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Biocoding technology constructed by readable tags with distinct signatures is a brand-new bioanalysis method to realize multiplexed identification and bio-information decoding. In this study, a novel fluorescence intensity coding technology termed Tetra-FICT was reported based on tetrahedron DNA nanostructure (TDN) carrier and Főrster Resonance Energy Transfer (FRET) effect. By modulating numbers and distances of Cy3 and Cy5 at four vertexes of TDN, different fluorescence intensities of twenty-six samples were produced at ∼565.0 nm (FI Cy3 ) and ∼665.0 nm (FI Cy5 ) by detecting fluorescence spectra. By developing an error correction mechanism, eleven codes were established based on divided intensity ranges of the final FI Cy3 together with FI Cy5 (Final-FI Cy3 &FI Cy5 ). These resulting codes were used to construct barcode probes, with three miRNA biomarkers (miRNA-210, miRNA-199a and miRNA-21) as cases for multiplexed bio-assay. The high specificity and sensitivity were also demonstrated for the detection of miRNA-210. Overall, the proposed Tetra-FICT enriched the toolbox of fluorescence coding, which could be applied to multiplexing biomarkers detection., Competing Interests: Declaration of competing interest We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The blood-brain barrier (BBB) is essential for maintaining central nervous system (CNS) stability, and neuroinflammation may cause the dysfunction of the BBB. MicroRNA-146a (miR-146a) is closely associated with neuroinflammation, which showed significant upregulation in response to lipopolysaccharide (LPS) induction. Elucidating the relationship between LPS-induced miR-146a expression and the BBB could decipher the mechanism of many neurological diseases. Here, we constructed an in vitro microfluidic human-BBB (μF-hBBB) chip consisting of human umbilical vein vascular endothelial cells (HUVECs) and human astrocyte (HAs) cells. A tetrahedral DNA framework (TDF-3MB) nanoprobe was used to label miR-146a in HUVECs on μF-hBBB chips before and after LPS induction, and the study revealed a significant increase in miR-146a expression after LPS induction. We believe that such a μF-hBBB chip is a promising in vitro platform for further use in understanding CNS diseases.

miRNA analysis has played an important role in precise diagnosis, treatment and prognosis of cancer, especially multiplexed miRNA imaging. In this work, a novel fluorescence emission intensity (FEI) encoding strategy was developed based on a tetrahedron DNA framework (TDF) carrier and the FRET effect between Cy3 and Cy5. Six FEI-encoded TDF (FEI-TDF) samples were constructed by tuning the labeling number of Cy3 and Cy5 at the vertexes of the TDF. For fluorescence characterization in vitro , distinct FEIs in the spectra and different colors under ultraviolet (UV) irradiation of FEI-TDF samples were observed. By dividing the ranges of FEIs of samples, the stability of FEIs was highly improved. Based on the ranges of FEIs in each sample, five codes with good discrimination were finally developed. Before the application of intracellular imaging, the excellent biocompatibility of the TDF carrier was proved by CCK-8 assay. The barcode probes based on samples 12, 21 and 11 were designed as example models to realize multiplexed imaging of miRNA-16, miRNA-21 and miRNA-10b in MCF-7 cells with obviously different fluorescence merged colors. FEI-TDFs provide a new research perspective for the development of fluorescence multiplexing strategies in the future.

Replication factor C 5 (RFC5) is involved in a variety of biological functions of cancer. However, the expression pattern of RFC5 and the underlying mechanisms in colorectal cancer (CRC) remain elusive. Here, we show that RFC5 is significantly upregulated in CRC tissues and cells. Patients with CRC and increased RFC5 levels have an unfavorable prognosis. RFC5 can promote the proliferation, migration, and invasion of CRC cells and inhibit the apoptosis of CRC cells. Additionally, upstream of RFC5, we constructed the competing endogenous RNA network and confirmed that RFC5 in this network was inhibited by miR-3614-5p by directly targeting its 3'-untranslated regions. We verified that circ_0038985, which is positively correlated with RFC5, directly targeted miR-3614-5p. Overexpression of circ_0038985 promoted CRC cell migration and invasion, and these effects were partially reversed by the reintroduction of miR-3614-5p. Moreover, we found that RFC5 may promote the vascular endothelial growth factor A (VEGFa)/vascular endothelial growth factor receptor 2 (VEGFR2)/extracellular signal-regulated protein kinase (ERK) pathway. The knockdown of RFC5 reduced CRC tumorigenesis in vivo. Collectively, these data demonstrate that the circ_0038985/miR-3614-5p/RFC5 axis plays a critical role in the progression of CRC, and RFC5 may promote CRC progression by affecting the VEGFa/VEGFR2/ERK pathway., (© 2023 Wiley Periodicals LLC.)

