Stress Induces Dynamic, Cytotoxicity-Antagonizing TDP-43 Nuclear Bodies via Paraspeckle LncRNA NEAT1-Mediated Liquid-Liquid Phase Separation.

Despite the prominent role of TDP-43 in neurodegeneration, its physiological and pathological functions are not fully understood. Here, we report an unexpected role of TDP-43 in the formation of dynamic, reversible, liquid droplet-like nuclear bodies (NBs) in response to stress. Formation of NBs alleviates TDP-43-mediated cytotoxicity in mammalian cells and fly neurons. Super-resolution microscopy reveals distinct functions of the two RRMs in TDP-43 NB formation. TDP-43 NBs are partially colocalized with nuclear paraspeckles, whose scaffolding lncRNA NEAT1 is dramatically upregulated in stressed neurons. Moreover, increase of NEAT1 promotes TDP-43 liquid-liquid phase separation (LLPS) in vitro. Finally, we discover that the ALS-associated mutation D169G impairs the NEAT1-mediated TDP-43 LLPS and NB assembly, causing excessive cytoplasmic translocation of TDP-43 to form stress granules, which become phosphorylated TDP-43 cytoplasmic foci upon prolonged stress. Together, our findings suggest a stress-mitigating role and mechanism of TDP-43 NBs, whose dysfunction may be involved in ALS pathogenesis.

LncRNA NEAT1 promotes proliferation, migration, and invasion of laryngeal squamous cell carcinoma cells through miR-411-3p/FZD3-mediated Wnt signaling pathway.

The lncRNA NEAT1 has been shown to promote the progression of several cancers, containing laryngeal squamous cell carcinoma (LSCC). However, the precise mechanism by which it promotes LSCC progression remains unclear. In this study, we verified the high expression of lncRNA NEAT1 in LSCC tissues and cells using RT-qPCR. Analysis of clinical data exhibited that high expression of lncRNA NEAT1 was associated with a history of smoking, worse T stage, lymph node metastasis, and later TNM stage in patients with LSCC. The promotion effect of lncRNA NEAT1 on LSCC cell proliferation, migration, invasion, and tumor growth in vivo was verified by CCK-8, plate clone formation, Transwell, and nude mouse tumorigenicity assays. Bioinformatics prediction and double luciferase reporter gene assay verified the binding of miR-411-3p to lncRNA NEAT1 and FZD3 mRNA, and inhibition of miR-411-3p reversed the inhibitory effect of lncRNA NEAT1 on FZD3 expression in LSCC cells. We also verified that lncRNA NEAT1-mediated FZD3 activation in the Wnt pathway affects LSCC development. In conclusion, we demonstrate that lncRNA NEAT1 promotes the progression of LSCC, and propose that the lncRNA NEAT1/miR-411-3p/FZD3 axis may be an effective target for LSCC therapy.

LncRNA NEAT1 inhibits apoptosis and autophagy of ovarian granulosa cells through miR-654/STC2-mediated MAPK signaling pathway.

Long non-coding RNA (lncRNA) anomalies cause early ovarian failure. LncRNA nuclear enriched abundant transcript 1 (NEAT1) was down-regulated in premature ovarian failure (POF) mice and connected to the illness, however, the mechanism remained unclear. The levels of gene and protein were measured by using quantitative real-time polymerase chain reaction, Western blot, and immunofluorescence. Follicle stimulating hormone (FSH), estradiol (E2), and luteinizing hormone (LH) levels were determined using enzyme-linked immunosorbent assay (ELISA). 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and flow cytometry were used to determine cell viability and apoptosis. The interaction of NEAT1, miR-654, and stanniocalcin-2 (STC2) was verified by dual-luciferase reporter assay or RNA binding protein immunoprecipitation (RIP) assays. The results showed NEAT1 and STC2 down-regulated, while miR-654 up-regulated in POF mice. Overexpression of NEAT1 reduced apoptosis and autophagy in cyclophosphamide (CTX)-treated ovarian granulosa cells (OGCs), and Bax, cleaved-caspase3, LC3B, LC3II/LC3I ratio were decreased and Bcl-2 and p62 were raised. NEAT1 suppressed miR-654 expression by directly targeting miR-654. The inhibition of NEAT1 overexpression on apoptosis and autophagy in OGCs was reversed by miR-654 mimics. STC2 was a target gene of miR-654, and miR-654 inhibitor reduced the apoptosis and autophagy by regulating the STC2/MAPK axis. To sum up, NEAT1 reduced miR-654 expression and modulated the STC2/MAPK pathway to decrease apoptosis and autophagy in POF, indicating a potential therapeutic target.

Icariin Regulates EMT and Stem Cell-Like Character in Breast Cancer through Modulating lncRNA NEAT1/TGFß/SMAD2 Signaling Pathway.

