lncRNA Maternally Expressed Gene 3 Research Articles

LncRNA MEG3 restrained the M1 polarization of microglia in acute spinal cord injury through the HuR/A20/NF-κB axis.

The M1 polarization of microglia and neuroinflammation restrict the treatment of acute spinal cord injury (ASCI), and long non‐coding ribonucleic acid (lncRNA) maternally expressed gene 3 (MEG3) expression is lessened in ASCI. However, the function and mechanism of lncRNA MEG3 in the M1 polarization of microglia and neuroinflammation in ASCI are unclear. The expressions of lncRNA MEG3 in ASCI mouse spinal cord tissues and lipopolysaccharide (LPS)‐treated primary microglia and BV2 cells were quantified through a quantitative real‐time polymerase chain reaction. In‐vitro assays were conducted to explore the function of lncRNA MEG3 in the M1 polarization of microglia and neuroinflammation in ASCI. RNA degradation, RNA immunoprecipitation, RNA pull‐down, cycloheximide‐chase, and ubiquitination analyses were carried out to probe into the mechanism of lncRNA MEG3 in the M1 polarization of microglia and neuroinflammation in ASCI. The lncRNA MEG3 expression was lessened in the ASCI mouse spinal cord tissues and LPS‐treated primary microglia and BV2 cells, and the overexpression of lncRNA MEG3 restrained the M1 polarization of microglia and the neuroinflammation by regulating the NF‐κB signaling pathway. For the investigation of the potential mechanism of such, the overexpression of lncRNA MEG3 restrained the M1 polarization of microglia through the HuR/A20/NF‐κB axis and boosted the motor function recovery and neuroinflammation relief in the mice with SCI. The overexpression of lncRNA MEG3 restrained the M1 polarization of microglia through the HuR/A20/NF‐κB axis.

LncRNA MEG3 up-regulates SIRT6 by ubiquitinating EZH2 and alleviates nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) is a global health threat. Here, we presented the significant role of a novel signaling axis comprising long non-coding RNA maternally expressed gene 3 (MEG3), enhancer of zeste homolog 2 (EZH2), and sirtuin 6 (SIRT6) in controlling lipid accumulation, inflammation, and the progression of NAFLD. Mice fed with high-fat diet (HFD) were established as in vitro and in vivo NAFLD models, respectively. Lipid accumulation was measured by oil red O staining and assays for triglycerides or cholesterol. Inflammation was examined by ELISA for pro-inflammatory cytokines. Gene expressions were examined by RT-qPCR or Western blot. Interactions between key signaling molecules were examined by combining expressional analysis, RNA immunoprecipitation, cycloheximide stability assay, co-immunoprecipitation, and chromatin immunoprecipitation. MEG3 level was reduced in FFA-challenged hepatocytes or liver from HFD-fed mice, and the reduction paralleled the severity of NAFLD in clinic. Overexpressing MEG3 suppressed FFA-induced lipid accumulation or inflammation in hepatocytes. By promoting the ubiquitination and degradation of EZH2, MEG3 upregulated SIRT6, an EZH2 target. SIRT6 essentially mediated the protective effects of MEG3 in hepatocytes. Consistently, overexpressing MEG3 alleviated HFD-induced NAFLD in vivo. By controlling the expressions of genes involved in lipid metabolism and inflammation, the MEG3/EZH2/SIRT6 axis significantly suppressed lipid accumulation and inflammation in vitro, and NAFLD development in vivo. Therefore, boosting MEG3 level may benefit the treatment of NAFLD.

Downregulation of the LncRNA MEG3 Promotes Osteogenic Differentiation of BMSCs and Bone Repairing by Activating Wnt/β-Catenin Signaling Pathway.

