly export a substantial amount of miRNAs into the culture medium and the export process is largely energy dependent. The exported miRNAs are found both within and outside of the 16.5 and 120 K centrifugation pellets which contain most of the known cell-derived vesicles, the microvesicles and exosomes. We have identified some candidate proteins involved in this system, and one of these proteins may also play a role in protecting extracellular miRNAs from degradation. Our results point to a hitherto unrecognized and uncharacterized miRNA trafficking system in mammalian cells that is consistent with the cell-cell communication hypothesis.
The microRNA spectrum in 12 body fluids Weber, Jessica A; Baxter, David H; Zhang, Shile ...
Clinical chemistry (Baltimore, Md.),
11/2010, Volume:
56, Issue:
11
Journal Article
Peer reviewed
Open access
MicroRNAs (miRNAs) are small, noncoding RNAs that play an important role in regulating various biological processes through their interaction with cellular messenger RNAs. Extracellular miRNAs in ...serum, plasma, saliva, and urine have recently been shown to be associated with various pathological conditions including cancer.
With the goal of assessing the distribution of miRNAs and demonstrating the potential use of miRNAs as biomarkers, we examined the presence of miRNAs in 12 human body fluids and urine samples from women in different stages of pregnancy or patients with different urothelial cancers. Using quantitative PCR, we conducted a global survey of the miRNA distribution in these fluids.
miRNAs were present in all fluids tested and showed distinct compositions in different fluid types. Several of the highly abundant miRNAs in these fluids were common among multiple fluid types, and some of the miRNAs were enriched in specific fluids. We also observed distinct miRNA patterns in the urine samples obtained from individuals with different physiopathological conditions.
MicroRNAs are ubiquitous in all the body fluid types tested. Fluid type-specific miRNAs may have functional roles associated with the surrounding tissues. In addition, the changes in miRNA spectra observed in the urine samples from patients with different urothelial conditions demonstrates the potential for using concentrations of specific miRNAs in body fluids as biomarkers for detecting and monitoring various physiopathological conditions.
We analyzed the whole-genome sequences of a family of four, consisting of two siblings and their parents. Family-based sequencing allowed us to delineate recombination sites precisely, identify 70% ...of the sequencing errors (resulting in > 99.999% accuracy), and identify very rare single-nucleotide polymorphisms. We also directly estimated a human intergeneration mutation rate of approximately 1.1 x 10⁻⁸ per position per haploid genome. Both offspring in this family have two recessive disorders: Miller syndrome, for which the gene was concurrently identified, and primary ciliary dyskinesia, for which causative genes have been previously identified. Family-based genome analysis enabled us to narrow the candidate genes for both of these Mendelian disorders to only four. Our results demonstrate the value of complete genome sequencing in families.
Biomolecular pathways and networks are dynamic and complex, and the perturbations to them which cause disease are often multiple, heterogeneous and contingent. Pathway and network visualizations, ...rendered on a computer or published on paper, however, tend to be static, lacking in detail, and ill-equipped to explore the variety and quantities of data available today, and the complex causes we seek to understand.
RCytoscape integrates R (an open-ended programming environment rich in statistical power and data-handling facilities) and Cytoscape (powerful network visualization and analysis software). RCytoscape extends Cytoscape's functionality beyond what is possible with the Cytoscape graphical user interface. To illustrate the power of RCytoscape, a portion of the Glioblastoma multiforme (GBM) data set from the Cancer Genome Atlas (TCGA) is examined. Network visualization reveals previously unreported patterns in the data suggesting heterogeneous signaling mechanisms active in GBM Proneural tumors, with possible clinical relevance.
Progress in bioinformatics and computational biology depends upon exploratory and confirmatory data analysis, upon inference, and upon modeling. These activities will eventually permit the prediction and control of complex biological systems. Network visualizations--molecular maps--created from an open-ended programming environment rich in statistical power and data-handling facilities, such as RCytoscape, will play an essential role in this progression.
Drug-induced liver injury is a frequent side effect of many drugs, constitutes a significant threat to patient health and has an enormous economic impact on health care expenditures. Numerous efforts ...have been made to identify reliable and predictive markers to detect the early signs of drug-induced injury to the liver, one of the most vulnerable organs in the body. These studies have, however, not delivered any more informative candidates than the serum aminotransferase markers that have been available for ≈30 years. Using acetaminophen overdose-induced liver injury in the mouse as a model system, we have observed highly significant differences in the spectrum and levels of microRNAs in both liver tissues and in plasma between control and overdosed animals. Based on our survey of microRNA expression among normal tissues, some of the microRNAs, like messenger RNAs, display restricted tissue distributions. A number of elevated circulating microRNAs in plasma collected from acetaminophen-overdosed animals are highly expressed in the liver. We have demonstrated that specific microRNA species, such as mir-122 and mir-192, both are enriched in the liver tissue and exhibit dose- and exposure duration-dependent changes in the plasma that parallel serum aminotransferase levels and the histopathology of liver degeneration, but their changes can be detected significantly earlier. These findings suggest the potential of using specific circulating microRNAs as sensitive and informative biomarkers for drug-induced liver injury.