Metastases and drug resistance are the major risk factors associated with breast cancer (BC), which is the most common type of tumor affecting females. Icariin (ICA) is a traditional Chinese medicine compound that possesses significant anticancer properties. Long non-coding RNAs (lncRNAs) are involved in a wide variety of biological and pathological processes and have been shown to modulate the effectiveness of certain drugs in cancer. The purpose of this study was to examine the potential effect of ICA on epithelial mesenchymal transition (EMT) and stemness articulation in BC cells, as well as the possible relationship between its inhibitory action on EMT and stemness with the NEAT1/transforming growth factor ß (TGFß)/SMAD2 pathway. The effect of ICA on the proliferation (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony assays), EMT (Western blotting, immunofluorescence, and wound healing), and stemness (mammosphere formation assays, Western blotting) of BC cells were examined. According to the findings, ICA suppressed the proliferation, EMT, and stem cell-like in MDA-MB-231 cells, and exerted its inhibitory impact by downregulating the TGFß/SMAD2 signaling pathway. ICA could significantly downregulate the expression of lncRNA NEAT1, and silencing NEAT1 enhanced the effect of ICA in suppressing EMT and expression of different stem cell markers. In addition, silencing NEAT1 was found to attenuate the TGFß/SMAD2 signaling pathway, thereby improving the inhibitory impact of ICA on stemness and EMT in BC cells. In conclusion, ICA can potentially inhibit the metastasis of BC via affecting the NEAT1/TGFß/SMAD2 pathway, which provides a theoretical foundation for understanding the mechanisms involved in potential application of ICA for BC therapy.

LncRNA NEAT1 in the pathogenesis of liver-related diseases.

Nuclear paraspeckle assembly transcript 1 (NEAT1) is a long noncoding RNA (lncRNA) that is widely expressed in a variety of mammalian cell types. Altered expression levels of the lncRNA NEAT1 have been reported in liver-related disorders including cancer, fatty liver disease, liver fibrosis, viral hepatitis, and hepatic ischemia. lncRNA NEAT1 mostly acts as a competing endogenous RNA (ceRNA) to sponge various miRNAs (miRs) to regulate different functions. In regard to hepatic cancers, the elevated expression of NEAT1 has been reported to have a relation with the proliferation, migration, angiogenesis, apoptosis, as well as epithelial-mesenchymal transition (EMT) of cancer cells. Furthermore, NEAT1 upregulation has contributed to the pathogenesis of other liver diseases such as fibrosis. In this review, we summarize and discuss the molecular mechanisms by which NEAT1 contributes to liver-related disorders including acute liver failure, nonalcoholic fatty liver disease (NAFLD), liver fibrosis, and liver carcinoma, providing novel insights and introducing NEAT1 as a potential therapeutic target in these diseases.

LncRNA-NEAT1 blocks the Wnt/ß-catenin signaling pathway by targeting miR-217 to inhibit trophoblast cell migration and invasion.

OBJECTIVE: This study aimed to study the correlation between preeclampsia (PE) and lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1), and to examine the molecular mechanisms behind the development of PE. METHODS: 30 PE and 30 normal pregnant women placental samples were assessed the levels of NEAT1 and miR-217 by quantitative real-time PCR (qRT-PCR). The trophoblast cell line HTR8/SVneo was used for silencing NEAT1 or miR-217 inhibitor in the absence or presence of an inhibitor and H2O2. Cell counting Kit 8 (CCK-8), flow cytometry, and Transwell were used to detect cell proliferation, apoptosis, migration, and invasion. Luciferase reporter gene assay was utilized to verify the binding between miR-217 and Wnt family member 3 (Wnt3), and between the miR-217 and NEAT1. Proteins related to the Wnt/ß-catenin signaling pathway were detected using western blotting. RESULTS: The PE group exhibited a significantly downregulated expression of miR-217 and a significantly upregulated expression of NEAT1. NEAT1 targeted miR-217, and Wnt is a miR-217 target gene. siRNA-NEAT1 inhibited the apoptosis of trophoblast cells, but promoted their invasion, migration, and proliferation. MiR-217 inhibitor could partially reverse the effects of siRNA-NEAT1. The expression of the Wnt/ß-catenin signaling pathway-related proteins, WNT signaling pathway inhibitor 1 (DKK1), cyclin-D1 and ß-catenin, was significantly increased after siRNA-NEAT1. CONCLUSIONS: NEAT1 could reduce trophoblast cell invasion and migration by suppressing miR-217/Wnt signaling pathway, leading to PE.

[Expression relationship and significance of NEAT1 and miR-27a-3p in serum and cerebrospinal fluid of patients with Alzheimer disease].

