Expression Levels Of microRNAs Research Articles

MiR‑137 is a diagnostic tumor‑suppressive miRNA that targets SPHK2 to promote M1‑type tumor‑associated macrophage polarization.

The present study investigated the expression level of microRNA (miR)-137 in glioma tissues and cell lines and explored its potential diagnostic significance as well as its function effects on glioma cells. miR-137 expression level was detected in glioma tissues using in situ hybridization, and in glioma cell lines using reverse transcription-quantitative PCR (RT-qPCR). The diagnostic significance of miR-137 in glioma was assessed using receiver operating characteristic curve analyses. Quantibody® Human Inflammation Array 1 was used to evaluate the impact of ectopic miR-137 expression on release of cytokines in glioma cell lines. IL-13, TNF-α and IFN-γ levels were detected using ELISA. To confirm that sphingosine kinase 2 (SPHK2) is a target of miR-137, RT-qPCR, western blot analysis and dual-luciferase assay were adopted. The results demonstrated that miR-137 expression was downregulated in both glioma tissues and cell lines. Downregulation of miR-137 was significantly associated with high grade gliomas. Additionally, it was found that overexpression of miR-137 reduced IL-13, but promoted TNFα and IFN-γ production. SPHK2 knockdown inhibited IL-13 release, promoted TNF-α and IFN-γ production. SPHK2 was a direct target of miR-137. Collectively, the results of the present study indicated that miR-137 expression plays a tumor-suppressive role in glioma. It is downregulated in glioma and may promote M1-type TAMs polarization, and may be a diagnostic biomarker and potential therapeutic strategy for glioma treatment in the future.

Research Progress of Maternal Metabolism on Cardiac Development and Function in Offspring.

The developmental origin of health and disease (DOHaD) hypothesis refers to the adverse effects of suboptimal developmental environments during embryonic and early fetal stages on the long-term health of offspring. Intrauterine metabolic perturbations can profoundly impact organogenesis in offspring, particularly affecting cardiac development and giving rise to potential structural and functional abnormalities. In this discussion, we contemplate the existing understanding regarding the impact of maternal metabolic disorders, such as obesity, diabetes, or undernutrition, on the developmental and functional aspects of the offspring's heart. This influence has the potential to contribute to the susceptibility of offspring to cardiovascular health issues. Alteration in the nutritional milieu can influence mitochondrial function in the developing hearts of offspring, while also serving as signaling molecules that directly modulate gene expression. Moreover, metabolic disorders can exert influence on cardiac development-related genes epigenetically through DNA methylation, levels of histone modifications, microRNA expression, and other factors. However, the comprehensive understanding of the mechanistic underpinnings of these phenomena remains incomplete. Further investigations in this domain hold profound clinical significance, as they can contribute to the enhancement of public health and the prevention of cardiovascular diseases.

The effect of trehalose administration on the serum expression levels of microRNAs associated with lipid metabolism and autophagy process in patients with myocardial infarction - the post-hoc analysis of the TREAT-IR trial.

IntroductionThis study aimed to evaluate the TRE-induced changes in the serum expression levels of miRNAs associated with lipids metabolism and autophagy process in MI patients to assess the potential protective effects of TRE in these patients.Material and methodsThis post hoc investigation was performed on serum samples obtained from a pilot randomized, double blind, placebo-controlled clinical trial that recruited 14 men (aged 18-80) who had MI and systemic inflammation. The patients were randomized in a 2:1 ratio to either TRE (15 g/week, intravenous (IV) ad-ministration) (N=10) or placebo groups (equal volume of saline 0.9 %) (N=4) for a period of 12 weeks. To measure the relative serum expression levels of miRNA-155, miRNA-221, and miRNA-33, the SYBR Green qPCR method was used.ResultsmiRNA-155 showed significantly higher serum expression levels in the TRE group (2.772±0.73; P=0.009) when compared to the placebo group. Also, significant reductions in miRNA-155 (0.171±0.03; P=0.016), miRNA-221 (0.116±0.07; P=0.013), and miRNA-33a (0.076±0.07; P=0.025) were observed in the placebo group at the end of the study. Nevertheless, the reduction (normalized to the baseline) of the serum expres-sion levels of miRNA-221 (FC:0.87±0.20 vs FC:0.18±0.08; P=0.009) and miRNA-33a (FC:0.73±0.22 vs FC:0.13±0.08; P=0.025) were significantly less in TRE group than in the placebo group.ConclusionsIV TRE administration did not reduce the expression of miRNA-221 and miRNA33a as much as placebo. Keeping a steady state of the serum expression levels of these miRNAs associated with lipoproteins metabolism and autophagy in the TRE group might have protective effects in patients with MI.

