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Diagnosis and treatment of tumor especially drug-resistant tumor remains a huge challenge, which requires intelligent nanomedicines with low toxic side effects and high efficacy. Herein, deformable smart DNA nanomachines are developed for synergistic intracellular cancer-related miRNAs imaging and chemo-gene therapy of drug-resistant tumors. The tetrahedral DNA framework (MA-TDNA) with fluorescence quenched component and five antennas is self-assembled first, and then DOX molecules are loaded on the MA-TDNAs followed by linking MUC1-aptamer and Mcl-1 siRNA to the antennas of MA-TDNA, so that the apt-MA-TDNA@DOX-siRNA (DNA nanomachines) is constructed. The DNA nanomachine can respond to two tumor-related miRNAs in vitro and in vivo, which can undergo intelligent miRNA-triggered opening of the framework, resulting in the "turn on" of the fluorescence for sensitively and specifically sensing intracellular miRNAs. Meanwhile, both miRNA-responded rapid release and pH-responded release of DOX are achieved for chemotherapy of tumor. In addition, the gene therapy of the DNA nanomachines is achieved due to the miRNA-specific capture and the RNase H triggered release of Mcl-1 siRNA. The DNA nanomachines intergrading both tumor imaging and chemo-gene therapy in single nanostructures realized efficient tumor-targeted, image-guided, and microenvironment-responsive tumor diagnosis and treatment, which provides a synergetic antitumor effect on drug-resistant tumor., (© 2024 Wiley‐VCH GmbH.)

Glaucoma, an insidious ophthalmic pathology, is typified by an aberrant surge in intraocular pressure (IOP) which culminates in the degeneration of retinal ganglion cells and optical neuropathy. The mitigation of IOP stands as the principal therapeutic strategy to forestall vision loss. The trabecular meshwork's (TM) integrity and functionality are pivotal in modulating aqueous humor egress. Despite their potential significance in glaucomatous pathophysiology, the implications of microRNAs (miRNAs) on TM functionality remain largely enigmatic. Transcriptomic sequencing was employed to delineate the miRNA expression paradigm within the limbal region of rodent glaucoma models, aiming to elucidate miRNA-mediated mechanisms within the glaucomatous milieu. Analytical scrutiny of the sequencing data disclosed 174 miRNAs with altered expression profiles, partitioned into 86 miRNAs with augmented expression and 88 with diminished expression. Notably, miRNAs such as hsa-miR-196b-5p were identified as having substantial expression discrepancies with concomitant statistical robustness, suggesting a potential contributory role in glaucomatous progression. Subsequent in vitro assays affirmed that miR-196b-5p augments the inflammatory cascade within immortalized human TM (iHTM) and glaucoma-induced human TM (GTM3) cells, concurrently attenuating cellular proliferation, motility, and cytoskeletal architecture. Additionally, miR-196b-5p implicates itself in the regulation of IOP and inflammatory processes in rodent models. At a mechanistic level, miR-196b-5p modulates its effects via the targeted repression of Nras (neuroblastoma RAS viral oncogene homolog). Collectively, these transcriptomic investigations furnish a comprehensive vista into the regulatory roles of miRNAs within the glaucomatous framework, and the identification of differentially expressed miRNAs alongside their targets could potentially illuminate novel molecular pathways implicated in glaucoma, thereby aiding in the development of innovative therapeutic avenues., 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. Published by Elsevier B.V.)

N6-methyladenosine (m 6 A) is a master driver of RNA function and implicates in the pathogenesis of renal injury. LncRNA SNHG14 is highly expressed in sepsis patients with acute kidney injury (AKI) and aggravates kidney cell dysfunction. This study aimed to explore whether demethylase FTO affect m 6 A methylation of SNHG14 in AKI injury and its underlying mechanism. The expression level of FTO was obviously downregulated in sepsis-associated AKI patients compared with normal controls. Mechanistically, FTO overexpression impeded SNHG14 expression by decreasing the stability of SNHG14 in an m6A-dependent manner in LPS-induced HK-2 cells. Additionally, FTO overexpression inhibited cell autophagy and apoptosis while promoting cell viability of LPS-induced HK-2 cells. Moreover, overexpression of FTO inhibited SNHG14 expression and autophagy in LPS-induced AKI mice. Functionally, SNHG14 acts as a competing endogenous RNA (ceRNA) via directly sponging miR-373-3p in LPS induced HK-2 cells. Additionally, miR-373-3p directly targets ATG7. Inhibition of SNHG14 suppresses NF-κB signaling pathway and production of inflammatory cytokines (TNF-α, IL-6, and IL-1β) via miR-373-3p/ATG7 in LPS-induced HK-2 cells. Furthermore, the SNHG14/miR-373-3p/ATG7 interaction network contributes to the regulatory effect of FTO on LPS-induced HK-2 cell viability, apoptosis and autophagy. These results suggested demethylase FTO suppressed the m 6 A modification of lncRNA SNHG14 and inhibits autophagy in LPS-induced AKI via regulating miR-373-3p/ATG7, which provided an important novel perspective for understanding sepsis-associated AKI and is conducive for developing new therapeutic targets and strategies., 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 The Authors. Published by Elsevier B.V. All rights reserved.)

The molecular heterogeneity of extracellular vesicles (EVs) and the co-isolation of physically similar particles, such as lipoproteins (LPs), confounds and limits the sensitivity of EV bulk biomarker characterization. Herein, we present a single-EV and particle (siEVP) protein and RNA assay ( siEVP PRA) to simultaneously detect mRNAs, miRNAs, and proteins in subpopulations of EVs and LPs. The siEVP PRA immobilizes and sorts particles via positive immunoselection onto micropatterns and focuses biomolecular signals in situ. By detecting EVPs at a single-particle resolution, the siEVP PRA outperformed the sensitivities of bulk-analysis benchmark assays for RNA and protein. To assess the specificity of RNA detection in complex biofluids, EVs from various glioma cell lines were processed with small RNA sequencing, whereby two mRNAs and two miRNAs associated with glioblastoma multiforme (GBM) were chosen for cross-validation. Despite the presence of single-EV-LP co-isolates in serum, the siEVP PRA detected GBM-associated vesicular RNA profiles in GBM patient siEVPs. The siEVP PRA effectively examines intravesicular, intervesicular, and interparticle heterogeneity with diagnostic promise., (© 2023 The Authors. Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)

Tumor-associated immune cells play a crucial role in cancer progression. Myeloid-derived suppressor cells (MDSCs), for example, are immature innate immune cells that infiltrate the tumor to exert immunosuppressive activity and protect cancer cells from the host's immune system and/or cancer-specific immunotherapies. While tumor-associated immune cells have emerged as a promising therapeutic target, efforts to counter immunosuppression within the tumor niche have been hampered by the lack of approaches that selectively target the immune cell compartment of the tumor, to effectively eliminate "tumor-protecting" immune cells and/or drive an "anti-tumor" phenotype. Here we report on a novel nanotechnology-based approach to target tumor-associated immune cells and promote "anti-tumor" responses in a murine model of breast cancer. Engineered extracellular vesicles (EVs) decorated with ICAM-1 ligands and loaded with miR-146a and Glut1 , were biosynthesized ( in vitro or in vivo ) and administered to tumor-bearing mice once a week for up to 5 weeks. The impact of this treatment modality on the immune cell compartment and tumor progression was evaluated via RT-qPCR, flow cytometry, and histology. Our results indicate that weekly administration of the engineered EVs ( i.e. , ICAM-1-decorated and loaded with miR-146a and Glut1 ) hampered tumor progression compared to ICAM-1-decorated EVs with no cargo. Flow cytometry analyses of the tumors indicated a shift in the phenotype of the immune cell population toward a more pro-inflammatory state, which appeared to have facilitated the infiltration of tumor-targeting T cells, and was associated with a reduction in tumor size and decreased metastatic burden. Altogether, our results indicate that ICAM-1-decorated EVs could be a powerful platform nanotechnology for the deployment of immune cell-targeting therapies to solid tumors.

Most cells can eliminate unstable or misfolded proteins through quality control mechanisms. In the inherited red blood cell disorder β-thalassemia, mutations in the β-globin gene (HBB) lead to a reduction in the corresponding protein and the accumulation of cytotoxic free α-globin, which causes maturation arrest and apoptosis of erythroid precursors and reductions in the lifespan of circulating red blood cells. We showed previously that excess α-globin is eliminated by Unc-51-like autophagy activating kinase 1 (ULK1)-dependent autophagy and that stimulating this pathway by systemic mammalian target of rapamycin complex 1 (mTORC1) inhibition alleviates β-thalassemia pathologies. We show here that disrupting the bicistronic microRNA gene miR-144/451 alleviates β-thalassemia by reducing mTORC1 activity and stimulating ULK1-mediated autophagy of free α-globin through 2 mechanisms. Loss of miR-451 upregulated its target messenger RNA, Cab39, which encodes a cofactor for LKB1, a serine-threonine kinase that phosphorylates and activates the central metabolic sensor adenosine monophosphate-activated protein kinase (AMPK). The resultant enhancement of LKB1 activity stimulated AMPK and its downstream effects, including repression of mTORC1 and direct activation of ULK1. In addition, loss of miR-144/451 inhibited the expression of erythroblast transferrin receptor 1, causing intracellular iron restriction, which has been shown to inhibit mTORC1, reduce free α-globin precipitates, and improve hematological indices in β-thalassemia. The beneficial effects of miR-144/451 loss in β-thalassemia were inhibited by the disruption of Cab39 or Ulk1 genes. Together, our findings link the severity of β-thalassemia to a highly expressed erythroid microRNA locus and a fundamental, metabolically regulated protein quality control pathway that is amenable to therapeutic manipulation., (© 2023 by The American Society of Hematology.)

