Nuclear Paraspeckle Assembly Transcript 1 Knockdown Research Articles

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.

MALAT1 and NEAT1 Are Neuroprotective During Hypoxic Preconditioning in the Mouse Hippocampus Possibly by Regulation of NR2B.

Wang L, Fu G, Han R, Fan P, Yang J, Gong K, Zhao Z, Zhang C, Sun K, Shao GMALAT1 and NEAT1 Are Neuroprotective during Hypoxic Preconditioning in the Mouse Hippocampus Possibly by Regulation of NR2B High Alt Med Biol. 00:000-000, 2024. Background: The regulation of noncoding ribonucleic acid (ncRNA) has been shown to be involved in cellular and molecular responses to hypoxic preconditioning (HPC), a situation created by the induction of sublethal hypoxia in the brain. The ncRNAs metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and nuclear paraspeckle assembly transcript 1 (NEAT1) are abundantly expressed in the brain, where they regulate the expression of various genes in nerve cells. However, the exact roles of MALAT1 and NEAT1 in HPC are not fully understood. Methods: A mouse model of acute repeated hypoxia was used as a model of HPC, and MALAT1 and NEAT1 levels in the hippocampus were measured using real-time polymerase chain reaction (PCR). The mRNA and protein levels of N-methyl-d-aspartate receptor subunit 2 B (NR2B) in the mouse hippocampus were measured using real-time PCR and western blotting, respectively. HT22 cells knocked-down for MALAT1 and NEAT1 were used for in vitro testing. Expression of NR2B, which is involved in nerve cell injury under ischemic and hypoxic conditions, was also evaluated. The levels of spectrin and cleaved caspase-3 in MALAT1 and NEAT1 knockdown HT22 cells under oxygen glucose deprivation/reperfusion (OGD/R) were determined by western blotting. Results: HPC increased the expression of MALAT1 and NEAT1 and decreased the expression of NR2B mRNA in the mouse hippocampus (p < 0.05). Knockdown of MALAT1 and NEAT1 increased both NR2B mRNA and protein levels nearly twofold and caused damage under OGD/R conditions in HT22 cells (p < 0.05). Conclusion: MALAT1 and NEAT1 exert neuroprotective effects by influencing the expression of NR2B.

Long non-coding RNA NEAT1 promotes aerobic glycolysis and progression of cervical cancer through WNT/β-catenin/PDK1 axis.

Cervical cancer is one of the most common gynecological cancers. Accumulated evidence shows that long non-coding RNAs (lncRNAs) play essential roles in cervical cancer occurrence and progression, but their specific functions and mechanisms remain to be further explored. The RT-qPCR assay was used to detect the expression of NEAT1 in cervical cancer tissues and cell lines. CCK-8, colony formation, flow cytometry, western blotting, and Transwell assays were used to evaluate the impact of NEAT1 on the malignant behavior of cervical cancer cells. Glucose consumption, lactate production, ATP levels, ROS levels, MMP levels, and the mRNA expressions of glycolysis-related genes and tricarboxylic acid cycle-related genes were detected to analyze the effect of NEAT1 on metabolism reprograming in cervical cancer cells. The expressions of PDK1, β-catenin and downstream molecules of the WNT/β-catenin signaling pathway in cervical cancer cells and tissues were detected by western blotting, RT-qPCR, immunofluorescence and immunohistochemistry assays. This study investigated the role and possible molecular mechanism of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in cervical cancer. Our results showed that NEAT1 was highly expressed in cervical cancer tissues and cell lines. Downregulation of NEAT1 inhibited the proliferation, migration, invasion and glycolysis of cervical cancer cells, while overexpression of NEAT1 led to the opposite effects. Mechanistically, NEAT1 upregulated pyruvate dehydrogenase kinase (PDK1) through the WNT/β-catenin signaling pathway, which enhanced glycolysis and then facilitated cervical cancer metastasis. Furthermore, NEAT1 maintained the protein stability of β-catenin but did not affect its mRNA level. We also excluded the direct binding of NEAT1 to the β-catenin protein via RNA pull-down assay. The suppressive impact of NEAT1 knockdown on cell proliferation, invasion, and migration was rescued by β-catenin overexpression. The WNT inhibitor iCRT3 attenuated the carcinogenic effect induced by NEAT1 overexpression. In summary, these findings indicated that NEAT1 may contribute to the progression of cervical cancer by activating the WNT/β-catenin/PDK1 signaling axis.