Background: Chronic obstructive pulmonary disease (COPD) and lung cancer is the most common respiratory diseases among smokers. Lung cancer is one of the most common malignancy. COPD patients are at high risk of lung cancer, but the specific pathogenesis is still unclear. In recent years, circRNAs have become the focus of lung cancer research. This study investigated the effect of CircTMEM30A on COPD and lung cancer and its possible molecular mechanisms., Methods: QPCR was used to detect the expression of CircTMEM30A in COPD patients, lung cancer patients, and COPD patients with lung cancer. CircTMEM30A was transfected with PPF, HFL1, A549 and NCI-H446 cells, and the transfection efficiency was verified by real-time quantitative PCR. The effects of CircTMEM30A on the proliferation, migration and invasion of fibroblasts and lung cancer cells were detected by MTT, Transwell and scratch tests. The target gene of CircTMEM30A was verified by luciferase reporter assay. Real-time quantitative PCR was used to detect the expression level of CircTMEM30A target gene., Results: CircTMEM30A was the most highly expressed in COPD patients with lung cancer. Overexpression of CircTMEM30A enhanced the effects of proliferation, migration and invasion of fibroblasts and lung cancer cells, down-regulated the expression of E-cadherin, up-regulated the expression of Vimentin, and promoted EMT. After knocking down CircTMEM30A, the migration and invasion ability of lung cancer cells was significantly reduced. Bioinformatics retrieval and luciferase reporter gene confirmed that CircTMEM30A could be bind to miR-130a, while TNFα gene was the target gene of miR-130a. Further studies showed that overexpression of miR-130a and knockdown of TNFα reversed the promoting effect of CircTMEM30A on lung cancer cells., Conclusions: CircTMEM30A was highly expressed in COPD patients with lung cancer, and miR-130a was low expressed in lung cancer cells. CircTMEM30A regulated the expression of TNFα through miR-130a, thereby affecting the progression of COPD and lung cancer. CircTMEM30A may be a therapeutic target for COPD patients with lung cancer.

microRNA (miRNA) is a kind of small non-coding RNA that has been regarded as potential biomarkers for cancers. Sensitive and specific detection of miRNA at low expression levels is highly desirable but remains challenging, especially for amplification-free and portable point of care (POC) diagnostics. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a has been recently discovered and used in the field of RNA detection. Nonetheless, most CRISPR/Cas13a-based methods were burdened with expensive equipment, time-consuming procedures, and complicated operations which were not suitable for POC analysis. In this work, we constructed a three-dimensional tetrahedral DNA framework based CRISPR-electrochemical biosensor (CRISPR-E). By combining tetrahedral DNA framework, CRISPR, and electrochemical biosensor, the process of activation, cleavage of Cas13a, and signal readout were all finished on the chip, and a simple, amplification-free and sensitive detection of miRNA-19b was realized. Under the optimal experimental conditions, a linear range from 10 pM to 10 4 pM with detection limit of 10 pM for miRNA-19b in buffer solution was achieved. Selectivity analysis indicated that our CRISPR-E had good distinguishing ability between miRNA-19b and miRNA-197. The results of miRNA-19b detection in mimic serum samples were consistent with that of the buffer solution. This all-on-chip strategy of our CRISPR-E is very suitable for POC testing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Background: Colorectal cancer (CRC) is one of the leading causes of cancer-related death worldwide. P21 activated kinase 4 (PAK4) and Breast cancer anti-estrogen resistance 3 (BCAR3) have been reported to be involved in numerous aspects in tumorous progression. In this study, we propose to screen multi-targeted microRNAs. (miRNAs), which simultaneously inhibit neoplastic evolution through suppressing the transcription of target genes., Methods: MTT and Colony formation assays measured cell's viability and proliferation. Scratch wound and Transwell assays detected the ability in migration and invasion for SW116 cells. The multi-targeted microRNAs of PAK4 and BCAR3 were predicted using bioinformatics analysis and verified by conducting dual luciferase reporter assay, western blot and qRT-PCR that could detect the expression levels of miR-199a/b-3p., Results: The knockdown of PAK4 significantly impeded proliferation and colony formation of SW1116 cells when the knockdown of BCAR3 hindered migration and invasion of SW1116 cells. MiR-199a/b-3p directly targeted the 3'-UTR of PAK4 and BCAR3, further effected proliferation, colony formation, migration, and invasion of SW1116 cells. PAK4 or BCAR3 overexpression could partially reversed inhibitory effects of miR-199a/b-3p., Conclusions: These results provided a new multi-targeted cite for cancerous suppressant to improve the prognosis of CRC inpatients., (© 2022. The Author(s).)

