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MicroRNAs (miRNAs) play a key role in physiological processes, and their dysregulation is closely related to various human diseases. Simultaneous detection of multiple miRNAs is pivotal to cancer diagnosis at an early stage. However, most multicomponent analyses generally involve multiple excitation wavelengths, which are complicated and often challenging to simultaneously acquire multiple detection signals. In this study, a convenient and sensitive sensor was developed to simultaneously detection of multiple miRNAs under a single excitation wavelength through the fluorescence resonance energy transfer between the carbon dots (CDs)/quantum dots (QDs) and graphene oxide (GO). A hybridization chain reaction (HCR) was triggered by miRNA-141 and miRNA-21, resulting in the high sensitivity with a limit of detection (LOD) of 50 pM (3σ/k) for miRNA-141 and 60 pM (3σ/k) for miRNA-21. This simultaneous assay also showed excellent specificity discrimination against the mismatch. Furthermore, our proposed method successfully detected miRNA-21 and miRNA-141 in human serum samples at a same time, indicating its diagnostic potential in a clinical setting., 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 © 2024 Elsevier B.V. All rights reserved.)

Challenges remained in precisely real-time monitoring of apoptotic molecular events at the subcellular level. Herein, we developed a new type of intelligent DNA biocomputing nanodevices (iDBNs) that responded to mitochondrial microRNA-21 (miR-21) and microRNA-10b (miR-10b) simultaneously which were produced during cell apoptosis. By hybridizing two hairpins (H1 and H2) onto DNA nanospheres (DNSs) that had been previously modified with mitochondria-targeted triphenylphosphine (TPP) motifs, iDBNs were assembled in which two localized catalytic hairpins self-assembly (CHA) reactions occurred upon the co-stimulation of mitochondrial miR-21 and miR-10b to perform AND logic operations, outputting fluorescence resonance energy transfer (FRET) signals for sensitive intracellular imaging during cell apoptosis. Owing to the spatial confinement effects of DNSs, it was discovered that iDBNs had a high efficiency and speed of logic operations by high local concentrations of H1 and H2, making the simultaneous real-time responses of mitochondrial miR-21 and miR-10b reliable and sensitive during cell apoptosis. These results demonstrated that iDBNs were simultaneously responsive to multiple biomarkers, which greatly improved the detection accuracy to identify the cell apoptosis, demonstrating that iDBNs are highly effective and reliable for the diagnosis of major disease and screening of anticancer drugs.

DNA logic computing has captured increasing interest due to its ability to assemble programmable DNA computing elements for disease diagnosis, gene regulation, and targeted therapy. In this work, we developed an aptamer-equipped high-integrated DNA biocomputing platform (HIDBP-A) with a dual-recognition function that enabled cancer cell targeting. Dual microRNAs were the input signals and can perform AND logic operations. Compared to the free DNA biocomputing platform (FDBP), the integration of all computing elements into the same DNA tetrahedron greatly improved logic computing speed and efficiency owing to the confinement effect reflected by the high local concentration of computing elements. As a proof of concept, the utilization of microRNA as the input signal was beneficial for improving the scalability and flexibility of the sequence design of the logic nano-platform. Given that the different microRNAs were over-expressed in cancer cells, this new HIDBP-A has great promise in accurate diagnosis and logic-controlled disease treatment., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Accurate detection and imaging of tumor-related microRNA (miRNA) in living cells hold great promise for early cancer diagnosis and prognosis. One of the challenges is to develop methods that enable the identification of multiple miRNAs simultaneously to further improve the detection accuracy. Herein, a simultaneous detection and imaging method of two miRNAs was established by using a programmable designed DNA tetrahedron nanostructure (DTN) probe that includes a nucleolin aptamer (AS1411), two miRNA capture strands, and two pairs of metastable catalytic hairpins at different vertexes. The DTN probe exhibited enhanced tumor cell recognition ability, excellent stability and biocompatibility, and fast miRNA recognition and reaction kinetics. It was found that the DTN probe could specifically enter tumor cells, in which the capture strand could hybridize with miRNAs and initiate the catalytic hairpin assembly (CHA) only when the overexpressed miR-21 and miR-155 existed simultaneously, resulting in a distinct fluorescence resonance energy transfer signal and demonstrating the feasibility of this method for tumor diagnosis.

