miR-1 as a Key Epigenetic Regulator in Early Differentiation of Cardiac Sinoatrial Region.

A large diversity of epigenetic factors, such as microRNAs and histones modifications, are known to be capable of regulating gene expression without altering DNA sequence itself. In particular, miR-1 is considered the first essential microRNA in cardiac development. In this study, miR-1 potential role in early cardiac chamber differentiation was analyzed through specific signaling pathways. For this, we performed in chick embryos functional experiments by means of miR-1 microinjections into the posterior cardiac precursors-of both primitive endocardial tubes-committed to sinoatrial region fates. Subsequently, embryos were subjected to whole mount in situ hybridization, immunohistochemistry and RT-qPCR analysis. As a relevant novelty, our results revealed that miR-1 increased Amhc1, Tbx5 and Gata4, while this microRNA diminished Mef2c and Cripto expressions during early differentiation of the cardiac sinoatrial region. Furthermore, we observed in this developmental context that miR-1 upregulated CrabpII and Rarß and downregulated CrabpI, which are three crucial factors in the retinoic acid signaling pathway. Interestingly, we also noticed that miR-1 directly interacted with Hdac4 and Calm1/Calmodulin, as well as with Erk2/Mapk1, which are three key factors actively involved in Mef2c regulation. Our study shows, for the first time, a key role of miR-1 as an epigenetic regulator in the early differentiation of the cardiac sinoatrial region through orchestrating opposite actions between retinoic acid and Mef2c, fundamental to properly assign cardiac cells to their respective heart chambers. A better understanding of those molecular mechanisms modulated by miR-1 will definitely help in fields applied to therapy and cardiac regeneration and repair.

Transcription of microRNAs is regulated by developmental signaling pathways and transcription factors.

In early embryonic development, the cross-regulation of transcription factors and signaling pathways are critical in mediating developmental and physiological processes. Additionally, many studies have shown the importance of post-transcriptional regulation of signaling and network components mediated by microRNAs (miRNAs); however, how miRNAs are transcriptionally regulated is poorly understood. miRNAs are critical fine-tuners of many biological processes and their dysregulation leads to a variety of diseases and developmental defects. Previously, we have shown that miRNAs are dynamically expressed throughout sea urchin development, suggesting that miRNAs are likely to be under transcriptional regulation. Here, we used pharmacological inhibitors, genetic constructs, and loss-of-function reagents to assess the impact of key signaling pathways (Wnt, Nodal, MAPK, Sonic Hedgehog, Delta/Notch, VEGF, and BMP) and transcription factors (Alx1, Ets1/2, and Tbr) on the transcript levels of the evolutionarily conserved miR-1, miR-31, miR-92 and miR-124; the invertebrate-specific miR-71; and the echinoderm-specific miR-2002, miR-2007, and miR-2012. We also used computational methods to identify potential transcription factor binding sites of these miRNAs. Lists of binding motifs for transcription factors (TFs) were acquired from the MEME-Suite Motif Database and used as inputs for the algorithm FIMO (Find Individual Motif Occurrences), which detects short nucleotide motifs within larger sequences. Based on experimental data on miRNA expression in conjunction with bioinformatic predictions, we propose that the transcription factors Tbr, Alx1, and Ets1 regulate SpmiR-1, SpmiR-31, and SpmiR-71, respectively. We additionally observed significant effects on miRNA levels as a result of perturbations to Wnt, Nodal, MAPK, and Sonic Hedgehog signaling pathways, while no significant change on miRNA levels were observed with perturbations to Delta/Notch, VEGF, or BMP signaling pathways. Overall, this study provides insights into the transcriptional regulation of miRNAs by signaling pathways and transcription factors and contribute to our overall understanding of the genetic regulation of developmental processes.

Long noncoding RNA AI504432 upregulates FASN expression by sponging miR-1a-3p to promote lipogenesis in senescent adipocytes.