CircRUNX1 drives the malignant phenotypes of lung adenocarcinoma through mediating the miR-5195-3p/HMGB3 network.

Circular RNAs (circRNAs) are key factors in the regulation of cancer progression. However, the role of circRUNX1 in lung adenocarcinoma (LUAD) progression is unclear. The expression levels of circRUNX1, microRNA (miR)-5195-3p, and high-mobility group protein B3 (HMGB3) were detected by quantitative real-time PCR. Cell proliferation, migration, invasion and apoptosis were analyzed by EdU staining, colony formation assay, transwell assay and flow cytometry. Protein levels were measured using western blot analysis. The interaction between miR-5195-3p and circRUNX1 or HMGB3 was verified by dual-luciferase reporter assay and RIP assay. Animal experiments were performed to investigate the role of circRUNX1 in LUAD tumorigenesis. We found that circRUNX1 was upregulated in LUAD tumor tissues and cells. CircRUNX1 knockdown suppressed LUAD cell proliferation and metastasis, while promoted apoptosis. In terms of mechanism, we found that circRUNX1 could sponge miR-5195-3p, and miR-5195-3p inhibitor abolished the regulation of circRUNX1 knockdown on LUAD cell proliferation, metastasis and apoptosis. In addition, miR-5195-3p could target HMGB3, and HMGB3 overexpression reversed the inhibition effect of miR-5195-3p on LUAD progression. Moreover, circRUNX1 knockdown reduced LUAD tumorigenesis. CircRUNX1 facilitated LUAD proliferation and metastasis by regulating the miR-5195-3p/HMGB3 axis, suggesting that it might be a possible therapeutic target for LUAD.

Profiling salivary miRNA expression levels in Fanconi anemia patients – a pilot study

The overarching goal of this study is to predict the risk of developing oral squamous cell carcinoma (OSCC) in Fanconi anemia (FA) patients. We have compared the microRNA (miRNA, miR) expression levels in saliva samples from FA patients (n = 50) who are at a low-moderate and/or high risk of developing OSCC to saliva samples from healthy controls (n = 16). The miRNA expression levels in saliva samples were quantified using qPCR. We observed that miR-744, miR-150-5P, and miR-146B-5P had the best discriminatory capacity between FA patients and controls, with an area under the curve (AUC) of 94.0%, 92.9% and 85.3%, respectively. Our data suggest that miR-1, miR-146B-5P, miR-150-5P, miR-155-5P, and miR-744 could be used as panel to predict the risk of developing OSCC in FA patients, with a 89.3% sensitivity and a 68.2% specificity (AUC = 81.5%). Our preliminary data support the notion that the expression levels of salivary miRNAs have the potential to predict the risk of developing OSCC in FA patients and in the future may reduce deaths associated with OSCC.

Relationship between miRNA-21, miRNA-155, and miRNA-182 expression and inflammatory factors in cerebrospinal fluid from patients with multiple sclerosis

BackgroundThe impact of microRNAs (miRNAs) on the differentiation and function of inflammatory cells is well-established. MiRNAs play a crucial role in modulating the expression of pro-inflammatory genes in neuronal cells as well. With this knowledge in mind, our study aimed to explore the relationship between the expression of miRNAs and inflammatory markers in the cerebrospinal fluid (CSF) of patients diagnosed with multiple sclerosis (MS). By investigating this relationship, we aimed to gain insights into the potential involvement of miRNAs in the regulation of inflammation in the context of MS. Materials and methodsThe expression levels of miRNA-21, miRNA-155, and miRNA-182 in cerebrospinal fluid (CSF) samples from multiple sclerosis (MS) patients and controls were determined by RT-PCR. CSF levels of the inflammatory cytokines IL-1β, IL-6, and TNF-α were measured by enzyme-linked immunosorbent assay (ELISA). In addition, high-sensitivity C-reactive protein (hs-CRP) levels were measured by quantitative turbidimetry. ResultsThe expression levels of microRNAs and inflammatory factors were found to be significantly higher in the CSF of MS patients compared to controls (P < 0.05). Receiver operating characteristics (ROC) analysis revealed that miRNA-21, miRNA-182, and miRNA-155 had a high area under the curve (AUC) in discriminating MS patients, with AUC values of 0.97 (P < 0.0001) for miRNA-21, 0.97 (P < 0.0001) for miRNA-182, and 0.96 (P < 0.0001) for miRNA-155. Notably, CSF miRNA-155 showed the highest accuracy in correctly diagnosing MS. Furthermore, a statistically significant relationship was observed between inflammatory cytokines and miRNA-21, miRNA-155 and miRNA-182. ConclusionOur results demonstrated that cerebrospinal fluid (CSF) levels of IL-1β, IL-6, TNF-α, hs-CRP and specific miRNAs serve as biomarkers for assessing central nervous system (CNS) inflammation and neurodegenerative processes in patients with multiple sclerosis (MS).