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.

LncRNA NEAT1 aggravates human microvascular endothelial cell injury by inhibiting the Apelin/Nrf2/HO-1 signalling pathway in type 2 diabetes mellitus with obstructive sleep apnoea

ABSTRACT Long noncoding RNAs (lncRNAs) regulate the progression of type 2 diabetes mellitus complicated with obstructive sleep apnoea (T2DM-OSA). However, the role of the lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in T2DM-OSA remains unknown. This study aimed to reveal the function of NEAT1 in T2DM-OSA and the underlying mechanism. KKAy mice were exposed to intermittent hypoxia (IH) or intermittent normoxia to generate a T2DM-OSA mouse model. HMEC-1 cells were treated with high glucose (HG) and IH to construct a T2DM-OSA cell model. RNA expression was detected by qRT-PCR. The protein expression of Apelin, NF-E2-related factor 2 (Nrf2), haem oxygenase-1 (HO-1), and up-frameshift suppressor 1 (UPF1) was assessed using western blot. Cell injury was evaluated using flow cytometry, enzyme-linked immunosorbent assay, and oxidative stress kit assays. RIP, RNA pull-down, and actinomycin D assays were performed to determine the associations between NEAT1, UPF1, and Apelin. NEAT1 expression was upregulated in the aortic vascular tissues of mice with T2DM exposed to IH and HMEC-1 cells stimulated with HG and IH, whereas Apelin expression was downregulated. The absence of NEAT1 protected HMEC-1 cells from HG- and IH-induced damage. Furthermore, NEAT1 destabilized Apelin mRNA by recruiting UPF1. Apelin overexpression decreased HG- and IH-induced injury to HMEC-1 cells by activating the Nrf2/HO-1 pathway. Moreover, NEAT1 knockdown reduced HG- and IH-induced injury to HMEC-1 cells through Apelin. NEAT1 silencing reduced HMEC-1 cell injury through the Apelin/Nrf2/HO-1 signalling pathway in T2DM-OSA. Abbreviations: LncRNAs, long non-coding RNAs; T2DM, type 2 diabetes mellitus; OSA, obstructive sleep apnoea; NEAT1, nuclear paraspeckle assembly transcript 1; IH, intermittent hypoxia; HMEC-1, human microvascular endothelial cells; HG, high glucose; Nrf2, NF-E2-related factor 2; UPF1, up-frameshift suppressor 1; HO-1, haem oxygenase-1; qRT-PCR, quantitative real-time polymerase chain reaction; ELISA, enzyme-linked immunosorbent assay; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; TNF-α, tumour necrosis factor-α; CCK-8, Cell Counting Kit-8; IL-1β, interleukin-1β; ROS, reactive oxygen species; MDA, malondialdehyde; SOD, superoxide dismutase; RIP, RNA immunoprecipitation; SD, standard deviations; GSH, glutathione; AIS, acute ischaemic stroke; HMGB1, high mobility group box-1 protein; TLR4, toll-like receptor 4.