A novel and reusable electrochemiluminescence biosensor was developed based on tetrahedral DNA (TDN) signal amplification for ultrasensitive detection of miRNA-27a. The flowered nickel-iron layered double hydroxide@AuNPs (NiFe-LDH@AuNPs) composites increase the amount of hairpin DNA fixed on the electrode. When miRNA is present, TDN-Ru(bpy) 3 2+ acts as an ECL probe, forming a stable sandwich structure with miRNA-27a and hairpin DNA through base complementation pairing, thus achieving miRNA detection. This biosensor has the characteristics of high sensitivity, excellent selectivity, and good reproducibility.

miRNAs alter significantly throughout pregnancy to support the development of the fetus. However, sensitive detection of miRNA remains a challenge. Herein, a reliable miRNA detection approach integrating self-assembly-triggered signal amplification and CRISPR-Cas12a-system cleavage-based color generation is described. The colorimetric approach contains three signal amplification processes. The first signal amplification is formed by the released miRNA in a chain extension process. The produced sequence that is similar to the target miRNA initiates the second signal recycle. Finally, CRISPR-Cas12a-based transcleavage on linker sequences induces the third signal amplification. The method exhibits high sensitivity and a low limit of detection of 254 aM, showing promising prospects in disease diagnosis.

The interconnection of microRNAs (miRNAs) and metal ions governs multiple biological processes in disease development and progression. However, developing multiplexed tools for dynamic imaging of these regulators remains a significant challenge. Herein, we report a conceptual approach for the design of an optically controlled DNA nanomachine by introducing a ternary DNAzyme-based, UV light-cleavable DNA scaffold and upconversion nanoparticle to the activatable hybrid chain reaction. We demonstrate that this nanomachine is capable of being effectively operated either in the presence of an endogenous miRNA target or the coexistence of intracellular Zn 2+ and external near-infrared light, resulting in enhanced fluorescence resonance energy transfer signals. With this design, the logic-gated imaging of endogenous miR-21 and Zn 2+ is demonstrated in living cells. More importantly, taking advantages of photoacoustic imaging modality, a combinational logic circuit (AND/OR) is constructed for the bioorthogonal cascade imaging of miR-21 and Zn 2+ in vivo , realizing dynamic monitoring of the correlation of miRNA and metal ions levels. Collectively, our results suggest that this conceptual design possesses the ability to expand the DNA nanomachine toolbox for visualizing a broad spectrum of interconnected molecules and thus provides new perspectives to improve the diagnostic and therapeutic outcomes.

Obesity is a major global health issue that contributes to the occurrence of metabolic disorders. Based on this fact, understanding the underlying mechanisms and to uncover promising therapeutic approaches for obesity have attracted intense investigation. Brown adipose tissue (BAT) can help burns excess calories. Therefore, promoting White adipose tissue (WAT) browning and BAT activation is an attractive strategy for obesity treatment. MicroRNAs (miRNAs) are small, non-coding RNAs, which are involved in regulation of adipogenic processes and metabolic functions. Evidence is accumulating that miRNAs are important regulators for both brown adipocyte differentiation and white adipocyte browning. Here we report that the expression of miR-669a-5p increases during the adipogenic differentiation of 3T3-L1 and C3H10T1/2 adipocytes. miR-669a-5p supplementation promotes adipogenic differentiation and causes browning of 3T3-L1 and C3H10T1/2 cells. Moreover, the expression of miR-669a-5p is upregulated in iWAT of mice exposed to cold. These data demonstrate that miR-669a-5p plays a role in regulating adipocyte differentiation and fat browning. Abbreviations : Acadl : long-chain acyl-Coenzyme A dehydrogenase; Acadm : medium-chain acyl-Coenzyme A dehydrogenase; Acadvl : very long-chain acyl-Coenzyme A dehydrogenase, very long chain; Aco2 : mitochondrial  aconitase 2; BAT: brown adipose tissue; Bmper : BMP-binding endothelial regulator; Cpt1-b :carnitine palmitoyltransferase 1b; Cpt2 : carnitine palmitoyltransferase 2; Crat : carnitine acetyltransferase; Cs : citrate synthase; C2MC: Chromosome 2 miRNA cluster; DMEM: Dulbecco's modified Eagle medium; eWAT: epididymal white adipose tissue; ETC: electron transport chain; FAO: fatty acid oxidation; Fabp4 :fatty acid binding protein 4; FBS: fetal bovine serum; Hadha : hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit alpha; Hadhb : hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit beta; HFD: high fat diet; Idh3a : isocitrate dehydrogenase 3 alpha; iWAT: inguinal subcutaneous white adipose tissue; Lpl : lipoprotein lipase; Mdh2 : malate dehydrogenase 2; NBCS: NewBorn Calf Serum; mt-Nd1 : mitochondrial NADH dehydrogenase 1; Ndufb8 :ubiquinone oxidoreductase subunit B8; Nrf1 : nuclear respiratory factor 1; Pgc1α : peroxisome proliferative activated receptor gamma coactivator 1 alpha; Pgc1b : peroxisome proliferative activated receptor, gamma, coactivator 1 beta; Pparγ : peroxisome proliferator activated receptor gamma; Prdm16 : PR domain containing 16; Rgs4 : regulator of G-protein signaling 4; Sdhb : succinate dehydrogenase complex, subunit B; Sdhc : succinate dehydrogenase complex, subunit C; Sdhd : succinate dehydrogenase complex, subunit D; Sh3d21 : SH3 domain containing 21; Sfmbt2 : Scm-like with four mbt domains 2; TG: triglyceride; TCA: tricarboxylic acid cycle; Tfam : transcription factor A, mitochondrial; TMRE: tetramethylrhodamine, methyl ester; Ucp1 : uncoupling protein 1; Uqcrc2 : ubiquinol cytochrome c reductase core protein 2; WAT: White adipose tissue.

Aims: Vascular smooth muscle cells are key participants in atherosclerosis. Circular RNA hsa_circ_0000345 (circ_0000345) and miR-647 are related to oxygenized low-density lipoprotein (ox-LDL)-induced arterial smooth muscle cell (ASMC) dysregulation. However, the relationship between circ_0000345 and miR-647 in ox-LDL-induced ASMC dysregulation is unclear., Methods: Relative levels of circ_0000345, miR-647, and PAP-associated domain containing 5 (PAPD5) mRNA in AS patient's serum and ox-LDL-induced ASMCs were detected via RT-qPCR. Gain-of-function experiments were utilized to analyze the effects of circ_0000345 upregulation on ox-LDL-induced cell proliferation, migration, invasion, and inflammatory response in ASMCs. The relationship between circ_0000345 or PAPD5 and miR-647 was validated by dual-luciferase reporter and RNA immunoprecipitation assays., Results: Circ_0000345 and PAPD5 were lowly expressed in AS patient's serum and ox-LDL-induced ASMCs, while miR-647 expression had an opposing trend. Mechanistically, circ_0000345 was verified as a miR-647 sponge, and miR-647 overexpression impaired the inhibitory effects of circ_0000345 upregulation on ox-LDL-induced ASMC proliferation, migration, invasion, and inflammatory response. Further experiments demonstrated that PAPD5 was a miR-647 target, and circ_0000345 adsorbed miR-647 to mediate PAPD5 expression. Also, PAPD5 inhibition relieved miR-647 silencing-mediated suppression on ox-LDL-induced ASMC proliferation, migration, invasion, and inflammatory response., Conclusions: Circ_0000345 elevated PAPD5 expression via acting as a miR-647 sponge, resulting in alleviating ox-LDL-induced ASMC dysregulation. The study highlighted the critical role of circ_0000345 in AS.

Background: The velvet antler is a complex mammalian bone organ with unique biological characteristics, such as regeneration. The rapid growth stage (RGS) is a special period in the regeneration process of velvet antler., Methods: To elucidate the functions of microRNAs (miRNAs) at the RGS of antler development in Gansu red deer ( Cervus elaphus kansuensis ), we used RNA sequencing (RNA-seq) to analyze miRNA expression profiles in cartilage tissues of deer antler tips at three different growth stages., Results: The RNA-seq results revealed 1,073 known and 204 novel miRNAs, including 1,207, 1,242, and 1,204 from 30-, 60-, and 90-d antler cartilage tissues, respectively. To identify key miRNAs controlling rapid antler growth, we predicted target genes of screened 25 differentially expressed miRNAs (DEMs) and specifically expressed miRNAs (SEMs) in 60 d and annotated their functions. The KEGG results revealed that target genes of 25 DEMs and 30 SEMs were highly classified in the "Metabolic pathways", "Pathways in cancer", "Proteoglycans in cancer" and "PI3K-Akt signaling pathway". In addition, a novel miRNA (CM008039.1_315920), highly enriched in "NF-kappa B signaling pathway", may need further study., Conclusions: The miRNAs identified in our study are potentially important in rapid antler growth. Our findings provide new insights to help elucidate the miRNA-mediated regulatory mechanisms involved during velvet antler development in C. elaphus kansuensis ., Competing Interests: The authors declare there are no competing interests., (©2022 Chen et al.)