Altered MicroRNA Maturation in Ischemic Hearts: Implication of Hypoxia on XPO5 and DICER1 Dysregulation and RedoximiR State.

Ischemic cardiomyopathy (ICM) is associated with abnormal microRNA expression levels that involve an altered gene expression profile. However, little is known about the underlying causes of microRNA disruption in ICM and whether microRNA maturation is compromised. Therefore, we focused on microRNA maturation defects analysis and the implication of the microRNA biogenesis pathway and redox-sensitive microRNAs (redoximiRs). Transcriptomic changes were investigated via ncRNA-seq (ICM, n = 22; controls, n = 8) and mRNA-seq (ICM, n = 13; control, n = 10). The effect of hypoxia on the biogenesis of microRNAs was evaluated in the AC16 cell line. ICM patients showed a reduction in microRNA maturation compared to control (4.30 ± 0.94 au vs. 5.34 ± 1.07 au, p ˂ 0.05), accompanied by a deregulation of the microRNA biogenesis pathway: a decrease in pre-microRNA export (XPO5, FC = -1.38, p ˂ 0.05) and cytoplasmic processing (DICER, FC = -1.32, p ˂ 0.01). Both processes were regulated by hypoxia in AC16 cells (XPO5, FC = -1.65; DICER1, FC = -1.55; p ˂ 0.01; Exportin-5, FC = -1.81; Dicer, FC = -1.15; p ˂ 0.05). Patients displayed deregulation of several redoximiRs, highlighting miR-122-5p (FC = -2.41, p ˂ 0.001), which maintained a good correlation with the ejection fraction (r = 0.681, p ˂ 0.01). We evidenced a decrease in microRNA maturation mainly linked to a decrease in XPO5-mediated pre-microRNA export and DICER1-mediated processing, together with a general effect of hypoxia through deregulation of biogenesis pathway and the redoximiRs.

Terfezia boudieri and Terfezia claveryi inhibit the LPS/IFN-γ-mediated inflammation in RAW 264.7 macrophages through an Nrf2-independent mechanism

Desert truffles have been used as traditional treatments for numerous inflammatory disorders. However, the molecular mechanisms underlying their anti-inflammatory effects in RAW 264.7 macrophages have yet to be fully elucidated. The present study investigated the anti-inflammatory activities of two main desert truffles, Terfezia boudieri and T. claveryi, and the underlying mechanisms associated with their anti-inflammatory activities in RAW 264.7 macrophages stimulated with lipopolysaccharide/interferon-gamma (LPS/IFN-γ). Our results demonstrated that treatment with T. boudieri and T. claveryi extracts effectively suppressed the inflammatory response in LPS/IFN-γ-stimulated RAW 264.7 macrophages. Specifically, T. boudieri extract was found to reduce the production of nitric oxide and inhibit the expression of various pro-inflammatory markers, including inducible nitric oxide synthase, cyclooxygenase-2 (COX-2), tumor necrosis factor-α, and interleukin-6 (IL-6) at both the mRNA and protein levels. Similarly, T. claveryi extract exhibited comparable inhibitory effects, except for the expression of IL-6 and COX-2 at the protein level, where no significant effect was observed. Moreover, both studied extracts significantly downregulated the microRNA expression levels of miR-21, miR-146a, and miR-155, suggesting that T. boudieri and T. claveryi suppress the inflammatory response in LPS/IFN-γ-stimulated RAW 264.7 cells through an epigenetic mechanism. Furthermore, our study reveals a new mechanism for the anti-inflammatory properties of desert truffle extracts. We show for the first time that Terfezia extracts do not rely on the nuclear factor erythroid 2-related factor 2 pathway, previously linked to anti-inflammatory responses. This expands our understanding of natural product anti-inflammatory mechanisms and could have important implications for developing new therapies. To account for differences in truffle effects, extracts prepared were subjected to secondary metabolites profiling using UPLC-MS. UPLC-MS led to the annotation of 87 secondary metabolites belonging to various classes, including amino acids, carbohydrates, alkaloids, amides, fatty acids, sterols, and phenolic compounds. Therefore, these results indicate that T. boudieri and T. claveryi exhibit anti-inflammatory activities through suppressing multiple inflammatory mediators and cytokines and may be potential anti-inflammatory agents.