Knockdown of NEAT1 prevents post-stroke lipid droplet agglomeration in microglia by regulating autophagy

BackgroundLipid droplets (LD), lipid-storing organelles containing neutral lipids like glycerolipids and cholesterol, are increasingly accepted as hallmarks of inflammation. The nuclear paraspeckle assembly transcript 1 (NEAT1), a long non-coding RNA with over 200 nucleotides, exerts an indispensable impact on regulating both LD agglomeration and autophagy in multiple neurological disorders. However, knowledge as to how NEAT1 modulates the formation of LD and associated signaling pathways is limited.MethodsIn this study, primary microglia were isolated from newborn mice and exposed to oxygen-glucose-deprivation/reoxygenation (OGD/R). To further explore NEAT1-dependent mechanisms, an antisense oligonucleotide (ASO) was adopted to silence NEAT1 under in vitro conditions. Studying NEAT1-dependent interactions with regard to autophagy and LD agglomeration under hypoxic conditions, the inhibitor and activator of autophagy 3-methyladenine (3-MA) and rapamycin (RAPA) were used, respectively. In a preclinical stroke model, mice received intraventricular injections of ASO NEAT1 or control vectors in order to yield NEAT1 knockdown. Analysis of readout parameters included qRT-PCR, immunofluorescence, western blot assays, and behavioral tests.ResultsMicroglia exposed to OGD/R displayed a temporal pattern of NEAT1 expression, peaking at four hours of hypoxia followed by six hours of reoxygenation. After effectively silencing NEAT1, LD formation and autophagy-related proteins were significantly repressed in hypoxic microglia. Stimulating autophagy in ASO NEAT1 microglia under OGD/R conditions by means of RAPA reversed the downregulation of LD agglomeration and perilipin 2 (PLIN2) expression. On the contrary, application of 3-MA promoted repression of both LD agglomeration and expression of the LD-associated protein PLIN2. Under in vivo conditions, NEAT1 was significantly increased in mice at 24 h post-stroke. Knockdown of NEAT1 significantly alleviated LD agglomeration and inhibited autophagy, resulting in improved cerebral perfusion, reduced brain injury and increased neurological recovery.ConclusionNEAT1 is a key player of LD agglomeration and autophagy stimulation, and NEAT1 knockdown provides a promising therapeutic value against stroke.Graphical abstract

NEAT1-SOD2 Axis Confers Sorafenib and Lenvatinib Resistance by Activating AKT in Liver Cancer Cell Lines.

This study investigated the effects of a long noncoding RNA, nuclear paraspeckle assembly transcript 1 (NEAT1) variant 1 (NEAT1v1) on drug resistance in liver cancer cell lines. NEAT1 knockdown activated mitogen-activated protein kinase (MAPK) signaling pathways, including MAPK kinase (MEK)/extracellular signal-regulated kinase (ERK), but suppressed AKT. Moreover, NEAT1 knockdown sensitized liver cancer cells to sorafenib and lenvatinib, both clinically used for treating hepatocellular carcinoma, whereas it conferred resistance to an AKT-targeted drug, capivasertib. NEAT1v1 overexpression suppressed MEK/ERK and activated AKT, resulting in resistance to sorafenib and lenvatinib and sensitization to capivasertib. Superoxide dismutase 2 (SOD2) knockdown reverted the effects of NEAT1v1 overexpression on the sensitivity to the molecular-targeted drugs. Although NEAT1 or SOD2 knockdown enhanced endoplasmic reticulum (ER) stress, concomitant with the suppression of AKT, taurodeoxycholate, an ER stress suppressor, did not restore AKT activity. Although further in vivo and clinical studies are needed, these results suggested that NEAT1v1 switches the growth modality of liver cancer cell lines from MEK/ERK-dependent to AKT-dependent mode via SOD2 and regulates sensitivity to the molecular-targeted drugs independent of ER stress.

The Interplay of NEAT1 and miR-339-5p Influences on Mesangial Gene Expression and Function in Various Diabetic-Associated Injury Models.

Mesangial cells (MCs), substantial cells for architecture and function of the glomerular tuft, take a key role in progression of diabetic kidney disease (DKD). Despite long standing researches and the need for novel therapies, the underlying regulatory mechanisms in MCs are elusive. This applies in particular to long non-coding RNAs (lncRNA) but also microRNAs (miRNAs). In this study, we investigated the expression of nuclear paraspeckle assembly transcript 1 (NEAT1), a highly conserved lncRNA, in several diabetes in-vitro models using human MCs. These cells were treated with high glucose, TGFβ, TNAα, thapsigargin, or tunicamycin. We analyzed the implication of NEAT1 silencing on mesangial cell migration, proliferation, and cell size as well as on mRNA and miRNA expression. Here, the miRNA hsa-miR-339-5p was not only identified as a potential interaction partner for NEAT1 but also for several coding genes. Furthermore, overexpression of hsa-miR-339-5p leads to a MC phenotype comparable to a NEAT1 knockdown. In-silico analyses also underline a relevant role of NEAT1 and hsa-miR-339-5p in mesangial physiology, especially in the context of DKD.