The accurate and effective imaging of tumor-related miRNA in living cells has been playing an increasingly important role in cancer imaging. However, due to the low miRNA content and complex intracellular microenvironment, the current imaging methods of miRNAs in living cells still have some limitations. In this work, we developed a designer nanoprobe of tetrahedral DNA framework (TDF) combined with MB (termed TDFM nanoprobe) for the efficient fluorescence imaging of tumor-related miRNA-214 in living cells. In cell-free experiments, we demonstrated that the TDFM nanoprobe has sensitive detection and good specificity by fluorescence measurements. Before the TDFM nanoprobe was used for intracellular miRNA-214 fluorescence imaging, we confirmed its intracellular stability and negligible cytotoxicity by a standard MTT assay. In intracellular imaging experiments, we observed the strong fluorescence signal exhibited by the cells incubated with the TDFM nanoprobe using confocal fluorescence microscopy, which indicated that the TDFM nanoprobe was suitable for detecting and imaging tumor-related miRNA-214 in living cells. Furthermore, under the optimal incubation conditions, we employed the TDFM nanoprobe to study differences in the expression levels of tumor-related miRNA-214 in human breast cancer cells (MCF-7) and human umbilical vein endothelial cells (HUVEC). The TDFM nanoprobe we designed shows great potential to be applied in the development of DNA nanodevices, providing an improved strategy for the fluorescence imaging of miRNAs in living cells.

In situ nondestructive bioanalysis of targets in nanoscale confined space, e.g. exosomes, poses a high challenge to analytical technologies, especially to molecular fluorescent probes, because it is required to enter the confined space to recognize the target, and maintain independent and stable signal output. The unexpected fluorescence quenching and fluorescence resonance energy transfer (FRET) caused by high-frequency Brownian motion and collision in confined space are the main limiting factors. Herein, we constructed a well-defined and programmable cubic DNA nanocage-based three-dimensional molecular beacon (ncMB), which successfully broke through the above dilemma, and realized the detection of miRNA in exosomes. Specifically, steric hindrance and electrostatic repulsion derived from the unique three-dimensional structure of ncMB result in a barrier between fluorescent probes, thus eliminating unexpected fluorescence quenching during single exosomal miRNA detection and unexpected FRET during dual exosomal miRNA detection. Benefiting from the excellent anti-fluorescence and anti-FRET performance of ncMB, compared with traditional molecular beacons (MB), the detected fluorescence signal in exosomes can be improved by an order of magnitude. Moreover, ncMB is proven to have powerful programmability and anti-interference capability. Overall, it is believed that the ncMB can eliminate the signal distortion that was usually associated with commonly used MB, especially in the confined space. The ncMB is considered as a powerful and versatile tool for accurate in situ signal output in exosomes and maybe other confined spaces., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Accurately detecting and quantifying tumor-related microRNAs (miRNAs) in living cells is of great value for early cancer diagnosis. Herein, we present poly-adenine (polyA)-mediated spherical nucleic acid (SNA) nanoprobes for intracellular miRNA imaging in living cells., Methods and Results: polyA-mediated spherical nucleic acid (pASNA) nanoprobes consist of gold nanoparticles (AuNPs) anchored with fluorophore-labeled DNA molecules pre-hybridized with recognition sequences and polyA tails. The detection performance for miRNAs in vitro was studied to confirm the feasibility of pASNA nanoprobes for imaging live cell miRNAs. Before the pASNA nanoprobes were used for imaging intracellular miRNAs in MCF-7, HeLa, and LO2 cells, the stability and non-cytotoxicity were investigated using Dnase I and a standard colorimetric CCK8 assay. Flow cytometry, qRT-PCR analyses were conducted to confirm the different expression levels of miR-155 in live cells. Results showed that the pASNA nanoprobes had good detection sensitivity and specificity, excellent stability, and low toxicity. After incubating with pASNA nanoprobes, noticeable fluorescence signal enhancement could be clearly observed in MCF-7 and HeLa cells but not LO2 cells by confocal microscopy. Flow cytometry analysis and qRT-PCR indicated that MCF-7 and HeLa cells had higher miR-155 expression levels compared to LO2 cells., Conclusions: The pASNA nanoprobes we developed had good sensitivity and specificity, excellent nuclease stability and low toxicity, thus representing a new approach to exquisitely reveal the distribution of endogenous miRNAs in live cells., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

