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• CircCUL3 overexpression drives cervical cancer progression. • CircCUL3 acts as a sponge for miR-223-3p to upregulate ATG7 and activate autophagy. • Chloroquine reverses circCUL3-induced effects by restoring miR-223-3p expression. Circular RNA (circRNA) has emerged as a pivotal regulatory factor in cancer biology, yet its exact role in cervical cancer remains incompletely understood. In this study, we investigated the functional role of circCUL3 in cervical cancer and explored its potential as a therapeutic target. Functional gain and loss experiments were conducted in Hela and Siha cell lines to elucidate the biological functions of circCUL3 in cervical cancer. The results revealed that circCUL3 overexpression significantly enhanced cell viability, migration, and invasion while suppressing apoptosis, while circCUL3 knockout displayed the opposite effects. Mechanistically, we identified hsa-miR-223-3p as a target of circCUL3, with its expression being negatively regulated by circCUL3. Furthermore, we discovered that circCUL3 could sequester miR-223-3p, leading to the upregulation of ATG7 expression, and this was linked to the regulation of autophagy in cervical cancer cells. In vivo validation using a xenograft mouse model further supported our in vitro findings. Notably, we found that chloroquine (CQ), an autophagy inhibitor, restored miR-223-3p expression and counteracted the oncogenic effect of circCUL3 overexpression. In conclusion, circCUL3 potentially contributes to the malignant progression of cervical cancer by acting as a sponge for miR-223-3p, resulting in the upregulation of ATG7 and the activation of autophagy. [ABSTRACT FROM AUTHOR]

Highlights • A sensitive self-enhanced rolling circle amplification strategy was proposed even if only two probes were involved. • A sensing platform is developed for intracellular miRNA imaging, displaying high sensitivity and desirable universality. • The measured data demonstrates its potential application in clinical diagnosis and other areas relying on miRNA analysis. Abstract MicroRNAs (miRs) are a class of non-coding, small RNA molecules that play important roles in gene expression. The aberrant expression of miRs is associated with various human diseases, and miRs have been regarded as biomarkers and therapeutic targets in cancer treatment. In this work, using miR-21 as model target species, an efficiently self-enhanced rolling circle amplification (seRCA)-based biosensing platform was developed for the ultrasensitive detection of cancer-related miRs. In the presence of target, a linear DNA template can be cyclized by ligase and initiate the first RCA (fRCA) reaction. Without any separation step and chemical modification, the fRCA products (fRCAp) are capable of activating the second exponential RCA and produce a much higher fluorescence signal, causing the self-enhancement effect of RCA reaction. Because two different signal amplification processes are introduced, there are two linear dynamic ranges together over 7 orders of magnitude. Target miR can be detected down to 1.0 fM with very high specificity, eliminating the interference from homologous miRs. If a fluorescently modified oligonucleotide probe is involved, the seRCA-based biosensing strategy is able to be employed for the intracellular imaging of cancer-related miR. Significantly, the comparative evaluation demonstrates that the proposed strategy exhibits the impressive capability to execute the miR imaging within cells higher than classical fluorescence in situ hybridization (FISH) method. The newly-developed powerful sensing strategy for the specific miR imaging would offer a unique opportunity to promote molecular biological research, early diagnosis and treatment of human cancers. [ABSTRACT FROM AUTHOR]