Development of theranostic nanosystems integrating cascaded surface-enhanced Raman scattering (SERS) imaging and gene silencing therapy for accurate cancer diagnosis and treatment is still a big challenge and rarely reported. Herein, a novel Au nanoparticles (AuNPs)-based theranostic nanosystem containing AuNP-Ys and AuNP-Ds for highly sensitive and specific cancer diagnosis and treatment was proposed for cascaded SERS imaging of intracellular cancer-related miR-106a and miR-106a-triggered DNAzyme-based dual gene-silencing therapy of cancer cells. The AuNP-Ys were prepared by modifying the AuNPs with specially designed Y-motifs, and the AuNP-Ds were obtained by colabeling Raman molecules and dsDNA linkers on AuNPs. When identifying the intracellular cancer-related miRNAs, the Y-motifs and dsDNA linkers undergoes miRNA-triggered ATP-driven conformational transitions and releases the miRNA for recycling, which results in the formation of AuNP network nanostructures to generate significantly enhanced SERS signals for sensitive identification of the cancer cells as well as the amplification and specific activation of DNAzymes to catalyze the Mg 2+ -assisted cleavage of the Survivin and c-Jun mRNAs for effective dual gene-silencing therapy of cancer cells. The AuNP-based theranostic nanosystem achieves the synergism of target-triggered SERS imaging and DNAzyme-based dual gene-silencing therapy with enhanced specificity, sensitivity, and curative effect, which can be a powerful tool for accurate diagnosis and efficient treatment of cancers.

Background: Prolonged forced abstinence from morphine can increase cue-induced cravings for the drug, contributing to a persistent vulnerability to relapse. Previous studies have identified the implications of aberrant microRNA (miRNA) regulation in the pathogenesis of morphine addiction, but the changes in miRNA expression during the incubation of morphine craving are still unknown., Methods: Nucleus accumbens (NAc)-specific altered miRNA transcriptomics was determined in a mouse model of cue-induced incubation of morphine craving following a next-generation sequencing method and verified by RT-qPCR. Bioinformatics analysis was performed to predict the target gene of selected miRNA, and the protein expression of the target gene was detected by western blot. A dual-luciferase assay was performed to confirm the binding sites, and gain- and loss-of-function strategy was applied to understand the mechanism of miRNA and its target gene., Results: The miR-592-3p observed to be downregulated in the NAc core was linked to the incubation of morphine craving, and a dual-luciferase assay was performed to confirm the binding sites of miR-592-3p in its target gene, tomoregulin-1 (TMEFF1). Also, gain- and loss-of-function analyses revealed that the inhibition of miR-592-3p expression in the NAc core negatively regulated TMEFF1 expression, thereby enhancing the incubation of morphine craving; however, the overexpression of miR-592-3p in the NAc core resulted in a decreased expression of TMEFF1, thereby reducing the incubation of morphine craving., Conclusion: Our findings demonstrated that miR-592-3p can improve the incubation of morphine craving by targeting TMEFF1, and thus, it holds a therapeutic potential to inhibit opioid craving., (© The Author(s) 2022. Published by Oxford University Press on behalf of CINP.)

To sensitively detect multiple and cross-species disease-related targets from a single biological sample in a quick and reliable manner is of high importance in accurately diagnosing and monitoring diseases. Herein, a surface-enhanced Raman scattering (SERS) sensor based on a functionalized multiple-armed tetrahedral DNA nanostructure (FMTDN) immobilized silver nanorod (AgNR) array substrate and Au nanoparticle (AuNP) SERS tags is constructed to achieve both multiplex detection and enhanced sensitivity using a sandwich strategy. The sensor can achieve single, dual, and triple biomarker detections of three lung cancer-related nucleic acid and protein biomarkers, i.e. , miRNA-21, miRNA-486 and carcinoembryonic antigen (CEA) in human serum. The enhanced SERS signals in multiplex detections are due to the DNA self-assembled AuNP clusters on the silver nanorod array during the assay, and the experimentally obtained relative enhancement factor ratios, 150 for AuNP dimers and 840 for AuNP trimers, qualitatively agree with the numerically calculated local electric field enhancements. The proposed FMTDN-functionalized AgNR SERS sensor is capable of multiplex and cross-species detection of nucleic acid and protein biomarkers with improved sensitivity, which has great potential for the screening and clinical diagnosis of cancer in the early stage.

The abnormal expression of miRNA-21 is often found in tumor specimens and cell lines, and thus, its specific detection is an urgent need for the diagnosis and effective therapy of cancers. In this contribution, we demonstrate a palindrome-based hybridization chain reaction (PHCR) upon the stimuli of a short oligonucleotide trigger to perform the autonomous assembly of cross-linked network structures (CNSs) for the amplification detection of miRNA-21 and sensitive fluorescence imaging of cancerous cells. The building blocks are only two palindromic hairpin-type DNA strands that are separately modified with different fluorophores (Cy3 and Cy5), which is easily combined with the catalytic hairpin assembly (CHA) technique that can further amplify the signal output. Utilizing the CHA-PHCR assay system, a small amount of miRNA-21 can activate many triggers via CHA and in turn induce the PHCR-based CNS assembly from more DNA building blocks, bringing Cy3 and Cy5 into close proximity to each other and generating ultrasensitive fluorescence resonance energy transfer signals. As a result, target miRNA can be quantitatively detected down to as low as 10 pM with high assay specificity. The coexisting nontarget miRNAs and other biomacromolecules do not interfere with signal transduction. The developed assay system is suitable for screening different expression levels of miRNA-21 in living cells by fluorescence imaging. The palindrome-based cross-linking assembly can enhance the intracellular stability of assembled nanostructures by at least fivefold and exhibit the good universality for the detection of other miRNAs. Moreover, cancerous cells can be distinguished from healthy cells, and the CHA-PHCR assay is in good accordance with the gold standard PCR method, indicating a promising platform for the diagnosis of human cancers and other genetic diseases.

Our previous experiments have confirmed that human milk oligosaccharides (HMO) and its main component 2'-fucosyllactose (2'-FL), as prebiotics, could effectively alleviate cow milk allergy by regulating the intestinal microecology. This study intended to further explore the molecular mechanism of HMO regulating intestinal immunity. The results of the allergic mouse model showed that oral administration of 2'-FL or HMO reduced β-lactoglobulin (β-LG)-induced serum-specific IgE secretion and mast cell degranulation, while reducing the inflammatory cytokines, TNF-α, IL-4, and IL-6 production and promoting the miR-146a expression. In vitro results further confirmed that 2'-FL or HMO treatment reduced allergen-induced secretion of iNOS, NO, pro-inflammatory cytokines and reactive oxygen species in RAW264.7 cells. At the same time, in contrast to the β-LG group, 2'-FL dose-dependently inhibited the TLR4/NF-κB inflammatory pathway and upregulated miR-146a expression, and the effect of the 2'-FL mid-dose group was similar to that of the HMO intervention group. In particular, adding miR-146a inhibitors to macrophages attenuated the inhibitory effect of 2'-FL on the expression of TRAF6 and IRAKI in the TLR4 pathway, suggesting that miR-146a might be involved in the immune regulation of 2'-FL. The above results indicated that 2'-FL had a similar effect to HMOs, and its effect of reducing β-LG allergy might be related to the regulation of miR-146a to inhibit TLR4/NF-κB signaling pathway., (Copyright © 2021 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Our previous study has confirmed that Lactobacillus acidophilus KLDS 1.0738 (La KLDS 1.0738) could alleviate β-lactoglobulin (β-Lg)-induced allergic inflammation. This study further explored its molecular regulation mechanism through an in vitro macrophage model. β-Lg-induced macrophages were treated with strains of viable or non-viable L. acidophilus and Toll-like receptor 4 (TLR4) inhibitor or miR-146a inhibitor. Our results revealed that β-Lg stimulation led to the increased expression of TLR4/NF-κB signal pathway in macrophages. Similar to TLR4 inhibitor treatment, La KLDS 1.0738 interventions significantly reduced the allergic inflammation by inhibition of TLR4 pathway, which was superior to the commercial L. acidophilus GMNL-185 strains (La GMNL-185) or the control, especially in living L. acidophilus-treated group. Furthermore, La KLDS 1.0738 strains could remarkably reduce transduction of TLR4 and inflammatory cytokine production, which was closely associated with up-regulation of miR-146a levels. MiR-146a inhibitor attenuated the alleviative effect of La KLDS 1.0738, indicating miR-146a might be a crucial mediator of L. acidophilus strains to reduce β-Lg-induced inflammation in macrophages through TLR4 pathway. In conclusion, these observations highlighted that probiotics might regulate host miRNA levels to down-regulate TLR4/NF-κB-dependent inflammation. PRACTICAL APPLICATIONS: Cow's milk allergy (CMA) is one of the most common diseases of food allergy, which has a high prevalence in infants and young children. La KLDS 1.0738 has been shown to be effective in alleviating β-Lg-induced allergic inflammation. Our study further found that treatment with La KLDS 1.0738 could suppress the TLR4/NF-κB signaling pathway via modulating miR-146a expression, thereby reducing the overexpression of downstream inflammatory factors. This study not only elucidates the specific pathway of La KLDS 1.0738 to relieve allergic inflammation, but also provides a new insight into the molecular mechanism for the remission and treatment of CMA by probiotics., (© 2021 Wiley Periodicals LLC.)