Aging affects lipid metabolism and can cause obesity as it is closely related to the disorder of many lipogenic regulatory factors. LncRNAs have been recognized as pivotal regulators across diverse biological processes, but their effects on lipogenesis in aging remain to be further studied. In this work, using RNA sequencing (RNA-Seq), we found that the expression of lncRNA AI504432 was significantly upregulated in the eWAT (epididymal white adipose tissue) of aging mice, and the knockdown of AI504432 notably reduced the expression of several adipogenic genes (e.g., Cebp/α, Srebp-1c, Fasn, Acaca, and Scd1) in senescent adipocytes. The bioinformatics investigation revealed that AI504432 possessed a binding site for miR-1a-3p, and the discovery was verified by the luciferase reporter assay. The expression of Fasn was increased upon the inhibition of miR-1a-3p but restored upon the simultaneous silencing of AI504432. Taken together, our results suggested that AI504432 controlled lipogenesis through the miR-1a-3p/Fasn signaling pathway. The findings may inspire new therapeutic approaches to target imbalanced lipid homeostasis due to aging.

Ame-miR-1-3p of bee venom reduced cell viability through the AZIN1/OAZ1-ODC1-polyamines pathway and enhanced the defense ability of honeybee (Apis mellifera L.).

Bee venom serves as an essential defensive weapon for bees and also finds application as a medicinal drug. MicroRNAs (miRNAs) serve as critical regulators and have been demonstrated to perform a variety of biological functions. However, the presence of miRNAs in bee venom needs to be confirmed. Therefore, we conducted small RNA sequencing and identified 158 known miRNAs, 15 conserved miRNAs and 4 novel miRNAs. It is noteworthy that ame-miR-1-3p, the most abundant among them, accounted for over a quarter of all miRNA reads. To validate the function of ame-miR-1-3p, we screened 28 candidate target genes using transcriptome sequencing and three target gene prediction software (miRanda, PITA and TargetScan) for ame-miR-1-3p. Subsequently, we employed real-time quantitative reverse transcription PCR (qRT-PCR), Western blot and other technologies to confirm that ame-miR-1-3p inhibits the relative expression of antizyme inhibitor 1 (AZIN1) by targeting the 3' untranslated region (UTR) of AZIN1. This, in turn, caused ODC antizyme 1 (OAZ1) to bind to ornithine decarboxylase 1 (ODC1) and mark ODC1 for proteolytic destruction. The reduction in functional ODC1 ultimately resulted in a decrease in polyamine biosynthesis. Furthermore, we determined that ame-miR-1-3p accelerates cell death through the AZIN1/OAZ1-ODC1-polyamines pathway. Our studies demonstrate that ame-miR-1-3p diminishes cell viability and it may collaborate with sPLA2 to enhance the defence capabilities of honeybees (Apis mellifera L.). Collectively, these data further elucidate the defence mechanism of bee venom and expand the potential applications of bee venom in medical treatment.

The Hansenula polymorpha mitochondrial carrier family protein Mir1 is dually localized at peroxisomes and mitochondria.

Peroxisomes are ubiquitous cell organelles involved in various metabolic pathways. In order to properly function, several cofactors, substrates and products of peroxisomal enzymes need to pass the organellar membrane. So far only a few transporter proteins have been identified. We analysed peroxisomal membrane fractions purified from the yeast Hansenula polymorpha by untargeted label-free quantitation mass spectrometry. As expected, several known peroxisome-associated proteins were enriched in the peroxisomal membrane fraction. In addition, several other proteins were enriched, including mitochondrial transport proteins. Localization studies revealed that one of them, the mitochondrial phosphate carrier Mir1, has a dual localization on mitochondria and peroxisomes. To better understand the molecular mechanisms of dual sorting, we localized Mir1 in cells lacking Pex3 or Pex19, two peroxins that play a role in targeting of peroxisomal membrane proteins. In these cells Mir1 only localized to mitochondria, indicating that Pex3 and Pex19 are required to sort Mir1 to peroxisomes. Analysis of the localization of truncated versions of Mir1 in wild-type H. polymorpha cells revealed that most of them localized to mitochondria, but only one, consisting of the transmembrane domains 3-6, was peroxisomal. Peroxisomal localization of this construct was lost in a MIR1 deletion strain, indicating that full-length Mir1 was required for the localization of the truncated protein to peroxisomes. Our data suggest that only full-length Mir1 sorts to peroxisomes, while Mir1 contains multiple regions with mitochondrial sorting information. Data are available via ProteomeXchange with identifier PXD050324.