Monitoring tumor biomarkers is crucial for cancer diagnosis, progression monitoring, and treatment. However, identifying single or multiple biomarkers with the same spatial locations can cause false-positive feedback. Herein, we integrated the DNA tetrahedron (DT) structures with logic-responsive and signal amplifying capability to construct transmembrane DNA logic nanodevices (TDLNs) for the in situ sequential imaging of transmembrane glycoprotein mucin 1 (MUC1) and cytoplasmic microRNA-21 (miR-21) to cell identifications. The TDLNs were developed by encoding two metastable hairpin DNAs (namely, H1 and H2) in a DT scaffold, in which the triggering toeholds of H1 for miR-21 were sealed by the MUC1-specific aptamer (MUC1-apt). The TDLNs not only had the function of signal amplification owing to the localized catalytic hairpin assembly (CHA) reaction through spatial constraints effect of DT structures but also performed an AND logic operation to output a green Cy3 signal in MCF-7 cells, where MUC1 protein and miR-21 were simultaneously expressed. These results showed that the newly developed TDLNs have better molecular targeting and recognition ability so as to be easily identify cell types and diagnose cancer early.

MicroRNAs (miRNAs) are found in extremely low concentrations in cells, so highly sensitive quantitation is a great challenge. Herein, a simple dual-amplification strategy involving target-activated catalytic hairpin assembly (CHA) coupled with multiple fluorophores concentrated on one X-shaped DNA is reported. In this strategy, four hairpin probes (H1, H2, H3, and H4) are modified with FAM and BHQ1 at both sticky ends, while a circulating hairpin probe (H0) is used to activate CHA circuits once it binds to complementary sequences in the target miR-21 (T). The powerful dual-amplification cascades in Förster resonance energy transfer (FRET)-based nonenzymatic nucleic acid circuits are triggered by T-H0-activated formation of the X-shaped DNA nanostructure, freeing T-H0 for the next CHA reaction cycle. CHA circuits increase the fluorescence due to the wide distance between FAM and BHQ1 in the formed X-shaped DNA nanostructure, resulting in signal amplification and highly sensitive detection of miR-21, with a limit of detection (LOD, 3σ) of 0.025 nM, which is 25.6 or 57.6 times lower than that obtained through a single-amplification strategy without multiple fluorophores on one X-shaped DNA or CHA circuit. Furthermore, this cascade reaction was completed in 45 min, effectively avoiding target degradation. This new enzyme-free signal amplification strategy holds promising potential for sensitively detecting different DNA or RNA sequences by simply adapting the fragment of the H0 sequence complementary to the target.