Sorafenib is a first-line drug for hepatocellular carcinoma (HCC). Autophagy has been shown to facilitate sorafenib resistance. miR-375 has been shown to be an inhibitor of autophagy. In this study, miR-375 and sorafenib were co-loaded into calcium carbonate nanoparticles with lipid coating (miR-375/Sf-LCC NPs). The nanoparticles had high loading efficiency and were ∼50 nm in diameter. Besides, the NPs could increase the stability and residence time of both drugs. Moreover, we demonstrated that autophagy was activated in HCC cells by sorafenib but not by miR-375/Sf-LCC NPs. In vitro , miR-375/Sf-LCC NPs exhibited pH-dependent drug release and potent cytotoxicity. In vivo , miR-375/Sf-LCC NPs increased miR-375 and sorafenib uptake in tumor (2 folds compared with Lipofectamine 2000-miR-375 and 2–5 folds compared with free sorafenib). Furthermore, miR-375/Sf-LCC NPs showed greatly enhanced therapeutic efficacy in an HCC xenograft model. These findings suggest that miR-375/Sf-LCC NPs may be a promising agent for the HCC therapy. Statement of Significance Hepatocellular carcinoma (HCC) is the most common primary liver tumor and the third leading cause of cancer mortality globally. In this manuscript, miR-375 and sorafenib were co-loaded into calcium carbonate nanoparticles with lipid coating (miR-375/Sf-LCC NPs) to treat HCC. We demonstrated that miR-375/Sf-LCC NPs can deliver sorafenib and miR-375 into HCC cells and tumor tissues, increase drug retention time in tumor, significantly inhibit autophagy and produce enhanced anti-tumor effect. [ABSTRACT FROM AUTHOR]

Chronic cerebral hypofusion (CCH) promotes hyperphosphorylation of tau proteins, a key neuropathological trait that reflects neurodegenerative disorders. Nimodipine, an L-type calcium channel antagonist, has been reported to show neuroprotective effects. In this study, we investigated the potential mechanism of nimodipine in tauopathies induced by CCH. MiR-132 is downregulated in tauopathies such as AD and directly targets tau mRNA to regulate its expression. Here, we report that CCH induced miR-132 deficiency and increased tau phosphorylation at Ser396 while tau expression was not influenced. Nimodipine treatment attenuated CCH induced tau phosphorylation by up-regulating expression of miR-132. Furthermore, nimodipine inhibited activation of GSK-3β and neuronal apoptosis induced by CCH. Interestingly, GSK-3βmRNA level negatively correlated with the expression of miR-132. These findings support a role for nimodipine inhibiting tau phosphorylation at Ser396 via miR-132/GSK-3β. Therefore, nimodipine may be a candidate for the treatment of tauopathy present in CCH. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is a prevalent and lethal disease that is characterized by drug resistance. Doxorubicin (DOX) is a widely used chemotherapeutic drug and miR-375 has been shown to be a tumor suppressor in HCC. Here, we reported that miR-375 and DOX co-loaded into lipid-coated calcium carbonate nanoparticles (LCC-DOX/miR-375 NPs), enhanced the anti-tumor effects through combination therapy, and were highly effective in reversing drug resistance in HCC. LCC-DOX/miR-375 NPs were prepared by a reverse microemulsions method. In vitro , LCC-DOX/miR-375 NPs exhibited enhanced intracellular accumulation, pH-sensitive DOX release and potent cytotoxicity. In vivo , LCC-DOX/miR-375 NPs showed efficient antitumor effect both in xenograft and primary HCC murine models. Our results showed that the LCC-DOX/miR-375 nanoparticles provide a novel strategy to overcome the drug resistance and promote addictive effect between miR-375 and DOX in HCC. [ABSTRACT FROM AUTHOR]