In this study, MicroRNA-210 (miR-210), which was previously proved to be a potential immunomodulator in various disease, attenuated mouse myocardium ischemia/reperfusion (I/R) injury. miR-210 was increased in cardiomyocytes exposed to hypoxia/reoxygenation (H/R). The expression of IL-6 and TNF-α in both serum and supernatant were reduced in miR-210 mimics groups. Mice were randomly divided into four groups, which were pre-treated with saline (sham and ischemia/reperfusion group), miR-210 mimics and miR-210 inhibitor treatments. Three days later, the mouse IR model was established by ischemia for 30 min, followed by reperfusion for 3 h. Myocardium and plasma were harvested and assessed. The myocardium histopathological changes were reduced in miR-210 mimics groups, and serum levels of Creatine kinase isoenzyme (CK-MB) and Lactate dehydrogenase (LDH) were significantly decreased compared with I/R groups. The protein expression of proinflammatory factor interleukin (IL)-1β and IL-6 were suppressed by the up-regulation of miR-210. The expression of miR-210 was negatively correlated with the expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). In conclusion, our study indicates that miR-210 protects heart from myocardium I/R injury via suppressing NF-κB signal pathway., (Copyright 2020 Biolife Sas. www.biolifesas.org.)

Rationale: Differential activation of macrophages correlates closely with tumor progression, and the epigenetic factor lysine demethylase 6B (KDM6B, previously named JMJD3) mediates the regulation of macrophage polarization through an unknown mechanism. Methods: We developed a suspension coculture system comprising breast cancer cells and macrophages and used RT-qPCR and western blotting to measure KDM6B expression. Bioinformatics and luciferase reporter assays were used to identify candidate microRNAs of cancer cells responsible for the downregulation of KDM6B. To determine if exosomes mediated the transfer of miR-138-5p between cancer cells to macrophages, we treated macrophages with exosomes collected from the conditioned medium of cancer cells. The effects of exosomal miR-138-5p on macrophage polarization were measured using RT-qPCR, flow cytometry, and chromatin immunoprecipitation assays. We employed a mouse model of breast cancer, metastatic to the lung, to evaluate the effects on tumor metastasis of macrophages treated with miR-138-5p-enriched exosomes. To develop a diagnostic evaluation index, the levels of exosomal miR-138-5p in samples from patients with breast cancer were compared to those of controls. Results: Coculture of breast cancer cells led to downregulation of KDM6B expression in macrophages. Cancer cell-derived exosomal miR-138-5p inhibited M1 polarization and promoted M2 polarization through inhibition of KDM6B expression in macrophages. Macrophages treated with exosomal miR-138-5p promoted lung metastasis, and the level of circulating exosomal miR-138-5p positively correlated with the progression of breast cancer. Conclusion: Our data suggest that miR-138-5p was delivered from breast cancer cells to tumor-associated macrophages via exosomes to downregulate KDM6B expression, inhibit M1 polarization, and stimulate M2 polarization. Therefore, exosomal miR-138-5p represents a promising prognostic marker and target for the treatment of breast cancer., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background: Basal cell carcinoma (BCC) is the most common form of skin carcinoma., Aim: To investigate the function of key micro(mi)RNAs and to explore the potential molecular mechanisms involved in BCC., Methods: The microarray dataset GSE34535, which comprises seven BCC samples and seven control samples, was downloaded from the Gene Expression Omnibus database. Differentially expressed miRNAs (DE-miRNAs) were identified. We collected tissue samples from 20 patients with BCC and 20 healthy controls (HCs), to compare the miR-18a expression in their tissue samples. Expression of miR-18a in A431 and HaCaT cells was also assayed. Following this, we upregulated and downregulated miR-18a expression in A431 cells to examine the effects on cell proliferation, migration and apoptosis. To further investigate the relative mechanism, the proteins LC3, Beclin 1, Akt and mammalian target of rapamycin (mTOR) were examined by quantitative real-time PCR and Western blotting. For further verification, we examined the expression of LC3 in the 20 BCC and 20 HC tissue samples., Results: In total, 19 DE-miRNAs (13 upregulated and 6 downregulated) that were common to the BCC and HC groups were identified. Levels of miR-18a were about three-fold higher in BCC tissues and A431 cells compared with their respective control groups. In vitro, downregulation of miR-18a was shown to inhibit cell proliferation and activate autophagy via the Akt/mTOR signalling pathway, while upregulation of miR-18a promoted proliferation of these cells. LC3 was decreased in BCC compared with HC tissue samples., Conclusions: Our data support an oncogenic role of miR-18a through a novel Akt/mTOR/Beclin 1/LC3 axis, and suggest that the antitumour effects of miR-18a inhibitor may make it suitable for BCC therapy., (© 2020 British Association of Dermatologists.)