A dynamically regulated microenvironment, which is mediated by crosstalk between adipocytes and neighboring cells, is critical for adipose tissue homeostasis and function. However, information on key molecules and/or signaling pathways regulating the crosstalk remains limited. In this study, we identify adipocyte miRNA-182-5p (miR-182-5p) as a crucial antiobesity molecule that stimulated beige fat thermogenesis by promoting the crosstalk between adipocytes and macrophages. miR-182-5p was highly enriched in thermogenic adipocytes, and its expression was markedly stimulated by cold exposure in mice. In contrast, miR-182-5p expression was significantly reduced in adipose tissues of obese humans and mice. Knockout of miR-185-5p decreased cold-induced beige fat thermogenesis whereas overexpression of miR-185-5p increased beiging and thermogenesis in mice. Mechanistically, miR-182-5p promoted FGF21 expression and secretion in adipocytes by suppressing nuclear receptor subfamily 1 group D member 1 (Nr1d1) at 5'-UTR, which in turn stimulates acetylcholine synthesis and release in macrophages. Increased acetylcholine expression activated the nicotine acetylcholine receptor in adipocytes, which stimulated PKA signaling and consequent thermogenic gene expression. Our study reveals a key role of the miR-182-5p/FGF21/acetylcholine/acetylcholine receptor axis that mediates the crosstalk between adipocytes and macrophages to promote beige fat thermogenesis. Activation of the miR-182-5p-induced signaling pathway in adipose tissue may be an effective approach to ameliorate obesity and associated metabolic diseases.

Parkinson's disease (PD) can lead to movement injury and cognitive dysfunction. Although advances have been made in attenuating PD, the effect of inhibiting the development of PD remains disappointing. Therefore, the present study aimed at investigating the etiology of Parkinson's disease and developing an alternative therapeutic strategy for patients with PD. A PD mouse model was established using an intraperitoneal injection of 1‑methyl‑4‑phenyl‑1,2,3,6‑tetrahydropyridine hydrochloride (MPTP‑HCl; 30 mg/kg/day for 5 days), and a PD cellular model was established by treating SH‑SY5Y cells with different concentrations of 1‑methyl‑4‑phenylpyridinium (MPP + ) for 24 h. The expression levels of circular RNA sterile α motif domain containing 4A (circSAMD4A) and microRNA (miR)‑29c‑3p in both midbrain tissues and SH‑SY5Y cells were detected via reverse transcription‑quantitative PCR. The interaction between circSAMD4A and miR‑29c‑3p was verified using a dual‑luciferase reporter experiment. Apoptosis‑, autophagy‑ and 5'AMP‑activated protein kinase (AMPK)/mTOR cascade‑associated proteins in midbrain tissues and SH‑SY5Y cells were detected using western blotting. Furthermore, TUNEL staining and flow cytometry were used to analyze cell apoptosis. It was found that circSAMD4A was upregulated, while miR‑29c‑3p was downregulated in both PD animal and cellular models. Moreover, circSAMD4A directly targeted and negatively regulated miR‑29c‑3p. Further studies identified that circSAMD4A knockdown inhibited MPTP‑ or MPP + ‑induced apoptosis and autophagy; however, these effects were abolished by an miR‑29c‑3p inhibitor. In addition, circSAMD4A knockdown repressed phosphorylated‑AMPK expression and increased mTOR expression in MPTP‑ or MPP + ‑induced PD models, the effects of which were reversed by a miR‑29c‑3p inhibitor. Collectively, these results suggested that circSAMD4A participated in the apoptosis and autophagy of dopaminergic neurons by modulating the AMPK/mTOR cascade via miR‑29c‑3p in PD.

Emerging evidence has shown that Long noncoding RNAs (LncRNAs) are aberrantly expressed and functionally involved in the development of neurodegenerative disorders. In this work, we investigated the regulatory effects of lncRNA of LINC01311 and its competing endogenous RNA target of hsa-miR-146a-5p in a cellular model of Alzheimer's disease (AD). SH-SY5Y cells were treated with synthetic Βeta-Amyloid Peptide (1-42) (AB1-42) in vitro to induce AD-like neural injuries. Expressions of LINC01311 and hsa-miR-146a-5p were monitored by qRT-PCR. LINC01311 was upregulated and hsa-miR-146a-5p downregulated to examine their functional regulations on AB1-42-induced apoptosis, proliferation slowdown, autophagy, and amyloid precursor protein (APP) accumulations. Hsa-miR-146a-5p was also overexpressed in LINC01311-upregulated SH-SY5Y cells to examine their correlated regulations on AB1-42-induced neural injuries. LINC01311 was downregulated whereas hsa-miR-146a-5p upregulated in AB1-42 treated SH-SY5Y cells. LINC01311 upregulation and hsa-miR-146a-5p downregulation protected AB1-42-induced apoptosis, proliferation slowdown, autophagy, and APP accumulations in SH-SY5Y cells. Hsa-miR-146a-5p overexpression reversed the protection of LINC01311 on AB1-42-induced neural injuries. Our work demonstrated that the epigenetic axis of LINC01311/hsa-miR-146a-5p was involved in the functional regulation of human-lineage neurons in a cellular model of AD, thus suggesting a clinical potential of exploring epigenetic network for treating AD patients., (© 2021 International Union of Biochemistry and Molecular Biology.)

MicroRNAs (miRNAs) emerge as important regulators for myocardial infarction (MI). However, the function of miR-708-3p during MI is unclear. H9c2 cells were cultured in a hypoxic environment and Sprague-Dawley rats experienced surgical ligation of the left anterior descending coronary artery to establish MI models. qPCR was used to measure the expression level of miR-708-3p and ADAM17 mRNA. ELISA was used to detect inflammatory cytokines TNF-α, IL-6, and IL-1β, and myocardial injury markers LDH, CK-MB, and cTnI. Cell apoptosis and viability were monitored by flow cytometry analysis and MTT assay. ADAM17 expression was detected by Western blot. Dual-luciferase reporter gene experiments were carried out to identify binding sites between miR-708-3p and ADAM17 3'UTR. In vivo, left ventricle functions and myocardial remodeling of the rats were measured by echocardiography. MiR-708-3p was found to be significantly decreased in H9c2 cells after hypoxia induction and in heart tissues of rats with MI or serum samples of patients with MI, while ADAM17 was upregulated. Overexpression of miR-708-3p inhibited inflammation and injury of H9c2 cells cultured in hypoxia and the heart of the rats with MI. ADAM17 was verified as a direct target of miR-708-3p and restoration of ADAM17 reversed the effects of miR-708-3p. MiR-708-3p alleviated the inflammation and injury of cardiomyocytes via targeting ADAM17.

MicroRNA-155 (miRNA-155) as a characteristic myeloma-associated biomarker exhibits significant potential application in the diagnosis of multiple myeloma (MM). In this paper, a novel type of molecular beacon (MB)-functionalized monolayer MoS₂ nanosheet probe was proposed as fluorescent probe for high-sensitive assays of miRNA-155that uses a duplexspecificnuclease (DSN) enzyme to amplify the fluorescence signal. The preparation and detection conditions of the fluorescent probes were optimized in some aspects, such as the concentration of MoS₂ (0.80 μ M) and DSN (0.2 U), and the incubation time of DSN (30 min). The probesexhibited a sensitive fluorescence response to miRNA-155 and the fluorescence signal of the assay was significantly amplified by the cleavage of DSN. The relationship between F/F 0 and logC miRNA follows a linear calibration curve, and the limit of detection (LOD) of miRNA-155 in 10% human serum is calculated to be 10.96 fM based on this relationship. The good performance and fluorescence amplification effect of the fluorescent probe were confirmed by studying the recovery of miRNA-155 in 10% human serum, which was ranged from 98.32% to 106.3% with a relative standard deviation of less than 4.14%. Besides, the high expression of miRNA-155 in clinic blood of MM patients was sensitively distinguished from healthy peoples by using the proposed probes. The proposed novel fluorescent probe based on the DSN can be used to detect miRNA-155 in human serum and provide a potential, convenient and reliable tool for diagnosis of MM.