Abstract: Background: MicroRNAs (miRNAs) are widely involved in the process of chronic heart failure (HF), which is characterized by reactivation of the fetal gene program. Here, we examined whether the serum expression levels of some HF-related miRNAs in adult HF patients would tend to revert to fetal levels. Methods and results: Serum was obtained from the peripheral venous blood of 22 HF patients, 18 asymptomatic controls, and the umbilical venous blood of 9 fetuses from 9 independent parturitions. Serum pools of the three groups were initially screened against 40 known HF-associated miRNAs via quantitative reverse transcriptase polymerase chain reaction. Twenty-seven miRNAs were stably expressed in the serum pools. Nine miRNAs showed similar expression levels in the HF and fetus groups compared to the controls, two of which (miR-210, miR-30a) were significantly up-regulated in both groups. These miRNAs showed high diagnostic accuracy and correlations with blood N-terminal prohormone of brain natriuretic peptide, identifying them as potential biomarkers for HF. Putative targets of the miRNAs were predicted with online software programs, and the Kyoto Encyclopedia of Genes and Genomes pathway analysis was employed to identify miRNA-regulated functional modules. In particular, miR-210 seemed to be more closely related than miR-30a to the pathological mechanisms of HF, including the calcium signaling, vascular smooth muscle contraction, transforming growth factor-β signaling, and aldosterone-regulated sodium reabsorption pathways. Conclusion: The serum expression levels of some HF-related miRNAs in HF patients tended towards fetal levels. Among them, miR-210 and miR-30a were elevated in the HF and fetus groups. [Copyright &y& Elsevier]

• miR-205 exerts hepatoprotective effect against alcohol-associated liver disease (ALD). • miR-205 targets and inhibits importinα5 expression. • Baicalin upregulates miR-205 expression. • Baicalin inhibits importinα5 to repress the activation of NF-κB signaling pathway. • This study provides better understanding of the hepatoprotective effect of baicalin in ALD. Recently, baicalin refers to flavonoid compound extracted from Scutellaria baicalensis Georgi has been indicated to hold promising therapeutic effects in alcohol-associated liver disease (ALD). However, knowledge of the molecular mechanisms for its hepatoprotective effect is still very limited. Evidence exists suggesting potential association between miR-205 and baicalin's function. Bioinformatic analysis and dual luciferase reporter assay were conducted to determine the binding affinity between miR-205 and importinα5. Our findings revealed that baicalin could alleviate ALD by raising the expression of miR-205. Additionally, miR-205 repressed NF-κB signaling pathway activation by binding to importinα5 to relieve ALD. Baicalin inhibited importinα5-mediated NF-κB signaling pathway to protect the liver against alcohol-induced injury, inflammation, oxidative stress and hepatocyte apoptosis. Taken conjointly, baicalin confers hepatoprotective effect against ALD through miR-205-mediated importinα5 inhibition via the NF-κB signaling pathway, highlighting a promising therapeutic target for ALD treatment with the help of traditional Chinese medicine. [ABSTRACT FROM AUTHOR]

• Introduce the characteristics of circRNAs and describe their biogenesis. • Provide an overview on the molecular functions of circRNAs in cancers. • Discuss the potential application of circRNAs as biomarkers. Circular RNAs (circRNAs), characteristic of covalently closed loops generated by back-splicing, are a subclass of single-stranded RNA molecules. Owing to the circular structures, circRNAs are protected from exonuclease-induced degradation, which makes them more stable compared with linear RNAs. With the development of high-throughput sequencing technology and bioinformatics, the roles of circRNAs in a variety of physiological and pathophysiological processes have been increasingly revealed. Although the functions of most circRNAs remain largely elusive, accumulating studies have identified them as microRNA(miRNA) sponges, protein regulators, transcriptional regulators, protein templates, and so forth. In this review, we briefly describe the biogenesis of circRNAs and provide an overview on their functions in cancers, including miRNA sponges, protein regulators, transcriptional regulators, protein templates. Furthermore, we discuss the potential application of circRNAs as biomarkers and give insight into future perspectives. [ABSTRACT FROM AUTHOR]