The current research aimed to investigate the effect of miR-7 targeting matrix metalloproteinase 14 (MMP-14) on homocysteine (Hcy)-induced rat cerebral artery vascular smooth muscle cells (VSMCs) proliferation, migration and inflammatory factor expression and its possible mechanism. The expression of miR-7 and MMP-14 in Hcy-induced VSMCs were detected by real-time fluorescent quantitative PCR (RT-qPCR) and Western blot. Methyl Thiazolyl Tetrazolium (MTT) method, Transwell assays and enzyme-linked immunosorbent assay (ELISA) were performed to detect the effect of miR-7 and MMP-14 expression on the proliferation and migration, as well as interleukin 6 (IL-6) and tumor necrosis factor ɑ (TNF-ɑ) expression of Hcy-induced VSMCs. The interaction between miR-7 and MMP-14 was detected by dual-luciferase reporter gene assay. Western blot was applied to analyse the effects of miR-7 and MMP-14 expression on the Toll-like receptor (TLR4)/nuclear transcription factor-KB (NF-κB) signaling pathway. The results showed that after induced by Hcy, the expression of miR-7 in VSMCs was significantly reduced, the expression of MMP-14 was significantly increased, and the cell viability, the number of migrating cells, IL-6 and TNF-ɑ expression were significantly increased (P<0.05). After overexpression of miR-7, the viability, migration cell numbers, IL-6 and TNF-ɑ expression of Hcy-induced VSMCs were significantly reduced (P<0.05). miR-7 directly binds to MMP-14 and negatively regulates the expression of MMP-14. After overexpression of miR-7, the levels of TLR4 and p-NF-κB p65 in VSMCs were significantly reduced (P<0.05); overexpression of MMP-14 could reduce the effect of miR-7 overexpression on TLR4 and p-NF-κB p65 expression in VSMCs (P<0.05). Overexpression of MMP-14 and/or activation of the TLR4/NF-κB signaling pathway could reverse the effect of miR-7 overexpression on the proliferation, migration and IL-6 and TNF-ɑ expression of Hcy-induced VSMCs (P<0.05). It is concluded that miR-7 can inhibit Hcy-induced rat cerebral artery VSMCs proliferation, migration, and inflammatory factor expression by targeting the regulation of MMP-14 expression and inhibiting the activation of the TLR4/NF-κB signaling pathway.

T-cell acute lymphoblastic leukemia (T-ALL) is a type of aggressive leukemia with inferior prognosis. Although activating mutations of NOTCH1 are observed in most T-ALL cases, these mutations alone are not sufficient to drive the full development of T-ALL. β-Arrestins (ARRB) are versatile and multifunctional adapter proteins that regulate diverse cellular functions, including promoting the development of cancer. However, the role of ARRBs in T-ALL has largely remained elusive. In this study, we showed that ARRB1 is expressed at low levels in assayed T-ALL clinical samples and cell lines. Exogenous ARRB1 expression inhibited T-ALL proliferation and improved the survival of T-ALL xenograft animals. ARRB1 facilitated NOTCH1 ubiquitination and degradation through interactions with NOTCH1 and DTX1. Mechanistically, the oncogenic miRNA (oncomiR) miR-223 targets the 3'-UTR of ARRB1 (BUTR) and inhibits its expression in T-ALL. Furthermore, overexpression of the ARRB1-derived miR-223 sponge suppressed T-ALL cell proliferation and induced apoptosis. Collectively, these results demonstrate that ARRB1 acts as a tumor suppressor in T-ALL by promoting NOTCH1 degradation, which is inhibited by elevated miR-223, suggesting that ARRB1 may serve as a valid drug target in the development of novel T-ALL therapeutics. Significance: These findings highlight a novel tumor suppressive function of the adaptor protein β-arrestin1 in T-ALL., (©2019 American Association for Cancer Research.)

Lung adenocarcinoma is the most prevalent type of lung cancer with a high incidence and mortality worldwide. Metastasis is the major cause of high death rate in lung cancer and the potential mechanism of lung adenocarcinoma metastasis remains indistinct. Emerging investigations have demonstrated that long noncoding RNA is a kind of non-protein coding RNA and plays a critical role in cancer progression and metastasis. TTN antisense RNA 1 (TTN-AS1) has been reported to promote cell growth and metastasis in cancer. However, the function of TTN-AS1 in lung adenocarcinoma is still to be illustrated. In this study, we observed that TTN-AS1 was upregulated in tissues and cells of lung adenocarcinoma and associated with poor overall survival. TTN-AS1 promoted cell proliferation, migration, invasion, and epithelial-mesenchymal transition in lung cancer. TTN-AS1 directly bound with miR-4677-3p and negatively regulated miR-4677-3p. MiR-4677-3p rescued the inhibitive impacts of TTN-AS1 knockdown on lung adenocarcinoma. Furthermore, zinc finger E-box binding homeobox 1 (ZEB1) was the target of miR-4677-3p, and TTN-AS1 modulated ZEB1 by competing for miR-4677-3p. TTN-AS1 drove the invasion and migration of lung adenocarcinoma cells by targeting the miR-4677-3p/ZEB1 axis. To sum up, our study offers insights into the mechanism of TTN-AS1 in lung adenocarcinoma metastasis and targeting the TTN-AS1/miR-4677-3p/ZEB1 axis may be the potential innovate therapeutic strategy for the patients with lung adenocarcinoma., (© 2019 Wiley Periodicals, Inc.)

