MICROCHIMICA ACTA
·
2020
·Vol. 187
·期 2
·1-8
·
IF 6.232
Li Ruiying,Liu Qiang,Jin Yan,Li Baoxin
The authors have incidentally found that the three tandem repeats of a 13-mer G-rich oligomer (with sequence 5′-TGG GAA GGG AGG G-3′; referred to as G3) can directly fold into a stable G3 trimer. The G3 trimer/hemin DNAzyme exhibits an about 3-fold higher peroxidase-mimicking activity compared to the conventional G3/hemin DNAzyme. Combining this finding with rolling circle amplification (RCA), a colorimetric assay was developed for sensitive and specific determination of microRNA. In this method, each cycle of RCA generates three catalytic units. This leads to a significant signal amplification of the RCA. Using let-7a as a model analyte, the colorimetric method (best performed at 420 nm) exhibits high sensitivity toward microRNA-let-7a with a 37 fM detection limit and an analytical range that covers 3 orders of magnitude. The method was applied to the determination of let-7a in some cell lysates.
This G-triplex trimer-based rolling circle amplification (RCA) method can produce three catalytic units per RCA cycle, which can significantly improve the amplification efficiency of RCA.
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TALANTA
·
2018
·
IF 4.244
Jing Bao,Changjun Hou,Yanan Zhao,Xintong Geng,Mickey Samalo,Huisi Yang,Minghong Bian,Danqun Huo
In present study, a sensitive and effective electrochemical microRNA (miRNA) sensing platform is successfully developed by integrating gold nanoparticles/polypyrrole-reduced graphene oxide (Au/PPy-rGO), catalyzed hairpin assembly (CHA) and hybridization chain reaction (HCR) multiple signal amplification strategy. Firstly, Au/PPy–rGO was employed onto a bare GCE by electrodeposition that can greatly enhanced conductivity and effectively immobilize probes. Then, the thiolated capture probes (SH-CP) were self-assembled on the Au/PPy–rGO modified GCE via Au-S bond. The target miRNA triggered the dynamic assembly of the two hairpin substrates (H1 and H2), leading to the cyclicality of the target miRNA and the formation of H1–H2 complexes without the assistance of enzyme. Subsequently, the newly emerging DNA fragment of H2 triggered the HCR when a mixture solution (hairpins H3 and H4) and produced dsDNA polymers. Finally, a substantial amount of methylene blue (MB) as signal indicator was intercalated into the minor groove of the long dsDNA polymers to achieve detected electrochemical signal. The fabricated sensor is able to detect miRNA-16 (model target) with concentration range from 10 fM to 5 nM with a low detection limit (LOD) of 1.57 fM (S/N = 3). Current research suggests that the developed multiple signal amplification platform has a great potential for the applications in the field of biomedical research and clinical analysis.
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JOURNAL OF CELLULAR PHYSIOLOGY
·
2018
·Vol. 234
·期 7
·11440-11450
·
IF 3.923
Zheng-Yu Liu,Hong-Wei Pan,Yan Cao,Jiao Zheng,Yu Zhang,Yi Tang,Jin He,Yong-Jun Hu,Chang-Lu Wang,Qiong-Chao Zou,Qing-Hua Fu,Le Zhang,Jian-Qiang Peng,Jing Ling,Ning Peng,Jing-Jing Rong,Zhao-Fen Zheng
microRNAs (miRs) are essential in the development of heart failure. The aim of this study is to investigate the effect of microRNA-330 (miR-330) on left ventricular remodeling via the TGF-β1/Smad3 signaling pathway by targeting the sex-determining region Y (SRY) in mice with myocardial ischemia–reperfusion injury (MIRI). Differentially expressed gene (DEG) in myocardial ischemia–reperfusion (IR) was screened out and the miR that targeted the DEG was also predicted and verified. A model of MIRI was established to detect the expression of miR-330, SRY, transforming growth factor-β (TGF-β1), and Sekelsky mothers against dpp3 (Smad3). To further investigate the role of miR-330 in MIRI with the involvement of SRY and TGF-β1/Smad3 signaling pathway, the modeled mice were treated with different mimic, inhibitor, or small interfering RNA (siRNA) to observe the changes of the related gene expression, as well as the myocardial infarction size and volume of myocardial collagen. SRY was screened out and verified as a target gene of miR-330. The MIRI mice showed enlarged myocardial infarction size, increased volume of myocardial collagen, increased expression of miR-330, TGF-β1 and Smad3, while decreased the expression of SRY. The MIRI mice treated with miR-330 inhibitor showed decreased myocardial infarction size, the volume of myocardial collagen, and expression of TGF-β1 and Smad3 but promoted expression of SRY. Our findings demonstrated that downregulated miR-330 could suppress left ventricular remodeling to inhibit the activation of the TGF-β1/Smad3 signaling pathway via negatively targeting of SRY in mice with MIRI. This can be a potential target in the strategy to attenuate patient suffering.