Multi-microRNA (miRNA) detection would greatly facilitate early diagnosis of colorectal cancer (CRC). Here a convenient cascade isothermal amplification approach incorporating a G-quadruplex molecular beacon (G4MB) was established for achieving one-pot detection of multiple CRC miRNAs (miRNA-21, miRNA-92a, miRNA-31); this strategy incorporated a Bsu DNA polymerase (Bsu pol)-induced strand-displacement reaction and a Lambda exonuclease (λexo)-aided recycling reaction. In the presence of target miRNA, the G-rich stem structure was opened and became available for hybridization with the primer to initiate synthesis of Bsu pol-catalyzed double-stranded DNA (dsDNA) that displaced the miRNA target and released it, allowing it to participate in subsequent amplification cycles. Meanwhile, the dsDNA was gradually digested into fragments by λexo from the 5′ phosphorylated end, releasing the newly synthesized DNA strand for participation in subsequent cycles that led to amplification of the fluorescent signal. This approach provided a low limit of detection (LOD) of zeptomolar-level, 85.8 zM, 77.6 zM, 78.9 zM for miRNA-21, miRNA-92a, miRNA-31, respectively. It could distinguish the mismatched targets and achieved three miRNA targets detection run in parallel in one-pot within 2 h. Thus, this fast, simple, and convenient strategy holds great promise as a clinical application for the detection of multiple miRNAs in clinical CRC samples. • Test to detect colorectal cancer-associated miRNAs in patient samples is described. • miRNA is detected via one-pot 2-enzyme reaction with G-quadruplex molecular beacon. • Testing is specific and sensitive, with a low limit of miRNA detection of zeptomolar level. • Multiple miRNAs are detected in one pot without interference among 3 miRNA tests. • One-pot test holds promise for diagnostic/prognostic colorectal patient screening. [ABSTRACT FROM AUTHOR]

With the use of DNA as building blocks, a variety of microRNA amplification-based sensing systems have been developed. Nevertheless, ultrasensitive, selective and rapid detection of microRNAs with a high signal-to-background ratio and point mutation discrimination ability remains a challenge. Herein, we propose a novel wheel drive-based DNA sensing system (NWDS) based on a self-assembled, self-quenched nanoprobe (SQP) to conduct highly specific and ultrasensitive one-step measurement of microRNAs. In this work, a signalling recognition DNA hairpin (DH) sequence with a self-complementary stem domain of 14 base pairs was used, which contained three functional regions, namely a recognition region for the target miRNA-21, a sticky region with 9 complementary nucleotides to the 3′terminus of a DNA wheel (DW) and a region for the hybridization with a quenching DNA primer (DP). The SQP was ingeniously self-assembled at room temperature by the DH and DP, which was capable of eliminating unwanted background signals. MiRNA-21 was employed as a target model to specifically activate the SQP, leading to specific hybridization between the HP and DW. With the assistance of a polymerase, an SQP-based wheel driving took place to induce hybridization/polymerization displacement cycles, initiating target recycling and DP displacement. As a result, a large amount of the newly formed hybrid SQP/DW accumulated to generate a substantially enhanced fluorescence signal. In this way, the newly proposed NWDS exhibits ultrasensitivity with a detection limit of 5.62 aM across a wide linear dynamic response range up to 200 nM, excellent selectivity with the capability to discriminate homologous miRNAs and one-base, two-base and three-base mismatched sequences, and an outstanding analytical performance in complex systems. In addition, the significant simultaneous advantages of one-step operation, rapid detection within 15 min and a high signal-to-background ratio of 26 offer a unique opportunity to promote the early diagnosis of cancer-related diseases and molecular biological analysis. We develop a novel wheel drive-based DNA sensing system (NWDS) based on self-assembly self-quenched nanoprobe (SQP) to conduct highly specific, ultrasensitive and rapidly one-step detection of microRNAs at the attomolar level. [Display omitted] • A NWDS strategy is newly-proposed for miRNAs detection, displaying ultrasensitivity and desirable stability. • The strategy with impressive specificity can discriminate single-base mutation miRNAs. • The one-step operation, quick detection and high signal-to-background ratio emphasize its potential applications in bioanalysis. [ABSTRACT FROM AUTHOR]