Left ventricular assist devices promote changes in the expression levels of platelet microRNAs.

MicroRNAs (miRs) emerged as promising diagnostic and therapeutic biomarkers in cardiovascular diseases. The potential clinical utility of platelet miRs in the setting of left ventricular assist device (LVAD) support is unexplored. We prospectively measured the expression levels of 12 platelet miRs involved in platelet activation, coagulation, and cardiovascular diseases in LVAD patients by quantitative real-time polymerase chain reaction. Data were longitudinally measured before LVAD implant and after 1, 6, and 12 months of LVAD support, and compared with those measured in healthy volunteers (controls). In silico analysis was also performed to identify pathways targeted by differentially expressed miRs. Data from 15 consecutive patients and 5 controls were analyzed. Pre-implant expression levels of platelet miR-126, miR-374b, miR-223, and miR-320a were significantly different in patients vs. controls. The expression levels of platelet miR-25, miR-144, miR-320, and miR-451a changed significantly over the course of LVAD support; in silico analysis revealed that these miRs are implicated in both cardiac- and coagulation-associated pathways. Furthermore, the patients who suffered from bleeding (n = 5, 33%) had significantly higher pre-implant expression levels of platelet miR-151a and miR-454 with respect to the patients who did not. The same miRs were also differentially expressed in bleeders following LVAD implantation early before the clinical manifestation of the events. This study provides a proof-of-concept evidence of significant modulation of platelet miRs expression driven by LVADs. The possible existence of a platelet miRs signature predictive of the development of bleeding events warrants further validation studies.

#4536 INFLUENCE OF REDUCTION IN THE MASS OF ACTIVE NEPRONS ON THE EXPRESSION OF MIRNA AND NFκB IN THE MYOCARDIA AND AORT OF WKY RATS

Abstract Background and Aims At present, there is no doubt about the relationship between the state of the kidneys and the cardiovascular system, which is reflected in the concept of the cardiorenal continuum. The emerging data suggest that miRNAs may be one of the links connecting the kidneys and the cardiovascular system. However, information about the role of epigenomic changes in the development of damage to the heart and blood vessels in chronic kidney disease remains not fully understood. Target. The aim of the work was to evaluate changes in the expression levels of microRNA-21, 133,203, as well as the nuclear transcription factor κB (NFκB) in the tissues of the myocardium and aorta in the development of remodeling of the heart and blood vessels with a chronic reduction in the mass of active nephrons in the experiment. Method Systolic blood pressure (BP, mm Hg), myocardial mass index (MMI, mg/g), relative expression levels of miRNA-21, -133, -203 and NFκB in myocardium and aorta were analyzed in rats Wistar-Kyoto (WKY) with 5/6 nephrectomy (Nx) and duration of experimental exposure 2 mo (n = 9). Controls were sham operated (SO) animals (n = 9). The reverse transcription reaction for ‘complementary’ DNA (cDNA) preparation was carried out under the StemLoop-technology separately for the studied miRNA. The following primers were used in PCR: microRNA-21–5'-GCCCGCTAGCTTATCAGACTGATG-3_, microRNA-133–5_-GCCCGCAGCTGGTAAAATGGAAC-3_, microRNA-203–5_-GCCGGTGAAATGTTTAGGACC-3_ and U6–5_-GCGCGTCGTGAAGCGTTC-3_, and common reverse 5_-GTGCAGGGTCCGAGGT-3. The semi-qualitative evaluation of the miRNA expression level (in relative units, RU) under the 2-(ΔΔCt) protocol at the laboratory referent (0.09) was used in calculations. NFκB expression was quantified using a StemLoop technology and RealTimePCR compared to the expression of the reference-gene GAPDH. Data were analyzed using Student's t-test and the Mann–Whitney, a P&amp;lt;0.05 was considered statistically significant. Results In rats WKY with an experimental decrease of the nephron mass in comparison to control, significantly higher levels (median [interquartile rang]) of blood pressure (145.5[135.0; 155.0] vs 117.5[110.0; 120.0] mm Hg; P&amp;lt;0.001) NFκB expression in the myocardium (1.25[0.87; 1.87] vs 0.89[0.31; 1.15] relative units; P&amp;lt;0.01) and aorta (0.87[0.57;1.63] vs 0.35[0.22; 0.87] relative units; P&amp;lt;0.05). The expression level of miRNA-21 in the myocardium increased in rats with NE (1.48[0.54; 3.73] vs 0.37[0.06; 1.62] relative units; P&amp;lt;0.01) relative to the SO group, but did not change significantly in the aorta (p&amp;gt;0.05). Relative expression levels of microRNA-133 and -203 both in the myocardium and in the aorta of WKY rats with NE were more than 10 times lower compared to those of the SO group (p&amp;lt;0.001). BMI in rats 2 months after NE increased relative to the SO group (p&amp;lt;0.05). Conclusion An experimental decrease in the number of functioning nephrons not only leads to an increase in blood pressure and myocardial mass in WKY rats, but also causes epigenomic changes in the myocardium and blood vessels, in particular, activation of signaling pathways associated with NFκB in the myocardium and aorta of rats. Possibly, it is accompanied by growth in expression of NFκB-associated proliferative, proinflammatory and profibrotic cytokines. It is possible that microRNA-21 is also involved in the formation of fibrosis and myocardial remodeling. However, specific mechanisms of miRNA involvement in the pathogenesis of heart and vascular remodeling require further study.