OBJECTIVE: To explore the expression relationship and significance of long chain non-coding RNA nuclear-enriched abundant transcript 1 (LncRNA NEAT1) and miR-27a-3p in serum and cerebrospinal fluid of patients with Alzheimer disease (AD). METHODS: Sixty-six AD patients received by the department of neurology of our hospital from October 2019 to September 2021 were gathered, according to the clinical dementia rating scale score, they were grouped into mild group (≤1 point, n=41) and moderate-to-severe group (>1 point, n=25). Another 66 cases of serum and cerebrospinal fluid samples from outpatient physical examination personnel were regarded as the control group. The general information on all subjects was recorded and cognition was assessed; real-time quantitative PCR was performed to measure the expression levels of miR-27a-3p and NEAT1 in serum and cerebrospinal fluid; enzyme-linked immunosorbent assay was performed to measure the protein levels of ß-amyloid precursor protein cleaving enzyme 1 (BACE1), ß-amyloid (Aß) 40 and Aß42 in cerebrospinal fluid; Spearman' s method was performed to analyze the correlation of serum miR-27a-3p and NEAT1 levels with mini-mental state examination (MMSE) and montreal cognitive assessment (MoCA) scores; Pearson method was performed to analyze the correlation between serum miR-27a-3p and NEAT1 levels and Aß deposition standard uptake value ratio (SUVR) and cerebrospinal fluid miR-27a-3p, NEAT1, BACE1, Aß42 and Aß40 levels. RESULTS: The MMSE score [21 (17, 25), 9(7, 11) vs. 27 (21, 34)], MoCA score [17 (12, 21), 10 (7, 13) vs. 27 (21, 31)], serum miR-27a-3p level (0.55±0.13, 0.46±0.06 vs. 0.97±0.22), cerebrospinal fluid miR-27a-3p (0.48±0.10, 0.35±0.10 vs. 1.03±0.31), Aß42 levels [(303.55±36.77) ng/L, (231.45±34.14) ng/L vs. (499.99±53.63) ng/L] and Aß42/Aß40 ratio (0.030±0.008, 0.022±0.007 vs. 0.048±0.010) of AD patients in mild group and moderate-to-severe group were all lower than those in the control group, and the moderate-to-severe group were lower than the mild group (all P < 0.05); the serum NEAT1 level (2.31±0.64, 3.13±0.76 vs. 1.05±0.20), SUVR (1.50±0.29, 1.76±0.52 vs. 0.74±0.15), and cerebrospinal fluid NEAT1 (3.51±1.24, 4.30±1.65 vs. 1.01±0.23) and BACE1 levels [(55.78±5.98) µg/L, (72.32±16.08) µg/L vs. (21.39±3.73) µg/L] were higher than those in the control group, and the moderate-to-severe group were higher than the mild group (all P < 0.05). Serum NEAT1 level in AD patients was positively correlated with SUVR, cerebrospinal fluid NEAT1 and BACE1 (r=0.350, 0.606, 0.341, P < 0.05), and negatively correlated with MMSE score and MoCA score (r=-0.473, -0.482, all P < 0.05); serum miR-27a-3p level was positively correlated with cerebrospinal fluid miR-27a-3p level, MMSE score and MoCA score (r=0.695, 0.424, 0.412, all P < 0.05), and negatively correlated with SUVR and cerebrospinal fluid BACE1 level (r=-0.521, -0.447, all P < 0.05). CONCLUSION: The expression trends of NEAT1 and miR-27a-3p in the serum and cerebrospinal fluid of AD patients are consistent, the level of NEAT1 is increased, and the level of miR-27a-3p is decreased. The levels of the two are negatively correlated, which is related to the degree of Aß deposition in the brain of AD patients and is involved in the progression of AD.

LncRNA NEAT1 accelerates renal fibrosis progression via targeting miR-31 and modulating RhoA/ROCK signal pathway.