Objective The aim of this study was to investigate the effect of miR-133a and miR-133b on regulatory T cell (Treg) differentiation in IgA nephropathy (IgAN) through targeting forkhead box P3 (FOXP3). Methods Peripheral blood mononuclear cells (PBMCs) were isolated from IgAN patients (n = 20) and healthy controls (n = 20). Percentage of Tregs defined as CD4 + CD25 + FOXP3 + T cells were determined by flow cytometry. The mRNA expression levels of miR-133a, miR-133b and FOXP3 were measured by real-time PCR. FOXP3 protein level was analyzed by western blotting. Results Tregs percentage in PBMCs of IgAN patients was significantly lower than that of healthy controls, whereas the expression levels of miR-133a and miR-133b in IgAN patients were dramatically higher than that in the control group. Treg percentage was negatively correlated with miR-133a and miR-133b expressions. Meanwhile, miR-133a and miR-133b modulated FOXP3 expression by detecting of its gene 3′-untranslated region. MiR-133a or miR-133b overexpression significantly decreased the % Tregs (CD4 + CD25 + FOXP3+) of the total CD4 + T cells while miR-133a or miR-133b knockdown led to an opposite effect. Moreover, FOXP3 levels in IgAN patients was significantly lower than that in the control group and was negatively correlated with miR-133a and miR-133b expression. Conclusion MiR-133a and miR-133b inhibited Treg differentiation in IgA nephropathy through targeting FOXP3. [ABSTRACT FROM AUTHOR]

We looked for differentially expressed MicroRNAs (miRNAs) in Immunoglobulin A nephropathy (IgAN). Forty-eight miRNAs were identified through the initial screening phase (2 IgAN pools vs. 1 normal control (NC) pool) using quantitative reverse transcription polymerase chain reaction (qRT-PCR) based Exiqon panel (miRCURY-Ready-to-Use-PCR-Human-panel-I + II-V 1 .M). By qRT-PCR, these miRNAs were further assessed in the training (32 IgAN VS. 31 NCs) and testing stages (51 IgAN VS. 51 NCs). The renal pathological lesions of patients with IgAN were evaluated according to Lee's grading system. We discovered a plasma miRNA signature including four up-regulated miRNAs (miR-148a-3p, miR-150-5p, miR-20a-5p and miR-425-3p) and the areas under the receiver operating characteristic (ROC) curve (AUC) were 0.80 and 0.76 for the training and testing stage, respectively. The expression of the four miRNAs in IgAN grade I–II subgroups (according to Lee's grading system) was obviously higher than that in IgAN grade III-V ( P  < .05). In summary, the plasma e xpression of miR-148a-3p, miR-150-5p, miR-20a-5p and miR-425-3p were up-regulated in patients with IgAN, especially the early-stage disease. Further studies are needed to explore the roles of the four miRNAs in the pathogenesis and progression of IgAN. [ABSTRACT FROM AUTHOR]

Fat-1 transgenic cattle have high levels of ω-3 fatty acids, which regulate several genes in fatty acid metabolism. In the current study, fibroblasts derived from in vivo fertilized (Ferti) and fat-1 transgenic (TG) Luxi cattle ( Bos taurus ), a local breed in China, were cultured and their miRNA expression was characterized. Expression of 352 known miRNAs differed in cells from Ferti and TG cattle: 83 miRNAs were found to be specifically expressed in cells from Ferti cattle while 23 miRNAs were found to be specifically expressed in cells from TG cattle. Novel differences in miRNA expression were also found in cells from Ferti and TG cattle. The identity of seven differentially expressed miRNAs was verified using quantitative real-time PCR, and target genes were identified computationally. GO and KEGG analysis revealed that these miRNAs were involved in seven major biological pathways, including metabolism, MAPK signaling, calcium signaling, purine metabolism, ubiquitin mediated proteolysis, pyrimidine metabolism, and the cell cycle. Overexpression of one of these miRNAs, miR-21-5p, was found to suppress expression of fibroblast growth factor 10 (FGF10) and adipose triglyceride lipase (ATGL) in fibroblasts from TG cattle and 3T3-L1 pre-adipocytes. Conversely, knockdown of miR-21-5p stimulated expression. Together, these results suggest that miRNAs potentially play a role in expression of lipogenic and lipolytic genes as well as in synthesis of ω-3 fatty acids facilitated by fat-1. [ABSTRACT FROM AUTHOR]