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.

LncRNA NEAT1 knockdown ameliorates LPS-induced human kidney injury by mediating the miR-330-5p/FOXO3 axis.

Sepsis is a systemic process with multiple inflammatory responses and organ injuries, particularly in the damage of the kidney. Recently, numerous studies suggest that long non-coding RNAs (lncRNAs) are involved in sepsis-related kidney injury. This study aimed to investigate the functional role and mechanism of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in sepsis-related kidney injury. Cell model of kidney injury was constructed in human kidney 2 (HK-2) cells with the treatment of lipopolysaccharide (LPS). The expression of NEAT1 was measured by quantitative real-time PCR (qRT-PCR). Cell viability was examined using CCK-8 assay. Flow cytometry was performed to detect cell apoptosis, and apoptosis-related proteins were quantified by western blot. The release of proinflammatory cytokines was assessed by ELISA. Oxidative stress was assessed by the levels of SOD and MDA using kits. The putative relationship between miR-330-5p and NEAT1 or FOXO3 was confirmed using dual-luciferase reporter assay, RIP assay and pull-down assay. The expression of NEAT1 was increased in LPS-treated HK-2 cells. LPS exposure promoted apoptotic rate, inflammatory responses and oxidative stress in HK-2 cells, which were largely ameliorated by NEAT1 knockdown. MiR-330-5p was verified as a target of NEAT1, and miR-330-5p inhibition reversed the effects of NEAT1 knockdown in LPS-treated HK-2 cells. Moreover, FOXO3 was a target of miR-330-5p, and miR-330-5p restoration-blocked cell apoptosis, inflammation and oxidative stress in LPS-treated HK-2 cells were recovered by FOXO3 overexpression. NEAT1 downregulation meliorated LPS-induced HK-2 cell injuries partly by regulating the miR-330-5p/FOXO3 pathway.

The role of long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) in chronic periodontitis progression

ABSTRACT Long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) is a novel pro-inflammatory factor in severe human diseases. Since inflammatory plays important roles in periodontitis progression, we aimed to explore the role of NEAT1 in chronic periodontitis (CP) in vitro. We established a periodontitis cell model was established by Porphyromonas gingivalis lipopolysaccharide (Pg-LPS)-induced periodontal ligament cells (PDLCs). Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to detect the expression of NEAT1, microRNA (miR)-200c-3p, and tumor necrosis factor receptor-associated factor 6 (TRAF6). Cell viability, inflammatory factors, and protein expression of Bcl-2, Bax, and TRAF6 were analyzed by MTT, enzyme-linked immunosorbent assay, and Western blot. The target relationships among NEAT1, miR-200c-3p, and TRAF6 were predicted by the StarBase/TargetScan software, and further validated by dual-luciferase reporter assay. In this research, NEAT1 is up-regulated in CP tissues and periodontitis model group. Silencing of NEAT1 and over-expression of miR-200c-3p enhanced cell viability and repressed apoptosis in the periodontitis model group. NEAT1 targets miR-200c-3p, and miR-200c-3p further targets TRAF6. MiR-200c-3p inhibitor or over-expression of TRAF6 reversed the promoting effect of NEAT1 knockdown on cell viability, and the inhibiting effects on inflammatory cytokines and cell apoptosis. Consequently, the silencing of NEAT1 inhibits inflammation and apoptosis via targeting miR-200c-3p/TRAF6 axis, thereby contributing to alleviate the progression of CP. This finding could provide an underlying target for the treatment of CP.