Background: Previous studies have demonstrated that long non-coding RNA maternally expressed gene 3 (MEG3) emerged as a key regulator in development and tumorigenesis. This study aims to investigate the function and mechanism of MEG3 in osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and explores the use of MEG3 in skull defects bone repairing. Methods: Endogenous expression of MEG3 during BMSCs osteogenic differentiation was detected by quantitative real-time polymerase chain reaction (qPCR). MEG3 was knockdown in BMSCs by lentiviral transduction. The proliferation, osteogenic-related genes and proteins expression of MEG3 knockdown BMSCs were assessed by Cell Counting Kit-8 (CCK-8) assay, qPCR, alizarin red and alkaline phosphatase staining. Western blot was used to detect β-catenin expression in MEG3 knockdown BMSCs. Dickkopf 1 (DKK1) was used to block wnt/β-catenin pathway. The osteogenic-related genes and proteins expression of MEG3 knockdown BMSCs after wnt/β-catenin inhibition were assessed by qPCR, alizarin red and alkaline phosphatase staining. MEG3 knockdown BMSCs scaffold with PHMG were implanted in a critical-sized skull defects of rat model. Micro-computed tomography(micro-CT), hematoxylin and eosin staining and immunohistochemistry were performed to evaluate the bone repairing. Results: Endogenous expression of MEG3 was increased during osteogenic differentiation of BMSCs. Downregulation of MEG3 could promote osteogenic differentiation of BMSCs in vitro. Notably, a further mechanism study revealed that MEG3 knockdown could activate Wnt/β-catenin signaling pathway in BMSCs. Wnt/β-catenin inhibition would impair MEG3-induced osteogenic differentiation of BMSCs. By using poly (3-hydroxybutyrate-co-3-hydroxyhexanoate, PHBHHx)-mesoporous bioactive glass (PHMG) scaffold with MEG3 knockdown BMSCs, we found that downregulation of MEG3 in BMSCs could accelerate bone repairing in a critical-sized skull defects rat model. Conclusions: Our study reveals the important role of MEG3 during osteogenic differentiation and bone regeneration. Thus, MEG3 engineered BMSCs may be effective potential therapeutic targets for skull defects.

Elevated Expression of lncRNA MEG3 Induces Endothelial Dysfunction on HUVECs of IVF Born Offspring via Epigenetic Regulation.

Cardiovascular dysfunction in children born after in vitro fertilization (IVF) has been of great concern, the potential molecular mechanisms for such long-term outcomes are still unknown. Here, we found that systolic blood pressure was a little higher in IVF born offspring at 2 years old compared to those born after being naturally conceived. Besides, the expression level of maternally expressed gene 3 (MEG3) was higher in human umbilical vein endothelial cells (HUVECs) from IVF offspring than that in spontaneously born offspring. Pearson correlation test showed that MEG3 relative expression is significantly related to the children's blood pressure (Coefficient = 0.429, P = 0.0262). Furthermore, we found decreased expression of endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) along with elevated expression of endothelial-1(ET1) in HUVECs from IVF offspring, accompanied by lower secretion of nitrite, VEGF, and higher secretion of ET1 in the umbilical cord serum of IVF offspring. Correlation analysis showed MEG3 expression highly correlated with ET1 and Nitrate concentration. With pyrosequencing technology, we found that elevated expression of MEG3 was the result of hypomethylation of the MEG3 promoter. Therefore, our results provide a potential mechanism addressing the high-risk of hypertension in IVF offspring via MEG3 epigenetic regulation.

Changes in Serum LncRNA MEG3/miR-181b and UCH-L1 Levels in Patients with Moderate and Severe Intracerebral Hemorrhage.

To study on the association of lnc-MEG3/miR-181b and UCH-L1 in the progression of cerebral hemorrhage. Further facilitation the development of novel strategies that use their potential as a therapeutic targets. In this study, we studied the lncRNA MEG3 and miR-181b expression in 30 patients with ICH who were admitted to the General Hospital of Xinjiang Military Command from January 2021 to May 2021 by quantitative polymerase chain reaction. Serum levels of UCH-L1 were detected by enzyme-linked immune sorbent assay, and disease severity was evaluated using the Glasgow Coma Scale. We also recorded ICH-related deaths in hospital. We found that lnc-MEG3 and UCH-L1 levels increased and miR-181b levels decreased in the serum of patients with ICH. lnc-MEG3, miR-181b, and UCH-L1 levels were also associated with the severity of the condition. Our data indicated that lnc-MEG3, miR-181b, and UCH-L1 are likely involved in the pathophysiology of ICH, and could form the basis of future studies on potential targets for the treatment of traumatic CNS injuries. UCH-L1 could also find application in ICH management. Further studies on the plasma lncRNA-MEG3, miR-181b, and UCH-L1 levels in patients with ICH could yield novel biological targets for the prediction and treatment of ICH.