Circulating miR-26b-5p and miR-451a as diagnostic biomarkers in medullary thyroid carcinoma patients

Purpose/methodsThe determination of tumour biomarkers is paramount to advancing personalized medicine, more so in rare tumours like medullary thyroid carcinoma (MTC), whose diagnosis is still challenging. The aim of this study was to identify non-invasive circulating biomarkers in MTC. To achieve this goal, paired MTC tissue and plasma extracellular vesicle samples were collected from multiple centres and microRNA (miRNA) expression levels were evaluated.ResultsThe samples from a discovery cohort of 23 MTC patients were analysed using miRNA arrays. Lasso logistic regression analysis resulted in the identification of a set of circulating miRNAs as diagnostic biomarkers. Among them, miR-26b-5p and miR-451a, were highly expressed and their expression decreased during follow-up in disease-free patients in the discovery cohort. Circulating miR-26b-5p and miR-451a were validated using droplet digital PCR in a second independent cohort of 12 MTC patients.ConclusionThis study allowed the identification and validation of a signature of two circulating miRNAs, miR-26b-5p and miR-451a, in two independent cohorts reporting a significant diagnostic performance for MTC. The results of this study offer advancements in molecular diagnosis of MTC proposing a novel non-invasive tool to use in precision medicine.

Identification of aberrantly expressed lncRNAs and ceRNA networks in multiple myeloma: a combined high-throughput sequencing and microarray analysis.

This study aimed to explore the potential effects of long non-coding RNAs (lncRNAs) in multiple myeloma (MM) patients using two detection methods: high-throughput sequencing and microarray. In this study, lncRNAs were detected in 20 newly diagnosed MM patients, with 10 patients analyzed by whole transcriptome-specific RNA sequencing and 10 patients analyzed by microarray (Affymetrix Human Clariom D). The expression levels of lncRNAs, microRNAs, and messenger RNAs (mRNAs) were analyzed, and the differentially expressed lncRNAs identified by both methods were selected. The significant differentially expressed lncRNAs were further validated using PCR. This study established the aberrant expression of certain lncRNAs involved in the occurrence of MM, with AC007278.2 and FAM157C showing the most significant differences. The top 5 common pathways identified by the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were the chemokine signaling pathway, inflammatory mediator regulation, Th17 cell differentiation, apoptosis, and NF-kappa B signaling pathway. Furthermore, three microRNAs (miRNAs) (miR-4772-3p, miR-617, and miR-618) were found to constitute competing endogenous RNA (ceRNA) networks in both sequencing and microarray analyses. By the combination analysis, our understanding of lncRNAs in MM will be increased significantly. More overlapping differentially expressed lncRNAs were found to predict therapeutic targets precisely.

Leucocyte-derived micro-RNAs as candidate biomarkers in Brugada syndrome.