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ACS Applied Materials & Interfaces
·
2015
·
IF 6.723
Sai Bi,Bao Xiu,Jiayan Ye,Ying Dong
Here we report a target-catalyzed DNA four-way junction (DNA-4WJ) on the basis of toehold-mediated DNA strand displacement reaction (TM-SDR), which is readily applied in enzyme-free amplified chemiluminescence resonance energy transfer (CRET) imaging of microRNA. In this system, the introduction of target microRNA-let-7a (miR-let-7a) activates a cascade of assembly steps with four DNA hairpins, followed by a disassembly step in which the target microRNA is displaced and released from DNA-4WJ to catalyze the self-assembly of additional branched junctions. As a result, G-quadruplex subunit sequences and fluorophore fluorescein amidite (FAM) are encoded in DNA-4WJ in a close proximity, stimulating a CRET process in the presence of hemin/K+ to form horseradish peroxidase (HRP)-mimicking DNAzyme that catalyzes the generation of luminol/H2O2 chemiluminescence (CL), which further transfers to FAM. The background signal is easily reduced using magnetic graphene oxide (MGO) to remove unreacted species through magnetic separation, which makes a great contribution to improve the detection sensitivity and achieves a detection limit as low as 6.9 fM microRNA-let-7a (miR-let-7a). In addition, four-input concatenated logic circuits with an automatic reset function have been successfully constructed relying on the architecture of the proposed DNA-4WJ. More importantly, DNA nanohydrogels are self-assembled using DNA-4WJs as building units after centrifugation, which are driven by liquid crystallization and dense packaging of building units. Moreover, the DNA nanohydrogels are readily functionalized by incorporating with aptamers, bioimaging agents, and drug loading sites, which thus are served as efficient nanocarriers for targeted drug delivery and cancer therapy with high loading capacity and excellent biocompatibility.
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Theranostics
·
2017
·
IF 8.766
Feifei Xu,Weixian Zhou,Jianxiang Cao,Qingqing Xu,Dechen Jiang,Yun Chen
The distorted and unique expression of microRNAs (miRNAs) in cancer makes them an attractive source of biomarker. There is much evidence indicating that a panel of miRNAs, termed “miRNA fingerprints”, is more specific and informative than an individual miRNA as biomarker. Thus, multiplex assays for simultaneous quantification of multiple miRNAs could be more potent in clinical practice. However, current available assays normally require pre-enrichment, amplification and labeling steps, and most of them are semi-quantitative or lack of multiplexing capability. In this study, we developed a quasi-targeted proteomics assay for multiplexed miRNA quantification by a combination of DNA-peptide probes and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Specifically, the signal of target miRNAs (i.e., miR-21, miR-let7a, miR-200c, miR-125a and miR-15b) was converted into the mass response of reporter peptides by hybridization of miRNAs with DNA-peptide probes and subsequent tryptic digestion to release the peptides. After a careful optimization of conditions related to binding, conjugation, hybridization and multiple reaction monitoring (MRM) detection, the assay was validated for each miRNA and the limit of quantification (LOQ) for all the miRNAs can achieve 1 pM. Moreover, crosstalk between DNA-peptide probes in multiplex assay was sophisticatedly evaluated. Using this quasi-targeted proteomics assay, the level of target miRNAs was determined in 3 human breast cell lines and 36 matched pairs of breast tissue samples. Finally, simplex assay and qRT-PCR were also performed for a comparison. This approach grafts the strategy of targeted proteomics into miRNA quantification and may offer a new way for multiplexed miRNA profiling.
