Non-coding RNA Nuclear Paraspeckle Assembly Research Articles

Silencing of lncRNA NEAT1 alleviates acute myocardial infarction by suppressing miR-450–5p/ACSL4-mediated ferroptosis

Ferroptosis is principally initiated by dysregulation of iron metabolism and excessive accumulation of ROS, which exacerbates myocardial injury during acute myocardial infarction (AMI). Previous studies have indeed demonstrated the significant involvement of long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) exerts its pleiotropic effects in the pathophysiology of myocardial infarction, heart failure and atherosclerosis by modulating inflammation, apoptosis, and oxidative stress. However, whether and how NEAT1 mediates myocardial ferroptosis remain unknown. In this study, we found that NEAT1 expression was significantly elevated in hypoxic HL-1 cells and AMI mice, while silencing of NEAT1 alleviated lipid peroxidation and myocardial ferroptosis both in vitro and in vivo. Mechanistically, NEAT1 directly sponged miR-450b-5p and negatively regulated its expression. In addition, miR-450b-5p directly targeted Acyl-CoA synthase long-chain family member 4 (ACSL4). Notably, inhibition of miR-450b-5p reversed the role of NEAT1 in AMI mice. Collectively, these findings newly illustrated that NEAT1 acts as a competitive endogenous RNA (ceRNA) of miR-450–5p in AMI. Especially, silencing of NEAT1 effectively ameliorated myocardium ischemia by suppression of ferroptosis via miR-450–5p/ACSL4 pathway, which providing a brand-new therapeutic strategy for myocardial ischemia injury.

METTL3 aggravates cell damage induced by Streptococcus pneumoniae via the NEAT1/CTCF/MUC19 axis.

Disruption of the alveolar barrier can trigger acute lung injury. This study elucidated the association of methyltransferase-like 3 (METTL3) with Streptococcus pneumoniae (SP)-induced apoptosis and inflammatory injury of alveolar epithelial cells (AECs). AECs were cultured and then infected with SP. Furthermore, the expression of METTL3, interleukin (IL)-10, IL-6, tumor necrosis factor-alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1), long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1), mucin 19 (MUC19), N6-methyladenosine (m6A), and NEAT1 after m6A modification were detected by qRT-PCR, Western blot, and enzyme-linked immunosorbent, m6A quantification, and methylated RNA immunoprecipitation-qPCR analyses, respectively. Moreover, the subcellular localization of NEAT1 was analyzed by nuclear/cytosol fractionation assay, and the binding between NEAT1 and CCCTC-binding factor (CTCF) was also analyzed. The results of this investigation revealed that SP-induced apoptosis and inflammatory injury in AECs and upregulated METTL3 expression. In addition, the downregulation of METTL3 alleviated apoptosis and inflammatory injury in AECs. METTL3-mediated m6A modification increased NEAT1 and promoted its binding with CTCF to facilitate MUC19 transcription. NEAT1 or MUC19 overexpression disrupted their protective role of silencing METTL3 in AECs, thereby increasing apoptosis and inflammatory injury. In conclusion, this is the first study to suggest that METTL3 aggravates SP-induced cell damage via the NEAT1/CTCF/MUC19 axis.

Isoform balance of the long noncoding RNA NEAT1 is regulated by the RNA-binding protein QKI, governs the glioma transcriptome, and impacts cell migration

The long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) is involved in a variety of human cancers. Two overlapping NEAT1 isoforms, NEAT1_1 and NEAT1_2, are produced through mutually exclusive alternative 3' end formation. Previous studies extensively investigated NEAT1 dysregulation in tumors, but often failed to achieve distinct quantification of the two NEAT1 isoforms. Moreover, molecular mechanisms governing the biogenesis of NEAT1 isoforms and the functional impacts of their dysregulation in tumorigenesis remain poorly understood. In this study, we employed an isoform-specific quantification assay and found differential dysregulation of NEAT1 isoforms in patient-derived glioblastoma multiforme cells. We further showed usage of the NEAT1 proximal polyadenylation site (PAS) is a critical mechanism that controls glioma NEAT1 isoform production. CRISPR-Cas9-mediated PAS deletion reduced NEAT1_1 and reciprocally increased NEAT1_2, which enhanced nuclear paraspeckle formation in human glioma cells. Moreover, the utilization of the NEAT1 PAS is facilitated by the RNA-binding protein quaking (QKI), which binds to the proximal QKI recognition elements. Functionally, we identified transcriptomic changes and altered biological pathways caused by NEAT1 isoform imbalance in glioma cells, including the pathway for the regulation of cell migration. Finally, we demonstrated the forced increase of NEAT1_2 upon NEAT1 PAS deletion is responsible for driving glioma cell migration and promoting the expression of genes implicated in the regulation of cell migration. Together, our studies uncovered a novel mechanism that regulates NEAT1 isoforms and their functional impacts on the glioma transcriptome, which affects pathological pathways of glioma, represented by migration.