Long noncoding RNA matrilineal expression gene 3 inhibits hepatocellular carcinoma progression by targeting microRNA-5195-3p and regulating the expression of forkhead box O1

ABSTRACT We investigated the effect of the long noncoding RNA (lncRNA) maternally expressed gene 3 (MEG3) on hepatocellular carcinoma (HCC) tumorigenesis and progression by targeting miR-5195-3p and transcription factor forkhead box O1 (FOXO1) to identify a novel target for HCC treatment. HCC clinical samples were collected, and cell counting kit-8 (CCK-8), and transwell migration and invasion assays were performed. Furthermore, interaction was detected via double luciferase reporter and RNA pull-down assays. MEG3, miR-5195-3p, and FOXO1 expression was determined by quantitative real-time polymerase chain reaction (RT-qPCR) and Western blotting. Xenograft tumor models were established to investigate the effect of MEG3 in vivo. Compared with normal tissues, MEG3 expression was significantly downregulated in HCC tissues. MEG3 overexpression inhibited the viability and migration of HCC cells. Double luciferase reporter and RNA pull-down assays confirmed the binding between MEG3 and miR-5195-3p as well as between miR-5195-3p and FOXO1. RT-qPCR and Western blotting results showed that MEG3 inhibited the expression of miR-5195-3p and promoted that of FOXO1. Additionally, MEG3 overexpression inhibited HCC tumorigenesis and progression in xenograft tumor models while depletion of MEG3 exerted the opposite way. Therefore, the lncRNA MEG3 inhibits HCC tumorigenesis and progression through the miR-5195-3p/FOXO1 signaling axis.

LncRNA MEG3 modulates hepatic stellate cell activation by sponging miR‑145 to regulate PPARγ.

It is important to determine the mechanism of liver fibrosis for targeted therapy and the development of targeted therapies for liver fibrosis may offer promise for patients with liver disease. Long non-coding RNAs (lncRNAs) serve a role in hepatic fibrosis. The lncRNA maternally expressed gene 3 (MEG3) has been confirmed to inhibit liver fibrosis. The present study investigated the role of the MEG3 in healthy patients and patients with liver fibrosis. The expression levels of MEG3 and microRNA (miR)-145 in the serum of healthy volunteers and patients with liver fibrosis and in LX-2 cells were detected using reverse transcription-quantitative PCR. A dual-luciferase reporter assay was used to determine the targeting relationship between MEG3 and miR-145, and the targeting relationship between miR-145 and peroxisome proliferator-activated receptor γ (PPARγ). The protein expression levels of PPARγ, α-smooth muscle actin (α-SMA) and collagen I (COL1A1) were detected using western blotting. The expression levels of α-SMA and COL1A1 were also determined using immunofluorescence. Finally, a Cell Counting Kit-8 assay was performed to assess the proliferative ability of LX-2 cells. A significantly reduced MEG3 expression level was demonstrated in serum from patients with liver fibrosis compared with serum from healthy controls. TGF-β1 induced a significantly decreased MEG3 expression level in LX-2 human hepatic stellate cells in vitro. The TGF-β1-induced increases in cell proliferation and α-SMA and COL1A1 protein expression levels were reversed following MEG3 overexpression. The results also demonstrated that MEG3 sponged miR-145 and competed endogenously with miR-145 to regulate PPARγ. In summary, the present study identified MEG3 as an anti-fibrotic lncRNA and provided new information regarding the role of MEG3 in liver fibrosis. MEG3 may therefore be a potential target in the treatment of liver fibrosis.

LncRNA MEG3 reverses CCl4-induced liver fibrosis by targeting NLRC5

Liver fibrosis is a persistent pathological repair of chronic liver injury, which is characterized by excessive deposition of collagen-dominated extracellular matrix (ECM). It is well known that hepatic fibrosis can be reversed in the absence of etiology. Studies have shown that long non-coding RNA (Lnc RNA) maternally expressed gene3 (MEG3) has strong effects on the activation of hepatic stellata cells (HSCs). However, the function of MEG3 in the reversal of liver fibrosis has not been studied. In this experiment, we studied the content expression, function, and part of the potential mechanism of MEG3 in reversing liver fibrosis. In in vivo and in vitro models, we found that MEG3 was down-regulated during the formation of liver fibrosis, while it was up-regulated during the reversal of liver fibrosis. Then, it was found that the silencing of MEG3 could gradually restore the activity of the inactivated LX-2 cells, Overexpression of MEG3 can inhibit the activation of LX-2 cells, accelerate the reversal of liver fibrosis. Through catRAPID analysis, it was found that NLR family CARD domain containing 5 (NLRC5) may be a target of MEG3. We found that, after MEG3 silencing, NLRC5 expression was upregulated in LX-2 cells in the reverse phase, while, after MEG3 overexpression, NLRC5 expression was decreased. Further, we verified that MEG3 can target NLRC5 through RNA pull down experiment. Therefore, MEG3 may inhibit the activation of hepatic stellate cells by targeting NLRC5, thus accelerating the reversal of hepatic fibrosis.

LncRNA MEG3, Acting as a ceRNA, Modulates RPE Differentiation Through the miR-7-5p/Pax6 Axis.