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ANALYST
·
2015
·Vol. 140
·期 13
·4566-4575
·
IF 4.107
Jacky F. C. Loo,S. S. Wang,F. Peng,J. A. He,L. He,Y. C. Guo,D. Y. Gu,H. C. Kwok,S. Y. Wu,H. P. Ho,W. D. Xie,Y. H. Shao,S. K. Kong
As in all RNA viruses, influenza viruses change and mutate constantly because their RNA polymerase has no proofreading ability. This poses a serious threat to public health nowadays. In addition, traditional pathogen-based detection methods may not be able to report an infection from an unknown type or a subtype of virus if its nucleotide sequence is not known. Because of these factors, targeting host microRNA signatures may be an alternative to classify infections and distinguish types of pathogens as microRNAs are produced in humans shortly after infection. Although this approach is in its infant stage, there is an urgent need to develop a rapid reporter assay for microRNA for disease control and prevention. As a proof of concept, we report herein for the first time a non-PCR MARS (MicroRNA-RNase-SPR) assay to detect the microRNA miR-29a-3p from human subjects infected with influenza virus H1N1 by surface plasmon resonance (SPR). In our MARS assay, RNase H is employed to specifically hydrolyze the RNA probes immobilized on the gold surface where they hybridize with its cognate target cDNAs miR-29a-3p, where it was formed from reverse transcription with mature miR-29a-3p specific stem-looped primers. After the digestion of the RNA probe by RNase H, the intact cDNA was released from the RNA–DNA hybrid and bound to a new RNA probe for another enzymatic reaction cycle to amplify signals. With assay optimization, the detection limit of our MARS assay for miR-29a-3p was found to be 1 nM, and this new assay could be completed within 1 hour without thermal cycling. This non-PCR assay with high selectivity for mature microRNA provides a new platform for rapid disease diagnosis, quarantine and disease control.
研究领域:
病毒学
·
分子生物学
·
诊断学
·
公共卫生
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MICROCHEMICAL JOURNAL
·
2026
·
IF 5.2
Junyi Wu,Yao Liu,Dongwei Zhu,Jianxing Xu,Jian Yun
A DSN-triggered Y-shaped dual-circular-probe DNA walker is developed for RCI-associated miR-29a detection. • The Y-shaped steric hindrance structure effectively suppresses non-specific DSN cleavage, ensuring ultralow background. • The cascade amplification strategy achieves a detection limit as low as 0.26 fM for miR-29a. • The ICP-MS-based Rh readout provides label-free, photobleaching-free, and robust quantification. • The method exhibits excellent single-base mismatch discrimination and reliable performance in serum and cell samples. Rotator cuff injury (RCI) is a prevalent musculoskeletal disorder with limited early diagnostic options, partly due to the lack of sensitive and non-invasive biomarkers. Magnetic resonance imaging (MRI), the current mainstream imaging tool for RCI, suffers from high cost, limited accessibility, and an inability to reflect molecular-level pathological changes before structural damage becomes evident. MicroRNAs (miRNAs) have shown promise as diagnostic indicators, yet their reliable detection in clinical contexts requires methods with ultrahigh sensitivity and single-base resolution. Here, we report a duplex-specific nuclease (DSN)-triggered DNA walker with dual circular probes for highly specific and ultrasensitive analysis of RCI-associated miR-29a by inductively coupled plasma mass spectrometry (ICP-MS). The DNA walker is assembled on magnetic beads, where the walking track comprises two circular probes (c1 and c2) forming a Y-shaped steric hindrance structure that effectively suppresses non-specific DSN cleavage, ensuring an ultralow background. Upon target miR-29a hybridization, DSN specifically cleaves the DNA–RNA hybrid, initiating a cascade amplification involving self-priming, nicking/polymerase cycling, rolling circle amplification, and DNAzyme-mediated cleavage of a rhodium-labeled substrate. The released rhodium is quantified by ICP-MS, providing a photobleaching-free readout. The proposed method achieves a detection limit as low as 0.26 fM with outstanding single-base mismatch discrimination, a wide dynamic range from 1 fM to 100 pM, excellent recoveries in serum (93.2–103.2%), and high correlation with RT-PCR (R 2 = 0.9912) in real cell line samples. This DSN-triggered DNA walker offers a robust, sensitive, and specific platform for RCI-related miRNA detection, holding great promise for early diagnosis and prognosis monitoring of rotator cuff injuries. Download: Download high-res image (78KB) Download: Download full-size image
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MOLECULES
·
2023