Long non-coding RNA NEAT1 exacerbates NLRP3-mediated pyroptosis in allergic rhinitis through regulating the PTBP1/FOXP1 cascade

BackgroundAllergic Rhinitis (AR) is a prevalent chronic non-infectious inflammation affecting the nasal mucosa. NLRP3-mediated pyroptosis of epithelial cells plays a pivotal role in AR pathogenesis. Herein, we evaluated the impact of the long non-coding RNA nuclear paraspeckle assembly transcript 1 (lncRNA NEAT1) on NLR family pyrin domain containing 3 (NLRP3)-mediated pyroptosis in AR. MethodsNasal inflammation levels in ovalbumin (OVA)-induced AR mice were assessed using HE staining, and NLRP3 expression was evaluated through immunohistochemistry. ELISA was utilized to detect OVA-specific IgE, IL-6, IL-5, and inflammatory cytokines (IL-1β, IL-18). Human nasal epithelial cells (HNEpCs) stimulated with IL4/IL13 were used to analyze the mRNA and protein levels of associated genes utilizing RT-qPCR and western blot, respectively. Cell viability and pyroptosis were assessed by CCK-8 and flow cytometry. The targeting relationship between NEAT1, PTBP1 and FOXP1 were analyzed by RIP and RNA pull down assays. FISH and IF analysis were performed to assess the co-localization of NEAT1 and PTBP1. ResultsIn both the AR mouse and cellular models, increased levels of NEAT1, PTBP1 and FOXP1 were observed. AR mice exhibited elevated inflammatory infiltration and pyroptosis, evidenced by enhanced expressions of OVA-specific IgE, IL-6, and IL-5, NLRP3, Cleaved-caspase 1, GSDMD-N, IL-1β and IL-18. Functional assays revealed that knockdown of PTBP1 or NEAT1 inhibited pyroptosis while promoting the proliferation of IL4/IL13-treated HNEpCs. Mechanistically, NEAT1 directly interacted with PTBP1, thereby maintaining FOXP1 mRNA stability. Rescue assays demonstrated that FOXP1 upregulation reversed the inhibitory effects of silencing NEAT1 or PTBP1 on IL4/IL13-stimulated pyroptosis activation in HNEpCs. ConclusionNEAT1 acts as a RNA scaffold for PTBP1, activating the PTBP1/FOXP1 signaling cascade, subsequently triggering NLRP3-mediated pyroptosis in HNEpCs, and ultimately promoting AR progression. These findings highlight some new insights into the pathogenesis of AR.

LncRNA NEAT1/miR-211/IL-10 Axis Regulates Inflammation of Peripheral Blood Mononuclear Cells in Acute Myocardial Infarction.

This study aimed to explore the expression of long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) in patients with acute myocardial infarction (AMI) and its inflammatory regulation mechanism through miR-211/interleukin 10 (IL-10) axis.A total of 75 participants were enrolled in this study: 25 healthy people in the control group, 25 patients with stable angina pectoris (SAP) in the SAP group, and 25 patients with AMI in the AMI group. Real-time qPCR was used to detect mRNA expression levels of NEAT1, miR-211, and IL-10. The interaction between miR-211, NEAT1, and IL-10 was confirmed by dual-luciferase reporter assay, and protein expression was detected using western blot.High expression of NEAT1 in peripheral blood mononuclear cells (PBMCs) of patients with AMI was negatively related to serum creatine kinase-MB (CK-MB), cardiac troponin I (cTnI), tumor necrosis factor-α (TNF-α), IL-6, and IL-1β and was positively correlated with left ventricular ejection fraction (LVEF). In THP-1 cells, miR-211 was confirmed to target and inhibit IL-10 expression. NEAT1 knockdown and miR-211-mimic markedly decreased IL-10 protein levels, whereas anti-miR-211 markedly increased IL-10 protein levels. Importantly, miR-211 level was negatively related to NEAT1 and IL-10 levels, whereas IL-10 level was positively related to the level of NEAT1 expression in PBMCs of patients with AMI.LncRNA NEAT1 was highly expressed in PBMCs of patients with AMI, and NEAT1 suppressed inflammation via miR-211/IL-10 axis in PBMCs of patients with AMI.