Background: The endothelium is the first line of defence against harmful microenvironment risks, and microRNAs (miRNAs) involved in vascular inflammation may be promising therapeutic targets to modulate atherosclerosis progression. In this study, we aimed to investigate the mechanism by which microRNA-216a (miR-216a) modulated inflammation activation of endothelial cells . Methods . A replicative senescence model of human umbilical vein endothelial cells (HUVECs) was established, and population-doubling levels (PDLs) were defined during passages. PDL8 HUVECs were transfected with miR-216a mimics/inhibitor or small interfering RNA (siRNA) of SMAD family member 7 (Smad7). Real-time PCR and Western blot assays were performed to detect the regulatory role of miR-216a on Smad7 and NF- κ B inhibitor alpha (I κ B α ) expression. The effect of miR-216a on adhesive capability of HUVECs to THP-1 cells was examined. MiR-216a and Smad7 expression in vivo were measured using human carotid atherosclerotic plaques of the patients who underwent carotid endarterectomy ( n = 41)., Results: Luciferase assays showed that Smad7 was a direct target of miR-216a. Smad7 mRNA expression, negatively correlated with miR-216a during endothelial aging, was downregulated in senescent PDL44 cells, compared with young PDL8 HUVECs. MiR-216a markedly increased endothelial inflammation and adhesive capability to monocytes in PDL8 cells by promoting the phosphorylation and degradation of I κ B α and then activating NF- κ B signalling pathway. The effect of miR-216a on endothelial cells was consistent with that blocked Smad7 by siRNAs. When inhibiting endogenous miR-216a, the Smad7/I κ B α expression was rescued, which led to decreased endothelial inflammation and monocytes recruitment. In human carotid atherosclerotic plaques, Smad7 level was remarkably decreased in high miR-216a group compared with low miR-216a group. Moreover, miR-216a was negatively correlated with Smad7 and I κ B α levels and positively correlated with interleukin 1 beta (IL1 β ) expression in vivo ., Conclusion: In summary, our findings suggest a new mechanism of vascular endothelial inflammation involving Smad7/I κ B α signalling pathway in atherosclerosis., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2020 Shujun Yang et al.)

Based on the abundance of taste compounds in leaves at different leaf positions on the same shoot, green tea made from one bud and one leaf, or even just one bud, has the best quality. To elucidate the mechanism underlying the regulation of the biosynthesis of these compounds, we profiled the metabolome, transcriptome, sRNA, degradome, and WGCNA using leaves from five leaf positions of shoots. Through this analysis, we found 139 miRNA-target pairs related to taste compound biosynthesis and 96 miRNA-target pairs involved in phytohormone synthesis or signal transduction. Moreover, miR166- HD-ZIP , miR169- NF-YA , IAA, ZA, ABA, and JA were positively related to the accumulation of gallated catechin, caffeine, and theanine. However, miR396- GRF , miR393- bHLH , miR156- SBP , and SA were negatively correlated with these compounds. Among these important pairs, the miR396- GRF and miR156- SBP pairs were further validated by using qRT-PCR, Northern blots, and cotransformation. This is the first report describing that miRNA-TF pairs and phytohormones might synergistically regulate the biosynthesis of taste compounds in the leaves of tea plants.

Main Conclusion: The roles of microRNA-mediated epigenetic regulation were highlighted in the bud dormancy-activity cycle, implying that certain differentially expressed miRNAs play crucial roles in apical bud burst, such as csn-miR319c/TCP2. microRNAs (miRNAs) are a class of small non-coding RNAs that regulate gene expression by targeting mRNA transcripts for cleavage or directing translational inhibition. To investigate whether miRNAs regulate bud dormancy-activation transition in tea plant, which largely affects the yield and price of tea products and adaptability of tea trees, we constructed small RNA libraries from three different periods of bud dormancy-burst transition. Through sequencing analysis, 262 conserved and 83 novel miRNAs were identified, including 118 differentially expressed miRNAs. Quantitative RT-PCR results for randomly selected miRNAs exhibited that our comprehensive analysis is highly reliable and accurate. The content of caffeine increased continuously from the endodormancy bud to flushing bud, and differentially expressed miRNAs coupling with their targets associated with bud burst were identified. Remarkably, csn-miR319c was downregulated significantly from the quiescent bud to burst bud, while its target gene CsnTCP2 (TEOSINTE BRANCHED/CYCLOIDEA/PROLIFERATING CELL FACTOR 2) displayed opposite expression patterns. Co-transformation experiment in tobacco demonstrated that csn-miR319c can significantly suppress the functions of CsnTCP2. This study on miRNAs and the recognition of target genes could provide new insights into the molecular mechanism of the bud dormancy-activation transition in tea plant.