LncRNA UCA1 promotes vasculogenic mimicry by targeting miR-1-3p in gastric cancer.

Long non-coding RNA (lncRNA) urothelial carcinoma-associated 1 (UCA1) has been implicated in several tumors. UCA1 promotes cell proliferation, migration and invasion of GC cells, but the molecular mechanism has not been fully elucidated. This study revealed the oncogenic effects of UCA1 on cell growth and invasion. Furthermore, UCA1 expression was significantly correlated with the overall survival of GC patients, and the clinicopathological indicators, including tumor size, depth of invasion, lymph node metastasis, and TNM stage. Additionally, miR-1-3p was identified as a downstream target of UCA1, which was negatively regulated by UCA1. MiR-1-3p inhibited cell proliferation and vasculogenic mimicry (VM), and induced cell apoptosis by upregulating BAX, BAD, and tumor suppressor TP53 expression levels. Moreover, miR-1-3p almost completely reversed the oncogenic effect caused by UCA1, including cell growth, migration and VM formation. This study also confirmed UCA1 promoted tumor growth in vivo. In this study, we also revealed the correlation between UCA1 and VM formation, which is potentially crucial for tumor metastasis. Meanwhile, its downstream target miR-1-3p inhibited VM formation in GC cells. In summary, these findings indicate that UCA1/miR-1-3p axis is potential target for GC treatment.

Long Non-coding RNA COX10-AS1 Promotes Glioma Progression by Competitively Binding miR-1-3p to Regulate ORC6 Expression.

Glioma is one of the most common and difficult to cure malignant primary tumors of the central nervous system. Long non-coding RNA (lncRNA) has been reported to play important functions in biological processes of many tumors, including glioma. In our study, we aimed to reveal the role and molecular mechanisms of lncRNA COX10-AS1 in regulating the progression of glioma. First of all, we showed that lncRNA COX10-AS1 was significantly increased in glioma tissues and cell lines, and high-expressed COX10-AS1 was associated with a poor prognosis in glioma patients. Moreover, through performing the functional experiments, including CCK-8, colony formation and Transwell assays, we confirmed that COX10-AS1 ablation curbed cell proliferation, migration and invasion in glioblastoma (GBM) cells. In addition, we uncovered that there existed a regulatory relationship that COX10-AS1 upregulated OCR6 by sponging miR-1-3p in GBM cells, and the following rescue assays demonstrated that both miR-1-3p downregulation and origin recognition complex subunit 6 (ORC6) overexpression rescued cell viability, migration and invasion in the COX10-AS1-deficient GBM cells. Consistently, we also verified that COX10-AS1 promoted tumorigenesis of the GBM cells in vivo through modulating the miR-1-3p/ORC6 axis. On the whole, our findings indicated a novel ceRNA pattern in which COX10-AS1 elevated OCR6 expression via sponging miR-1-3p, therefore boosting tumorigenesis in glioma, and we firstly discussed the underlying mechanisms by which the COX10-AS1/miR-1-3p/ORC6 axis affected the progression of glioma.

Identification of key miRNAs in unilateral mastication-induced disruption of cartilage homeostasis.