The association between the expression level of nuclear paraspeckle assembly transcript 1 and the survival rate of head and neck cancer patients after treatment

Background/purposeThe long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) exhibits diverse and complicated functions in cancer progression. Despite reports suggesting both tumor-suppressive and oncogenic effects in various cancers, its specific role in head and neck squamous cell carcinoma (HNSCC) remains unclear. This study aimed to investigate the association between NEAT1 expression levels and survival outcomes in HNSCC patients. Materials and methodsPaired tissue samples of tumor and non-cancerous matching tissues (NCMT) from 92 HNSCC patients were collected. NEAT1 expression was analyzed using RT-qPCR. Clinical characteristics, treatment received, and survival rates of the patients were assessed to determine the correlation with NEAT1 expression and explore its association with alcohol, betel quid, and cigarette use. Additionally, we examined the effect of arecoline on NEAT1 expression in normal human oral keratinocytes (NHOK) and fibroblasts (NHOF). ResultsThe study revealed a significant downregulation of NEAT1 expression in oral cancer tissues compared to NCMT. Meanwhile, arecoline increased NEAT1 expression in NHOK and NHOF cells. However, patients with downregulated NEAT1 expression exhibited higher overall survival rates, particularly in those who did not receive chemotherapy or radiotherapy. ConclusionNEAT1 expression levels are associated with survival outcomes in HNSCC patients, with upregulated expression indicating a worse prognosis, suggesting this lncRNA might contribute to cancer aggressiveness, especially in the absence of active treatment. These findings indicate NEAT1 may serve as a potential prognostic biomarker in HNSCC, but further research is required to elucidate its role in cancer progression and its potential as a therapeutic target.

NEAT1 regulates VSMC differentiation and calcification in as long noncoding RNA NEAT1 enhances phenotypic and osteogenic switching of vascular smooth muscle cells in atherosclerosis via scaffolding EZH2.

Atherosclerosis (AS) is a significant contributor to cardio-cerebrovascular ischemia diseases, resulting in high mortality rates worldwide. During AS, vascular smooth muscle cells (VSMCs) play a crucial role in plaque formation by undergoing phenotypic and osteogenic switching. Long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) has previously been identified as a nuclear regulator that promotes tumorigenesis and metastasis, but its role in regulating VSMCs in AS remains unclear. Our study aimed to investigate the biological functions and specific mechanisms of NEAT1 in regulating VSMCs in AS. We found that NEAT1 was upregulated in the aortas of AS mouse models and dedifferentiated primary VSMCs. Silencing NEAT1 in vitro attenuated the proliferation, migration, and osteogenic differentiation of VSMCs, while NEAT1 overexpression had the opposite effect. Furthermore, NEAT1 promoted VSMC osteogenic differentiation and vascular calcification in both in vivo and in vitro vascular calcification models. We also discovered that NEAT1 directly activates enhancer of zeste homolog 2 (EZH2), an epigenetic enzyme that suppresses the expression of senescence- and antimigration-related genes, by translocating it into the nucleus. CUT&Tag assay revealed that NEAT1 guides EZH2 to the promoters of senescence-related genes (P16, P21, and TIMP3), methylating local histones to reduce their transcription. Our findings suggest that NEAT1 functions in AS by modulating the epigenetic function of EZH2, which enhances the proliferation, migration, and osteogenic differentiation of VSMCs. This study provides new insights into the molecular mechanisms underlying the pathogenesis of AS and highlights the potential of NEAT1 as a therapeutic target of AS.NEW & NOTEWORTHY Our study demonstrates that the upregulation of long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) promotes proliferation and migration during phenotypic switching of vascular smooth muscle cells in atherosclerosis. We also provide in vivo and in vitro evidence that NEAT1 accelerates vascular calcification. Our findings identified the direct interaction between enhancer of zeste homolog 2 (EZH2) and NEAT1 during atherosclerosis. NEAT1 is necessary for EZH2 to translocate from the cytoplasm to the nucleus, where EZH2 epigenetically inhibits the expression of genes related to senescence and antimigration.