Risk stratification for sudden cardiac death in patients with Brugada syndrome remains a major challenge. Contemporary risk prediction models have only modest predictive value. The aim of this study was to assess the role of micro-RNAs from peripheral blood as candidate biomarkers in Brugada syndrome. In this prospective study, Brugada patients and unaffected control individuals were enrolled for analysis of leucocyte-derived microRNAs (miRNAs) levels. Expression levels of 798 different circulating miRNAs were analysed on the NanoString® nCounter platform. All results were cross-validated by using a quantitative polymerase chain reaction. Micro-RNA expression levels of Brugada patients were compared with clinical data. A total of 21 definite Brugada patients (38% with a history of ventricular arrhythmia or cardiac arrest) and 30 unaffected control individuals were included in the study. Micro-RNA analysis showed a distinct expression profile in Brugada patients with 42 differentially expressed markers (38 up-regulated, 4 down-regulated miRNAs). The symptom status of Brugada patients was associated with a distinct miRNA signature. Micro-RNAs 145-5p and 585-3p were significantly up-regulated in symptomatic Brugada patients (P = 0.04). Incorporating miRNAs 145-5p and 585-3p into a multivariable model demonstrated significantly increased symptom prediction (area under the curve = 0.96; 95% confidence interval: 0.88-1.00). Brugada patients display a distinct miRNA expression profile compared with unaffected control individuals. There is also evidence that certain miRNAs (miR-145-5p and miR-585-3p) are associated with the symptom status of Brugada patients. The results suggest the principal utility of leucocyte-derived miRNAs as prognostic biomarkers for Brugada syndrome.

Evaluation of microRNA expression levels during the craving period of patients diagnosed with severe alcohol use disorder

ABSTRACT Background The aim of this study is to compare the serum MicroRNA (miRNA) expression levels on the first day of hospitalization and at the end of the first month of treatment in patients with alcohol use disorder and severe craving symptoms. Through the obtained data, biological agents targeting microRNA for alcohol craving may be developed. Methods The volunteer group consists of four patients diagnosed with severe alcohol use disorder. Sociodemographic and Clinical Information Form, DSM-5 Structured Clinical Interview-Clinical Version (SCID-5-CV), Penn Alcohol Craving Scale (PACS) were used as clinical assessment tools. Peripheral blood samples were obtained at the time of application and on the 30th day. miRNA isolation was performed. The expression levels of 2578 different mature human miRNAs were measured. Results The miRNAs showing statistically significant differences in peripheral blood samples taken on the 1st and 30th days from alcohol users are as follows: miR-19a-3p, miR-6849-5p, miR-6726-5p, miR-6848-5p, miR-186-5p, miR-6889-5p, miR-933, miR-370-3p, miR-5739, miR-103a-2-5p, miR-6860, miR-1254, miR-1260a, miR-3921, miR-127-3p, miR-8064, miR-6870-5p, miR-4688 Conclusions A significant difference was found between the 1st and 30th days in 18 of 2578 miRNAs scanned. It is aimed to investigate these 18 miRNAs in larger sample groups during the alcohol craving period.

МикроРНК: взгляд клинициста на состояние проблемы. Часть 2. МикроРНК в качестве биомаркера

Cardiovascular diseases are the main cause of death and disability in economically developed countries in the world. In response to the impact of various factors, the structure and function of several types of cells changes, contributing to the occurrence and progression of cardiovascular diseases. Search for sensitive and specific biomarkers is one of the most important problems in the field of diagnosis of cardiovascular diseases. In the last decade, microRNAs have more often been considered as potential biomarkers of a wide range of cardiovascular diseases, such as myocardial infarction, ischemic heart disease, heart failure, hypertension, diabetes mellitus and obstructive sleep apnoea. Early diagnosis of these diseases is essential to initiate immediate treatment, which can lead to improved outcomes. MicroRNAs are endogenous small (21-23 nucleotides) ribonucleotides involved in the regulation of protein synthesis from amino acids based on matrix RNA. MicroRNAs are involved in the regulation of expression of the majority (&gt;60%) of genes encoding proteins, mainly due to its suppression, modulate numerous signaling pathways and cellular processes and participate in intercellular communication. There are different advantages of these biomarkers: low invasiveness during research, stability, resistance to destructive factors, for example, freeze-thaw cycles, enzymes in the blood. Some microRNAs are expressed everywhere, while others are specific to certain tissues and/or stages of development of the organism. At the same time, microRNAs were detected in various biological fluids: blood plasma, urine, seminal fluid, saliva, breast milk. Changes in both the amount and functional activity of microRNAs can lead to the development of various diseases. In the cardiovascular system, microRNAs control the functions of various cells, such as cardiomyocytes, endothelial cells, smooth muscle cells and fibroblasts. Thus, due to the stability of microRNAs, the tissue­specific nature of expression and secretion into specific fluids, it becomes possible to consider them as an attractive diagnostic. It is also particularly important that the expression levels of certain microRNAs reflect not only the presence of diseases in the early stages, but also the dynamic development of diseases in the later stages. This review presents the features of various microRNAs as biomarkers and their influence on some molecular pathways underlying cardiovascular diseases and describes the significant potential of supplementing traditionally used markers in clinical practice with microRNA biomarkers. Prospects for the development and limitations of the use of microRNAs as potential biomarkers are also described.