Renal fibrosis is the final pathway for chronic kidney disease to end-stage renal failure. Noncoding RNAs have been reported to play a crucial role in renal fibrosis. Here, the effects of long noncoding RNA (lncRNA) nuclear-enriched abundant transcript 1 (NEAT1) and miR-31 on renal fibrosis and their regulatory mechanism were evaluated. RT-qPCR was used to assess NEAT1, miR-31, and RhoA levels. Western blot was performed to analyze the expression of fibrosis markers, RhoA, rho-related kinase (ROCK1), and connective tissue growth factor (CTGF). RNA immunoprecipitation (RIP), fluorescence in situ hybridization (FISH), and luciferase reporter assays verified the interaction between miR-31 and NEAT1 or RhoA. Renal fibrosis and injury were observed by Masson and hematoxylin and eosin (H&E) staining. The expression level of inflammatory cytokines was detected by ELISA. Immunohistochemistry (IHC) was performed to examine the expression levels of α-smooth muscle actin (α-SMA) and RhoA in renal tissues. We showed that NEAT1 was highly expressed, whereas miR-31 was decreased in renal fibrosis. NEAT1 was found to directly bind miR-31 to positively regulate RhoA expression. Furthermore, NEAT1 silencing inhibited renal fibrosis and inflammation and suppressed the RhoA/ROCK1 signaling pathway. However, knockdown of miR-31 could reverse these effects. NEAT1 silencing or overexpression of miR-31 alleviated renal fibrosis in vivo. In conclusion, NEAT1 accelerates renal fibrosis progression via negative regulation of miR-31 and the activation of RhoA/ROCK1 pathway, thereby upregulating the expression level of CTGF, providing a theoretical basis for treatment and prognostic evaluation of renal fibrosis.

LncRNA NEAT1 regulates HCV-induced Hepatocellular carcinoma by modulating the miR-9-BGH3 axis.

Chronic hepatitis C virus (HCV) infection is a leading cause of end-stage liver diseases, such as fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Several cellular entities, including paraspeckles and their related components, are involved in viral pathogenesis and cancer progression. NEAT1 lncRNA is a major component of paraspeckles that has been linked to several malignancies. In this study, analysis of the Cancer Genome Atlas (TCGA) database and validation in HCV-induced HCC tissue and serum samples showed significantly high expression of NEAT1 in patients with liver cancer. Moreover, we found that NEAT1 levels increased upon HCV infection. To further understand the mechanism of NEAT1-induced HCC progression, we selected one of its targets, miR-9-5 p, which regulates BGH3 mRNA levels. Interestingly, miR-9-5 p levels were downregulated upon HCV infection, whereas BGH3 levels were upregulated. Additionally, partial NEAT1 knockdown increased miR-9-5 p levels and decreased BGH3 levels, corroborating our initial results. BGH3 levels were also upregulated in HCV-induced HCC and TCGA tissue samples, which could be directly correlated with NEAT1 levels. As a known oncogene, BGH3 is directly linked to HCC progression mediated by NEAT1. We also found that NEAT1 levels remained upregulated in serum samples from patients treated with direct-acting antivirals (DAA), indicating that NEAT1 might be a molecular trigger that promotes HCC development. Collectively, these findings provide molecular insights into HCV-induced HCC progression via the NEAT1-miR-9-BGH3 axis.

RBM10 regulates alternative splicing of lncRNA Neat1 to inhibit the invasion and metastasis of NSCLC.

BACKGROUND: Non-small cell lung cancer (NSCLC) accounts for more than 85% of the total cases with lung cancer. NSCLC is characterized by easy metastasis, which often spreads to bones, brains and livers. RNA-binding motif protein 10 (RBM10) is an alternative splicing (AS) regulator frequently mutated in NSCLC. We found that there were multiple peak binding sites between RBM10 and long non-coding RNA nuclear enriched abundant transcript 1 (LncRNA Neat1) by crosslinking-immunprecipitation and high-throughput sequencing (Clip-Seq). LncRNA Neat1 plays an indispensable role in promoting cancer in a variety of tumors and produces two splicing variants: Neat1_1 and Neat1_2. This study aims to explore the mechanism of RBM10 and LncRNA Neat1 in invasion and metastasis of NSCLC. METHODS: Through histological and cytological experiments, we assessed the expression level of RBM10 protein expression. The interaction between RBM10 and Neat1 was evaluated via Clip-Seq and RNA immunoprecipitation assay. The effect of RBM10 on Neat1 and its splicing variants was identified by RT-qPCR. The effect of RBM10 and Neat1 on invasive and metastasis phenotypes of NSCLC was analyzed using transwell invasion assay and scratch test. Additionally, downstream signaling pathway of RBM10 were identified by immunofluorescence and western blot. RESULTS: RBM10 exhibited low levels of expression in NSCLC tissues and cells. RBM10 inhibited the invasion and metastasis of NSCLC and recruited Neat1 and Neat1_2. Overexpression of RBM10 simultaneously inhibited Neat1 and Neat1_2, and promoted the expression of Neat1_1. On the other hand, silencing RBM10 promoted Neat1 and Neat1_2, and inhibited the expression of Neat1_1. From this, we concluded that RBM10 regulated AS of Neat1, and the tumor-promoting effect of Neat1 was mainly attributed to Neat1_2. RBM10 had a negative correlation with Neat1_2. In addition, RBM10 upregulated the expression of PTEN and downregulated the phosphorylation of PI3K/AKT/mTOR through Neat1_2, which ultimately inhibited the invasion and metastasis of NSCLC. CONCLUSION: The RBM10 regulated AS of Neat1 to cause the imbalance of Neat1_1 and Neat1_2, and RBM10 suppressed the activation of the PTEN/PI3K/AKT/mTOR signal by downregulating Neat1_2, finally affected the invasion and metastasis of NSCLC.