Knockdown of lncRNA NEAT1 suppresses streptococcus pneumoniae-induced ferroptosis in alveolar epithelial cells by regulating the Nrf2-GPX4 pathway

ObjectivesStreptococcus pneumoniae (SP) is a major cause of community-acquired pneumonia. Ferroptosis pitches in pneumonia. Long noncoding RNA nuclear paraspeckle assembly transcript 1 (lncRNA NEAT1) regulates ferroptosis in various cells. Therefore, this study probed the mechanism of lncRNA NEAT1 on SP-induced ferroptosis in AECs. MethodsSerum lncRNA NEAT1 level in 36 streptococcus pneumonia patients were retrospectively detected, with its correlations with inflammatory factor (TNF-α/IL-1β/IL-6) levels analyzed. Human pulmonary alveolar epithelial cells (HPAEpiC) were transfected with sh-NEAT1 and induced by SP. Cell viability was evaluated by CCK-8. Lactate dehydrogenase (LDH) activity was assessed. Iron content, and levels of TNF-α/IL-1β/IL-6/IL-10/lncRNA NEAT1/lipid peroxidation products [malondialdehyde (MDA)/glutathione (GSH)/reactive oxygen species/(ROS)]/ferroptosis-related proteins [Cyclooxgenase 2 (COX2)/recombinant solute carrier family 7 member 11 (SLC7A11)/total nuclear factor erythroid 2-related factor 2 (Nrf2)/cytoplasmic Nrf2 (C-Nrf2)/nuclear Nrf2 (N-Nrf2)/GPX4)] were determined by kit/ELISA/RT-qPCR/kits/Western blot. Nrf2 nuclear translocation was detected by immunofluorescence assay. On top of lncRNA NEAT1 knockdown, SP-induced HPAEpiC were treated with ML385. ResultsSerum lncRNA NEAT1 level was elevated in streptococcus pneumonia patients, and were positively interrelated with TNF-α/IL-1β/IL-6 levels. SP promoted cell HPAEpiC injury and inflammatory response, and up-regulated lncRNA NEAT1 level. LncRNA NEAT1 knockdown suppressed HPAEpiC injury/inflammatory response (reduced LDH activity and TNF-α/IL-1β/IL-6 levels, elevated IL-10) and suppressed ferroptosis (decreased iron/MDA/ROS contents and COX2 level, increased GSH/SLC7A11), facilitated Nrf2 nuclear translocation, and up-regulated GPX4. Nrf2-GPX4 pathway inhibition annulled NEAT1 knockdown-mediated improvement on SP-induced HPAEpiC ferroptosis/injury/inflammatory response. ConclusionsLncRNA NEAT1 knockdown suppressed SP-induced HPAEpiC ferroptosis by activating Nrf2-GPX4 pathway, thereby alleviating cell injury and inflammatory response.

NEAT1_1 confers gefitinib resistance in lung adenocarcinoma through promoting AKR1C1-mediated ferroptosis defence

Gefitinib is one of the most extensively utilized epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) for treating advanced lung adenocarcinoma (LUAD) patients harboring EGFR mutation. However, the emergence of drug resistance significantly compromised the clinical efficacy of EGFR-TKIs. Gaining further insights into the molecular mechanisms underlying gefitinib resistance holds promise for developing novel strategies to overcome the resistance and improve the prognosis in LUAD patients. Here, we identified that the inhibitory efficacy of gefitinib on EGFR-mutated LUAD cells was partially dependent on the induction of ferroptosis, and ferroptosis protection resulted in gefitinib resistance. Among the ferroptosis suppressors, aldo-keto reductase family 1 member C1 (AKR1C1) exhibited significant upregulation in gefitinib-resistant strains of LUAD cells and predicted poor progression-free survival (PFS) and overall survival (OS) of LUAD patients who received first-generation EGFR-TKI treatment. Knockdown of AKR1C1 partially reversed drug resistance by re-sensitizing the LUAD cells to gefitinib-mediated ferroptosis. The decreased expression of miR-338-3p contributed to the aberrant upregulation of AKR1C1 in gefitinib-resistant LUAD cells. Furthermore, upregulated long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1_1 (NEAT1_1) sponged miR-338-3p to neutralize its suppression on AKR1C1. Dual-luciferase reporter assay and miRNA rescue experiment confirmed the NEAT1_1/miR-338-3p/AKR1C1 axis in EGFR-mutated LUAD cells. Gain- and loss-of-function assays demonstrated that the NEAT1_1/miR-338-3p/AKR1C1 axis promoted gefitinib resistance, proliferation, migration, and invasion in LUAD cells. This study reveals the effects of NEAT1_1/miR-338-3p/AKR1C1 axis-mediated ferroptosis defence in gefitinib resistance in LUAD. Thus, targeting NEAT1_1/miR-338-3p/AKR1C1 axis might be a novel strategy for overcoming gefitinib resistance in LUAD harboring EGFR mutation.

NEAT1 repression by MED12 creates chemosensitivity in p53 wild-type breast cancer cells.