OBJECTIVE: The present study identified potentially pivotal miRNAs contributing to chondrogenic differentiation in temporomandibular joint suffering abnormal stress. MATERIALS AND METHODS: Sprague-Dawley rats were randomly divided into control and experimental unilateral mastication (EUM) group. Bone micro-structure parameters was detected by micro-CT, and FGF-1 and MMP-1 expression was examined by immunohistochemistry. Differentially expressed miRNAs of bilateral condyle cartilage were screened via miRNA microarray at 4- and 8-week EUM, then further verified using quantitative reverse-transcription PCR. Over-expression of five differentially expressed miRNAs in chondrocytes was triggered by transfecting miRNA mimics. The expression of MMP-13, Col-II, OPN, and Runx2 was verified by western blotting. RESULTS: Expressions of FGF-1 and MMP-1 in right condyles gradually increased from 2 to 6 weeks after EUM. A total of 20 differentially expressed miRNAs were regulated by EUM, which related to cell proliferation, invasion, and osteoblast differentiation pathways. The over-expression of miR-148a-3p and miR-1-3p led to down-regulation of Col-II, while MMP-13 and Runx2 were up-regulated by induction of hypotrophic differentiation or IL-1ß stimulation. These findings suggested that miR-148a-3p and miR-1-3p promote chondrogenic differentiation. CONCLUSIONS: Several pivotal miRNAs were found to be related to chondrogenic differentiation, which provides novel insight into pathogenic mechanisms of cartilage homeostasis.

The important role of miR-1-3p in cancers.

Cancer is a malignant tumor that seriously threatens human life and health. At present, the main treatment methods include surgical resection, chemotherapy, radiotherapy, and immunotherapy. However, the mechanism of tumor occurrence and development is complex, and it produces resistance to some traditional treatment methods, leading to treatment failure and a high mortality rate for patients. Therefore, exploring the molecular mechanisms of tumor occurrence, development, and drug resistance is a very important task. MiRNAs are a type of non-coding small RNA that regulate a series of biological effects by binding to the 3'-UTR of the target mRNA, degrading the mRNA, or inhibiting its translation. MiR-1-3p is an important member of them, which is abnormally expressed in various tumors and closely related to the occurrence and development of tumors. This article introduces miR-1-3p from multiple aspects, including its production and regulation, role in tumor occurrence and development, clinical significance, role in drug resistance, and approaches for targeting miR-1-3p. Intended to provide readers with a comprehensive understanding of the important role of miR-1-3p in tumors.

MiR-1a-3p Inhibits Apoptosis in Fluoride-exposed LS8 Cells by Targeting Map3k1.

Dental fluorosis is a common chemical disease. It is currently unclear how fluorosis occurs at the molecular level. We used miRNA-seq to look at the differences between miRNAs in the cell line of ameloblasts LS8 that had been treated with 3.2 mmol/L NaF. We also performed gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. miR-1a-3p levels were significantly lower in mouse LS8 cells treated with 3.2 mmol/L NaF, and miR-1a-3p-targeted genes were significantly enriched in the MAPK pathway. LS8 cells were divided into four groups: control, NaF, NaF+miR-1a-3p mimics, and NaF+miR-1a-3p mimics normal control groups. Cellular morphology was observed by an inverted microscope, and the proliferation activity of LS8 cells was assessed by Cell Counting Kit-8 (CCK-8). Using the real-time quantitative polymerase chain reaction (RT-qPCR), transcription levels of miR-1a-3p and Map3k1 were detected. The expressions of Bax, Bcl-2, Map3k1, p38MAPK, ERK1/2, p-p38MAPK, and p-ERK1/2 were measured by Western blot. After bioinformatics analysis, we used a luciferase reporter assay (LRA) to validate the target of miR-1a-3p, showing that miR-1a-3p could inhibit apoptosis while increasing proliferation in fluoride-exposed LS8 cells. Generally, miR-1a-3p might directly inhibit Map3k1, reduce MAPK signal pathway activation, and promote phosphorylation. Thus, our findings revealed that the interaction of miR-1a-3p with its target gene Map3k1 and MAPK signal pathway might decrease the apoptosis of LS8 cells treated with 3.2 mmol/L NaF.

miRNAs: novel noninvasive biomarkers as diagnostic and prognostic tools in neonatal sepsis.