Implication of long non-coding RNA NEAT1 in the pathogenesis of bacterial meningitis-induced blood-brain barrier damage.

Blood-brain barrier (BBB) damage is closely related to various neurological disorders, including bacterial meningitis (BM). Determining a reliable strategy to prevent BBB damage in the context of infection would be highly desirable. In the present study, we investigated the implications of the long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) in moderating BBB damage. In vitro BBB models were developed by co-culturing hCMEC/D3 cells with glioma cells, whereupon the glioma-exposed endothelial cells (GECs) were treated with a series of mimics, inhibitors, overexpression plasmids, and shRNAs for evaluating whether NEAT1, microRNA-135a (miR-135a) and hypoxia-inducible factor 1α (HIF1α) mediated BBB integrity and permeability. Furthermore, the in vivo biological function of NEAT1 was validated in a mouse model of BBB damage. NEAT1 and HIF1α were determined to be up-regulated, while miR-135a was under-expressed in GECs. As demonstrated by chromatin immunoprecipitation and dual-luciferase reporter assays, NEAT1 could bind to miR-135a, and HIF1α was confirmed as a target of miR-135a. Either overexpression of NEAT1 or depletion of miR-135a impaired the integrity and augmented the permeability of BBB. However, HIF1α silencing could reverse the BBB damage induced by NEAT1 overexpression or by inhibition of miR-135a. In vivo experiments substantiated that knockdown of NEAT1 could alleviate BBB damage in living mice. Hence, NEAT1 knockdown prevents BBB disruption and exerts promise as a potential target for BM treatment.

LncRNA NEAT1 aggravates lipopolysaccharide-induced acute lung injury by regulating the miR-98-5p/TLR4 axis.

Although long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) was reported to be associated with acute lung injury (ALI), its specific mechanism has not been well studied. Mouse and cell ALI models were constructed by lipopolysaccharide (LPS). Cell viability was evaluated by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide assay. Cell death was evaluated by lactate dehydrogenase release (LDH) detection kit and flow cytometry. The levels of cytokines in lung tissues lysates were detected by quantitative real-time PCR (qRT-PCR) and enzyme-linked immunosorbentassay (ELISA). The expression of apoptosis-related markers was detected by Western blot. The relationship between NEAT1, miR-98-5p, and toll-like receptor 4 (TLR4) was determined by bioinformatics prediction, luciferase reporter assay, and RNA immunoprecipitation (RIP) assay. Rescue experiments were performed to determine the role of NEAT1/miR-98-5p/TLR4 in ALI. NEAT1 was significantly upregulated during ALI both in vitro and in vivo. NEAT1 knockdown efficiently attenuated LPS-induced ALI and reduced LPS-induced elevation of cytokines both in vitro and in vivo. NEAT1 negatively regulated miR-98-5p by directly sponging it, and TLR4 was a target of miR-98-5p. MiR-98-5p inhibition or TLR4 overexpression could obviously attenuate the protective effects of NEAT1 knockdown in LPS-treated A549 cells. Our study demonstrated that NEAT1 knockdown alleviated LPS-induced ALI by targeting the miR-98-5p/TLR4 axis.

Long Noncoding RNA NEAT1 Knockdown Ameliorates 1-Methyl-4-Phenylpyridine–Induced Cell Injury Through MicroRNA-519a-3p/SP1 Axis in Parkinson Disease