Breast cancer is often treated with chemotherapy. However, the development of chemoresistance results in treatment failure. Long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) has been shown to contribute to chemoresistance in breast cancer cells. In studying the transcriptional regulation of NEAT1 using multi-omics approaches, we showed that NEAT1 is up-regulated by 5-fluorouracil in breast cancer cells with wild-type cellular tumor antigen p53 but not in mutant-p53-expressing breast cancer cells. The regulation of NEAT1 involves mediator complex subunit 12 (MED12)-mediated repression of histone acetylation marks at the promoter region of NEAT1. Knockdown of MED12 but not coactivator-associated arginine methyltransferase 1 (CARM1) induced histone acetylation at the NEAT1 promoter, leading to elevated NEAT1 mRNAs, resulting in a chemoresistant phenotype. The MED12-dependent regulation of NEAT1 differs between wild-type and mutant p53-expressing cells. MED12 depletion led to increased expression of NEAT1 in a wild-type p53 cell line, but decreased expression in a mutant p53 cell line. Chemoresistance caused by MED12 depletion can be partially rescued by NEAT1 knockdown in p53 wild-type cells. Collectively, our study reveals a novel mechanism of chemoresistance dependent on MED12 transcriptional regulation of NEAT1 in p53 wild-type breast cancer cells.

Mitochondria-targeting nanozyme alleviating temporomandibular joint pain by inhibiting the TNFα/NF-κB/NEAT1 pathway.

Inflammatory cytokines that are secreted into the spinal trigeminal nucleus caudalis (Sp5C) may augment inflammation and cause pain associated with temporomandibular joint disorders (TMD). In a two-step process, we attached triphenylphosphonium (TPP) to the surface of a cubic liposome metal-organic framework (MOF) loaded with ruthenium (Ru) nanozyme. The design targeted mitochondria and was designated Mito-Ru MOF. This structure scavenges free radicals and reactive oxygen species (ROS) and alleviates oxidative stress. The present study aimed to investigate the effects and mechanisms by which Mito-Ru MOF ameliorates TMD pain. Intra-temporomandibular joint (TMJ) injections of complete Freund's adjuvant (CFA) induced inflammatory pain for ≥10 d in the skin areas innervated by the trigeminal nerve. Tumor necrosis factor-alpha (TNF-α), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), long non-coding RNA nuclear paraspeckle assembly transcript 1 (lncRNA NEAT1), and ROS also have been proved to be significantly upregulated in the Sp5C of TMD mice. Moreover, a single Mito-Ru MOF treatment alleviated TMD pain for 3 d and downregulated TNF-α, NF-κB, lncRNA NEAT1, and ROS. NF-κB knockdown downregulated NEAT1 in the TMD mice. Hence, Mito-Ru MOF inhibited the production of ROS and alleviated CFA-induced TMD pain via the TNF-α/NF-κB/NEAT1 pathway. Therefore, Mito-Ru MOF could effectively treat the pain related to TMD and other conditions associated with severe acute inflammatory activation.

Triptolide improves Alzheimer's disease by regulating the NF‑κB signaling pathway through the lncRNA NEAT1/microRNA 361‑3p/TRAF2 axis.

Alzheimer's disease (AD) is the most common type of dementia and is a serious social and medical problem threatening human health. The present study investigated the effect and underlying action mechanism of triptolide (Tri) on AD progression. Reverse transcription-quantitative PCR and western blotting analysis were used to determine the changes in RNA expression and levels of NF-κB signaling pathway proteins before and after lipopolysaccharide (LPS) induction. Nucleocytoplasmic separation experiments determined the intracellular localization of long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1). A dual-luciferase assay was used to analyze the binding between NEAT1 and microRNA (miRNA/miR)-361 or tumor necrosis factor receptor-associated factor 2 (TRAF2) and miR-361-3p and RNA pull-down was used to analyze the binding between NEAT1 and miR-361-3p. Cell Counting Kit-8, flow cytometry and ELISA were used to detect the effects of interaction between Tri and NEAT1/miR-361-3p/TRAF2 on cell viability, apoptosis and inflammatory factor levels, respectively. The results showed that LPS-mediated human microglial clone 3 cell line (HMC3) viability decreased and apoptosis and inflammatory factors (IL-1β, IL-6, IL-18 and TNF-α) increased. Tri inhibited LPS-mediated effects in a dose-dependent manner by downregulating NEAT1 expression. NEAT1 is highly expressed in the cytoplasm and reduces the transcription and translation of downstream TRAF2 by acting as a competitive endogenous RNA that adsorbs miR-361-3p. LPS-mediated HMC3 cell injury, inflammation and activation of NF-κB signaling were partially reversed in presence of Tri. The miR-361-3p mimic promoted the Tri effect and overexpression of (ov)-NEAT1 partially reversed the Tri-miR-361-3p combined effect. The effects of ov-NEAT1 were partially attenuated by small interfering (si)-TRAF2. Overall, Tri inhibited the LPS-induced decrease in viability, increase in apoptosis and inflammation and activation of NF-κB signaling in HMC3 cells. Tri regulation affected the NEAT1/miR-361-3p/TRAF2 axis. These findings suggested a potential therapeutic role for Tri in the clinical management of AD by modulating this molecular axis.