The study aimed to assess the diagnostic and prognostic value of 3 specific microRNAs (miRNAs) in early-onset neonatal sepsis (NS). We examined miR-1, miR-124, and miR-34a in 70 NS patients upon admission and compared them to 70 healthy controls by RT-PCR. The main finding of the study was the difference in miRNA expression levels between NS patients and controls. Higher expression levels of miR-1 and miR-124 were significantly associated with NS, while miR-34a expression was reduced. Among the studied miRNAs, miR-34a exhibited the highest specificity (97%) as a confirmatory test for NS. In the multivariate model, miR-1 and miR-124 were found to be significant predictors of disease progression or mortality. Overall, the study suggests that miR-1, miR-124, and miR-34a could serve as potential biomarkers for diagnosing and predicting outcomes in early-onset NS.

Tumor-derived exosomes promote macrophages M2 polarization through miR-1-3p and regulate the progression of liver cancer.

Hepatic carcinoma is one of the most life-threatening malignancies in the world. In the clinic, it is urgent to establish a clear mechanism of hepatic carcinoma development as the basis for intervention and treatment. The purpose of this study was to explore the regulatory effect of tumor-derived exosomes on the progression of hepatocellular carcinoma.qPCR was used to detect the expression of miR-1-3p. CCk-8 and EdU staining were used to detect the proliferation and activity of hepatocellular carcinoma cells under different conditions. Transwell assay was used to detect migration and invasion of hepatocellular carcinoma cells. The morphology and size of exosomes were detected by transmission electron microscope and nanoparticle tracking analysis. Western blot was used to detect the expression of markers of exosomes. Immunofluorescence staining was used to explore the location of exosomes in hepatocellular carcinoma cells.The results showed that the expression of miR-1-3p was significantly reduced in hepatocellular carcinoma cells, and the exosomes transfected with miR-1-3p could enter macrophages and express miR-1-3p in large quantities. Macrophages polarized to M2 type under the action of miR-1-3p. Polarized M2 macrophages further down-regulated the proliferation, migration and invasion of Huh-7 cells.In summary, miR-1-3p can enter macrophages through exosomes and affect their polarization, thus affecting the growth of hepatic carcinoma cells. miR-1-3p may be a potentially effective target for regulating liver cancer progression.

Anti-inflammatory, Anti-fibrotic and Pro-cardiomyogenic Effects of Genetically Engineered Extracellular Vesicles Enriched in miR-1 and miR-199a on Human Cardiac Fibroblasts.

RATIONALE: Emerging evidence indicates that stem cell (SC)- derived extracellular vesicles (EVs) carrying bioactive miRNAs are able to repair damaged or infarcted myocardium and ameliorate adverse remodeling. Fibroblasts represent a major cell population responsible for scar formation in the damaged heart. However, the effects of EVs on cardiac fibroblast (CFs) biology and function has not been investigated. OBJECTIVE: To analyze the biological impact of stem cell-derived EVs (SC-EVs) enriched in miR-1 and miR-199a on CFs and to elucidate the underlying molecular mechanisms. METHODS AND RESULTS: Genetically engineered human induced pluripotent stem cells (hiPS) and umbilical cord-derived mesenchymal stem cells (UC-MSCs) expressing miR-1 or miR-199a were used to produce miR-EVs. Cells and EVs were thoughtfully analyzed for miRNA expression using RT-qPCR method. Both hiPS-miRs-EVs and UC-MSC-miRs-EVs effectively transferred miRNAs to recipient CFs, however, hiPS-miRs-EVs triggered cardiomyogenic gene expression in CFs more efficiently than UC-MSC-miRs-EVs. Importantly, hiPS-miR-1-EVs exhibited cytoprotective effects on CFs by reducing apoptosis, decreasing levels of pro-inflammatory cytokines (CCL2, IL-1ß, IL-8) and downregulating the expression of a pro-fibrotic gene - α-smooth muscle actin (α-SMA). Notably, we identified a novel role of miR-199a-3p delivered by hiPS-EVs to CFs, in triggering the expression of cardiomyogenic genes (NKX2.5, TNTC, MEF2C) and ion channels involved in cardiomyocyte contractility (HCN2, SCN5A, KCNJ2, KCND3). By targeting SERPINE2, miR-199a-3p may reduce pro-fibrotic properties of CFs, whereas miR-199a-5p targeted BCAM and TSPAN6, which may be implicated in downregulation of inflammation. CONCLUSIONS: hiPS-EVs carrying miR-1 and miR-199a attenuate apoptosis and pro-fibrotic and pro-inflammatory activities of CFs, and increase cardiomyogenic gene expression. These finding serve as rationale for targeting fibroblasts with novel EV-based miRNA therapies to improve heart repair after myocardial injury.

