scholarly journals Small RNA sequencing of extracellular vesicles identifies circulating miRNAs related to inflammation and oxidative stress in HIV patients

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Sukrutha Chettimada ◽  
David R. Lorenz ◽  
Vikas Misra ◽  
Steven M. Wolinsky ◽  
Dana Gabuzda
Keyword(s):  
Oxidative Stress ◽  
Rna Sequencing ◽  
Extracellular Vesicles ◽  
Small Rna ◽  
Stress Responses ◽  
Hiv Positive ◽  
Small Rna Sequencing ◽  
And Oxidative Stress ◽  
Potential Biomarkers ◽  
Hiv Negative

Abstract Background Extracellular vesicles (EVs) are nano-sized particles secreted by most cells. EVs carry nucleic acids that hold promise as potential biomarkers in various diseases. Human immunodeficiency virus type 1 (HIV) infects CD4+ T cells and induces immune dysfunction, inflammation, and EV secretion, but little is known about EV small RNA cargo in relation to immune dysregulation in HIV-infected individuals. Here, we characterize small RNA carried by circulating EVs in HIV-positive subjects on antiretroviral therapy (ART) relative to uninfected controls by next-generation RNA sequencing. Results Plasma EVs isolated from HIV-positive and HIV-negative subjects in test (n = 24) and validation (n = 16) cohorts were characterized by electron microscopy, nanoparticle tracking analysis, and immunoblotting for exosome markers. EVs were more abundant in plasma from HIV-positive compared to HIV-negative subjects. Small RNA sequencing of plasma EVs in the test cohort identified diverse small RNA species including miRNA, piRNA, snRNA, snoRNA, tRNA, and rRNA, with miRNA being the most abundant. A total of 351 different miRNAs were detected in plasma EVs, with the top 50 miRNAs accounting for 90% of all miRNA reads. miR-26a-5p was the most abundant miRNA, followed by miR-21-5p and miR-148-3p. qRT-PCR analysis showed that six miRNAs (miR-10a-5p, − 21-5p, −27b-3p, − 122-5p, −146a-5p, − 423-5p) were significantly increased in plasma EVs from HIV-positive compared to HIV-negative subjects in the validation cohort. Furthermore, miR-21-5p, −27b-3p, −146a-5p, and − 423-5p correlated positively with metabolite markers of oxidative stress and negatively with anti-inflammatory polyunsaturated fatty acids. Over-representation and pathway enrichment analyses of miRNAs and their target genes predicted functional association with oxidative stress responses, interferon gamma signaling, Toll-like receptor signaling, TGF beta signaling, and Notch signaling. Conclusions HIV-positive individuals on ART have increased abundance of circulating EVs carrying diverse small RNAs, with miRNAs being the most abundant. Several miRNAs associated with inflammation and oxidative stress are increased in circulating EVs of HIV-positive individuals, representing potential biomarkers of targetable pathways that contribute to disease pathogenesis.

scholarly journals Integrated transcriptome and small RNA sequencing analyses reveal a drought stress response network in Sophora tonkinensis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Ying Liang ◽  
Kunhua Wei ◽  
Fan Wei ◽  
Shuangshuang Qin ◽  
Chuanhua Deng ◽  
...  
Keyword(s):  
Drought Stress ◽  
Rna Sequencing ◽  
Small Rna ◽  
Stress Responses ◽  
Molecular Mechanisms ◽  
Differentially Expressed ◽  
Small Rna Sequencing ◽  
Severe Drought ◽  
A Genome ◽  
Sophora Tonkinensis

Abstract Background Sophora tonkinensis Gagnep is a traditional Chinese medical plant that is mainly cultivated in southern China. Drought stress is one of the major abiotic stresses that negatively impacts S. tonkinensis growth. However, the molecular mechanisms governing the responses to drought stress in S. tonkinensis at the transcriptional and posttranscriptional levels are not well understood. Results To identify genes and miRNAs involved in drought stress responses in S. tonkinensis, both mRNA and small RNA sequencing was performed in root samples under control, mild drought, and severe drought conditions. mRNA sequencing revealed 66,476 unigenes, and the differentially expressed unigenes (DEGs) were associated with several key pathways, including phenylpropanoid biosynthesis, sugar metabolism, and quinolizidine alkaloid biosynthesis pathways. A total of 10 and 30 transcription factors (TFs) were identified among the DEGs under mild and severe drought stress, respectively. Moreover, small RNA sequencing revealed a total of 368 miRNAs, including 255 known miRNAs and 113 novel miRNAs. The differentially expressed miRNAs and their target genes were involved in the regulation of plant hormone signal transduction, the spliceosome, and ribosomes. Analysis of the regulatory network involved in the response to drought stress revealed 37 differentially expressed miRNA-mRNA pairs. Conclusion This is the first study to simultaneously profile the expression patterns of mRNAs and miRNAs on a genome-wide scale to elucidate the molecular mechanisms of the drought stress responses of S. tonkinensis. Our results suggest that S. tonkinensis implements diverse mechanisms to modulate its responses to drought stress.