Long non-coding RNA nuclear paraspeckle assembly transcript 1 downregulation protects lens epithelial cells from oxidative stress-induced apoptosis by regulating the microRNA-124-3p/death-associated protein kinase 1 axis in age-related cataract.

Oxidative stress plays a significant role in cataract development. It causes the apoptosis of lens epithelial cells (LECs), resulting in lens opacification and accelerating cataract progression. Long non-coding RNAs (lncRNAs) and microRNAs have been linked to cataract development. Notably, lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) is involved in LEC apoptosis and cataract formation. However, the molecular mechanism by which NEAT1 causes age-related cataracts remains unknown. In this study, LECs (SRA01/04) were exposed to 200μM H2O2 to generate an in vitro cataract model. The apoptosis and viability of cells were determined using flow cytometry and 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assays, respectively. Additionally, western blotting and quantitative polymerase chain reaction were used to determine the miRNA and lncRNA expression levels. When LECs were treated with hydrogen peroxide, lncRNA NEAT1 expression levels were significantly upregulated, which contributed to LEC apoptosis. Notably, lncRNA NEAT1 suppressed the expression of miR-124-3p, a critical regulator of apoptosis, whereas NEAT1 inhibition increased miR-124-3p expression and alleviated apoptosis. However, this effect was reversed when miR1243p expression was inhibited. Additionally, the miR1243p mimic effectively inhibited the death-associated protein kinase 1 (DAPK1) expression and apoptosis of LECs, while the DAPK1 mimic reversed these effects. In conclusion, our findings indicate that the lncRNA NEAT1/miR-124-3p/DAPK1 signaling loop is involved in the regulation of LEC apoptosis induced by oxidative stress, which can be exploited to develop potential treatment strategies for age-related cataracts.

Long Non-Coding RNA Nuclear Paraspeckle Assembly Transcript 1: A Regulator of Trophoblast Cells in the Progression of Gestational Diabetes Mellitus

The potential function of NEAT1 in the progression of Gestational diabetes mellitus (GDM) and the molecular mechanisms are explored. NEAT1 levels in multiple types of cell lines and placental tissues collected from healthy, GDM and preeclampsia pregnancies were detected by quantitative real-time polymerase chain reaction (qRT-PCR). After intervening NEAT1 level in HTR-8/SVneo cells, proliferative, migratory and apoptotic potentials were examined by cell counting kit-8 (CCK-8), 5-Ethynyl-2′-deoxyuridine (EdU), transwell assay and flow cytometry. The regulatory effect of NEAT1 on the transcriptional activity of NSD1 was predicted by bioinformatic analysis and further confirmed by dual-luciferase reporter assay. In addition, histone modifications of NEAT1 on NSD1 transcription were examined by chromatin immunoprecipitation (ChIP). NEAT1 was upregulated in placental tissues collected from GDM patients. Overexpression of NEAT1 stimulated proliferative and migratory potentials, but inhibited apoptosis in HTR-8/SVneo cells. Knockdown of NEAT1 had the opposite outcomes. NEAT1 was able to regulate the transcriptional activity of NSD1 through histone modifications on H3K27Me3 and H3K27Cro. NEAT1 is upregulated in GDM cases, which triggers proliferative and migratory potentials in trophoblast cells mainly through regulating transcriptional activity of NSD1 via histone modification.

NEAT1 Regulates Calcium Oxalate Crystal-Induced Renal Tubular Oxidative Injury via miR-130/IRF1.

