Detection of Atherosclerosis by Small RNA-Sequencing Analysis of Extracellular Vesicle Enriched Serum Samples

Abstract

Atherosclerosis can occur throughout the arterial vascular system and lead to various diseases. Early diagnosis of atherosclerotic processes and of individual disease patterns would be more likely to be successful if targeted therapies were available. For this, it is important to find reliable biomarkers that are easily accessible and with little inconvenience for patients. There are many cell culture, animal model or tissue studies that found biomarkers at the microRNA (miRNA) and mRNA level describing atherosclerotic processes. However, little is known about their potential as circulating and liquid biopsy markers in patients. In this study, we examined serum-derived miRNA – profiles from 129 patients and 28 volunteers to identify potential biomarkers. The patients had four different atherosclerotic manifestations: abdominal aneurysm (n = 35), coronary heart disease (n = 34), carotid artery stenosis (n = 24) and peripheral arterial disease (n = 36). The samples were processed with an extracellular vesicle enrichment protocol, total-RNA extraction and small RNA-sequencing were performed. A differential expression analysis was performed bioinformatically to find potentially regulated miRNA biomarkers. Resulting miRNA candidates served as a starting point for an overrepresentation analysis in which relevant target mRNAs were identified. The Gene Ontology database revealed relevant biological functions in relation to atherosclerotic processes. In patients, expression of specific miRNAs changed significantly compared to healthy volunteers; 27 differentially expressed miRNAs were identified. We were able to detect a group-specific miRNA fingerprint: miR-122-5p, miR-2110 and miR-483-5p for abdominal aortic aneurysm, miR-370-3p and miR-409-3p for coronary heart disease, miR-335-3p, miR-381-3p, miR493-5p and miR654-3p for carotid artery stenosis, miR-199a-5p, miR-215-5p, miR-3168, miR-582-3p and miR-769-5p for peripheral arterial disease. The results of the study show that some of the identified miRNAs have already been associated with atherosclerosis in previous studies. Overrepresentation analysis on this data detected biological processes that are clearly relevant for atherosclerosis, its development and progression showing the potential of these miRNAs as biomarker candidates. In a next step, the relevance of these findings on the mRNA level is to be investigated and substantiated.

Circulating microRNA signatures associated with disease severity and outcome in COVID-19 patients

“…In the literature, miR-320 family members were found as upregulated in plasma of patients with COVID-19 and especially in those with severe COVID-19 compared to those with moderate disease ( 17 ). High levels of miR-320b and miR-483-5p were also associated with increased risk of in-hospital mortality in COVID-19 patients ( 41 ) and upregulated in extracellular vesicles-enriched sera of atherosclerotic patients, indicating a possible role in vascular and endothelial injury ( 42 ). In our study, the levels of serum miR-320 family showed a positive correlation with inflammatory and tissue injury biomarkers, suggesting a role for these miRNAs in inflammatory response.…”

“…Circulating isomiR signatures in COVID-19 patients according to disease severity -.2IsomiRs are miRNA isoforms produced during miRNA maturation that differ from their canonical counterpart in length at their 3' or 5' end and/or in their internal sequence. IsomiRs have variable expression in different organs and tissue types and may differ in their targeted mRNA spectrum (42). Since small RNA sequencing can identify isomiRs, we searched for DE isomiRs in our dataset.…”

Expression profiles and functions of ferroptosis-related genes in intimal hyperplasia induced by carotid artery ligation in mice

“…It is evident that miRNA-335-3p plays an important role because it regulates the expression of three important genes simultaneously. The available literature indicates that there are few studies on miR-335-3p, especially in the cardiovascular field, which mainly focus on cardiac development ( Kay and Soltani, 2019 ), pulmonary hypertension ( Fan and Fan, 2020 ), and atherosclerosis ( Hildebrandt and Kirchner, 2021 ). Moreover, Sun et al found that the overexpression of miR-185-5p could suppress the proliferation and migration of VSMCs by targeting FRS2 ( Sun and Li, 2021 ).…”

Applications of Extracellular Vesicles in Abdominal Aortic Aneurysm

“…Diverse miRNAs can be selectively packed into EVs and transferred from cells to cells to modulate disease-related processes. Recent research enriched EVs from serum of AAA patients and performed small RNA-sequencing to identify potential miRNA biomarkers ( 29 ). It turns out that the expressions of miR-122-5p, miR-2110 and miR-483-5p are upregulated in AAA patients' serum.…”