Parkinson disease is a neurodegenerative disease and is characterized by resting tremor, dementia, and gait disorder. Previous studies have indicated that long noncoding RNA participates in the regulation of the pathogenesis of Parkinson disease. The study aimed to reveal the effects of long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) on 1-methyl-4-phenylpyridine (MPP+)-induced human neuroblastoma cell injury and the underlying mechanism. The expressions of NEAT1, microRNA (miR)-519a-3p, and transcription factor specific protein 1 (SP1) were detected by quantitative real-time polymerase chain reaction. The protein expressions of SP1 and inflammation-related factors were determined by Western blot. Cell viability was determined by 3-(4,5-dimethylthazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptosis was investigated by flow cytometry analysis. The targeting relationship between miR-519a-3p and NEAT1 or SP1 was predicted by starBase online database and verified by a dual-luciferase reporter assay. NEAT1 and SP1 expressions were significantly upregulated, whereas miR-519a-3p was downregulated in MPP+-treated neuroblastoma cells in a dose-and time-dependent manner when compared with control groups. NEAT1 knockdown restrained MPP+-induced repression of cell viability and promotion of cell apoptosis and inflammation. Additionally, NEAT1 served as a sponge of miR-519a-3p and regulated MPP+-caused cell injury by interacting with miR-519a-3p. Also, SP1, a target gene of miR-519a-3p, rescued miR-519a-3p-mediated actions under MPP+ treatment. Importantly, NEAT1 stimulated SP1 expression through interaction with miR-519a-3p. NEAT1 silencing protected against MPP+-induced neuroblastoma cell injury by regulating the miR-519a-3p/SP1 pathway. This finding provides a novel direction for the development of therapeutic strategies for Parkinson disease.

Long non‑coding RNA nuclear paraspeckle assembly transcript 1 regulates ionizing radiation‑induced pyroptosis via microRNA‑448/gasdermin E in colorectal cancer cells.

Pyroptosis is mediated by gasdermins and serves a critical role in ionizing radiation (IR)-induced damage in normal tissues, but its role in cancer radiotherapy and underlying mechanisms remains unclear. Long non-coding (lnc) RNAs serve important roles in regulating the radiosensitivity of cancer cells. The present study aimed to investigate the mechanistic involvement of lncRNAs in IR-induced pyroptosis in human colorectal cancer HCT116 cells. LncRNA, microRNA (miR)-448 and gasdermin E (GSDME) levels were evaluated using reverse transcription-quantitative polymerase chain reaction. Protein expression and activation of gasdermins were measured using western blotting. The binding association between miR-448 and GSDME was assessed using the dual-luciferase reporter assay. Pyroptosis was examined using phase-contrast microscopy, flow cytometry, Cell Counting Kit-8 assay and lactate dehydrogenase release assay. IR dose-dependently induced GSDME-mediated pyroptosis in HCT116 cells. GSDME was identified as a downstream target of miR-448. LncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) was upregulated in response to IR and enhanced GSDME expression by negatively regulating miR-448 expression. Notably, NEAT1 knockdown suppressed IR-induced pyroptosis, full-length GSDME expression and GSDME cleavage compared with that in irradiated cells. In addition, NEAT1 knockdown rescued the IR-induced decrease in cell viability in HCT116 cells. The findings of the present study indicated that lncRNA NEAT1 modulates IR-induced pyroptosis and viability in HCT116 cells via miR-448 by regulating the expression, but not activation of GSDME. The present study provides crucial mechanistic insight into the potential role of lncRNA NEAT1 in IR-induced pyroptosis.

NEAT1 silencing alleviates pulmonary arterial smooth muscle cell migration and proliferation under hypoxia through regulation of miR‑34a‑5p/KLF4 invitro.

Pulmonary arterial hypertension (PAH) is a severe vascular disease that adversely affects patient health and can be life threatening. The present study aimed to investigate the detailed role of nuclear paraspeckle assembly transcript1 (NEAT1) in PAH. Using RT‑qPCR, the expression levels of NEAT1, microRNA (miR)‑34a‑5p, and Krüppel‑like factor4 (KLF4) were detected in both hypoxia‑treated pulmonary arterial smooth muscle cells (PASMCs) and serum from PAH patients. Then, the interactions among miR‑34a‑5p, NEAT1, and KLF4 were evaluated by dual‑luciferase reporter assay. The detailed role of the NEAT1/miR‑34a‑5p/KLF4 axis in PAH pathogenesis was further explored using MTT, Transwell, and western blot assays. The results revealed that NEAT1 targeted miR‑34a‑5p and miR‑34a‑5p targeted KLF4. In hypoxia‑treated PASMCs and serum from PAH patients, high NEAT1 and KLF4 expression levels and low miR‑34a‑5p expression were observed. The proliferation and migration of hypoxia‑treated PASMCs were reduced by transfection with sh‑NEAT1 or miR‑34a‑5p mimics. The suppressive effects of NEAT1 knockdown on the proliferation and migration of hypoxia‑treated PASMCs were reversed by knock down of miR‑34a‑5p expression and increased KLF4 expression. NEAT1 was not only highly expressed in the serum of PAH patients but its silencing also alleviated PAH by regulating miR‑34a‑5p/KLF4 invitro. The present study highlighted a potential new therapeutic target and diagnostic biomarker for PAH.