Objectives Circulating miRNAs have been reported to have potential in detecting various diseases. However, few studies explored differentially expressed miRNAs in plasma of patients with pulmonary embolism (PE). Our study is to identify plasma miRNAs which can serve as potential biomarkers of PE. Materials and methods Exiqon miRCURY Ready-to-Use PCR Human panel I + II V1.M was conducted to identify differently expressed miRNAs in pooled plasma samples of PE patients compared with normal controls. Expressions of identified miRNAs were assessed in 37 PE patients as well as matched normal individuals followed by validation on six Beagle dogs by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results Twelve miRNAs were identified from the screening phase. Moreover, miR-134, previously reported related with PE, and hypoxia-induced miR-210 were also submitted to the validation phase. Only miR-28-3p was found significantly elevated in the plasma of PE patients. Compared with the level of plasma miR-28-3p of the dogs before PE, the elevated miR-28-3p did not alter significantly at 1, 2, 4 and 6 h after PE. The area under the receiver operating characteristic (ROC) curve of plasma miR-28-3p was 0.792 (95% confidence interval: 0.689–0.896). KEGG pathway analysis showed that miR-28-3p might involve in PE related pathways such as inositol phosphate metabolism and phosphatidylinositol signaling system. Conclusion Our study indicated that elevated plasma miR-28-3p could be used as a non-invasive and stable biomarker in the detection of PE. Further researches on the miRNA are warranted. [ABSTRACT FROM AUTHOR]

Objective To comparatively analyze the human microRNAomes between normal pregnant and miscarriage deciduas by an in-depth sequencing of microRNA (miRNA); and to specifically examine miRNA-199b-5p and serum/glucocorticoid regulated kinase 1 ( SGK1 ) in vivo and in vitro for their possible roles in pregnancy maintenance. Design Samples of deciduas from 6–8-week spontaneous miscarriages and normal pregnant women were irrespectively collected and comparatively analyzed by miRNA sequencing. The miR-199b-5p and SGK1 expressions were validated in vivo and in vitro. Setting University research and clinical institutes. Patient(s) In this experimental study, samples of deciduas were obtained from October 2011 to April 2012 from 29 women with spontaneous miscarriages and 35 normal pregnant women (control group) who underwent pregnancy termination at 6–8 weeks at our university gynecology unit. Intervention(s) Endometrial biopsies, cell transfection, and production of an miR-199b-5p transgenic mouse model. Main Outcome Measure(s) In-depth sequencing of the miRNAome on human deciduas was performed for statistically significant differences in miRNA expression. Expression levels of SGK1 were detected by quantitative polymerase chain reaction and immunoblotting (Western blot) in vitro while miR-199b-5p is overexpressed or knockdown in miR-199b-5p transgenic mice. Result(s) Expression of the 1,921 known miRNAs was analyzed in the study. In aborted deciduas, 0.57% of the miRNAs were expressed abundantly (>10,000 transcripts per million) and represented 86.38% of all the miRNA reads. Six miRNAs were down-regulated (let-7a-5p, let-7f-5p, let-7g-5p, let-7e-5p, let-7d-5p, and miR-98), whereas miR-199b-5p was significantly up-regulated. Overexpression or knockdown of miR-199b-5p in HEK293T and Ishikawa cells decreased or increased SGK1 expression. Furthermore, overexpression of miR-199b-5p in human endometrial stromal cells or in transgenic mouse decreased SGK1 expression at the mRNA and protein levels, respectively. Conclusion(s) Among the miRNAomes, the abundant expression of the let-7 members was decreased in aborted samples, whereas miR-199b-5p expression was consistently increased. A significant inverse correlation was found between miR-199b-5p and SGK1 in vivo and in vitro. [ABSTRACT FROM AUTHOR]