·Vol. 28
·期 5
·2149
·
IF 4.6
Zhe Chen,Zhuoyi Wang,Yihua Yuan,Bo Liu,Jiangbo Yu,Zhiwen Wei,Keming Yun
DNA self-assembled fluorescent nanoprobes have been developed for bio-imaging owing to their high resistance to enzyme degradation and great cellular uptake capacity. In this work, we designed a new Y-shaped DNA fluorescent nanoprobe (YFNP) with aggregation-induced emission (AIE) characteristic for microRNA imaging in living cells. With the modification of the AIE dye, the constructed YFNP had a relatively low background fluorescence. However, the YFNP could emit a strong fluorescence due to the generation of microRNA-triggered AIE effect in the presence of target microRNA. Based on the proposed target-triggered emission enhancement strategy, microRNA-21 was detected sensitively and specifically with a detection limit of 122.8 pM. The designed YFNP showed higher bio-stability and cell uptake than the single-stranded DNA fluorescent probe, which has been successfully applied for microRNA imaging in living cells. More importantly, the microRNA-triggered dendrimer structure could be formed after the recognition of target microRNA, achieving a reliable microRNA imaging with a high spatiotemporal resolution. We expect that the proposed YFNP will become a promising candidate for bio-sensing and bio-imaging.Keywords:DNA fluorescent nanoprobes;Y-shaped;aggregation-induced emission;microRNA;live-cell imaging
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Bioengineered
·
2022
·
IF 6.832
Mingming Ma,Qiao Luo,Lijing Fan,Weilong Li,Qiang Li,Yu Meng,Chen Yun,Hongwei Wu,Yongping Lu,Shuang Cui,Fanna Liu,Bo Hu,Baozhang Guan,Huanhuan Liu,Shengling Huang,Wenxue Liang,Stanislao Morgera,Bernhard Krämer,Shaodong Luan,Lianghong Yin,Berthold Hocher
Acute kidney injury (AKI) is a susceptible factor for chronic kidney disease (CKD). There is still a lack of effective prevention methods in clinical practice. This study investigated the protective effect of the urinary exosomes from premature infants on cisplatin-induced acute kidney injury. Here we isolated exosomes from the fresh urine of premature infants. A C57BL/6 mice model of cisplatin-induced acute kidney injury was given 100 ug urinary exosomes 24 hours after model establishment. The kidneys were collected for pathological examination and the evaluation of renal tubular damage and apoptosis. In the in vitro experiment, human renal cortex/proximal tubular cells (HK-2) were induced by cisplatin to assess the effect of the urine exosomes from premature infants. Exosome microRNA (miRNA) sequencing technology was applied to investigate the miRNAs enriched in exosomes and the dual-luciferase gene reporter system to examine the targeting relationship of the miRNA with target genes. The results indicated that the urinary exosomes could decrease the serum creatinine level and the apoptosis of renal tubular cells, and reduce mice mortality. In addition, miR-30a-5p was the most abundant miRNA in the exosomes. It protected HK-2 cells from cisplatin-induced apoptosis by targeting and down-regulating the mitogen-activated protein kinase 8 (MAPK8). Together, our findings identified that the urinary exosomes derived from premature infants alleviated cisplatin-induced acute kidney injury and inhibited the apoptosis of HK-2 via miR-30a-5p, which could target MAPK8. These findings implied that urinary exosomes from premature infants riched in miR-30a-5p might become a potential treatment for AKI.
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MOLECULAR AND CELLULAR BIOCHEMISTRY
·
2022
·Vol. 477
·期 5
·1439-1451
·
IF 3.842
Li Jie,Chen Rongchang,Lu Yongzhen,Zeng Yuwei
This study investigated the regulatory effects of microRNA-1278 (miR-1278) on airway inflammation, airway reconstruction, and the proliferation and apoptosis of airway smooth muscle cells (ASMCs) induced by transforming growth factor β1 (TGF-β1). The results showed that miR-1278 was upregulated in the blood and lung tissues (LTs) of patients with asthma compared with that in healthy volunteers; miR-1278 expression was also upregulated in asthmatic mice, and miR-1278 inhibition improved the LTs of asthmatic mice. Moreover, miR-1278 inhibited inflammation in asthmatic mice and counteracted the effect of TGF-β1 of induced proliferation and reduced apoptosis in ASMCs. DLRA indicated that miR-1278 targeted the 3′-UTR of Src-homology 2-containing phosphatase 1 (SHP-1). Furthermore, miR-1278 promoted ASMC proliferation, in which TGF-β1 played an important role by regulating the SHP-1/STAT3 signaling pathway. In conclusion, this study showed that miR-1278 played a critical role in the processes of airway remodeling and reduction of apoptosis by targeting SHP-1.