Aims: Calcium oxalate (CaOx) crystal deposition induces damage to the renal tubular epithelium, increases epithelial adhesion, and contributes to CaOx nephrocalcinosis. The long noncoding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) is thought to be involved in this process. In this study, we aimed to investigate the mechanism by which NEAT1 regulates renal tubular epithelium in response to inflammatory and oxidative injury triggered by CaOx crystals. Results: As CaOx crystals were deposited in mouse kidney tissue, the expression of NEAT1 was significantly elevated and positively correlated with interferon regulatory factor 1 (IRF1), Toll-like receptor 4 (TLR4), and NF-κB. NEAT1 targets and inhibits miR-130a-3p as a competitor to endogenous RNA. miR-130 binds to and exerts inhibitory effects on the 3'-untranslated region of IRF1. After transfected with silence-NEAT1, IRF1, TLR4, and NF-κB were also variously inhibited, and oxidative damage in renal calcinosis was subsequently attenuated. When we simultaneously inhibited NEAT1 and miR-130, renal tubular injury was exacerbated. Innovation and Conclusion: We found that the lncRNA NEAT1 can enhance IRF1 signaling through targeted repression of miR-130a-3p and activate TLR4/NF-κB pathways to promote oxidative damage during CaOx crystal deposition. This provides an explanation for the tubular epithelial damage caused by CaOx crystals and offers new ideas and drug targets for the prevention and treatment of CaOx nephrocalcinosis. Antioxid. Redox Signal. 38, 731-746.

Impaired autophagy flux by lncRNA NEAT1 is critical for inflammation factors production in human periodontal ligament stem cells with nicotine treatment.

Periodontitis is the top reason for tooth loss, and smoking significantly increases severe periodontitis risk. Defective autophagy has been reported to play a vital role in periodontitis. This study aimed to elucidate the relationship between autophagy and inflammation factors production in nicotine-treated periodontal ligament stem cells (PDLSCs) and the underlying mechanism. In this study, transmission electron microscopy, immunofluorescence, and the mCherry-GFP-LC3 plasmid were used to study autophagy flux. The gene levels of inflammation factors and long noncoding RNA nuclear paraspeckle assembly transcript 1 (lncRNA NEAT1) were detected by quantitative real-time PCR (qRT-PCR). Western blot was performed to assess the protein levels of autophagic markers and α7 nicotinic acetylcholine receptor (α7nAChR). We found that nicotine impaired autophagosome-lysosome fusion and lysosome functions to block autophagy flux, contributing to inflammatory factors production in nicotine-treated PDLSCs. Moreover, nicotine upregulated NEAT1 by activating α7nAChR. NEAT1 decreased autophagy flux by downregulating syntaxin 17 (STX17). Our data indicate that NEAT1-decreased autophagy flux is pivotal for inflammation factors production in nicotine-treated PDLSCs.

Long non-coding RNA NEAT1 promotes angiogenesis in hepatoma carcinoma via the miR-125a-5p/VEGF pathway.

The study’s purpose was to investigate the biological function of long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) in hepatoma carcinoma (HCC). HCC tissues and cells exhibited increased levels of NEAT1 and decreased levels of miR-125a-5p. Reduction in the expression of NEAT suppressed HepG2 cell proliferation and increased apoptosis. This was accompanied by suppression of the AKT/mTOR and ERK pathways, while the opposite was observed for miR-125a-5p. Angiogenesis assay results indicated that NEAT was proangiogenic. A dual-luciferase reporter assay indicated that NEAT1 was bound to miR-125a-5p and miR-125a-5p was bound to vascular endothelial growth factor (VEGF). The proangiogenic effects of NEAT and its stimulation of AKT/mTOR and ERK were reversed by miR-125a-5p. The anti-angiogenic effects of miR-125a-5p and its inhibitory effect on AKT/mTOR and ERK pathways were reversed by co-incubation with VEGF. The conclusion was that NEAT1 enhances angiogenesis in HCC by VEGF via a competing endogenous RNA (ceRNA) of miR-125a-5p that regulates AKT/mTOR and ERK pathways.

Withdrawal: "Long Non-coding RNA NEAT1 Drives the Development of Polycystic Ovary Syndrome via Sponging Multiple MicroRNAs" by Xia Sang and Yuzhen Zhang.

A line of evidences have validated that multiple microRNAs (including miR-16, miR-483 and miR-324-3p) were abnormally expressed in granulosa cells, which may be closely related with the pathogenesis of polycystic ovary syndrome (PCOS). Long-chain non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) has been proved to participate in the progression of various human diseases via regulating microRNAs, but its function in PCOS are not yet clear. In this work, RT-PCR was performed to detect NEAT1 expression in PCOS tissues, and human ovarian granulosa cell line KGN were taken as the model to construct the cell line with high expression and low expression of NEAT1. CCK-8 and flow cytometry were carried out to monitor cells proliferation and apoptosis, respectively. Besides, we performed luciferase reporter gene assay and RNA binding protein immunoprecipitation (RIP) assay to verify the interaction between NEAT1, miR-16, miR-483 and miR-324-3p. We demonstrated that NEAT1 was highly expressed in PCOS tissues and cells. Over-expressed NEAT1 promoted the proliferation and inhibited the apoptosis. Moreover, NEAT1 was negatively correlated with the expression levels of miR-16, miR-483 and miR-324-3p. Dual luciferase analysis and RIP assay confirmed that NEAT1 can specifically bind to miR-16, miR-483 and miR-324-3p, by which NEAT1 can reduce their expression. In conclusion, NEAT1 can promote the proliferation of ovarian granulosa cells and arrest apoptosis via impeding expressions of miR-16, miR-483 and miR-324-3p. Our research would shed new light on the molecular mechanism of ovarian granulosa cell disorder. This article is protected by copyright. All rights reserved.