Accumulated evidence indicated that long non-coding RNAs (lncRNAs) involves in numerous biological and pathological processes, including age-related macular degeneration (AMD). Dysfunction and dedifferentiation of retinal pigment epithelium (RPE) cells have been demonstrated to be one of the crucial factor in AMD etiology. Herein, we aim to investigate the essential role of lncRNA maternally expressed gene 3 (MEG3) in AMD progression. Expression patterns of MEG3 were measured in dysfunctional REP cells exposed with H2O2 or TNF-α using qRT-PCR assay. Specifically, the intercellular distribution of MEG3 in REP cells was further explored using the subcellular fraction detection. Relative expression of RPE markers or RPE dedifferentiation-related markers was determined using qRT-PCR and western blot analysis, respectively. Immunofluorescence staining was performed to examine the expressions of RPE markers ZO-1 and β-catenin. Concentration of vascular endothelial growth factor (VEGFA) in the supernatant was detected using ELISA kit. Luciferase reporter assay was performed to verify the MEG3/miR-7-5p/Pax6 regulatory network, which was further determined in in vitro studies. MEG3 expression was significantly decreased in H2O2 or TNF-α-treated REP cells, and it was upregulated along with RPE differentiation. Reduced MEG3 expression resulted in RPE dedifferentiation, which was indicated by decreased expressions of RPE markers, accumulated mitochondrial reactive oxygen species, and reduced VEGFA. Mechanistically, MEG3 functioned as a sponge for miR-7-5p to restore the expression of Pax6. Our study demonstrated that MEG3 exerts a protective role against AMD by maintaining RPE differentiation via miR-7-5p/Pax6 axis, suggesting a protective therapeutic target in AMD treatment.

Long Non-coding RNA MEG3 Promotes Renal Tubular Epithelial Cell Pyroptosis by Regulating the miR-18a-3p/GSDMD Pathway in Lipopolysaccharide-Induced Acute Kidney Injury.

Background and Objective: Acute kidney injury (AKI) is a complication of sepsis. Pyroptosis of gasdermin D (GSDMD)-mediated tubular epithelial cells (TECs) play important roles in pathogenesis of sepsis-associated AKI. Long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3), an imprinted gene involved in tumorigenesis, is implicated in pyroptosis occurring in multiple organs. Herein, we investigated the role and mechanisms of MEG3 in regulation of TEC pyroptosis in lipopolysaccharide (LPS)-induced AKI.Materials and Methods: Male C57BL/6 mice and primary human TECs were treated with LPS for 24 h to establish the animal and cell models, respectively, of sepsis-induced AKI. Renal function was assessed by evaluation of serum creatinine and urea levels. Renal tubule injury score was assessed by Periodic acid-Schiff staining. Renal pyroptosis was assessed by evaluating expression of caspase-1, GSDMD, and inflammatory factors IL-1β and IL-18. Cellular pyroptosis was assessed by analyzing the release rate of LDH, expression of IL-1β, IL-18, caspase-1, and GSDMD, and using EtBr and EthD2 staining. MEG3 expression in renal tissues and cells was detected using RT-qPCR. The molecular mechanisms of MEG3 in LPS-induced AKI were assessed through bioinformatics analysis, RNA-binding protein immunoprecipitation, dual luciferase reporter gene assays, and a rescue experiment.Results: Pyroptosis was detected in both LPS-induced animal and cell models, and the expression of MEG3 in these models was significantly up-regulated. MEG3-knockdown TECs treated with LPS showed a decreased number of pyroptotic cells, down-regulated secretion of LDH, IL-1β, and IL-18, and decreased expression of GSDMD, compared with those of controls; however, there was no difference in the expression of caspase-1 between MEG3 knockdown cells and controls. Bioinformatics analysis screened out miR-18a-3P, and further experiments demonstrated that MEG3 controls GSDMD expression by acting as a ceRNA for miR-18a-3P to promote TECs pyroptosis.Conclusion: Our study demonstrates that lncRNA MEG3 promoted renal tubular epithelial pyroptosis by regulating the miR-18a-3p/GSDMD pathway in LPS-induced AKI.

LncRNA MEG3 Involved in NiO NPs-Induced Pulmonary Fibrosis via Regulating TGF-β1-Mediated PI3K/AKT Pathway.