Although many tools have been developed to analyze small RNA sequencing (sRNA-Seq) data, it remains challenging to accurately analyze the small RNA population, mainly due to multiple sequence ID ...assignment caused by short read length. Additional issues in small RNA analysis include low consistency of microRNA (miRNA) measurement results across different platforms, miRNA mapping associated with miRNA sequence variation (isomiR) and RNA editing, and the origin of those unmapped reads after screening against all endogenous reference sequence databases. To address these issues, we built a comprehensive and customizable sRNA-Seq data analysis pipeline-sRNAnalyzer, which enables: (i) comprehensive miRNA profiling strategies to better handle isomiRs and summarization based on each nucleotide position to detect potential SNPs in miRNAs, (ii) different sequence mapping result assignment approaches to simulate results from microarray/qRT-PCR platforms and a local probabilistic model to assign mapping results to the most-likely IDs, (iii) comprehensive ribosomal RNA filtering for accurate mapping of exogenous RNAs and summarization based on taxonomy annotation. We evaluated our pipeline on both artificial samples (including synthetic miRNA and Escherichia coli cultures) and biological samples (human tissue and plasma). sRNAnalyzer is implemented in Perl and available at: http://srnanalyzer.systemsbiology.net/.
Information theory-based measures of variable dependency (previously published) have been implemented into a software package, MIST. The design of the software and its potential uses are described, ...and a demonstration is presented in the discovery of modifier alleles of the
ApoE
gene in affecting Alzheimer's disease (AD) by analyzing the UK Biobank dataset. The modifier genes uncovered overlap strongly with genes found to be associated with AD. Others include many known to influence AD. We discuss a range of uses of the dependency calculations using MIST that can uncover additional genetic effects in similar complex datasets, like higher degrees of interaction and phenotypic pleiotropy.
Extracellular RNAs (exRNAs) in biofluids have attracted great interest as potential biomarkers. Although extracellular microRNAs in blood plasma are extensively characterized, extracellular messenger ...RNA (mRNA) and long non‐coding RNA (lncRNA) studies are limited. We report that plasma contains fragmented mRNAs and lncRNAs that are missed by standard small RNA‐seq protocols due to lack of 5′ phosphate or presence of 3′ phosphate. These fragments were revealed using a modified protocol (“phospho‐RNA‐seq”) incorporating RNA treatment with T4‐polynucleotide kinase, which we compared with standard small RNA‐seq for sequencing synthetic RNAs with varied 5′ and 3′ ends, as well as human plasma exRNA. Analyzing phospho‐RNA‐seq data using a custom, high‐stringency bioinformatic pipeline, we identified mRNA/lncRNA transcriptome fingerprints in plasma, including tissue‐specific gene sets. In a longitudinal study of hematopoietic stem cell transplant patients, bone marrow‐ and liver‐enriched exRNA genes were tracked with bone marrow recovery and liver injury, respectively, providing proof‐of‐concept validation as a biomarker approach. By enabling access to an unexplored realm of mRNA and lncRNA fragments, phospho‐RNA‐seq opens up new possibilities for plasma transcriptomic biomarker development.
Synopsis
A modified RNA‐seq method (Phospho‐RNA‐seq) revealed a new population of mRNA/lncRNA fragments in plasma, including ones that track with disease. This opens up new possibilities for disease detection via RNA profiling of plasma and other biofluids.
Phospho‐RNA‐seq reveals a large population of mRNA and long non‐coding RNA fragments in human plasma, which are missed by standard small RNA‐seq protocols that depend on target RNAs having a 5′ P and 3′ OH.
Accurate detection of plasma mRNA and lncRNA fragments requires a stringent bioinformatic analysis pipeline to avoid false positive alignments to mRNA and lncRNA genes.
Phospho‐RNA‐seq identified ensembles of tissue‐specific transcripts in plasma of hematopoietic stem cell transplant patients, which show co‐expression patterns that vary dynamically and track with pathophysiological processes.
By enabling access to an unexplored space of extracellular mRNA and lncRNA fragments, phospho‐RNA‐seq opens up new possibilities for monitoring health and disease via transcriptome fragment profiling of plasma and potentially other biofluids.
A modified RNA‐seq method reveals a large population of mRNA/lncRNA fragments in plasma that are missed by standard small RNA‐seq protocols including ones that are associated with disease.
To develop a map of cell-cell communication mediated by extracellular RNA (exRNA), the NIH Extracellular RNA Communication Consortium created the exRNA Atlas resource (https://exrna-atlas.org). The ...Atlas version 4P1 hosts 5,309 exRNA-seq and exRNA qPCR profiles from 19 studies and a suite of analysis and visualization tools. To analyze variation between profiles, we apply computational deconvolution. The analysis leads to a model with six exRNA cargo types (CT1, CT2, CT3A, CT3B, CT3C, CT4), each detectable in multiple biofluids (serum, plasma, CSF, saliva, urine). Five of the cargo types associate with known vesicular and non-vesicular (lipoprotein and ribonucleoprotein) exRNA carriers. To validate utility of this model, we re-analyze an exercise response study by deconvolution to identify physiologically relevant response pathways that were not detected previously. To enable wide application of this model, as part of the exRNA Atlas resource, we provide tools for deconvolution and analysis of user-provided case-control studies.
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•The exRNA Atlas provides access to human exRNA profiles and web-accessible tools•Atlas analysis reveals six exRNA cargo types present across five human biofluids•Five of the cargo types associate with specific vesicular and non-vesicular carriers•These findings and resources empower studies of extracellular RNA communication
An extracellular RNA atlas from five human biofluids (serum, plasma, cerebrospinal fluid, saliva, and urine) reveals six extracellular RNA cargo types, including both vesicular and non-vesicular carriers.
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