The Profiling of Apoptosis-Associated MicroRNA Expressions of the Lung Adenocarcinoma.

The most common histopathological subtype of lung cancer is adenocarcinoma. MicroRNAs are a class of non-coding RNAs that play roles in the regulation of gene expression. MicroRNAs affecting apoptosis may have different roles in lung adenocarcinoma development, progression, and differentiation. The objective of this study is to profile all known microRNAs linked to apoptosis in normal and lung cancer tissue using real-time polymerase chain reaction. Tissues with adenocarcinoma and healthy tissues were taken from the same lung. The degree of differentiation of all tumors was determined. Expressions of 84 apoptosis-associated microRNAs in both tissues were analyzed by real-time polymerase chain reaction array. Eleven patients and 22 samples were included in the study. In the comparison of expression levels of apoptosis-associated microRNAs in normal and adenocarcinoma tissue, miR-134, miR-183-5p, miR-192-5p, miR-193b-3, miR-194-5p, miR-200c-3, miR212-3p, miR-25-3p, miR-449a, and miR-9-5p showed significant difference in downregulation. Receiver operating characteristic curve analysis of 10 identified microRNAs was performed, and cut-off values, sensitivity, and specificity were determined. No significant difference was found between microRNA expression levels in adenocarcinoma tissues classified as moderate-well to poorly differentiated. Differently, downregulated expressed apoptosis-associated microRNAs were detected in lung adenocarcinoma tissues. MicroRNAs can be used as biomarkers in the diagnosis in lung adenocarcinoma. The expression of microRNAs linked to apoptosis should be investigated in different lung cancer histological subtypes in order to identify potential biomarkers.

Scoparone inhibits breast cancer cell viability through the NF‑κB signaling pathway.

Scoparone (SCO) is a compound found in the stems and leaves of Artemisia capillaris. The pharmacological uses of SCO include significant hypotensive, cholagogic, anti-inflammatory, analgesic, lipid-lowering, anti-asthmatic and anti-coagulant effects. The present study aimed to verify the anticancer potential of SCO in breast cancer (BC) cells and its underlying molecular mechanism. Cell Counting Kit-8 and flow cytometry were used to analyze the effects of SCO on cell viability and apoptosis. Nucleocytoplasmic separation was used to analyze the location of the long non-coding RNA (lncRNA) small nucleolar RNA host gene 12 (SNHG12) in BC cells. Reverse transcription-quantitative PCR was used to analyze the effect of SCO on the expression levels of SNHG12, microRNA (miRNA/miR)-140-3p and tumor necrosis factor receptor associated factor 2 (TRAF2). Western blotting was used to analyze the protein expression levels of TRAF2 and downstream nuclear factor κB (NF-κB) signaling pathways. The results demonstrated that SCO had a time- and dose-dependent inhibitory effect on the viability of BC cells, and that the upregulated lncRNA SNHG12 in BC cells was inhibited by SCO. SNHG12, which was primarily expressed in the cytoplasm, acted as a competing endogenous RNA, sponged miR-140-3p and inhibited the expression of miR-140-3p. The transcriptional activity and translational level of TRAF2, a downstream target of miR-140-3p, decreased following the SCO-mediated suppression of SNHG12 expression. As an upstream effector, TRAF2 activity reduction mediated the inhibition of NF-κB signaling, decreased the viability and migration of BC cells, and promoted BC cell apoptosis. In conclusion, SCO-induced inhibition of viability and promotion of apoptosis in BC cells are achieved through the inhibition of NF-κB signaling, which is associated with regulation of the SNHG12/miR-140-3p/TRAF2 axis. This understanding provides new drug candidates for the treatment of BC and a theoretical basis for biology.

Circular RNA DHTKD1 targets miR‑338‑3p/ETS1 axis to regulate the inflammatory response in human bronchial epithelial cells.