Long noncoding RNA maternally expressed gene 3 (MEG3) involves in fibrotic diseases, but its role in nickel oxide nanoparticles (NiO NPs)-induced pulmonary fibrosis remains unclear. The present study aimed to explore the relationships among MEG3, transforming growth factor-β1 (TGF-β1) and phosphoinositide 3-kinase (PI3K)/AKT pathway in NiO NPs-induced pulmonary fibrosis. Wistar rats were intratracheally instilled with NiO NPs twice a week for 9 weeks, and human lung adenocarcinoma epithelial cells (A549 cells) were exposed to NiO NPs for 24 h. The pathological alterations and increased hydroxyproline indicated that NiO NPs caused pulmonary fibrosis in rats. The up-regulated type I collagen (Col-I) suggested that NiO NPs-induced collagen deposition in A549 cells. Meanwhile, NiO NPs could significantly down-regulate MEG3, up-regulate TGF-β1 and activate PI3K/AKT signaling pathway both in vivo and in vitro. However, we found that the PI3K/AKT pathway activated by NiO NPs could be suppressed by 10 μM TGF-β1 inhibitor (SB431542) in A549 cells. The protein markers (Col-I, Fibronectin, and alpha-smooth muscle actin) of collagen deposition up-regulated by NiO NPs were reduced by 10 μM PI3K inhibitor (LY294002). Furthermore, we further found that overexpressed MEG3 inhibited the expression of TGF-β1, resulting in the inactivation of PI3K/AKT pathway and the reduction of collagen formation. In summary, our results validated that MEG3 could arrest NiO NPs-induced pulmonary fibrosis via inhibiting TGF-β1-mediated PI3K/AKT pathway.

LncRNA MEG3 Downregulation Relieves Intracerebral Hemorrhage by Inhibiting Oxidative Stress and Inflammation in an miR-181b-Dependent Manner.

BackgroundThis study was designed to illustrate the effects and latent mechanism of lncRNA maternally expressed gene 3 (MEG3) on intracerebral hemorrhage (ICH)-induced brain injury.Material/MethodsAn ICH rat model was generated to determine the role of lncRNA MEG3 in ICH. The interaction between lncRNA MEG3 and microRNA (miR)-181b were confirmed by Starbase and dual-luciferase reporter assay. One hour (h) or 3 days after ICH stimulation, rat neurological injury was evaluated by modified Neurological Severity Score (mNSS). Brain water content and cell apoptosis were assessed using brain edema assessment and flow cytometry (FCM), respectively. Caspase3 activity was also determined. Enzyme-linked immunosorbent assay (ELISA) was applied to evaluate the levels of pro-inflammatory cytokines. Moreover, the representative biomarkers of oxidative stress were evidenced using detection kits.ResultsThe lncRNA MEG3 level in ICH rat brain tissues was higher than that in the sham group. miR-181b was a direct target of lncRNA MEG3 and it was downregulated in brain tissues of ICH rats. Notably, we found that neurobehavioral scores, brain water content, and neuronal apoptosis were decreased and caspase3 activity was reduced in MEG3-shRNA-treated ICH rats, while we observed the opposite result in ICH+MEG3-shRNA+miR-181b inhibitor rats. Further analyses revealed that MEG3-shRNA inhibited inflammatory cytokines release and reduced oxidative stress. All these results were reversed by miR-181b inhibitor. In addition, MEG3-shRNA activated the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway, which was reversed by miR-181b inhibitor.ConclusionsMEG3-shRNA restrained oxidative stress and inflammation following ICH in an miR-181b-dependent manner.

Long non-coding RNA maternally expressed gene 3 affects cell proliferation, apoptosis and migration by targeting the microRNA-9-5p/midkine axis and activating the phosphoinositide-dependent kinase/AKT pathway in hepatocellular carcinoma.

Long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) is a tumor suppressor in several cancers, such as glioma, prostate cancer and esophageal cancer. However, the role of MEG3 in hepatocellular carcinoma (HCC) and the related molecular mechanisms are not well understood. The present study aimed to determine the biological function of MEG3 in regulating HCC cell viability, apoptosis and migration. In addition, the interaction between MEG3, microRNA (miR)-9-5p and Midkine (MDK), and the activation of the phosphoinositide-dependent kinase (PDK)/AKT pathway in HCC cell line MHCC-97L were examined. Luciferase reporter assays, reverse transcription-quantitative PCR and western blotting were used to determine the interaction between MEG3, miR-9-5p and MDK and the activation of the PDK/AKT pathway. Cell viability was determined by the CCK8 assay and the cell cycle analysis using flow cytometry analysis. Cell apoptosis was examined by flow cytometry analysis and caspase 3/9 activity. Wound healing assays and western blotting were used to investigate cell migration. The present study demonstrated that MEG3 suppressed HCC cell viability and migration, and induced cell apoptosis. In addition, it was also found that MEG3 targets the miR-9-5p/MDK axis and modulates the PDK/AKT pathway in HCC. In conclusion, the findings of the present study demonstrated that lncRNA MEG3 affects HCC cell viability, apoptosis and migration through its targeting of miR-9-5p/MDK and regulation of the PDK/AKT pathway. The MEG3/miR-9-5p/MDK axis may be a potential therapeutic target in HCC.