Acetylation of H3K27 activated lncRNA NEAT1 and promoted hepatic lipid accumulation in non-alcoholic fatty liver disease via regulating miR-212-5p/GRIA3.

Non-alcoholic fatty liver disease (NAFLD) was a world-wide health burden. H3K27 acetylation, long non-coding RNA (lncRNA), and miRNA were all implicated in NAFLD regulation, yet the detailed regulatory mechanism was not well understood. LncRNA NEAT1, miR-212-5p, and GRIA3 expression were detected both in high fatty acid-treated hepatocytes cells and NAFLD patients. Lipid droplets were stained and analyzed by oil red O staining. Expression of fatty acid synthase (FASN), acetyl-CoA carboxylase (ACC), and GRIA3 was detected by qRT-PCR and western blot. RNA level of lncRNA NEAT1 and miR-212-5p was analyzed by qRT-PCR. The binding sequences of lncRNA NEAT1/miR-212-5p and miR-212-5p/GRIA3 were predicted bioinformatically and validated through luciferase assay. ChIP was performed to analyze H3K27 acetylation on the promoter of lncRNA NEAT1. LncRNA NEAT1 and GRIA3 was upregulated, while miR-212-5p was downregulated in NAFLD patients. FFA promoted lncRNA NEAT1 and GRIA3 expression while suppressing miR-212-5p and promoted lipid accumulation as indicated by increased oil red O staining and FAS and ACC expression. ChIP indicated enrichment of H3K27 on NEAT1 promoter. Inhibition of H3K27 acetylation suppressed lncRNA NEAT1 level. Luciferase results indicated direct interaction of NEAT1/miR-212-5p (which was confirmed by RIP) and miR-212-5p/GRIA3. LncRNA NEAT1 knockdown upregulated miR-212-5p level and inhibited FFA-induced lipid accumulation while suppressing GRIA3 expression. Such function was antagonized by miR-212-5p inhibition and GRIA3 knockdown counteracted with miR-212-5p inhibition. H3K27 acetylation was enriched within the promoter of lncRNA NEAT1 and promoted lncRNA NEAT1 transcription. LncRNA NEAT1 could then interact with miR-212-5p and suppress its cellular concentration.

ALKBH5 promotes the progression of infantile hemangioma through regulating the NEAT1/miR-378b/FOSL1 axis.

Our work aims to investigate long non-coding RNA (lncRNA) N6-methyladenosine (m6A) modification and its role in infantile hemangioma (IH). The mRNA and protein expression levels were assessed using quantitative real-time polymerase chain reaction, western blot and immunohistochemistry. Me-RIP assay was performed to evaluate lncRNA NEAT1 m6A levels. Cell proliferation, migration and invasion were evaluated using cell counting kit-8 assay, transwell migration and invasion assay, respectively. Photo-activatable ribonucleoside-enhanced crosslinking and immunoprecipitation assay was conducted to verify the binding relationship between lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) and ALKBH5 (an RNA demethylase). The binding relationship between lncRNA NEAT1, microRNA (miR)-378b and FOS-like antigen 1 (FOSL1) was verified using dual-luciferase reporter gene assay and/or RNA immunoprecipitation assay. ALKBH5, lncRNA NEAT1 and FOLS1 expression was elevated in IH tissues, while miR-378b was downregulated. ALKBH5 knockdown suppressed cell proliferation, migration and invasion of IH cells, while promoting cell apoptosis. ALKBH5 promoted lncRNA NEAT1 expression by reducing the m6A modification of lncRNA NEAT1. In addition, miR-378b was the target of lncRNA NEAT1, and its overexpression reversed the promotion effect of lncRNA NEAT1 overexpression on IH cell tumor-like behaviors. Moreover, FOLS1 was the target of miR-378b, and its overexpression reversed the inhibitory effect of miR-378b overexpression on IH cell tumor-like behaviors in vitro. ALKBH5 might have great potential as therapeutic target for IH, since ALKBH5 silencing suppressed IH progression by regulation of the NEAT1/miR-378b/FOSL1 axis.

Silencing lncRNA NEAT1 reduces nonalcoholic fatty liver fat deposition by regulating the miR-139-5p/c-Jun/SREBP-1c pathway.