The role of miR1 and miR133a in new-onset atrial fibrillation after acute myocardial infarction.

BACKGROUND: The development of new-onset atrial fibrillation (NOAF) after acute myocardial infarction (AMI) is a clinical complication that requires a better understanding of the causative risk factors. This study aimed to explore the risk factors and the expression and function of miR-1 and miR-133a in new atrial fibrillation after AMI. METHODS: We collected clinical data from 172 patients with AMI treated with emergency percutaneous coronary intervention (PCI) between October 2021 and October 2022. Independent predictors of NOAF were determined using binary logistic univariate and multivariate regression analyses. The predictive value of NOAF was assessed using the area under the receiver operating characteristic (ROC) curve for related risk factors. In total, 172 venous blood samples were collected preoperatively and on the first day postoperatively; the expression levels of miR-1 and miR-133a were determined using the polymerase chain reaction. The clinical significance of miR-1 and miR-133a expression levels was determined by Spearman correlation analysis. RESULTS: The Glasgow prognostic score, left atrial diameter, and infarct area were significant independent risk factors for NOAF after AMI. We observed that the expression levels of miR-1 and miR-133a were significantly higher in the NOAF group than in the non-NOAF group. On postoperative day 1, strong associations were found between miR-133a expression levels and the neutrophil ratio and between miR-1 expression levels and an increased left atrial diameter. CONCLUSIONS: Our findings indicate that the mechanism of NOAF after AMI may include an inflammatory response associated with an increased miR-1-related mechanism. Conversely, miR-133a could play a protective role in this clinical condition.

Clinical value of serum miR-1-3p as a potential circulating biomarker for abdominal aortic aneurysm.

BACKGROUND: Although abdominal aortic aneurysm (AAA) is associated with life-threatening complications, there are still limited reliable biomarkers for diagnostic purpose. MicroRNAs (miRNAs) have been proposed as the potential diagnostic and risk stratification markers of AAA patients, and we aim to evaluate the serum level of miR-1-3p and its diagnostic value in AAA. METHODS: This study included 200 AAA patients and 200 controls. Demographic data and clinical information were collected from the subjects' medical records. Individual image analyses of AAA morphology were determined based on computed tomography angiography (CTA). The levels of serum miRNA expression were detected by quantitative real-time PCR. Bioinformatics tools were used to identify the target genes of miR-1-3p and their potential biological functions were further enriched. RESULTS: Serum miR-1-3p levels in the AAA group were significantly lower when compared with those in the control group in overall and subgroup comparisons. It was negatively related to WBC, CRP, maximal aneurysm diameter, area, and volume in AAA patients. Circulating miR-1-3p levels could significantly discriminate between AAA patients and healthy individuals with an area under the curve (AUC) of 0.672 (95% CI = 0.619-0.724, p < 0.001), a sensitivity of 84.5% and a specificity of 45.5%. Serum miR-1-3p was associated with a reduced risk of AAA even after adjustment for possible risk factors (OR = 0.440 per unit increase, 95% CI = 0.301-0.643, p < 0.001). And low levels of serum miR-1-3p could significantly elevate the risk of AAA in both univariate and multivariate logistic regression analyses with ORs of 4.076 and 4.136, respectively (all p < 0.001). Further GO enrichment analysis revealed that miR-1-3p was mainly involved in negative regulation of apoptotic process, sprouting angiogenesis, angiogenesis, positive regulation of blood vessel endothelial cell migration, positive regulation of cell proliferation, regulation of cell shape, etc. CONCLUSIONS: MiR-1-3p can be used as a promising circulating biomarker in the development of AAA, and it may participate in multiple biological processes to play a crucial role in AAA pathogenesis.