LncRNA NEAT1 promotes hypoxia-induced inflammation and fibrosis of alveolar epithelial cells via targeting miR-29a/NFATc3 axis.

The objective of the present study was to explore the function and mechanism of long noncoding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) in pulmonary fibrosis (PF) progression. HPAEpic cells and A549 cells were exposed to hypoxic conditions to establish an in vitro model. Cell apoptosis was detected by TUNEL assay, and inflammatory cytokine levels were detected by ELISA. Gene and protein expression levels were identified by qRT-PCR and Western blot assays, respectively. The interaction among NEAT1, miR-29a, and NFATc3 was identified by dual-luciferase reporter and RNA pull-down assays. In hypoxia-treated cells, hypoxia markers (HIF-1α and HIF-2α), cytokines (TNF-α, IL-1β, and IL-6) and fibrotic markers (α-SMA, collagen I and collagen III) were significantly enhanced. Consistently, the expression levels of NEAT1 and NFATc3 were increased, but miR-29a was decreased in hypoxia-stimulated cells. Knockdown of NEAT1 significantly decreased cell apoptosis and the releases of TNF-α, IL-1β, and IL-6 as well as reduced the levels of α-SMA, collagen I, and collagen III. Moreover, NEAT1 positively regulated NFATc3 expression by directly targeting miR-29a. Functional experiments showed that the anti-apoptotic, anti-inflammatory, and anti-fibrotic effects mediated by NETA1 silencing were impeded by miR-29a inhibition or NFATc3 overexpression in hypoxia-stimulated HPAEpic and A549 cells. Collectively, these data demonstrated that NEAT1 knockdown inhibited hypoxia-induced cell apoptosis, inflammation, and fibrosis by targeting the miR-29a/NFATc3 axis in PF, suggesting that NEAT1 might be a potential therapeutic target for relieving PF progression.

Mechanism of miR-340–5p in laryngeal cancer cell proliferation and invasion through the lncRNA NEAT1/MMP11 axis

ObjectivesLaryngeal cancer (LC), with a relatively rare diagnosis, is a primary malignancy originating from laryngeal mucosa. This study investigated the mechanisms of microRNA (miR)- 340–5p in LC cell proliferation and invasion. MethodsThe expression patterns of miR-340–5p, long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1), and matrix metallopeptidase 11 (MMP11) in LC cells, tissues, and para-carcinoma tissues, and human bronchial epithelial cells (HBEC) were examined via RT-qPCR. The effects of elevating or silencing miR-340–5p on LC cell proliferation and invasion were examined. The subcellular localization of lncRNA NEAT1 was determined. The binding relations among miR-340–5p, lncRNA NEAT1, and MMP11 were verified. Functional rescue experiments were designed to verify the functions of lncRNA NEAT1 and MMP11 on LC cell proliferation and invasion. Nude-mouse tumor models were established to assess the role of miR-340–5p in LC in vivo. ResultsmiR-340–5p was under-expressed in LC, and miR-340–5p overexpression repressed LC cell proliferation and invasion. Mechanically, miR-340–5p decreased lncRNA NEAT1 stability via directly binding to lncRNA NEAT1 and thus declined lncRNA NEAT1 expression in LC cells, while lncRNA NEAT1 accelerated MMP11 transcription via binding to heat shock factor 1 (HSF1). Overexpression of lncRNA NEAT1 or MMP11 reversed the repression of miR-340–5p overexpression on LC cell proliferation and invasion. In vivo, miR-340–5p overexpression repressed the tumor growth. ConclusionmiR-340–5p overexpression reduced lncRNA NEAT1 stability via binding to lncRNA NEAT1, which declined lncRNA NEAT1 expression and reduced the binding of lncRNA NEAT1 to HSF1 to further inhibit MMP11 transcription, thus repressing LC cell proliferation and invasion.