INTRODUCTION AND OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD) starts with the abnormal accumulation of lipids in the liver. Long noncoding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) was reported to modulate hepatic metabolic homeostasis in NAFLD. However, little is known about the molecular mechanisms of NAFLD. MATERIALS AND METHODS: To establish a NAFLD cellular model, HepG2 cells and LO2 cells were treated with 1 mM free fatty acids (FFAs) for 24 h. NEAT1, miRNA (miR)-139-5p, c-Jun and sterol-regulatory element binding protein-1c (SREBP-1c) were evaluated using qPCR. The protein levels of c-Jun, SREBP1c, acetyl-CoA carboxylase (ACC) and fatty acid synthetase (FAS) were determined using western blotting. Moreover, Oil Red O staining was employed to assess lipid accumulation. In addition, a kit assay was performed to evaluate TG levels. Finally, the interactions among NEAT1, miR-139-5p, c-Jun and SREBP1c were identified by dual luciferase reporter gene assay. RESULTS: NEAT1, c-Jun and SREBP1c expression was markedly elevated, while miR-139-5p expression was reduced in the NAFLD cellular model. NEAT1 knockdown restrained lipid accumulation in the NAFLD cellular model by directly targeting miR-139-5p. Moreover, miR-139-5p overexpression suppressed lipid accumulation by directly suppressing c-Jun expression. In addition, c-Jun silencing suppressed lipid accumulation by directly targeting SREBP1c. Finally, miR-139-5p inhibition mitigated the inhibitory effect of sh-NEAT1 on lipid accumulation. CONCLUSION: NEAT1 aggravated FFA-induced lipid accumulation in hepatocytes by regulating the c-Jun/SREBP1c axis by sponging miR-139-5p, indicating the potential of NEAT1 as a promising therapeutic target for NAFLD.

LncRNA NEAT1 ameliorate ischemic stroke via promoting Mfn2 expression through binding to Nova and activates Sirt3.

BACKGROUND: Recent studies revealed that long non-coding RNAs (lncRNAs) have significant roles in regulating the pathogenesis of ischemia stroke, and oxygen-glucose deprivation/reoxygenation (OGD/R)-induced cell apoptosis. Aberrant expression of NEAT1 was found after the injury of ischemia-reperfusion, but the mechanism was not fully understood. METHODS: The expression of NEAT1 and Mfn2 were detected in BV-2 and N2a cell with or without OGD/R-induced by qRT-PCR. Inflammatory cytokines secretion was detected by enzyme-linked immunosorbent assay (ELISA). The oxidative stress was evaluated by the examination of ROS, MDA and SOD levels. Flow cytometry and apoptosis marker detection by western blot were performed to examined apoptosis. RESULTS: The expression of NEAT1 and Mfn2 were decreased in OGD/R-induced cell model. Overexpression of NEAT1 or Mfn2 reduced oxidative stress and apoptosis by OGD/R-induced in neuronal cells, while knockdown of Sirt3 reversed the protective effect of NEAT1 and Mfn2. NEAT1 stabilized Mfn2 mRNA via recruiting Nova. NEAT1 alleviates the oxidative stress and apoptosis by OGD/R-induced via activating Sirt3. CONCLUSION: LncRNA NEAT1 stabilizes Mfn2 mRNA via recruiting Nova, therefore increase the expression of Mfn2 and alleviates ischemia-reperfusion induced oxidative stress and apoptosis via Mfn2/Sirt3 pathway.

Tetrandrine-induced downregulation of lncRNA NEAT1 inhibits rheumatoid arthritis progression through the STAT3/miR-17-5p pathway.

BACKGROUND: The inhibitory effect of Tetrandrine (Tet) on rheumatoid arthritis (RA) is well established. However, its exact molecular mechanism remains unknown. METHODS: RT-qPCR coupled with western blotting was employed to analyze the expression of NEAT1, miR-17-5p, and STAT3 in RA tissues and/or RA-fibroblast-like synoviocytes (RA-FLS) treated with 3 µmol/L of Tet for 48 h. Cell Counting Kit-8 assay and flow cytometry were performed to assess RA-FLS proliferation and apoptosis. Luciferase reporter assays were used to validate the interactions between miR-17-5p and STAT3 or NEAT1. RESULTS: The expression of NEAT1 decreased in a time-dependent manner upon Tet treatment. Tet significantly inhibited RA-FLS proliferation and triggered apoptosis by downregulating NEAT1 expression. Additionally, NEAT1 directly targeted miR-17-5p to upregulate STAT3 expression. Tet-induced low NEAT1 expression impaired RA-FLS growth by targeting miR-17-5p and inhibiting STAT3. CONCLUSION: Tet exerts its inhibitory role in RA progression by regulating the NEAT1/miR-17-5p/STAT3 pathway.

Gastrodin Alleviates Cerebral Ischaemia/Reperfusion Injury by Inhibiting Pyroptosis by Regulating the lncRNA NEAT1/miR-22-3p Axis.