336 Small RNA Sequencing of Glioblastoma Multiforme Extracellular Vesicles

Neurosurgery ◽  
2016 ◽  
Vol 63 ◽  
pp. 198 ◽  
Author(s):  
Tristan de Mooij ◽  
Brandon A. McCutcheon ◽  
Alexey A. Leontovich ◽  
Ian F. Parney
Keyword(s):  
Glioblastoma Multiforme ◽  
Rna Sequencing ◽  
Extracellular Vesicles ◽  
Small Rna ◽  
Small Rna Sequencing

scholarly journals Small RNA sequencing reveals the role of pearl millet miRNAs and their targets in salinity stress responses

2020 ◽  
Vol 132 ◽  
pp. 395-402
Author(s):  
Harshraj Shinde ◽  
Ambika Dudhate ◽  
Lakshay Anand ◽  
Daisuke Tsugama ◽  
Shashi K. Gupta ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Pearl Millet ◽  
Salinity Stress ◽  
Small Rna ◽  
Stress Responses ◽  
Small Rna Sequencing

scholarly journals Salivary microRNAs identified by small RNA sequencing and machine learning as potential biomarkers of alcohol dependence

Epigenomics ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 739-749 ◽  
Author(s):  
Andrew J Rosato ◽  
Xiaochun Chen ◽  
Yoshiaki Tanaka ◽  
Lindsay A Farrer ◽  
Henry R Kranzler ◽  
...  
Keyword(s):  
Machine Learning ◽  
Alcohol Dependence ◽  
Rna Sequencing ◽  
Small Rna ◽  
Small Rna Sequencing ◽  
Potential Biomarkers

scholarly journals High-Throughput and Automated Acoustic Trapping of Extracellular Vesicles to Identify microRNAs With Diagnostic Potential for Prostate Cancer

2021 ◽  
Vol 11 ◽  
Author(s):  
Anson Ku ◽  
Jacob Fredsøe ◽  
Karina D. Sørensen ◽  
Michael Borre ◽  
Mikael Evander ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Rna Sequencing ◽  
Extracellular Vesicles ◽  
Small Rna ◽  
Rna Extraction ◽  
Clinical Samples ◽  
Small Rna Sequencing ◽  
High Grade ◽  
Enrichment Technique ◽  
Diagnostic Potential

Molecular profiling of extracellular vesicles (EVs) offers novel opportunities for diagnostic applications, but the current major obstacle for clinical translation is the lack of efficient, robust, and reproducible isolation methods. To bridge that gap, we developed a microfluidic, non-contact, and low-input volume compatible acoustic trapping technology for EV isolation that enabled downstream small RNA sequencing. In the current study, we have further automated the acoustic microfluidics-based EV enrichment technique that enables us to serially process 32 clinical samples per run. We utilized the system to enrich EVs from urine collected as the first morning void from 207 men referred to 10-core prostate biopsy performed the same day. Using automated acoustic trapping, we successfully enriched EVs from 199/207 samples (96%). After RNA extraction, size selection, and library preparation, a total of 173/199 samples (87%) provided sufficient materials for next-generation sequencing that generated an average of 2 × 106 reads per sample mapping to the human reference genome. The predominant RNA species identified were fragments of long RNAs such as protein coding and retained introns, whereas small RNAs such as microRNAs (miRNA) accounted for less than 1% of the reads suggesting that partially degraded long RNAs out-competed miRNAs during sequencing. We found that the expression of six miRNAs was significantly different (Padj < 0.05) in EVs isolated from patients found to have high grade prostate cancer [ISUP 2005 Grade Group (GG) 4 or higher] compared to those with GG3 or lower, including those with no evidence of prostate cancer at biopsy. These included miR-23b-3p, miR-27a-3p, and miR-27b-3p showing higher expression in patients with GG4 or high grade prostate cancer, whereas miR-1-3p, miR-10a-5p, and miR-423-3p had lower expression in the GG4 PCa cases. Cross referencing our differentially expressed miRNAs to two large prostate cancer datasets revealed that the putative tumor suppressors miR-1, miR-23b, and miR-27a are consistently deregulated in prostate cancer. Taken together, this is the first time that our automated microfluidic EV enrichment technique has been found to be capable of enriching EVs on a large scale from 900 μl of urine for small RNA sequencing in a robust and disease discriminatory manner.

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