Nonalcoholic fatty liver disease (NAFLD) can progress gradually to liver failure, early warning of which is critical for improving the cure rate of NAFLD. In situ imaging and monitoring of overexpressed miR-21 is an advanced strategy for NAFLD diagnosis. However, this strategy usually suffers from the high background imaging in living cells owing to the complexity of the biological system. To overcome this problem, herein, we have developed a one-donor-two-acceptor nanoprobe by assembling gold nanoparticles (AuNPs) coupled with BHQ2 (AuBHQ) and quantum dots (QDs) through DNA hybridization for imaging of miR-21 in living cells. The fluorescence of QDs was quenched up to 82.8% simultaneously by the AuNPs and the BHQ2 via nanometal surface energy transfer and fluorescence resonance energy transfer, reducing the background signals for target imaging. This low background fluorescent nanoprobe was successfully applied for imaging the target miR-21 in nonalcoholic fatty liver cells by catalyzing the disassembly of QDs with the AuBHQ and the fluorescence recovery of QDs. In addition, the sensitivity of this nanoprobe has also been enhanced toward detecting miR-21 in the range of 2.0-15.0 nM with the detection limit (LOD, 3σ) of 0.22 nM, which was 13.5 times lower than that without BHQ2. The proposed approach provides a new way for early warning, treatments, and prognosis of NAFLD.

Background: 'Fengzao' is an early-ripening bud mutant of 'Kyoho', which matures nearly 30 days earlier than 'Kyoho'. To gain a better understanding of the regulatory role of miRNAs in early-ripening of grape berry, high-throughput sequencing approach and quantitative RT-PCR validation were employed to identify miRNAs at the genome-wide level and profile the expression patterns of the miRNAs during berry development in 'Kyho' and 'Fengzao', respectively., Results: Nine independent small RNA libraries were constructed and sequenced in two varieties from key berry development stages. A total of 108 known miRNAs and 61 novel miRNAs were identified. Among that, 159 miRNAs identified in 'Fengzao' all completely expressed in 'Kyoho' and there were 10 miRNAs specifically expressed in 'Kyoho'. The expression profiles of known and novel miRNAs were quite similar between two varieties. As the major differentially expressed miRNAs, novel_144, vvi-miR3626-3p and vvi-miR3626-5p only expressed in 'Kyoho', vvi-miR399b and vvi-miR399e were down-regulated in 'Fengzao', while vvi-miR477b-3p up-regulated in 'Fengzao'. According to the expression analysis and previous reports, miR169-NF-Y subunit, miR398-CSD, miR3626-RNA helicase, miR399- phosphate transporter and miR477-GRAS transcription factor were selected as the candidates for further investigations of miRNA regulation role in the early-ripening of grape. The qRT-PCR analyses validated the contrasting expression patterns for these miRNAs and their target genes., Conclusions: The miRNAome of the grape berry development of 'Kyoho', and its early-ripening bud mutant, 'Fengzao' were compared by high-throughput sequencing. The expression pattern of several key miRNAs and their target genes during grape berry development and ripening stages was examined. Our results provide valuable basis towards understanding the regulatory mechanisms of early-ripening of grape berry.