Cerebral ischaemia/reperfusion (I/R) injury-induced irreversible brain injury is a major cause of mortality and functional impairment in ageing people. Gastrodin (GAS), derived from the traditional Chinese herbal medicine Tianma, has been reported to inhibit the progression of stroke, but the mechanism whereby GAS modulates the progression of cerebral I/R remains unclear. The middle cerebral artery occlusion method was used as a model of I/R in vivo. Rats were pretreated with GAS by intraperitoneal injection 7 days before I/R surgery and were then treated with GAS for 7 days after I/R surgery. Additionally, an oxygen-glucose deprivation/reoxygenation model using neuronal cells was established in vitro to simulate I/R injury. 2,3,5-Triphenyltetrazolium chloride and Nissl staining were used to evaluate infarct size and neuronal damage, respectively. Lactate dehydrogenase release and cell counting kit-8 assays were used to assess neuronal cell viability. Enzyme-linked immunosorbent assay, qPCR, flow cytometry and western blotting were performed to analyse the expression levels of inflammatory factors (IL-1ß, IL-18), lncRNA NEAT1, miR-22-3p, NLRP3 and cleaved caspase-1. Luciferase reporter experiments were performed to verify the association between lncRNA NEAT1 and miR-22-3p. The results indicated that GAS could significantly improve the neurological scores of rats and reduce the area of cerebral infarction. Meanwhile, GAS inhibited pyroptosis by downregulating NLRP3, inflammatory factors (IL-1ß, IL-18) and cleaved caspase-1. In addition, GAS attenuated I/R-induced inflammation in neuronal cells through the modulation of the lncRNA NEAT1/miR-22-3p axis. GAS significantly attenuated cerebral I/R injury via modulation of the lncRNA NEAT1/miR-22-3p axis. Thus, GAS might serve as a new agent for the treatment of cerebral I/R injury.

LncRNA NEAT1 promotes airway smooth muscle cell inflammation by activating the JAK3/STAT5 pathway through targeting of miR-139.

Background Asthma is a chronic inflammatory heterogeneous respiratory disease. Previous studies showed that the lncRNA NEAT1 (nuclear paraspeckle assembly transcript 1) might play an important role in the pathogenesis of asthma, but its potential mechanism in airway smooth muscle cell (ASMC) inflammation remains largely unknown and needs further investigation.Methods We performed cellular immunofluorescence to identify the features of ASMCs and detected the expression levels of lncRNA NEAT1, miR-139, TNF-α, IL-6, IL-8 and IL-1ß by quantitative real-time PCR (Q-PCR) and ELISA. Western blotting (WB) was used to measure the protein expression of the related genes, and bioinformatics as well as dual luciferase assays were used to validate the interaction between lncRNA NEAT1 and miR-139 and the interaction between miR-139 and the 3'-UTR of JAK3.Results The expression of lncRNA NEAT1 was increased in the ASMCs of asthma patients, but miR-139 was decreased. Overexpression of lncRNA NEAT1 promoted the expression of the inflammatory cytokines such as TNF-α, IL-6, IL-8 and IL-1ß in ASMCs. LncRNA NEAT1 was able to target miR-139 to activate the JAK3/STAT5 signaling pathway and induced the expression of these inflammatory cytokines in ASMCs. Overexpression of miR-139 or suppression of the JAK3/STAT5 signaling pathway reversed the inflammatory effect of lncRNA NEAT1.Conclusion LncRNA NEAT1 played a pivotal role in ASMC inflammation and exerted its function through the miR-139/JAK3/STAT5 signaling network.

Long non-coding RNA NEAT1 and its targets (microRNA-21 and microRNA-125a) in rheumatoid arthritis: Altered expression and potential to monitor disease activity and treatment outcome.

BACKGROUND: The present study aimed to explore the association of long non-coding RNA nuclear-enriched abundant transcript 1 (lnc-NEAT1) with inflammation, disease activity, treatment outcome, and its targets (microRNA [miR]-21 and miR-125a) in patients with rheumatoid arthritis (RA). METHODS: Peripheral blood mononuclear cells were sampled from 130 RA patients at baseline, week (W) 6, and W12, as well as from 60 healthy controls (HCs) after enrollment. Meanwhile, the expressions of lnc-NEAT1, miR-21, and miR-125a were detected by reverse transcription-quantitative polymerase chain reaction. RESULTS: lnc-NEAT1 was elevated, but miR-21 and miR-125a were declined in RA patients compared with HCs (all p < 0.001); meanwhile, lnc-NEAT1 was negatively correlated with miR-21 and miR-125a (both p < 0.05) in RA patients. Besides, elevated lnc-NEAT1 but declined miR-21 and miR-125a were correlated with erythrocyte sedimentation rate (ESR) and C-reactive protein and the 28-joint Disease Activity-ESR score (all p < 0.05) in RA patients. Moreover, lnc-NEAT1 was declined from baseline to W12 in RA patients (p < 0.001). Additionally, lnc-NEAT1 at W12 was declined in response patients compared with non-response patients (p = 0.006), and also decreased in remission patients compared with non-remission patients (p < 0.001). CONCLUSION: lnc-NEAT1 and its targets (miR-21 and miR-125a) correlate with RA risk and disease activity, and declined lnc-NEAT1 associates with better treatment outcome to some extent in RA patients, suggesting that lnc-NEAT1 might be a potential biomarker to monitor disease activity and treatment outcome in RA.