Long Noncoding RNA Maternally Expressed Gene 3 Targets miR-30b and Regulates the AKT Serine/Threonine Kinase 1/Phosphoinositide 3-Kinase Signaling Pathway of H2O2-Induced Proliferation, Apoptosis, and Oxidative Stress in Retinal Ganglion Cells

Oxidative stress is an important factor affecting retinal ganglion cell (RGC) apoptosis. RGC apoptosis is the main pathophysiological feature of visual impairment as a result of glaucoma. Recently, it has been found that long noncoding RNA (lncRNA) and microRNAs are involved in RGC apoptosis. Here, the function of lncRNA maternally expressed gene 3 (MEG3) and miR-30b in H2 O2-induced RGC proliferation, apoptosis, and oxidative stress was investigated. The expression levels of MEG3 and miR-30b were detected by RT-PCR; the effects of MEG3 and miR-30b on the proliferation and apoptosis of RGCs were observed by flow cytometry; the levels of apoptosis-related proteins and AKT/PI3K signal pathway proteins were detected by protein immunoassay; and the regulation of miR-34a by pvt1 was verified by in vivo and in vitro experiments. The expression of MEG3 and miR-30b increased and decreased significantly in RGCs treated by H2O2. MEG3 expression decreased, apoptosis level-related proteins decreased, the apoptosis rate reduced, and the activity of MDA and SOD decreased. When the expression of miR-34a was inhibited, the proliferation rate of RGCs increased, the apoptosis rate decreased, and the level of apoptosis-related proteins decreased, which reversed MEG3’s effect on RGC apoptosis and proliferation. Furthermore, pvt1 could bind the miR-30b promoter and regulate it with in vitro expression and in vivo expression. Besides, we found that miR-30b can regulate the AKT/PI3K signaling pathway and participate in cell apoptosis and hyperplasia in stress response. LncRNA MEG3 targets miR-30b and regulates the AKT/PI3K signaling pathway on H2 O2-induced cell apoptosis, hyperplasia, and oxidative stress of RGCs.

Implication of LncRNAs MEG3 and LINC00305 in Pathophysiological Mechanisms associated with Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a joint destructive disorder with great morbidity. Early diagnosis and early and effective therapy may prevent joint damage and lead to better long-term results. Therefore, reliable biomarkers and outcome measures are needed. Complex interplay between multiple risk factors contributes to RA. These risk factors include environmental factors, genetic factors and epigenetic modifications. Long non-coding RNAs (lncRNAs) are involved in these epigenetic mechanisms. Accumulating evidence has shown that lncRNAs participate in the processes of inflammation, aberrant proliferation, apoptosis and angiogenesis. They also play roles in autoimmune diseases, such as SLE, Sjogren syndrome, RA and multiple sclerosis. Maternally expressed gene 3 (MEG3) is a maternally expressed lncRNA of the imprinted DLK1–MEG3 locus located on human 14q32 chromosome. It functions as a tumor suppressor. Decreased MEG3 expression has been observed in various human cancers, both type 1 and type 2 diabetes, osteoarthritis and in RA. Decreased serum levels of lncRNA MEG3 in RA could be attributed to MEG3 promoter hypermethylation induced by hypoxia. This downregulation was associated with increased inflammation, cell proliferation and cell invasion and decreased apoptosis. Long intergenic non-Protein coding RNA 00305 (LINC00305) was identified as a pro-inflammatory atherosclerosis-associated lncRNA. RA was associated with increased LINC00305 which served as a regulator of inflammatory, hypoxic, invasive, apoptotic and proliferative mechanisms associated with RA.

The lncRNA MEG3/miR-16-5p/VGLL4 regulatory axis is involved in etoposide-induced senescence of tumor cells.