The differential diagnosis of Parkinson's diseases (PD) is challenging, especially in the early stages of the disease. We developed a microRNA profiling strategy for exosomal miRNAs isolated from cerebrospinal fluid (CSF) in PD and AD. Sixteen exosomal miRNAs were up regulated and 11 miRNAs were under regulated significantly in PD CSF when compared with those in healthy controls (relative fold > 2, p < 0.05). MiR-1 and miR-19b-3p were validated and significantly reduced in independent samples. While miR-153, miR-409-3p, miR-10a-5p, and let-7g-3p were significantly over expressed in PD CSF exosome. Bioinformatic analysis by DIANA-mirPath demonstrated that Neurotrophin signaling, mTOR signaling, Ubiquitin mediated proteolysis, Dopaminergic synapse, and Glutamatergic synapse were the most prominent pathways enriched in quantiles with PD miRNA patterns. Messenger RNA (mRNA) transcripts [amyloid precursor protein (APP), α-synuclein (α-syn), Tau, neurofilament light gene (NF-L), DJ-1/PARK7, Fractalkine and Neurosin] and long non-coding RNAs (RP11-462G22.1 and PCA3) were differentially expressed in CSF exosomes in PD and AD patients. These data demonstrated that CSF exosomal RNA molecules are reliable biomarkers with fair robustness in regard to specificity and sensitivity in differentiating PD from healthy and diseased (AD) controls.

Background: The biological processes and molecular mechanisms underlying miR-107 remain unclear in gastric cancer(GC). In this study, we aimed to investigate the expression, biological functions and mechanisms of miR-107 in GC., Methods: Quantitative real-time RT-PCR was used to test miR-107 expression. MTT and colony formation assays were conducted to explore the potential function of miR-107 in human GC cell line SGC7901. The target gene was determined by bioinformatic algorithms, dual luciferase reporter assay, RT-PCR and Western blot., Results: Expression of miR-107 was significantly elevated in GC cell line than that in gastric epithelial cell line(p = 0.012). We found that miR-107 inhibitor transfection significantly decreased the proliferation of GC cell line, and clone formation rate of miR-107 inhibitor transfected group was significantly lower than that of control group. Luciferase assays using a reporter carrying a putative miR-107 target site in the 3'untranslated region (3'-UTR) of cyclin dependent kinase 8 (CDK8) revealed that miR-107 directly targets CDK8. The expression level of CDK8 mRNA and protein in miR-107 inhibitor transfected GC cell line was significantly decreased compared with control group., Conclusion: Our findings indicate that miR-107 is upregulated in GC and affects the proliferation of GC cells, partially through the regulation of CDK8., Virtual Slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000&#95;2014&#95;164.

microRNAs (miRNAs) are short noncoding RNAs that negatively regulate gene expression. Although recent evidences have been indicated that their aberrant expression may play an important role in cancer stem cells, the mechanism of their deregulation in neoplastic transformation of liver cancer stem cells (LCSCs) has not been explored. In our study, the HCC model was established in F344 rats by DEN induction. The EpCAM(+) cells were sorted out from unfractionated fetal liver cells and liver cancer cells using the FACS analysis and miRNA expression profiles of two groups were screened through microarray platform. Gain-of-function studies were performed in vitro and in vivo to determine the role of miR-92b on proliferation and differentiation of the hepatic progenitors. In addition, luciferase reporter system and gene function analysis were used to predict miR-92b target. we found that miR-92b was highly downregulated in EpCAM(+) fetal liver cells in expression profiling studies. RT-PCR analysis demonstrated reverse correlation between miR-92b expression and differentiation degree in human HCC samples. Overexpression of miR-92b in EpCAM(+) fetal liver cells significantly increased proliferation and inhibited differentiation as well as in vitro and in vivo studies. Moreover, we verified that C/EBPß is a direct target of miR-92b and contributes to its effects on proliferation and differentiation. We conclude that aberrant expression of miR-92b can result in proliferation increase and differentiation arrest of hepatic progenitors by targeting C/EBPß.