Effect Of Long Non-coding RNA Research Articles

Overexpression of lncRNA HULC Attenuates Myocardial Ischemia/reperfusion Injury in Rat Models and Apoptosis of Hypoxia/reoxygenation Cardiomyocytes via Targeting miR-377-5p through NLRP3/Caspase‑1/IL‑1β Signaling Pathway Inhibition

ABSTRACT Objective Acute myocardial infarction (AMI) is characterized by myocardial tissue necrosis and activation of inflammatory response. This study aims to elucidate the potential mechanism underlying the protective effects of long non-coding RNA (lncRNA) highly up-regulated in liver cancer (HULC) against myocardial ischemia/reperfusion (I/R) injury in rat models and apoptosis of cardiomyocytes. Methods We firstly established rat models of myocardial I/R injury and rat cardiomyocyte (H9c2 cells) models of hypoxia/reoxygenation (H/R) injury. Sprague-Dawley (SD) neonatal rats were randomized into four groups: sham, I/R, I/R+ microRNA (miR) −377-5p mimic, and I/R+ miR-377-5p antagomir, respectively. Then, histopathological examination was applied. Apoptosis was evaluated by transferase-mediated dUTP nick end labeling (TUNEL) staining. Cell vitality was measured using MTT assay. The concentrations of creatine kinase MB (CK-MB), cardiac troponin I (cTnI), interleukin (IL) −6 (IL-6), and tumor necrosis factor-α (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). The expression of Cleaved-Caspase-3, Caspase-3, NOD-like receptor P3 (NLRP3), Caspase-1, and IL-1β was analyzed by immunohistochemical (IHC) or Western blot analysis. Results We found that HULC was downregulated and miR-377-5p was upregulated in IR-injured myocardial tissue and the H/R-induced H9c2 cell. Overexpression of miR-377-5p increased myocardial dysfunction and apoptosis and activated formation and secretion of IL-6 and TNF-α. The preprocessing of miR-377-5p silencing emerged opposite results. Strikingly, dual luciferase reporter assay showed that HULC was a sponge of miR-377-5p. Subsequently, mechanism experiments revealed that NLRP3/Caspase‑1/IL‑1β was a target axis of miR-377-5p. In vitro, the protective effect of HULC overexpression on H9c2 cell viability and inflammation was offset by miR-377-5p silencing. Finally, rescue assay suggested that HULC-miR-377-5p -NLRP3/Caspase‑1/IL‑1β axis regulated the apoptosis and inflammation of H/R-induced H9c2 cells. Conclusions Overall, these results indicate that the protective effect of HULC against myocardial I/R injury and H/R cardiomyocyte apoptosis partially relies on the inhibition of NLRP3/Caspase‑1/IL‑1β signaling pathway.

Silencing long non-coding RNA H19 combined with paclitaxel inhibits nasopharyngeal carcinoma progression

PurposeThis study aimed to study the effect of long non-coding RNA (lncRNA) H19 on proliferation, apoptosis and chemosensitivity of nasopharyngeal carcinoma (NPC) cells. MethodsNP69 and HNE3, C666-1, SUNE1, 6–10B and 5–8F cell lines were selected to detect the expression of lncRNA H19 via RT-qPCR. LncRNA H19 was overexpressed or silenced for exploring the regulatory effect of lncRNA H19 in cell proliferation, clone formation, apoptosis and drug resistance through CCK-8, clone formation experiment and flow cytometry respectively. The tumorigenic effect of lncRNA H19 silencing was verified by xenograft tumor in nude mice. LncRNA H19 was significantly up-regulated in NPC cells. ResultsSilencing lncRNA H19 inhibited the proliferation of NPC C666-1 cells and promoted apoptosis, while overexpression of lncRNA H19 promoted the proliferation of NPC C666-1 cells and inhibited apoptosis. Knockdown of lncRNA H19 in drug-resistant cells remarkably reduced their drug resistance, and overexpression of lncRNA H19 in parental cells significantly reduced their drug sensitivity. Silencing lncRNA H19 inhibits tumor growth in vivo, and silencing lncRNA H19 combined with paclitaxel can enhance tumor inhibition in vivo. ConclusionsIn NPC cells, lncRNA H19 was up-regulated, lncRNA H19 inhibited the proliferation and chemosensitivity of NPC cells, promoted apoptosis, and silencing lncRNA H19 combined with paclitaxel could enhance tumor inhibition in vivo.