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ACS Applied Materials & Interfaces
·
2023
·
IF 9.5
Linlin Ren,Chengfang Jiang,Yuqi Zhang,Mengmeng Li,Yan Zhang,Xinli Shi,Qi Wang,Shusheng Zhang,Xinyue Song
DNA nanomachines could initiate the cascade reaction in an autonomous mode under the drive of triggers, which achieve the signal amplification for the bioimaging of intracellular biomarkers. Compared with the “always-on” nanomachine that possibly produces false-positive signals, a controllable nanomachine with the on-site activation could be better for accurate tumor imaging and precise tumor therapy. Till now, the endogenous and exogenous triggers have been developed to design the controllable nanosensors. However, their combinations to develop feasible DNA nanomachines have been rarely studied. Herein, we constructed a near-infrared (NIR)-light-controlled DNA nanomachine that was first activated by the NIR light and then induced a target-triggered amplification process under the drive of an endogenous stimulus. Owing to adenosine-5′-triphosphate (ATP) having much higher concentration in cancer cells than that in healthy cells and the extracellular fluid, the obtained DNA nanomachine was selectively activated in cancer cells with inhibited interference signals from the surrounding healthy tissues. With obvious advantages including the exogenous NIR light initiation, the selective activation by the target microRNA, and the sensitive acceleration by the ATP-induced strand recycling reaction, the constructed nanomachine could be used to image the intracellular microRNA with increased sensitivity. Besides, after modifying the DNA sequence with the photosensitizer molecules, the obtained nanomachine could perform the selective photodynamic therapy on the tumor sections with the outstandingly decreased side effects. Thus, we hope the designed nanomachine could provide some important hints to design feasible nanomachines for accurate tumor diagnosis and precise tumor therapy.
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EXPERIMENTAL EYE RESEARCH
·
2025
·
IF 2.7
Yarong Yan,Shulei Xing,Jinghua Liu,Xinlin Yan,Yi Guan,Zhixin Jiang,Wei Zhang,Xuan Li
Lycium barbarum glycopeptide (LbGp), derived from the Chinese medicinal plant Lycium barbarum, has demonstrated anti-inflammatory properties; however, its precise role and mechanism in corneal repair following injury remain elusive. The present research investigated the mechanisms and effects of LbGp on corneal repair following alkali burn injury using in vivo mouse models of corneal alkali burn and in vitro human keratocyte fibrosis models. Corneal inflammation, opacity, and epithelial defects were assessed via a slit lamp microscope. Results showed that LbGp-treated mice exhibited reduced edema, accelerated re-epithelialization, and decreased corneal opacity compared to the phosphate-buffered saline (PBS)-treated controls. Proteomic analysis revealed altered proteins enriched in the extracellular matrix among the control, injury, and LbGp treatment groups. Moreover, LbGp significantly attenuated TGFβ-1-induced myofibroblasts transdifferentiation from keratocytes. Consistently, LbGp treatment inhibited the upregulation of fibrosis markers (αSMA, fibronectin, and collagen III) at both the protein and mRNA levels after corneal alkali burns. LbGp also effectively suppressed the activation of the NF-κB/NLRP3/IL-1β signaling pathway and neutrophil infiltration following corneal alkali burn injury. Additionally, miR-21 was upregulated in TGFβ-1-stimulated keratocytes and in the alkali-burned mouse cornea. LbGp decreased miR-21 expression, while increasing expression of its target, Smad7, thereby dampening the TGFβ/Smad2/3 signaling pathway. This research demonstrates that LbGp promotes corneal healing by inhibiting inflammation and fibrosis after alkali burns, suggesting its potential as a supplementary therapy for corneal injury repair.
研究领域:
药理学
·
眼科
·
分子生物学
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