Review of Disease-Specific microRNAs by Strategically Bridging Genetics and Epigenetics in Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) is one of the most prevalent human malignancies and a global health concern with a poor prognosis despite some therapeutic advances, highlighting the need for a better understanding of its molecular etiology. The genomic landscape of OSCC is well-established and recent research has focused on miRNAs, which regulate gene expression and may be useful non-invasive biomarkers or therapeutic targets. A plethora of findings regarding miRNA expression have been generated, posing challenges for the interpretation and identification of disease-specific molecules. Hence, we opted to identify the most important regulatory miRNAs by bridging genetics and epigenetics, focusing on the key genes implicated in OSCC development. Based on published reports, we have developed custom panels of fifteen major oncogenes and five major tumor suppressor genes. Following a miRNA/target gene interaction analysis and a comprehensive study of the literature, we selected the miRNA molecules which target the majority of these panels that have been reported to be downregulated or upregulated in OSCC, respectively. As a result, miR-34a-5p, miR-155-5p, miR-124-3p, miR-1-3p, and miR-16-5p appeared to be the most OSCC-specific. Their expression patterns, verified targets, and the signaling pathways affected by their dysregulation in OSCC are thoroughly discussed.

MALAT1/ mir-1-3p mediated BRF2 expression promotes HCC progression via inhibiting the LKB1/AMPK signaling pathway.

BACKGROUND: The long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been reported to play a vital role in the occurrence and development of various tumors. However, the underlying mechanism of MALAT1 in hepatocellular carcinoma (HCC) has not been thoroughly elucidated. METHODS: The expression levels of MALAT1 in HCC tissues and different cell lines were detected by qRT-PCR. Antisense oligonucleotides (ASO)-MALAT1 transfected cells were used to explore the biological effects of MALAT1 in HCC cells by cell counting kit 8 (CCK-8), colony formation, transwell, wound healing, and flow cytometry analysis. Western blotting was performed to measure AMPK and apoptosis-related protein levels. Dual-luciferase reporter assay was performed to verify the relationship between MALAT1 and its specific targets. RESULTS: We found that MALAT1 was upregulated in HCC, and MALAT1 knockdown in HCC cells inhibited cell proliferation, migration, and invasion and inhibited apoptosis in vitro. Further studies demonstrated that MALAT1 positively regulated the expression of transcription factor II B­related factor 2 (BRF2), which was associated with tumor recurrence, large tumor size, and poor prognosis in HCC. Mechanistically, MALAT1 was found to act as a competitive endogenous RNA to sponge has-miR-1-3p, which upregulated BRF2 expression. Knockdown of BRF2 inhibited the progression of HCC by activating the LKB1/AMPK signaling pathway. Overexpression of BRF2 reversed the inhibitory effect of MALAT1 knockdown on HCC cell viability. Moreover, ASO targeting MALAT1 inhibited the growth of xenograft tumors. CONCLUSIONS: Our results demonstrate a novel MALAT1/miR-1-3p/BRF2/LKB1/AMPK regulatory axis in HCC, which may provide new molecular therapeutic targets for HCC in the future.

Cripto Is Targeted by miR-1a-3p in a Mouse Model of Heart Development.

During cardiac differentiation, numerous factors contribute to the development of the heart. Understanding the molecular mechanisms underlying cardiac development will help combat cardiovascular disorders, among the leading causes of morbidity and mortality worldwide. Among the main mechanisms, we indeed find Cripto. Cripto is found in both the syncytiotrophoblast of ampullary pregnancies and the inner cell mass along the primitive streak as the second epithelial-mesenchymal transformation event occurs to form the mesoderm and the developing myocardium. At the same time, it is now known that cardiac signaling pathways are intimately intertwined with the expression of myomiRNAs, including miR-1. This miR-1 is one of the muscle-specific miRs; aberrant expression of miR-1 plays an essential role in cardiac diseases. Given this scenario, our study aimed to evaluate the inverse correlation between Cripto and miR-1 during heart development. We used in vitro models of the heart, represented by embryoid bodies (EBs) and embryonic carcinoma cell lines derived from an embryo-derived teratocarcinoma in mice (P19 cells), respectively. First, through a luciferase assay, we demonstrated that Cripto is a target of miR-1. Following this result, we observed that as the days of differentiation increased, the Cripto gene expression decreased, while the level of miR-1 increased; furthermore, after silencing miR-1 in P19 cells, there was an increase in Cripto expression. Moreover, inducing damage with a cobra cardiotoxin (CTX) in post-differentiation cells, we noted a decreased miR-1 expression and increased Cripto. Finally, in mouse cardiac biopsies, we observed by monitoring gene expression the distribution of Cripto and miR-1 in the right and left ventricles. These results allowed us to detect an inverse correlation between miR-1 and Cripto that could represent a new pharmacological target for identifying new therapies.