LncRNA NEAT1 promotes glioma cancer progression via regulation of miR-98-5p/BZW1.

Background: Glioma is the most common malignant tumor in the human central nervous system. Long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) promotes oncogenesis in various tumors. In the present study, we aimed to examine the role of NEAT1 in altering the properties of gliomas.Methods: Quantitative real-time PCR technology was used to determine the expression levels of relevant genes in tumor tissues and cell lines. The protein expression levels were validated by Western blotting. Cell counting kit-8 (CCK-8) and colony formation assays were used to test the cell proliferation ability. A luciferase reporter assay was used to determine the interactions of the genes. Tumor xenografts were used to detect the role of NEAT1 in gliomas in vivo.Results: We demonstrated that NEAT1 up-regulated glioma cells and negatively correlated with miR-98-5p in glioma tissues. A potential binding region between NEAT1 and miR-98-5p was confirmed by dual-luciferase assays. NEAT1 knockdown inhibited glioma cell proliferation. The inhibition of miR-98-5p rescued the knockdown of NEAT1 in glioma cells. Basic leucine zipper and W2 domain containing protein 1 (BZW1) was identified as a direct target of miR-98-5p. We also identified that BZW1 was positively correlated with NEAT1 in glioma tissues. NEAT1 knockdown inhibited glioma cell proliferation in vivo via miR-98-5p/BZW1.Conclusion: Our results suggest that NEAT1 plays an oncogenic function in glioma progression. Targeting NEAT1/miR-98-5p/BZW1 may be a novel therapeutic treatment approach for glioma patients.

LncRNA NEAT1 Enhances Glioma Progression via Regulating the miR-128-3p/ITGA5 Axis

Accumulating evidences indicate that long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) promotes the progression of glioma. In this study, we postulated that NEAT1 may act as a miR-128-3p sponge. Relative levels of NEAT1 and miR-128-3p expression in human glioma samples and GBM cells were detected using quantitative real-time PCR. By means of CCK-8 assays, transwell assays, and flow cytometric analysis, the biological functions of miR-128-3p and NEAT1 were investigated in U87MG and U251MG human GBM cell lines with stable miR-128-3p and NEAT1 knockdown or overexpression. The luciferase reports, RNA pull-down assay, and RNA immunoprecipitation assay were conducted to determine the relevance of NEAT1 and miR-128-3p in glioma. As a result, high expression of NEAT1 and lack of miR-128-3p were observed in glioma specimens and cells. By binding to anti-oncogene miR-128-3p in the nucleus, NEAT1 enhanced tumorigenesis and glioma development. Further experiments suggested that ITGA5 expression was increased in glioma tissues and was found to be connected with miR-128-3p. Additionally, NEAT1 facilitated ITGA5 expression via competitively binding to miR-128-3p. For this reason, ITGA5 would not be decomposed by miR-128-3p and could activate FAK signaling pathway, thereby promoting cell growth. Collectively, these results indicated that the NEAT1/miR-128-3p/ITGA5 axis was involved in glioma initiation and progression, and might offer a potential novel strategy for treatment of glioma.

LncRNA NEAT1 Knockdown Alleviates Lipopolysaccharide-Induced Acute Lung Injury by Modulation of miR-182-5p/WISP1 Axis.