Acute myeloid leukemia (AML) is a hematological malignancy with a low cure rate, especially in the elderly. Previous studies have shown that long non-coding RNA (lncRNA) may be an important factor in the pathogenesis of hematological malignancies, including acute myeloid leukemia (AML). However, the biological roles and clinical significances of most lncRNAs in AML are not fully understood. LncRNA CD27 Antisense RNA 1 (CD27-AS1), as a member of lncRNA family, has rare reports on its function. In present study, we found that the expression of CD27-AS1 examined by quantitative real-time PCR was markedly increased in the AML patients (N = 40) compared with healthy volunteers (N = 40). The overall survival time was significantly shorter in patients with higher CD27-AS1 expression than that in patients with lower CD27-AS1 (P < 0.01). Furthermore, downregulation of CD27-AS1 in AML cells suppressed proliferative ability, arrested cell cycle in G0/G1 phase, and induced apoptosis. However, CD27-AS1 overexpression further enhanced the malignant phenotype of AML cells. Additionally, CD27-AS1 was proved to increase PBX3 expression through sponging miR-224-5p. CD27-AS1 knockdown blocked the MAPK signaling through PBX3 silencing and further inhibited the cell growth of AML cells. Taken together, we demonstrate that CD27-AS1 may be a potential prognostic biomarker of AML, and our finding also provides a new insight for non-coding RNA-based therapeutic intervention of AML.

Background: Alzheimer's disease (AD) is a common neurodegenerative disease, which is characterized by aberrant accumulation of amyloid-β (Aβ) and neuroinflammation. The purpose of this study was to explore the regulatory effects of long non-coding RNA (lncRNA) MAGI2-AS3 and microRNA-374b-5p (miR-374b-5p) on Aβ-induced neurotoxicity and neuroinflammation, as well as the relationship between MAGI2-AS3 and miR-374b-5p in AD patients., Methods: A luciferase reporter assay was used to analyze the interaction between MAGI2-AS3 and miR-374b-5p and between miR-374b-5p and beta-site amyloid precursor protein cleaving enzyme 1 (BACE1). SH-SY5Y and BV2 cells treated with Aβ25-35 were used to mimic neuronal injury and neuroinflammation in AD pathogenesis. Cell viability was evaluated using a MTT assay, and pro-inflammatory cytokine levels were measured using ELISA kits. MAGI2-AS3 and miR-374b-5p expression was examined using quantitative real-time PCR., Results: BACE1 served as a target gene of miR-374b-5p, and MAGI2-AS3 could sponge miR-374b-5p. The expression of MAGI2-AS3 was increased, and miR-374b-5p was decreased in both SH-SY5Y and BV2 cells exposed to Aβ25-35. MAGI2-AS3 reduction enhanced neuronal viability and attenuated neuroinflammation in AD cell models, and miR-374b-5p overexpression led to same effects, but miR-374b-5p inhibition reversed these effects. Serum MAGI2-AS3 and miR-374b-5p levels in AD patients were negatively correlated and correlated with disease severity., Conclusion: The findings indicated that the MAGI2-AS3/miR-374b-5p axis regulates Aβ-induced neurotoxicity in SH-SY5Y cells and neuroinflammation in BV2 cells. The MAGI2-AS3/miR-374b-5p axis may provide novel biomarkers and therapeutic targets for AD., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Diabetes mellitus (DM) has been one of the largest health concerns of the 21st century due to the serious complications associated with the disease. Therefore, it is essential to investigate the pathogenesis of DM and develop novel strategies to reduce the burden of diabetic complications. Sirtuin 1 (SIRT1), a nicotinamide adenosine dinucleotide (NAD + )-dependent deacetylase, has been reported to not only deacetylate histones to modulate chromatin function but also deacetylate numerous transcription factors to regulate the expression of target genes, both positively and negatively. SIRT1 also plays a crucial role in regulating histone and DNA methylation through the recruitment of other nuclear enzymes to the chromatin. Furthermore, SIRT1 has been verified as a direct target of many microRNAs (miRNAs). Recently, numerous studies have explored the key roles of SIRT1 and other related epigenetic mechanisms in diabetic complications. Thus, this review aims to present a summary of the rapidly growing field of epigenetic regulatory mechanisms, as well as the epigenetic influence of SIRT1 on the development and progression of diabetic complications, including cardiomyopathy, nephropathy, and retinopathy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The handling editor declared a shared affiliation with several of the authors JW(a), JW(b), MX, YG, YT, and JG at time of review., (Copyright © 2021 Wang(a), Wang, Wang(b), Xiao, Guo, Tang, Zhang and Gu.)

Background and Objective: Angiogenesis is the most important repair process of tissues subjected to ischemic injury. The present study aims to investigate whether the pro-angiogenic effect of Qiliqiangxin prescription (QL) is mediated through miR-21 signaling., Methods: Cardiac microvascular endothelial cells (CMECs) were isolated and cultured from 2-3 weeks old SD rats by the method of planting myocardium tissues. The purity was identified by CD31 immunofluorescence staining. CMECs were then cultured under 1% O2 hypoxia or normoxia condition for 24h in the presence or absence of QL pretreatment (QL, 0.5mg/ml, 24h). The mimics and inhibitors of miR-21 were transfected into CMECs. miR-21, HIF-1α, and VEGF expressions of CMECs were then detected by qRT-PCR and/or Western blot. The proliferation, migration, and tube formation functions of CMECs were assessed using the BrdU assay, wound healing test, and tube formation assay, respectively., Results: The results showed that compared with the control group, hypoxia significantly upregulated the expression of miR-21 and impaired CMECs proliferation, migration, and tube formation functions. Compared with the hypoxia group, QL further upregulated miR-21, HIF-1α, and VEGF expressions, and improved cell proliferation, migration, and tube formation of hypoxic CMECs. These effects of QL were abolished by a knockdown of miR-21. Conversely, treatment with miR-21 mimics further enhanced QL induced changes in hypoxic CMECs., Conclusion: Results indicate that the pro-angiogenesis effects of QL on hypoxic CMECs are mediated by activating miR-21 and its downstream HIF-1α/VEGF pathway possibly., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The proliferation and migration of vascular smooth muscle cells (VSMCs) are one of the key regulatory links of atherosclerosis (AS). Long noncoding RNAs (lncRNAs) are emerging as key regulators in AS development. In this study, we first assessed the expression level of smooth muscle and endothelial cell-enriched migration/differentiation-associated lncRNA (SENCR) in the plasma of patients with coronary heart disease (CHD) and its predictive and diagnostic value. Second, we investigated the role of SENCR in the regulation network of human aortic-VSMCs (HA-VSMCs) proliferation and migration and determined its downstream regulatory mechanism. The results showed that SENCR was downregulated in the peripheral blood of CHD, and negatively related to the Gensini score. SENCR was enriched in HA-VSMCs and mainly distributed in cytoplasm. Overexpression of SENCR significantly inhibited HA-VSMCs proliferation, migration, and block cell cycle, while the knockdown of SENCR had the opposite effects. Moreover, bioinformatics analysis and luciferase reporter assay demonstrated that miR-4731-5p could directly bind to SENCR. Besides, we proved that FOXO3a inhibited HA-VSMCs proliferation and migration by binding to the 3'-untranslated region of miR-4731-5p. In summary, our research suggested that SENCR affects HA-VSMCs proliferation and migration via regulating the miR-4731-5p/FOXO3a pathway., (© 2020 Wiley Periodicals LLC.)