Tea, made from leaves of Camellia sinensis, has long been consumed worldwide for its unique taste and aroma. Terpenoids play important roles not only in tea beverage aroma formation, but also in the productivity and quality of tea plantation due to their significant contribution to light harvesting pigments and phytohormones. To date, however, the regulation of terpenoid synthase genes remains unclear. Herein, the analyses of metabolomics, sRNAs, degradome, and transcriptomics were performed and integrated for identifying key regulatory miRNA-target circuits on terpenoid biosynthesis in leaf tissues over five different months in which the amount of terpenoids in tea leaves varies greatly. Four classes of miRNA-TF pairs that might play a central role in the regulation of terpenoid biosynthesis were also uncovered. Ultimately, a hypothetical model was proposed that mature miRNAs maintained by light regulator at both the transcriptional and posttranscriptional levels negatively regulate the targets to control terpenoid biosynthesis.

Study Question: Is it possible to improve fibrosis in endometriosis by microRNA-214 delivery in exosomes?, Summary Answer: Upregulation of miR-214 may inhibit fibrogenesis and its delivery by exosomes derived from ectopic endometrial stromal cells (ESCs), offers an alternative therapeutic approach for endometriosis fibrosis., What Is Known Already: Fibrosis is the primary pathological feature of endometriosis. MiR-214 plays an important role in fibrotic disease. Connective tissue growth factor (CTGF) is a critical fibrogenic mediator of miR-214. The expression of miR-214 is decreased in ectopic ESCs compared with normal ESCs. miRNAs are a natural cargo of exosomes and these could be exploited as carriers of miRNA in replacement therapy., Study Design, Size, Duration: Paired eutopic and ectopic endometrial tissue samples were obtained from 10 women with ovarian endometrioma. ESCs and epithelial cells from both were cultured in vitro. RT-PCR, western blot and immunohistochemistry were used to study the effect of transfection with miR-214 mimics on CTGF expression and fibrogenesis respectively, with and without TGFβ stimulation. Exosomes were isolated from ectopic ESCs and Endometrioma tissue was isolated from four patients, dispersed an injected (ip) into nude mice and allowed to implant. The mice were treated with miR-214-enriched exosomes or controls to confirm the effect of inhibiting CTGF overexpression on endometriosis fibrosis., Participants/materials, Setting, Methods: The primary ectopic ESCs were transfected with miR-214 mimics. The levels of miR-214, CTGF and fibrotic markers were measured by RT-PCR and Immunohistochemistry. A mouse model of endometriosis was established by ip injection of human ectopic endometrial tissues into nude mice. MiR-214-enriched exosomes were injected into the mice and endometriotic lesions were measured on Day 28. Changes in fibrosis of the endometriotic implants were studied by histopathological staining., Main Results and the Role of Chance: CTGF and fibrotic markers upregulation in endometriosis is associated with a reciprocal down-regulation of miR-214. By using miR-214 mimics and antagomirs to investigate expression of fibrotic markers, we found that increased production of miR-214 reduced Collagen αI and CTGF expression in endometriosis stromal and endometrial epithelial cells in response to fibrosis-inducing stimuli (P < 0.001 versus non-treatment). Ectopic ESCs yielded nano-sized exosomes which expressed miR-214. Loading exosomes with miR-214 mimics and injecting them into an experimental endometriosis mouse model resulted in a decrease in the expression of fibrosis-associated proteins (P < 0.001 versus PBS control group)., Large Scale Data: N/A., Limitations, Reasons for Caution: We only isolated exosomes from ectopic ESCs, whether this is the optimum source requires further study., Wider Implications of the Findings: Upregulation of miRNA-214 potentially offers an alternative therapeutic approach for endometriosis fibrosis., Study Funding/competing Interest(s): This work was supported by grants from the National Natural Science Foundation of China (Grant no. 81771549 Jinwei Miao). The authors declare that there is no conflict of interest.