Accumulating evidence has demonstrated the vital roles of long non-coding RNAs (lncRNAs) in acute lung injury (ALI). In this study, we aimed to explore the effect of Nuclear Paraspeckle Assembly Transcript 1 (NEAT1) on ALI development. The ALI mice and cell models were constructed using lipopolysaccharide (LPS)-induced method. The concentrations of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) were measured by enzyme-linked immunosorbent assay (ELISA). The levels of TNF-α mRNA, IL-6 mRNA, IL-1β mRNA, NEAT1, miR-182-5p, and WNT-inducible secreted protein 1 (WISP1) mRNA were determined by quantitative real-time polymerase chain reaction (qRT-PCR) assay. Cell viability was evaluated by Cell Counting Kit-8 (CCK-8) assay. The level of lactate dehydrogenase (LDH) and the activity of caspase-3 were measured by specific kits. The interaction between miR-182-5p and NEAT1 or WISP1 was investigated by dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. Protein levels were measured by Western blot assay. NEAT1 level was elevated in LPS-induced ALI mice and LPS-stimulated MH-S cells. LPS treatment repressed MH-S cell viability and promoted apoptosis and inflammation, while NEAT1 silencing restored the impacts. For mechanism analysis, NEAT1 was identified as the sponge for miR-182-5p to positively regulate WISP1 expression. Moreover, NEAT1 knockdown could accelerate cell viability and inhibit cell apoptosis and inflammation in LPS-induced MH-S cells by elevating miR-182-5p and decreasing WISP1 in LPS-exposed MH-S cells. In addition, NEAT1 deficiency blocked the activation of NF-κB pathway caused by LPS in MH-S cells. NEAT1 overexpression restrained cell viability and facilitated cell apoptosis and inflammation in LPS-exposed MH-S cells through miR-182-5p/WISP1 axis.

Long non‑coding RNA NEAT1 regulates glioma cell proliferation and apoptosis by competitively binding to microRNA‑324‑5p and upregulating KCTD20 expression.

Previous studies have demonstrated that long non-coding RNAs (lncRNAs) serve a key role in the development and progression of several types of cancer, including glioma. The lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) contributes to cancer growth through its effects on cell proliferation, migration, invasion and drug resistance. However, the exact regulatory mechanisms via which NEAT1 acts in glioma are unclear. In the present study, the expression levels and function of NEAT1 in glioma tissues and cell lines were examined in vitro and in vivo. By reverse transcription-quantitative PCR and fluorescence in situ hybridization analysis, NEAT1 expression was upregulated in glioma tissues compared with in adjacent normal brain tissues, and elevated NEAT1 levels were associated with poor prognosis. Cell Counting Kit-8, colony formation, ethynyldeoxyuridine, flow cytometry and western blotting assays were performed to detect the effects of NEAT1 on cell biological behavior. Knockdown of NEAT1 in glioma cell lines was associated with cell cycle arrest at the G0/G1 phase, decreased proliferation and elevated apoptosis in vitro, and resulted in reduced tumor growth and increased survival in a mouse xenograft model of glioma. Using bioinformatics analysis, RNA immunoprecipitation experiments and luciferase reporter assays, it was demonstrated that NEAT1 may competitively bind to microRNA (miR)-324-5p, thus blocking its interaction with target mRNAs. Potassium channel tetramerization protein domain containing 20 (KCTD20) was identified as a specific miR-324-5p target. Accordingly, the inhibition of NEAT1 resulted in the downregulation of KCTD20 through competitive binding with miR-324-5p, decreased cell proliferation and increased apoptosis. Concomitant NEAT1 knockdown and inhibition of miR-324-5p partially reversed the effects of NEAT1 knockdown on cell proliferation and apoptosis, and further regulated KCTD20 expression. Collectively, the present findings demonstrated that NEAT1 acted as a competing endogenous RNA for miR-324-5p, and identified the NEAT1/miR-324-5p/KCTD20 axis as a novel regulatory axis and a potential therapeutic target for human glioma.