lncRNA NEAT1 promotes the Taxol resistance of breast cancer via sponging the miR-23a-3p-FOXA1 axis.

Breast cancer is the most prevalent malignancy among women worldwide. Paclitaxel (Taxol) is a widely applied chemotherapeutic agent against breast cancer. Although Taxol therapy has achieved improvements recently, development of chemoresistance of breast cancer patients is a major obstacle, leading to therapeutic failure. Long non-coding RNAs (lncRNAs) play pivotal roles in tumorigenesis and progresses of breast cancer. However, the biological roles and molecular targets of lncRNA NEAT1 in Taxol-resistant breast cancer remain unclear. Here, we report that NEAT1 is significantly upregulated in breast tumors and cell lines. In addition, silencing NEAT1 effectively sensitizes breast cancer cells to Taxol. Bioinformatical analysis and luciferase assay demonstrated that miR-23a-3p could be sponged and downregulated by NEAT1. We demonstrated that miR-23a-3p was downregulated and functioned as a tumor suppressor in breast cancer. Furthermore, in the established Taxol-resistant MDA-MB-231 breast cancer cell line, we detected significantly increased NEAT1 expression and downregulated miR-23a-3p expression. Importantly, FOXA1 was identified and validated as a direct target of miR-23a-3p in breast cancer cells. Rescue experiments demonstrated that the restoration of miR-23a-3p in NEAT1-overexpressing Taxol-resistant breast cancer cells successfully overcame the NEAT1-promoted Taxol resistance. Taken together, our results revealed the clinical roles and molecular mechanisms for the NEAT1-mediated chemoresistance, providing new insights into the development of non-coding RNA-based therapeutic strategies for enhancing the anti-cancer effects of traditional chemotherapeutic drugs.

Baicalin enhances the proliferation and invasion of trophoblasts and suppresses vascular endothelial damage by modulating long non-coding RNA NEAT1/miRNA-205-5p in hypertensive disorder complicating pregnancy.

AIM: Trophoblastic and vascular endothelial injuries were closely associated with the pathogenesis of hypertensive disorder complicating pregnancy (HDCP). The present study was designed to determine the functional role of baicalin in the proliferation and invasion of trophoblasts and vascular endothelial injury. METHODS: Ang II was adopted to stimulate HTR-8/SVneo and human umbilical vein endothelial cells (HUVECs). Cell viability was examined by CCK-8 assay. Flow cytometry and TUNEL staining determined cell apoptosis. Invasive ability of HTR-8/SVneo cells was measured by transwell assay. In vitro angiogenesis of HUVECs was assessed by Tube formation assay. In addition, the production of reactive oxygen species (ROS) was determined by DCFH-DA staining. Furthermore, long non-coding RNA (lncRNA) NEAT1 and miRNA-205-5p levels were detected using real-time quantitative polymerase chain reaction and the binding relationship between lncRNA NEAT1 and miRNA-205-5p was verified by dual-luciferase reporter assay. Moreover, interactions among lncRNA NEAT1, miRNA-205-5p, and MMP9 or vascular endothelial growth factor (VEGF) were confirmed by RNA immunoprecipitation assay. RESULTS: Baicalin visibly improved cell viability, reduced the apoptosis of Ang II-stimulated HTR-8/SVneo and HUVEC cells, and repressed overproduction of ROS. Additionally, baicalin promoted the invasion of Ang II-stimulated HTR-8/SVneo cells and induced a stronger in vitro angiogenesis of Ang II-stimulated HUVECs. What's more, baicalin upregulated lncRNA NEAT1 expression and downregulated miR-205-5p expression. LncRNA NEAT1 sponged miR-205-5p and inhibited the combination of miR-205-5p and MMP9 or VEGF. CONCLUSIONS: Baicalin can facilitate the proliferation and invasion of trophoblasts and alleviate vascular endothelial damage by upregulating lncRNA NEAT1 to impede the interaction between miR-205-5p and MMP9 or VEGF.