Background: Numerous microRNAs (miRNAs) have been investigated in the progression of Alzheimer's disease (AD). The purpose of this study was to analyze the expression of miR-433 and its diagnostic value in patients with AD, and to explore the neuroprotective effect of miR-433 in amyloid β (Aβ)-treated SH-SY5Y and SK-N-SH cells., Methods: AD patients and AD cell model that established by Aβ treatment were used in this study. Quantitative real-time PCR was used to measure the expression of miR-433. The diagnostic value of miR-433 was evaluated using the receiver operating characteristic analysis. MTT assay was used to examine the viability of Aβ-treated SH-SY5Y and SK-N-SH cells. Bioinformatics and luciferase activity analyses were used to confirm the target gene that might be involved in the mechanisms of miR-433 in AD., Results: Expression levels of miR-433 were decreased in AD patients and cells compared with the corresponding controls. The decreased miR-433 expression levels in serum and cerebrospinal fluid (CS) were positively correlated with MMSE scores and had relatively high diagnostic accuracy in AD patients. The gain-of-function experiments found that the overexpression of miR-433 could rescue the Aβ-induced inhibition in neuronal viability in SH-SY5Y and SK-N-SH cells. The luciferase activity results showed that JAK2 was a target gene of miR-433 in neuronal cells., Conclusion: All the data of this study showed that miR-433 serves as a candidate diagnostic biomarker for AD patients, and may have the potential as a novel therapeutic target by ameliorating Aβ-induced neurotoxicity., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: Aerobic glycolysis is a unique tumor cell phenotype considered as one of the hallmarks of cancer. Aerobic glycolysis can accelerate tumor development by increasing glucose uptake and lactate production. In the present study, lactate dehydrogenase A (LDHA) is significantly increased within glioma tissue samples and cells, further confirming the oncogenic role of LDHA within glioma., Methods: Hematoxylin and eosin (H&E) and immunohistochemical (IHC) staining were applied for histopathological examination. The protein levels of LDHA, transporter isoform 1 (GLUT1), hexokinase 2 (HK2), phosphofructokinase (PFK) in target cells were detected by Immunoblotting. The predicted miR-9 binding to lncRNA Annexin A2 Pseudogene 2 (ANXA2P2) or the 3' untranslated region (UTR) of LDHA was verified using Luciferase reporter assay. Cell viability or apoptosis were examined by MTT assay or Flow cytometry. Intracellular glucose and Lactate levels were measured using glucose assay kit and lactate colorimetric assay kit., Results: The expression of ANXA2P2 showed to be dramatically upregulated within glioma tissue samples and cells. Knocking down ANXA2P2 within glioma cells significantly inhibited cell proliferation and aerobic glycolysis, as manifested as decreased lactate and increased glucose in culture medium, and downregulated protein levels of glycolysis markers, GLUT1, HK2, PFK, as well as LDHA. miR-9 was predicted to target both lncRNA ANXA2P2 and LDHA. The overexpression of miR-9 suppressed the cell proliferation and aerobic glycolysis of glioma cells. Notably, miR-9 could directly bind to LDHA 3'UTR to inhibit LDHA expression and decrease the protein levels of LDHA. ANXA2P2 competitively targeted miR-9, therefore counteracting miR-9-mediated repression on LDHA. Within tissues, miR-9 exhibited a negative correlation with ANXA2P2 and LDHA, respectively, whereas ANXA2P2 and LDHA exhibited a positive correlation with each other., Conclusions: In conclusion, ANXA2P2/miR-9/LDHA axis modulates the aerobic glycolysis progression in glioma cells, therefore affecting glioma cell proliferation., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The long noncoding RNA T cell receptor gamma locus antisense RNA 1 (TRG-AS1) plays an important role in glioblastoma progression. The objective of this study was to determine the expression status of TRG-AS1 in tongue squamous cell carcinoma (TSCC). The regulatory effects of TRG-AS1 depletion on the malignant processes of TSCC cells were illustrated both in vitro and in vivo . Additionally, the precise molecular mechanisms through which TRG-AS promotes TSCC oncogenicity were investigated. TRG-AS1 expression in TSCC tissues and cell lines was detected using reverse transcription-quantitative PCR. Functional experiments including Cell Counting Kit-8 assay, flow cytometric apoptotic assay, migration and invasion assays, and xenograft tumor model analysis were conducted to severally determine the effects of TRG-AS1 on TSCC cell proliferation, apoptosis, migration, and invasion in vitro and tumor growth in vivo . Herein, TRG-AS1 was highly expressed in TSCC and closely associated with advanced TNM stage, high lymph node metastasis, and poor overall survival. Functionally, TRG-AS1 depletion suppressed TSCC cell proliferation, migration, and invasion in vitro ; promoted cell apoptosis; and attenuated tumor growth in vivo . Mechanistically, TRG-AS1 served as a molecular sponge for microRNA-543 (miR-543), thereby contributing to the increased expression of Yes-associated protein 1 (YAP1) - a miR-543 target. Rescue experiments confirmed that miR-543 inhibition or YAP1 overexpression abrogated the anticancer effects of TRG-AS1 silencing in TSCC cells. In conclusion, TRG-AS1 aggravates TSCC malignancy by regulating the miR-543/YAP1 axis. Identification of the TRG-AS1/miR-543/YAP1 regulatory pathway may provide novel insights into TSCC diagnosis, prognosis, and therapy.

MicroRNAs regulate and control the growth and development of cells and can play the role of oncogenes and tumor suppressor genes, which are involved in the occurrence and development of cancers. In this study, DNA fragments obtained by target-induced rolling-circle amplification were constructed to complement with self-cleaving deoxyribozyme (DNAzyme) and release fluorescence biomolecules. This sensing approach can affect multiple signal amplification permitting fluorescence detection of microRNAs at the pmol L -1 level hence affording a simple, highly sensitive, and selective low cost detection platform.

Shikonin is an anti-inflammatory agent extracted from natural herbs. The aim of this study is to explain the treatment effects and mechanism of Shikonin in acute lung injury induced by sepsis. In this study, first, we evaluate different Shikonin concentrations for the anti-inflammation of acute lung injury induced by sepsis in an in vivo study. On the basis of the results, we confirm that 50.0 mg/kg was the best therapeutic Shikonin concentration. As a second step, we discuss the mechanism of Shikonin by a vitro cell experiment. Finaly, we validate that Shikonin has effective treatment effects on acute lung injury via regulation of microRNA-140-5p/toll-like receptor 4 (miRNA-140-5p/TLR4) in the in vivo study. The results of vitro and vivo study showed that Shikonin could improve acute lung injury induced by sepsis. The mechanism might be correlation miRNA-140-5p expression increasing, and regulated targeted gene TLR4, with TLR4 expression depressing, the downstream myeloid differentiation protein 88 and nuclear factor κB proteins expression were suppressed. In conclusion, Shikonin improved sepsis induced lung injury by regulation miRNA-140-5p/TLR4., (© 2019 Wiley Periodicals, Inc.)

Circular RNAs (circRNAs) are endogenous non-coding RNAs that form a covalently closed continuous loop, are widely distributed and play important roles in a series of developmental processes. In plants, an increasing number of studies have found that circRNAs can regulate plant metabolism and are involved in plant responses to biotic or abiotic stress. Acting as miRNA decoys is a critical way for circRNAs to perform their functions. Therefore, we developed GreenCircRNA-a database for plant circRNAs acting as miRNA decoys that is dedicated to providing a plant-based platform for detailed exploration of plant circRNAs and their potential decoy functions. This database includes over 210 000 circRNAs from 69 species of plants; the main data sources of circRNAs in this database are NCBI, EMBL-EBI and Phytozome. To investigate the function of circRNAs as competitive endogenous RNAs, the possibility of circRNAs from 38 plants to act as miRNA decoys was predicted. Moreover, we provide basic information for the circRNAs in the database, including their locations, host genes and relative expression levels, as well as full-length sequences, host gene GO (Gene Ontology) numbers and circRNA visualization. GreenCircRNA is the first database for the prediction of circRNAs that act as miRNA decoys and contains the largest number of plant species. Database URL: http://greencirc.cn., (© The Author(s) 2020. Published by Oxford University Press.)

Background: In recent years, accumulating studies have found that circular RNA (circRNA) exerts a great effect on tumor progression. Circ&#95;0000215, a novel circRNA, remains largely unknown in terms of its effect and mechanism in glioma., Method: Quantitative real-time polymerase chain reaction (qRT-PCR) was carried out to detect the expressions of circ&#95;0000215, miR-495-3p and CXCR2 in human glial cell line HEB and glioma cell lines (A172, U251, U87, SHG-44, LN-18), human glioma tissues and adjacent healthy tissues. Gain- and loss-assays of circ&#95;0000215 were conducted. Cell proliferation ability was detected via the CCK8 assay, and cell invasion ability was examined by Transwell assay. CXCR2 expression was evaluated via RT-PCR and Western blot. Moreover, bioinformatics was applied to analyze the targeting molecules of circ&#95;0000215 and CXCR2. Verification of the relationship between these molecules were supported through the dual-luciferase reporter gene and RNA immunocoprecipitation (RIP) assay., Results: Circ&#95;0000215 and CXCR2 were remarkably upregulated in glioma tissues and cells. Overexpression of circ&#95;0000215 notably promoted the proliferation, invasion and epithelial-mesenchymal transition (EMT) but inhibited apoptosis of glioma cells, while knocking down circ&#95;0000215 had the opposite effects. Additionally, miR-495-3p, a sponge RNA of circ&#95;0000215, inhibited the growth, invasion and EMT of glioma cells. Mechanistically, miR-495-3p targeted CXCR2 and negatively regulated CXCR2/PI3K/Akt pathway. However, the effects of miR-495-3p were all dampened by overexpression of circ&#95;0000215., Conclusion: These data demonstrated that circ&#95;0000215 functions as a competitive endogenous RNA by sponging miR-495-3p, thus accelerating glioma progression through CXCR2 axis.

Evidence has shown that hypoxia promotes esophageal squamous cell carcinoma (ESCC) growth and metastasis, but the molecular mechanisms underlying that response remain poorly understood. MicroRNAs (miRNAs) are post-transcriptional regulators that participate in various cancer-related processes. Here, we demonstrated that hypoxia along with hypoxia-inducible factor 1α significantly increased expression of miR-10b-3p. Inhibition of miR-10b-3p weakened the effects of hypoxia on ESCC cell proliferation, migration and invasion, while miR-10b-3p overexpression had the opposite effects. Mechanistically, miR-10b-3p acted as cancer-promoting gene by targeting testis specific 10. Using a xenograft model, we observed that administration of miR-10b-3p agomir to tumors enhanced their growth and metastasis in vivo. These findings verified the potent regulatory role played by hypoxia-induced miR-10b-3p expression in ESCC progression. These results suggest that miR-10b-3p may be a useful therapeutic target for treating ESCC.