The senescence of tumor cells is an important tumor suppressor mechanism. The present study aimed to investigate the role of long non-coding RNA (lncRNA) MEG3 (maternally expressed gene 3) in the senescence process of tumor cells and its potential molecular mechanism by competitively binding with microRNA miR-16-5p to regulate the expression of VGLL4 (encoding vestigial like family member 4). We used etoposide to construct senescence models of tumor cells. The degree of cellular senescence was detected by senescence-associated β-galactosidase, cell cycle and senescence-associated secretory phenotype. The expression of lncRNA MEG3, miR-16-5p and VGLL4 in senescent or non-senescent cells was evaluated using a quantitative real-time reverse transcriptase-PCR (qRT-PCR) or western blotting. Dual luciferase reporter assays were used to detect the binding of miR-16-5p to lncRNA MEG3 and VGLL4. The mRNA and protein expression levels of senescence-related markers (p53, p21 and p16) were detected using qRT-PCR or western blotting. Compared to the control group, the expression of lncRNA MEG3 and VGLL4 was significantly up-regulated in senescent cells. Knockdown of lncRNA MEG3 and VGLL4 reduced the degree of senescence and the expression of p21 and p16. lncRNA MEG3 interfered with the expression of miR-16-5p in senescent A549 and MCF-7 cells. The expression of VGLL4 was regulated by miR-16-5p in senescent A549 and MCF-7 cells. lncRNA MEG3 participated in the senescent progress of tumor cells induced by etoposide via the miR-16-5p/VGLL4 axis. The present study has confirmed the regulatory role of the lncRNA MEG3/miR-16-5p/VGLL4 axis in the low-dose etoposide-induced tumor cell senescence model, which has potential clinical application with respect to treating malignant tumors.

DNMT1-mediated lncRNA MEG3 methylation accelerates endothelial-mesenchymal transition in diabetic retinopathy through the PI3K/Akt/mTOR signaling pathway.

Diabetic retinopathy (DR) is one of the serious complications that occurs in diabetic patients that frequently causes blindness. Long noncoding RNAs (lncRNAs) have been associated with DR pathology. This study aimed to determine the underlying mechanism of lncRNA maternally expressed gene 3 (MEG3) in association with DNA methyltransferase 1 (DNMT1) in the endothelial-mesenchymal transition (endMT) that occurs in DR. A rat model of DR was induced by streptozotocin (STZ) injection, and a high-glucose (HG)-induced cell model was established by exposing microvascular endothelial cells obtained from retina of rats to HG. Subsequently, MEG3 was overexpressed in rat and cell models to characterize its impact on endMT in DR and the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway. Furthermore, the methylation level of MEG3 promoter region was determined with the application of methylation-specific polymerase chain reaction, followed by chromatin immunoprecipitation assay for methyltransferase enrichment. Finally, we examined the regulation of DNMT1 on MEG3 methylation and endMT in the HG-induced cell model. The results obtained revealed downregulated MEG3 expression in DR rat and cell models. Overexpressed MEG3 was shown to suppress endMT in DR rat and cell models through the inhibition of the PI3K/Akt/mTOR signaling pathway. Notably, DNMT1 could promote MEG3 promoter methylation to inhibit MEG3 expression by recruiting methyltransferase, which activated the PI3K/Akt/mTOR signaling pathway to accelerate endMT in DR. These findings further highlighted the inhibitory effect of MEG3 on endMT in DR, thus presenting a novel therapeutic target candidate for DR treatment.

Association of the Plasma Long Non-coding RNA MEG3 With Parkinson's Disease.

Objective: To investigate the expression level of the maternally expressed gene-3 (MEG3) of the free long non-coding RNA (lncRNAs) in the plasma of Parkinson's disease (PD) patients and its relationship with the disease.Methods: Thirty PD patients (PD group) who treated at Xuanwu Hospital of Capital University of Medical Sciences between January 2017 and December 2019 were selected as the research objects and 30 healthy subjects were enrolled in the study during the same period as the control group. Cognitive function was assessed according to the Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) were used to evaluate cognitive function, Non-Motor Symptoms Scale (NMSS) was used to evaluate severity of non-motor symptoms. The relative expression of lncRNAs MEG3 in plasma was measured by PCR, and the levels of neuron-specific enolase (NSE), nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) in plasma were measured by ELISA, and the relationship with these all indexes was analyzed.Results: The NMSS score of PD group was significantly higher than that of the control group, while the MMSE and MoCA scores were significantly lower than that of the control group (P < 0.05); The relative expression of lncRNAs MEG3, NGF and BDNF levels of PD group were significantly lower than that of the control group, and NSE level was significantly higher than that of the control group (P < 0.05); The H&Y stage and NMSS score in PD group were negatively correlated with the relative expression of lncRNAs MEG3, the levels of NGF and BDNF (P < 0.05), and positively correlated with NSE (P < 0.05); The MMSE and MoCA scores in PD group were positively correlated with the relative expression of lncRNAs MEG3, NGF, BDNF levels (P < 0.05), and negatively correlated with NSE (P < 0.05); The relative expression of lncRNAs MEG3 in PD group was positively correlated with NGF, BDNF levels (P < 0.05), and negatively correlated with NSE (P < 0.05).Conclusion: The expression of lncRNAs MEG3 in the plasma of PD patients was downregulated compared to that of healthy control subjects, and its expression level was closely related to the aggravation of non-motor symptoms, cognitive decline, and PD stage. These associations may reflect the synergism of the increase of NSE and decrease of NGF and BDNF levels, highlighting plasma lncRNA MEG3 as a new candidate biomarker of PD.