Vascular endothelial senescence contributes to atherosclerosis and coronary artery disease (CAD), but the mechanisms are yet to be clarified. We identified that microRNA-216a (miR-216a) significantly increased in senescent endothelial cells. The replicative senescence model of human umbilical vein endothelial cells (HUVECs) was established to explore the role of miR-216a in endothelial ageing and dysfunction. Luciferase assay indicated that Smad3 was a direct target of miR-216a. Stable expression of miR-216a induced a premature senescence-like phenotype in HUVECs with an impairment in proliferation and migration and led to an increased adhesion to monocytes by inhibiting Smad3 expression and thereafter modulating the degradation of NF-κB inhibitor alpha (IκBα) and activation of adhesion molecules. Conversely, inhibition of endogenous miR-216a in senescent HUVECs rescued Smad3 and IκBα expression and inhibited monocytes attachment. Plasma miR-216a was significantly higher in old CAD patients (>50 years) and associated with increased 31% risk for CAD (odds ratio 1.31, 95% confidence interval 1.03-1.66; P = .03) compared with the matched healthy controls (>50 years). Taken together, our data suggested that miR-216a promotes endothelial senescence and inflammation as an endogenous inhibitor of Smad3/IκBα pathway, which might serve as a novel target for ageing-related atherosclerotic diseases., (© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Objective: Cisplatin is an important chemotherapeutic agent frequently used in the treatment of ovarian cancer. However, resistance to cisplatin is an obstacle to the treatment of ovarian cancer. Recently, many studies have demonstrated that microRNAs (miRNAs) are involved in the drug resistance of ovarian cancer cells. In this study, we explored the role of miR-503 in cisplatin-resistant ovarian cancer., Materials and Methods: To investigate the relationship between miR-503 expression and the sensitivity of ovarian cancer cells to cisplatin, the cells were transfected with miR-503 mimics/inhibitors. The relative expression of miR-503 RNA and its targeted gene PI3K mRNA were detected by real-time PCR (RT-PCR). Western blot was used to measure relevant protein levels. Flow cytometry and CCK-8 assay were used to analyze cell proliferation and apoptosis., Results: MiR-503 expression was significantly downregulated in cisplatin-resistant ovarian cancer cell line SKOV3/DDP compared with parental SKOV3. Over-expression and knock-down of miR-503 partially regulated apoptotic activity and changed the cisplatin resistance of ovarian cancer cells. In exploring the underlying mechanisms of miR-503 in ovarian cancer cells' resistance to cisplatin, we found that miR-503 can directly target PI3K p85 and participates in the regulation of the PI3K/Akt signaling pathway. In vivo, miR-503 agomirs combined with cisplatin treatment significantly reduced the growth of tumors compared with cisplatin alone., Conclusions: Our data suggest that miR-503 might be a sensitizer to cisplatin treatment in ovarian cancer by targeting PI3K p85, thus giving a new insight into developing therapeutic strategies to overcome cisplatin resistance in ovarian cancer.

Objective: To detect the change in miRNA-210 expression of cardiomyocytes under hypoxia/reoxygenation status. Also, the effect of miR-210 on the apoptosis of cardiomyocytes induced by oxygen-glucose deprivation/reperfusion (OGD/R) and its mechanism through establishing the OGD/R injury model of primary cardiomyocytes in this experiment were investigated., Materials and Methods: The cell model of OGD/R injury was established. The cell apoptosis in each group was detected by methyl thiazolyl tetrazolium (MTT) assay and detection of Caspase-3 activity. The change in miR-210 expression in each group was detected by Real-time fluorescence quantitative polymerase chain reaction (PCR). The high-expression and low-expression miR-210 models were established through the transient transfection of miR-210 mimic and inhibitor to detect the relevant indexes of cell apoptosis. At the same time, changes in mRNA and protein expressions of E2F3 were detected by RT-PCR and Western blotting, respectively. The E2F3 overexpression vector was constructed, and the overexpression vector plasmid and miR-210 mimic were jointly transfected into the cells to detect the relevant indexes of cell apoptosis., Results: After OGD/R treatment, the activity of Caspase-3 was increased, the survival of cardiomyocytes was significantly inhibited and the expression level of miR-210 was up-regulated in OGD/R injury. Transfection of miR-210 mimic for miR-210 overexpression could alleviate the OGD/R-induced cardiomyocyte injury, while the decrease of miR-210 expression could aggravate the apoptosis of cardiomyocytes. In addition, the high expression of miR-210 could inhibit the protein expression of E2F3, and co-transfection of E2F3 plasmid and miR-210 mimic could reverse the inhibiting effect of miR-210 on the apoptosis of cardiomyocytes., Conclusions: We confirmed that miR-210 can inhibit the OGD/R-induced apoptosis of cardiomyocytes, and miR-210, as an upstream factor, plays a protective role in cardiomyocytes through directly inhibiting the protein expression of its target gene E2F3.