Cysticercus pisiformis-derived novel-miR1 targets TLR2 to inhibit the immune response in rabbits.

Cysticercosis pisiformis, a highly prevalent parasitic disease worldwide, causes significant economic losses in the rabbit breeding industry. Previous investigations have identified a novel microRNA, designated as novel-miR1, within the serum of rabbit infected with Cysticercus pisiformis. In the present study, we found that C. pisiformis-derived novel-miR1 was released into the rabbit serum via exosomes. Through computational analysis using TargetScan, miRanda, and PITA, a total of 634 target genes of novel-miR1 were predicted. To elucidate the functional role of novel-miR1, a dual-luciferase reporter assay was utilized and demonstrated that novel-miR1 targets rabbit Toll-like receptor 2 (TLR2). Rabbit peripheral blood lymphocytes (PBLCs) were transfected with novel-miR1 mimic and mimic NC, and the in vitro experiments confirmed that novel-miR1 suppressed the expression of pro-inflammatory cytokines such as TNF-α, IL-1ß, and IL-6 through the nuclear factor kappa B (NF-κB) pathway. In vivo experiments demonstrated that novel-miR1 was significantly upregulated during the 1-3 months following infection with C. pisiformis in rabbits. Notably, this upregulation coincided with a downregulation of TLR2, P65, pP65, TNF-α, IL-1ß, and IL-6 in PBLCs. Collectively, these results indicate that the novel-miR1 derived from C. pisiformis inhibited the rabbits' immune response by suppressing the NF-κB-mediated immune response. This immune modulation facilitates parasite invasion, survival, and establishment of a persistent infection.

miR-1-3p Inhibits Osteosarcoma Cell Proliferation and Cell Cycle Progression While Promoting Cell Apoptosis by Targeting CDK14 to Inactivate Wnt/Beta-Catenin Signaling.

Osteosarcoma (OS) is a common bone malignancy and is diagnosed frequently in children and young adults. According to previous RNA sequencing, miR-1-3p is downregulated in OS clinical samples. Nevertheless, the functions of miR-1-3p in OS cell process and the related mechanism have not been revealed yet. In the current study, miR-1-3p expression in OS tissues and cells were evaluated using quantitative polymerase chain reaction. CCK-8 assays were conducted to measure OS cell viability in response to miR-1-3p overexpression. Colony forming assays and EdU staining were conducted for measurement of cell proliferation, and flow cytometry analysis was performed to determine cell apoptosis and cell cycle progression. Protein levels of apoptotic markers, beta-catenin, and Wnt downstream targets were quantified using western blotting. The binding relation between miR-1-3p and cyclin dependent kinase 14 (CDK14) was validated utilizing luciferase reporter assays. Experimental results revealed that miR-1-3p expression was decreased in OS tissues and cells. Additionally, miR-1-3p inhibited cell proliferation and cell cycle progression while enhancing OS cell apoptosis. Moreover, miR-1-3p directly targeted CDK14 and inversely regulated CDK14 expression in OS cells. Furthermore, miR-1-3p inactivated the Wnt/beta-catenin signaling. CDK14 overexpression partially rescued the inhibitory impact of miR-1-3p on OS cell growth. Overall, miR-1-3p inhibits OS cell proliferation and cell cycle progression while promoting cell apoptosis by targeting CDK14 and inactivating the Wnt/beta-catenin signaling.