Vitamin D ameliorated endothelial cell damage induced by diabetes via regulation of lncRNA MEG3 in human umbilical vein endothelial cells

IntroductionThe purpose of this study was to investigate long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) effects in vitamin D (Vit. D) treatment in endothelial cell damage induced by diabetes.Material and methodsWe used human umbilical vein endothelial cells (HUVECs) as a research objective in our study and used high glucose in a diabetic cell model. We evaluated cell apoptosis by flow cytometry, inflammatory factors (IL-6, IL-1β and TNF-α) concentrations by ELISA assay, relative gene and protein expression by RT-qPCR and WB assay, and NF-κB(p65) nuclear volume by cellular immunofluorescence.ResultsCompared with the NC (normal control) group, the cell apoptosis rate was significantly increased, inflammatory factor (IL-6, IL-1β and TNF-α) concentrations were significantly up-regulated, lncRNA MEG3 gene expression was significantly depressed, Toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear factor B p65 (NF-κB(p65)) gene and protein expression levels were significantly increased and NF-κB(p65) nuclear volume was significantly up-regulated (p &lt; 0.001, respectively). With Vit. D supplementation, compared with the Model group, Vit. D improved endothelial cell damage induced by diabetes, while lncRNA MEG3 was significantly increased and the TLR4/MyD88/NF-κB(p65) pathway was significantly depressed dose-dependently (all p &lt; 0.05). With sh-MEG3 transfection, the Vit. D treatment effects were significantly reduced.ConclusionsVit. D improved endothelial cell damage induced by diabetes via lncRNA MEG3 up-regulation in vitro study.

Long non-coding RNA maternally expressed gene regulates cigarette smoke extract induced lung inflammation and human bronchial epithelial apoptosis via miR-149-3p.

Chronic obstructive pulmonary disease (COPD) has become a significant public health risk. Long non-coding RNAs (lncRNAs) have been identified as important factors involved in the proliferation, apoptosis and inflammatory cytokine expression of lung cells. Peripheral blood samples from 66 subjects (18 non-smokers, 24 smokers without COPD and 28 smokers with COPD) and HBE135-E6E7 cell treated with cigarette smoke extract (CSE) or not were used as the research object. The aim of the present study was to investigate the underlying mechanism of lncRNA maternally expressed gene 3 (MEG3) in COPD. Following transfection with microRNA (miR)-149-3p mimics, miR-negative control mimics, miR-149-3p inhibitor, miR-negative control inhibitor, small interfering (si)RNA targeting MEG3 (si-MEG3) and si-negative control (si-NC), levels of MEG3 and microRNA (miR)-149-3p were detected using reverse transcription-quantitative PCR, Proliferation and apoptosis were examined using the Cell Counting Kit-8 and flow cytometry assays, respectively. Enzyme-linked immunosorbent assay (ELISA) was performed to detect the expression of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). Protein levels of B-cell lymphoma-2 (Bcl-2), cleaved-caspase-3, cleaved-caspase-9, phosphorylated (p)-p65, total (t)-p65, p-lkBα and t-lkBα were measured by western blotting. Luciferase assay was conducted to examine the relationship between MEG3 and miR-149-3p. LncRNA MEG3 was highly expressed, whereas miR-149-3p expression was downregulated in smokers with COPD peripheral blood samples, compared with non-smokers and smokers without COPD samples. Compared with untreated human bronchial epithelial (HBE) cells, MEG3 expression was increased in cigarette smoke extract (CSE)-treated HBE cells. Compared with CSE-treated HBE cells transfected with si-NC, MEG3 knockdown promoted cell proliferation and inhibited apoptosis in CSE-treated HBE cells transfected with si-MEG3, and it also decreased the levels of IL-6, TNF-α, Bcl-2 and increased cleaved-caspase-3 and cleaved-caspase-9 in CSE-treated HBE cells transfected with si-MEG3. The luciferase assay demonstrated that miR-149-3p has target sites for MEG3. MEG3 was demonstrated to regulate the NF-κB signaling pathway by sponging miR-149-3p in CSE-treated HBE cells. In conclusion, these findings suggested that MEG3 promoted proliferation and inhibited apoptosis by regulating the NF-κB signal pathway via miR-149-3p in CSE-treated HBE cells. These results provide an insight for further verification and understanding of the molecular basis of COPD.