Simultaneous detection of multiple biomarkers has important prospects in the biomedical field. In this work, we demonstrated a novel strategy for the detection of multiple microRNAs (miRNAs) based on gold nanoparticles (Au NPs) and polyadenine (polyA) mediated nanoscale molecular beacon (MB) probes (denoted p-nanoMBs). Novel fluorescent labeled p-nanoMBs bearing consecutive adenines were designed, of which polyA served as an effective anchoring block binding to the surface of Au NPs, and the appended hairpin block formed an upright conformation that favored the hybridization with targets. Using the co-assembling method and the improved hybridization conformation of the hairpin probes, we achieved high selectivity for specifically distinguishing DNA targets from single-base mismatched DNA targets. We also realized multicolor detection of three different synthetic miRNAs in a wide dynamic range from 0.01 nM to 200 nM with a detection limit of 10 pM. What's more, we even detected miRNAs in a simulated serum environment, which indicated that our method could be used in complex media. Compared with the traditional method, our strategy provides a promising alternative method for the qualitative and quantitative detection of miRNAs.

A novel biosensor platform was developed for detection of microRNAs (miRNAs) based on graphene quantum dots (GQDs) and pyrene-functionalized molecular beacon probes (py-MBs). Pyrene was introduced to trigger specifically fluorescence resonance energy transfer (FRET) between GQDs and fluorescent dyes labeled on py-MBs, and the unique fluorescent intensity change produced a novel signal for detection of the target. The platform realized detection of miRNAs in a wide range from 0.1 nM to 200 nM with great discrimination abilities, as well as multidetection of different kinds of miRNAs, which paved a brand new way for miRNA detection based on GQDs.

Competing endogenous RNA (ceRNA) interactions form a multilayered network that regulates gene expression in various biological pathways. Recent studies have demonstrated novel roles of ceRNA interactions in tumorigenesis, but the dynamics of the ceRNA network in cancer remain unexplored. Here, we examine ceRNA network dynamics in prostate cancer from the perspective of alternative cleavage and polyadenylation (APA) and reveal the principles of such changes. Analysis of exon array data revealed that both shortened and lengthened 3'UTRs are abundant. Consensus clustering with APA data stratified cancers into groups with differing risks of biochemical relapse and revealed that a ceRNA subnetwork enriched with cancer genes was specifically dysregulated in high-risk cancers. The novel connection between 3'UTR shortening and ceRNA network dysregulation was supported by the unusually high number of microRNA response elements (MREs) shared by the dysregulated ceRNA interactions and the significantly altered 3'UTRs. The dysregulation followed a fundamental principle in that ceRNA interactions connecting genes that show opposite trends in expression change are preferentially dysregulated. This targeted dysregulation is responsible for the majority of the observed expression changes in genes with significant ceRNA dysregulation and represents a novel mechanism underlying aberrant oncogenic expression.

MicroRNAs (miRNAs) are a class of endogenous small non-coding RNAs that regulate gene expression by post-transcriptional repression of messenger RNA. The echinoderm, Strongylocentrotus nudus, is an excellent model organism for studying development and commercially important as a food source. However, to date no miRNAs have been reported to modulate sex gonad differentiation in S. nudus. In this study, we constructed two small RNA libraries from male and female S. nudus gonad respectively for Solexa sequencing. A total of 184 miRNAs including 60 known and 124 novel miRNAs were identified from the two libraries. Furthermore, the nucleotide bias and end variation of the known miRNAs were also analyzed. In addition, 67 differently expressed of the 86 co-expressed and 98 gender-specific (47 male-specific and 51 female-specific) miRNAs that may be involved in sexual differentiation were found by comparing the miRNA expression profiles in the two libraries. This study reveals the first miRNA profile related to the gonad differentiation of the S. nudus. This study gives a first insight into sex differences in miRNA expression of sea urchin which could facilitate studies of the reproductive organ-specific roles of miRNAs., (Copyright © 2014 Elsevier Inc. All rights reserved.)

microRNAs (miRNAs) are a family of important small non-coding RNA molecules, which participate in the post transcriptional gene regulation. In this review, the numbers and chromosomal distribution of chicken miRNAs, and the regulation and function of chicken miRNAs in immune, embryo development and virus infection were reviewed. Additionally, the applications of chicken miRNAs were also discussed briefly. We hope it can provide references for further study and use of miRNAs in poultry husbandry fields.