Diabetic retinopathy (DR) is a leading cause of new-onset blindness. Recent studies showed that protecting retinal ganglion cells (RGCs) from high glucose-induced injury is a promising strategy for delaying DR. This study is to investigate the role of miR-145-5p in high glucose-induced RGC injury. Here, RGCs were randomly divided into low glucose and high glucose groups. PCR assay showed miR-145-5p was significantly upregulated in high glucose group. Transfection of miR-145-5p inhibitor decreased pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) levels, elevated cell viability and proliferation, as well as suppressed cell apoptosis by ELISA, MTT, EdU proliferation, colony formation and flow cytometry assays, respectively. Moreover, dual-luciferase reporter assay confirmed FGF5 as a target gene of miR-145-5p. FGF5 knockdown could partially reverse the protective effects of miR-145-5p on RGC-5 cells. In conclusion, our results demonstrated that inhibition of miR-145-5p might be a neuroprotective target for diabetes mellitus-related DR. Abbreviations: DR: diabetic retinopathy; RGCs: retinal ganglion cells; miR-145-5p: microRNA-145-5p; TNF-α: tumor necrosis factor-α; IL-6: interleukin-6; FGF: fibroblast growth factor; ATCC: American Type Culture Collection; WT: wild type; MUT: mutant type.

Oxaliplatin is a third-generation derivative of platinum that is effective in the treatment of multiple solid tumors. However, it can cause peripheral neuropathic pain, and the molecular mechanisms of this effect remain unknown. We induced a model of peripheral neuropathic pain in rats by intraperitoneally injecting them with oxaliplatin twice a week for 4.5 weeks. We found that both the mRNA and protein expression levels of Na v 1.6 (encoded by the gene Scn8a) increased while the miR-30b-5p (shorthand for miR-30b) expression decreased in the dorsal root ganglion (DRG) of treated rats. Using TargetScan and miRanda predictive software, we discovered that Scn8a was a major target of miR-30b. Moreover, we found that miR-30b negatively regulated Scn8a by binding to the Scn8a 3'UTR in PC12 cells. In addition, Na v 1.6 and miR-30b were colocalized in the DRG neurons of naive rats. Overexpression of miR-30b using an miR-30b agomir attenuated neuropathic pain induced by oxaliplatin and inhibited both the mRNA and protein expression levels of Na v 1.6 both in vitro and in vivo. Conversely, the inhibition of miR-30b with an miR-30b antagomir resulted in neuropathic pain and an increase in the expression of Na v 1.6. More importantly, overexpression of miR-30b inhibited the proliferation of LS-174t cells (Colorectal cancer cells). These data suggest that miR-30b contributes to oxaliplatin-induced chronic neuropathic pain through Na v 1.6 downregulation and could be a novel therapeutic target for the treatment of oxaliplatin-induced neuropathic pain as a side effect of chemotherapy in cancer patients., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

In general, it is extremely problematic to discriminate between monozygotic twins (MZTs), who share the same genomic DNA sequence, using traditional DNA-based identification methods such as short tandem repeat profiling. MicroRNAs (miRNAs) have shown potential in forensic applications owing to their low molecular weight, abundant and tissue-specific expression. In this study, we utilized massively parallel sequencing technology to perform genome-wide profiling of miRNAs in the blood from four pairs of healthy MZTs. On average, 158 miRNAs were detected in each individual and 14% of which were differentially expressed within each pair of MZTs. The miRNAs with the most significant differences in expression between the twins were confirmed using real-time polymerase chain reaction. Our results demonstrated that miRNAs have potential for use as molecular markers in MZTs discrimination., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Long noncoding BRAF-activated noncoding RNA has been reported to be tightly associated with tumorigenesis and development in various types of cancers. However, the expression, biological function, and modulatory mechanism of BRAF-activated noncoding RNA in pancreatic cancer remained unclear. In the present work, we explored the carcinogenic activity and underlying mechanism of BRAF-activated noncoding RNA on pancreatic cancer in vitro . We identified that BRAF-activated noncoding RNA was upregulated in pancreatic cancer tissues and cell lines, and BRAF-activated noncoding RNA was related to tumor metastasis and stage. BRAF-activated noncoding RNA reinforces proliferation, invasion, and migration in PANC-1 and SW1990 cells. Moreover, miR-195-5p was downregulated in both PC tissues and cell lines. Our results based on luciferase reporter, RIP-Ago2 and qRT-PCR assays, showed that miR-195-5p was a direct target of BRAF-activated noncoding RNA. Furthermore, miR-195-5p inhibitor abrogated the effects of short-interfering BRAF-activated noncoding RNA on PANC-1 and SW1990 cell growth and invasion in vitro . We further identified that BRAF-activated noncoding RNA played a vital role in activating the Wnt/β-catenin pathway by sponging miR-195-5p. Collectively, our study showed that BRAF-activated noncoding RNA promotes pancreatic cancer tumorigenesis through miR-195-5p/Wnt/β-catenin axis may serve as a potential target for diagnostics and therapeutics in pancreatic cancer.

The sodium channel 1.7 (Nav1.7), which is encoded by SCN9A gene, is involved in neuropathic pain. As crucial regulators of gene expression, many miRNAs have already gained importance in neuropathic pain, including miR-182, which is predicted to regulate the SCN9A gene. Nav1.7 expression in L4-L6 dorsal root ganglions (DRGs) can be up regulated by spared nerve injury (SNI), while miR-182 expression was down regulated following SNI model. Exploring the connection between Nav1.7 and miR-182 may facilitate the development of a better-targeted therapy. In the current study, direct pairing of miR-182 with the SCN9A gene was verified using a luciferase assay in vitro. Over-expression of miR-182 via microinjection of miR-182 agomir reversed the abnormal increase of Nav1.7 at both mRNA and protein level in L4-6 DRGs of SNI rats, and significantly attenuated the hypersensitivity to mechanical stimulus in the rats. In contrast, administration of miR-182 antagomir enhanced the Nav1.7 expression at both mRNA and protein level in L4-6 DRGs, companied with the generation of mechanical hypersensitivity in naïve rats. Collectively, we concluded that miR-182 can alleviate SNI- induced neuropathic pain through regulating Nav1.7 in rats.

The present study aimed to explore the effects and underlying mechanisms of microRNA (miR)‑23a on pancreatic cancer (PC) cells progression. Reverse transcription‑quantitative polymerase chain reaction and western blot analysis were used to detect the mRNA and protein miR‑23a and PLK‑1 level. Cell viability, cell cycle, migration and invasion assasy, and in vivo tumorigenicity assay were used to investigate the effects of miR‑204. Further luciferase reporter assay was used to explore the mechanisms contributing to miR‑204 effects. It was observed that miR‑23a expression was upregulated and negatively associated with polo‑like kinase‑1 (PLK‑1) expression in human PC tissues. PLK‑1 was a direct target of miR‑23a in PC cells. Functional analysis demonstrated that miR‑23a overexpression suppressed cell proliferation, inhibited cell migration and invasion and promoted cell apoptosis in vitro. When PC cells were transfected with has‑miR‑23a PLK‑1 was downregulated and its downstream molecules were deregulated, including decreased expression of B‑cell lymphoma‑2, cyclin B1 and vimentin, and increased expression of Bax and E‑cadherin. The inhibitory effect of miR‑23a on PC cell progression was observed in vivo tumor xenografts. The results of the study suggest that miR‑23a inhibits PC cell progression by directly targeting PLK‑1‑associated signaling and promoting miR‑23a expression may be a potential method for treating patients with PC.

MicroRNAs (miRNAs/miRs) serve a role as important regulators in cardiac hypertrophy. The present study aimed to reveal the differential expression profile of miRNAs between young and aging spontaneously hypertensive rats (SHRs) and studied the functional annotation of predicted targets. Briefly, 3‑month‑old and 12‑month‑old SHRs (n=3/group) were subjected to echocardiography, histopathological analysis and dihydroethidium staining. Subsequently, small RNA sequencing and data processing was conducted to identify the differentially expressed miRNAs between these two groups. Eight significantly upregulated miRNAs were validated by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR), followed by in silico target gene prediction. Functional annotation analysis of the predicted targets was performed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. As a result, significantly impaired left ventricular diastolic function was detected in the 12‑month‑old SHRs, alongside increased myocyte cross‑sectional area and percentage area of fibrosis, elevated reactive oxygen species production and reduced microvessel density (P<0.05). Compared with the 3‑month‑old SHRs, 21 miRNAs were significantly upregulated and five miRNAs were downregulated in 12‑month‑old rats (P<0.05). Eight upregulated, remodeling‑associated miRNAs, including rno‑miR‑132‑3p, rno‑miR‑182, rno‑miR‑208b‑3p, rno‑miR‑212‑3p, rno‑miR‑214‑3p, rno‑miR‑218a‑5p, rno‑miR‑221‑3p and rno‑miR‑222‑3p, underwent bioinformatics analysis. The target genes were significantly enriched in 688 GO terms and 39 KEGG pathways, including regulation of peptidyl‑tyrosine phosphorylation, regulation of protein serine/threonine kinase activity, adrenergic signaling in cardiomyocytes, ErbB signaling pathway, mTOR signaling pathway, FoxO signaling pathway, Ras signaling pathway, insulin secretion, adipocytokine signaling pathway, HIF‑1 signaling pathway, Rap1 signaling pathway, VEGF signaling pathway and TNF signaling pathway. Collectively, the present study identified a dysregulated miRNA profile in aging SHRs, which targeted numerous signaling pathways associated with cardiac hypertrophy, autophagy, insulin metabolism, angiogenesis and inflammatory response.