Metformin inhibits cervical cancer cell proliferation by modulating PI3K/Akt-induced major histocompatibility complex class I-related chain A gene expression

BackgroundRecent studies have shown that the classic hypoglycemic drug metformin inhibits tumor growth; however, the underlying mechanism remains unclear. We previously showed that metformin disrupts the sponge effect of long non-coding RNA MALAT1/miR-142-3p to inhibit cervical cancer cell proliferation. In this study, we interrogated the ability of metformin to modulate the anti-tumor immune response in cervical cancer.MethodsThe cell counting kit-8 assay was used to detect the viability of cervical cancer cells. Flow cytometry assays were performed to measure cell apoptosis and cell cycle. Lactate dehydrogenase (LDH) cytotoxicity assay was used to detect NK Cell Cytotoxicity. Relative protein levels were determined by immunoblotting and relative gene levels were determined by quantitative real-time PCR. Tumor Xenograft Modeling was used to evaluate the effect of metformin in vivo.ResultsMetformin inhibited cervical cancer cell proliferation, cervical cancer xenograft growth, expression of PCNA, p-PI3K and p-Akt. Moreover metformin induced cervical cancer cell apoptosis and caused cancer cell cycle arrest. In addition, metformin upregulated the expression of DDR-1 and p53 in human cervical cancer cells. Furthermore, metformin also regulated the mRNA and protein expression of MICA and HSP70 on the surface of human cervical cancer cells via the PI3K/Akt pathway, enhancing NK cell cytotoxicity.ConclusionsIn conclusion, our results suggest that metformin may be used as immunopotentiator to inhibit cervical cancer progression and may be considered a viable candidate for combination therapy with immunotherapy.

Regulation of lncRNA-H19/miR-140-5p in cartilage matrix degradation and calcification in osteoarthritis.

To study the effect of long non-coding RNA H19/microRNA (miR)-140-5p regulatory axis on the degradation and calcification of osteoarthritis (OA) cartilage matrix. Twenty cases of clinical OA cartilage tissue and healthy cartilage tissue were collected. Construction of small interfering RNA vector (si-H19) and negative control (si-Control), miR-140-5p mimic and negative control mimic-NC, miR-140-5p inhibitor, and negative control inhibitor-NC transfected human chondrocytes HC-A cells. Chondrocytes were categorized into classes of si-H19, si-Control, mimic, mimic-NC, inhibitor, NC inhibitor, inhibitor + si-H19, NC inhibitor + si-H19. The expression levels of H19, miR-140-5p, alkaline phosphatase (ALP), osteocalcin (OCN), and bone sialoprotein (BSP) in cartilage tissue and chondrocytes were detected using real-time fluorescent quantitative PCR (RT-qPCR). In each group of chondrocytes, Western blotting was used to detect the levels of matrix metalloproteinase (MMP)- 1, MMP-13, and type II α1 collagen fibers (COL2A1). CCK-8 was used to detect the cell proliferation rate, and flow cytometry was used to detect the cell apoptosis rate. H19 was up-regulated in OA cartilage tissue (P<0.05); Silencing H19 not only prevented chondrocyte apoptosis but also facilitated chondrocyte proliferation (both P<0.05), down-regulated MMP1 and MMP-13 expression levels (both P<0.05), and up-regulated COL2A1 expression rates (P<0.05). Also, silenced H19 inhibited the mRNA level of ALP, OCN, and BSP and ALP activity of chondrocytes (all P<0.05). There was a negative correlation between miR-140-5p and H19 in OA cartilage tissue and chondrocytes (r=0.98, P<0.001). Silenced miR-140-5p significantly reversed the inhibitory effects of silenced H19 on matrix degradation and calcification markers in chondrocytes (all P<0.05). The present study showed that the H19/ miR-140-5p regulatory axis could affect the degradation and calcification of the cartilage matrix in OA cells.

The long noncoding RNA H19 attenuates force-driven cartilage degeneration via miR-483–5p/Dusp5

Developmental dysplasia of the hip (DDH) is a common hip disease characterized by abnormal development of the acetabulum and femoral head. In most cases, DDH ultimately leads to osteoarthritis. Anomalous biomechanical force plays an important role in cartilage degeneration in DDH. However, in addition to mechanical wear, the underlying molecular mechanisms in cartilage degeneration in DDH remain unclear. This study analyzed the effect of long noncoding RNA (lncRNA)-H19 on DDH cartilage degradation. To elucidate the specific role of lncRNA H19, we established an intermittent cyclic mechanical stress (ICMS) cell force model to simulate abnormal biomechanical environment in vitro. Then, the roles of lncRNA-H19 were also determined in vivo by establishing a model of swaddling DDH. We observed that patients with DDH possessed low levels of lncRNA-H19, COL2A1, and Aggrecan but high levels of MMP3 and Adamts5. The same results were also obtained in a DDH rat model. Furthermore, the data suggested that ICMS promoted cartilage degeneration and caused reorientation of the cytoskeleton, and lncRNA H19 helped inhibit cartilage degeneration. Bioinformatics analysis and lncRNA sequencing were performed, and luciferase assays showed that lncRNA H19 and Dusp5 are both direct targets of miR-483–5p. Moreover, Dups5 plays a negative role in ICMS-induced cartilage degradation by activating the Erk and p38 pathways. In vivo, lncRNA H19 had protective effects on the swaddling DDH model. These findings indicate that lncRNA-H19 played a positive role in cartilage degradation in DDH through the lncRNA H19/miR-483–5p/Dusp5 axis.

H19/miR-152-3p/TCF4 axis increases chemosensitivity of gastric cancer cells through suppression of epithelial-mesenchymal transition.