Asthma is a chronic inflammatory airway disease and the airway epithelium is involved in airway inflammation and innate immunity. However, whether circular RNA (circRNA) is involved in the pathogenesis of asthma remains unclear. The present study aimed to determine the functions and molecular mechanisms of circRNA targeting dehydrogenase E1 (circDHTKD1) in the inflammation response of human bronchial epithelial cells. BEAS-2B cells were stimulated with lipopolysaccharide (LPS) to establish a model of in vitro airway inflammation. Cell viability was assessed using Cell Counting Kit-8 assay. CircDHTKD1 was characterised by nucleocytoplasmic isolation and Sanger sequencing. The RNA expression levels of circDHTKD1, microRNA (miR)-338-3p and potential ERK pathway downstream genes were evaluated by reverse transcription-quantitative polymerase chain reaction. Western blot analysis was performed to measure associated protein levels. The levels of inflammatory cytokines were detected by ELISA. The interaction between circDHTKD1 and miR-338-3p was confirmed by dual-luciferase reporter assay. circDHTKD1 expression was significantly upregulated by LPS treatment, whereas miR-338-3p expression was decreased. Furthermore, circDHTKD1 directly targeted miR-338-3p, which negatively regulated expression of E26 transformation specific-1 (ETS1). Inflammatory cytokine and ETS1 expression levels decreased following transfection with small interfering RNA targeting circDHTKD1 or miR-338-3p mimics. In addition, co-transfection with miR-338-3p inhibitor reversed the effects caused by circDHTKD1 knockdown. The knockdown of ETS1 in LPS-induced BEAS-2B cells resulted in decreased cytokine production and inhibition of the ERK signalling pathway. Overall, these results suggested that the knockdown of circDHTKD1 alleviated the LPS-induced production of inflammatory cytokines and activation of the ERK pathway in BEAS-2B cells through the miR-338-3p/ETS1 axis. In summary, circDHTKD1 exacerbated LPS-triggered inflammation responses in BEAS-2B cells by regulating ETS1 expression by interacting with miR-338-3p, suggesting that circDHTKD1 may serve as a potential therapeutic target against asthma.

Predictive assessment of micro RNA expression level and number of nucleolar organizer regions with follicular thyroid tumor of undetermined malignant potential

Introduction This article presents the current views on follicular thyroid tumor of undetermined malignant potential (FTUMP) taking into account the principles of WHO classification. Molecular genetic study with identification of mutation of some genes as well as determination of expression level of some microRNAs are given as methods for detection of potential predictors of oncodiagnostics. It was shown that the state of nucleus organizer regions is a marker of cell proliferation rate.The aim of the investigation was to estimate prognostic significance of expression level of some microRNAs and amount of regions of the nucleus organizers in the thyroid tissue at FTUMP to objectify approaches to development of adequate surgical tactics.Materials and methods Profile, microRNA expression level by real time PCR and regions of the nucleus organizers were studied in 98 cases of FTUMP which made up 4.75% of all cases of thyroid tumors operated on during four years.Results In 29 (29.6%) cases the spectrum of microRNA typical for malignant neoplasms was registered in the tissue of the FTUMP. In the remaining 69 (70.4%) cases out of 98 observations the spectrum and the expression level of microRNA corresponded to the follicular adenoma. Statistically significant differences in the average number of AgNORs and correlation between the cell area and the number of AgNORs granules were found within the formed subgroups.Discussion Increase of the mean number of AgNORs and the ratio of the nucleus area to the number of silver nitrate granules in the 2nd subgroup in comparison with the 1st one, as well as the appearance of the type II cells testifies to the appearance of the proliferating tumor cells in a new tumor growth and can be considered as prognostic criteria of tumor malignization. These results correlate with the findings of molecular-genetic study. Conclusion The data of the literature and the results obtained by studying the profile and the expression level of a number of microRNAs, morphometric characteristics and counting of thyrocyte nucleus organizer regions may serve as additional criteria for evaluation of the proliferative activity of the follicular epithelium in thyroid neoplasms, in particular to clarify the malignancy potential. Preoperatively diagnosed conditionally malignant FTUMP will allow to justify the necessity and amount of surgical intervention, and if a complex of criteria of conditionally benign FTUMP is revealed, to stick to the waiting tactics and refuse surgical intervention.

Single cell quantification of microRNA from small numbers of non-invasively sampled primary human cells

Expression levels of microRNAs (miRNAs) in single cells are low and conventional miRNA detection methods require amplification that can be complex, time-consuming, costly and may bias results. Single cell microfluidic platforms have been developed; however, current approaches are unable to absolutely quantify single miRNA molecules expressed in single cells. Herein, we present an amplification-free sandwich hybridisation assay to detect single miRNA molecules in single cells using a microfluidic platform that optically traps and lyses individual cells. Absolute quantification of miR-21 and miR-34a molecules was achieved at a single cell level in human cell lines and validated using real-time qPCR. The sensitivity of the assay was demonstrated by quantifying single miRNA molecules in nasal epithelial cells and CD3+ T-cells, as well as nasal fluid collected non-invasively from healthy individuals. This platform requires ~50 cells or ~30 µL biofluid and can be extended for other miRNA targets therefore it could monitor miRNA levels in disease progression or clinical studies.