Colorectal cancer (CRC) is the most frequently diagnosed malignancy of the gastrointestinal tract. The dysregulation of microRNAs (miRNAs/miRs) has been reported in the majority of types of human cancer, and is correlated with tumorigenesis and tumor development. Abnormal expression of miR‑663 has been observed in various types of human cancer. However, little is known about its role in CRC. Therefore, the aim of the present study was to clarify the expression and potential role of miR‑663, and its underlying molecular mechanism in CRC. It was observed that miR‑663 was markedly downregulated in CRC tissues and cell lines. Decreased miR‑663 expression levels in CRC tissues were correlated with tumor, node, metastasis stage and lymph node metastasis. Functional assays revealed that upregulation of miR‑663 inhibited cell proliferation and invasion in CRC. Further molecular mechanism assays demonstrated the fascin (FSCN1) was a target gene of miR‑663. In addition, FSCN1 was increased and negatively correlated with miR‑663 expression in CRC tissues. FSCN1 underexpression mimicked the tumor suppressive functions induced by miR‑663 overexpression on CRC cell proliferation and invasion. Collectively, the present study presented evidence that miR‑663 may act as a tumor suppressor in CRC by directly targeting FSCN1, which may lead to a potential therapeutic strategy focusing on miR‑663 and FSCN1 for patients with this disease.

BACKGROUND The objective of this study was to identify the pathway responsible for ventricular remodeling. MATERIAL AND METHODS We collected remodeling myocardium tissue (n=18) and control myocardium tissue (n=22), and detected the expression of 4 miRNAs in these 2 groups using real-time PCR. We then searched the miRNA database online to find the candidate genes of miR-93. Real-time PCR and Western blot analysis were used to confirm the regulatory relationship. RESULTS We found that only miR-93 was decreased in remodeling myocardium tissue, and validated CCND1 to be the direct target gene of miR-93, with the "seed sequence" located within the 3'-UTR of the target gene via luciferase reporter assay system. Furthermore, we established the negative regulatory relationship between miR-93 and CCND1 by determining the relative luciferase activity of cells transfected with wild-type or mutant 3'-UTR of CCND1. We also found that The CCND1 protein and mRNA expression level of HL-1 cells treated with 50 nM miR-93 mimics were apparently lower than the scramble control, and those of the cells treated with 100 nM miR-93 mimics and CCND1 siRNA (100 nM) were even lower than those in the 50 nM treatment group. Meanwhile, cells transfected with miR-93 mimics (50 nM) showed evidently downregulated viability when compared with the scramble controls, while cells transfected with (100 nM) and CCND1 siRNA (100nM) showed even lower viability. CONCLUSIONS We showed that CCND1 is a direct target of miR-93, and the dysregulation of the miR-93/CCND1 signaling pathway is responsible for the development of ventricular remodeling.

Rapeseed (Brassica napus) is an important cash crop considered as the third largest oil crop worldwide. Rapeseed oil contains various saturation or unsaturation fatty acids, these fatty acids, whose could incorporation with TAG form into lipids stored in seeds play various roles in the metabolic activity. The different fatty acids in B. napus seeds determine oil quality, define if the oil is edible or must be used as industrial material. miRNAs are kind of non-coding sRNAs that could regulate gene expressions through post-transcriptional modification to their target transcripts playing important roles in plant metabolic activities. We employed high-throughput sequencing to identify the miRNAs and their target transcripts involved in fatty acids and lipids metabolism in different development of B. napus seeds. As a result, we identified 826 miRNA sequences, including 523 conserved and 303 newly miRNAs. From the degradome sequencing, we found 589 mRNA could be targeted by 236 miRNAs, it includes 49 novel miRNAs and 187 conserved miRNAs. The miRNA-target couple suggests that bna-5p-163957&#95;18, bna-5p-396192&#95;7, miR9563a-p3, miR9563b-p5, miR838-p3, miR156e-p3, miR159c and miR1134 could target PDP, LACS9, MFPA, ADSL1, ACO32, C0401, GDL73, PlCD6, OLEO3 and WSD1. These target transcripts are involving in acetyl-CoA generate and carbon chain desaturase, regulating the levels of very long chain fatty acids, β-oxidation and lipids transport and metabolism process. At the same, we employed the q-PCR to valid the expression of miRNAs and their target transcripts that involve in fatty acid and lipid metabolism, the result suggested that the miRNA and their transcript expression are negative correlation, which in accord with the expression of miRNA and its target transcript. The study findings suggest that the identified miRNA may play important role in the fatty acids and lipids metabolism in seeds of B. napus., (Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.)

Hematopoietic stem cells (HSCs), which are localized in the bone marrow of adult mammals, come from hematopoietic endothelium during embryonic stages. Although the basic processes of HSC generation and differentiation have been described in the past, the epigenetic regulation of embryonic hematopoiesis remains to be fully described. Here, by utilizing an in vitro differentiation system of mouse embryonic stem cells (ESCs), we identified more than 20 microRNAs that were highly enriched in embryonic hematopoietic cells, including some (e.g. miR-10b, miR-15b, and miR-27a) with previously unknown functions in blood formation. Luciferase and gene expression assays further revealed combinational binding and regulation of these microRNAs by key transcription factors in blood cells. Finally, bioinformatics and functional analyses supported an interactive regulatory control between transcription factors and microRNAs in hematopoiesis.

"Metabolic memory" is identified as a phenomenon that previous exposure to hyperglycemia results in the long-lasting deleterious effects on cardiovascular events. More and more researches show that epigenetics plays an important role in the pathogenesis of metabolic memory. It remains unclear whether microRNA (miRNA) dysfunctions participate in the event. In this study, the miRNA arrays on human aortic endothelial cells were adopted to seek the miRNAs that may be involved in the metabolic memory and were verified in vivo and in vitro. Sixteen miRNAs were found differentially expressed. Among these miRNAs, the expressions of miR-125b, miR-146a-5p, and miR-29a-3p were associated with persistent impaired endothelial function and altered proinflammatory gene expressions, including nuclear factor-κB (NF-κB) subunit p65. Direct inhibition of miR-125b expression or increased miR-146a-5p expression blunted NF-κB signals and improved the endothelial function. Luciferase reporter assays confirmed the biochemical relationship for miR-125b targeting on TNF-α-induced protein 3 and miR-146a-5p targeting on TNF receptor-associated factor 6 and IL-1 receptor-associated kinase 1 during the activation of NF-κB pathway. Thus, our findings demonstrate that glucose induced changes in miR-125b and miR-146a-5p are related to the long-lasting activation of NF-κB pathway and contribute to follow-up metabolic memory.

Purpose: Ketamine is widely used in pediatric anesthesia. Recent studies have demonstrated that excessive application of ketamine leads to cortical neurodegeneration in neonatal brains. The present study aims to characterize the functional role of neuronal microRNA, miR-124, in regulating ketamine-induced neurotoxicity in mouse hippocampus., Methods: Real-time quantitative PCR (RT-PCR) was used to examine the effect of high-dosage ketamine on the expression of miR-124 in murine hippocampus in vitro. Downregulation of hippocampal miR-124 was achieved by lentivirual transfection, and its effects on protecting ketamine-induced hippocampal neurodegeneration were examined both in vitro and in vivo., Results: Hippocampal miR-124 was upregulated by ketamine treatment. Knocking down miR-124 in vitro reduced ketamine-induced apoptosis in hippocampal CA1 neurons, upregulated AMPA receptors phosphorylation and activated the protein kinase C/extracellular signal-regulated kinases (PKC/ERK) pathway. In the in vivo Morris water maze test, following ketamine-induced hippocampal neurodegeneration, mice subjected to hippocampal miR-124 inhibition showed improved memory performance., Conclusions: Our study demonstrated that miR-124 played an important role in regulating ketamine-induced hippocampal neurodegeneration. Inhibiting miR-124 may provide a molecular target to improve memory performance in both human and animals suffering from overanesthetizing-related neurotoxicity.

Expression of the transmembrane-4-L-six-family-1 (TM4SF1) is high in human pancreatic cancer cells, but the underlying mechanism remains unclear. In this study, we aimed to identify and characterize microRNAs that regulate TM4SF1 expression in PC cells. Western blot analysis and quantitative polymerase chain reaction were used to detect TM4SF1 and hsa-miR-141 levels in four PC cell lines. SW1990 and BxPc-3 cells were transfected with the inhibitor miR-141, the inhibitor negative control, the miR-141 mimic and the mimic negative control; and cell invasion, migration, proliferation, cell cycle progression and apoptosis were detected by Transwell, MTT and flow cytometry assays, respectively. The miR-141 levels negatively correlated with the TM4SF1 protein levels in PC cells. The TM4SF1 protein levels were lower in the 141M group but higher in the 141I group, although the TM4SF1 mRNA levels had no significant changes, compared to the negative controls. Luciferase assays demonstrated that hsa-miR-141 directly targeted the 3'-untranslated region of the TM4SF1 gene. In addition, miR-141 downregulated TM4SF1 expression to inhibit invasion and migration of PC cells but had no effects on cell proliferation, cell cycle progression or apoptosis. TM4SF1 is a direct target of miR-141. Our findings that TM4SF1 expression was inhibited by miR-141 provide new insights into the oncogenic mechanism of TM4SF1 and suggest that miR-141 represents a novel molecular target for PC therapy.