BackgroundAdriamycin (ADM) is a drug commonly used for treating gastric cancer (GC). However, chemoresistance presents an obstacle to achieving successful outcomes in patients treated with chemotherapy. This study aimed to explore the effect of long non-coding RNA (lncRNA) H19 on chemoresistance in GC and its potential molecular mechanism.MethodsThe expression of H19 was detected in GC tissues and cell lines. The interaction between miR-152 with H19 and transcription factor 4 (TCF4) was validated by luciferase reporter assay. CCK-8 and flow cytometry were employed to measure cell viability and apoptosis, respectively. The levels of epithelial-mesenchymal transition (EMT) markers (E-cadherin, N-cadherin, Slug, Snail, and Twist) were detected by Western blot assay. A mouse model was established by subcutaneously injecting MGC-803 cells stably transfected with sh-H19 to investigate the effect of H19/miR-152-3p/TCF4 axis on ADM in vivo.ResultsIn vitro, we observed that H19 was overexpressed in GC tissues and cell lines. After knockdown of H19, the IC50 of ADM was decreased and cell apoptosis rates increased in both BGC-823ADM and MGC-803ADM cells. Furthermore, H19 shRNA was found to inhibit epithelial-mesenchymal transition (EMT), and induction of EMT counteracted the inhibitory effect of H19 shRNA on chemoresistance of GC cells. miR-152 was a target of H19, and its expression was downregulated in GC tissues and cell lines. Furthermore, the expression of TCF4 was negatively regulated by miR-152 but positively regulated by H19. In vivo, the data indicated that H19 shRNA enhanced the chemosensitivity of GC tumor cells to ADM through sponging miR-152 from TCF4, resulting in the suppression of EMT.ConclusionsThe results of this study elucidated that H19 was overexpressed in GC tissues and cell lines, and knockdown of lncRNA H19 increased the chemosensitivity of GC cells to ADM via sponging miR-152 from TCF4. The H19/miR-152/TCF4 axis may provide a new perspective for treating GC.

LncRNA DANCR-miR-758-3p-PAX6 Molecular Network Regulates Apoptosis and Autophagy of Breast Cancer Cells.

ObjectiveThis study set out to probe into the effects of long non-coding RNA (LncRNA) differentiation antagonizing non-protein coding RNA (DANCR) on apoptosis and autophagy of breast cancer (BC) cells.MethodsThe expression levels of DANCR, miR-758-3p and paired box 6 (PAX6) in BC tissues and cell lines were detected. The transcription and protein levels of PAX6, apoptosis-related factors (caspase-3, caspase-9, Bax/Bcl-2), and autophagy-related factors (LC3B, Atg5, Beclin-1) in BC cells were detected. The cell proliferation, apoptosis, autophagy and the regulatory relationship between genes and target genes were analyzed.ResultsDANCR and PAX6 were up-regulated in BC tissues and cell lines, while miR-758-3p was opposite. Down-regulating DANCR inhibited the malignant proliferation of BC cells and also promoted apoptosis and autophagy, which showed that caspase-3, caspase-9, Bax/Bcl-2, LC3B, Atg5 transcription and protein levels increased, while Beclin-1 transcription and protein levels decreased. DANCR regulated miR-758-3p in a targeted manner, and its over-expression could weaken the anti-cancer effect of miR-758-3p on BC cells. In addition, miR-758-3p also directly targeted PAX6, and knocking down its expression could weaken the inhibitory effect of down-regulating PAK6 on BC cell apoptosis and autophagy. We also found that DANCR acted as a competitive endogenous RNA sponge miR-758-3p, thus regulating the PAX6 expression.ConclusionDANCR-miR-758-3p-PAX6 molecular network plays a key regulatory role in BC cell apoptosis and autophagy, which may provide reference for treating patients.

Long Noncoding RNA RMRP Suppresses the Tumorigenesis of Hepatocellular Carcinoma Through Targeting microRNA-766.

PurposeThis study aimed to explore the regulatory effect of long noncoding RNA (lncRNA) ribonuclease mitochondrial RNA processing gene (RMRP) on hepatocellular carcinoma (HCC).MethodsThe expression of RMRP in HCC tissues and cell lines was assessed by qRT-PCR. Kaplan–Meier method was utilized to analyze the correlation between RMRP expression and the survival of HCC patients. MHCC97H and HuH7 cells were transfected with pcDNA3.1-RMRP or pcDNA3.1, respectively. MTT and flow cytometry assays were conducted to examine the proliferation and apoptosis of HCC cells, respectively. The migration and invasion of HCC cells were assessed using wound healing and transwell assays, respectively. StarBase3.0 and dual-luciferase reporter gene assay were used to identify the target relationship between miR-766 and RMRP. A xenografted tumor model was established in rats to evaluate the effect of RMRP in vivo.ResultsRMRP was down-regulated in HCC tissues and cells. Low expression of RMRP was correlated with poor survival of HCC patients. The A495 value and colony number were significantly decreased in pcDNA3.1-RMRP-transfected MHCC97H and HuH7 cells. The apoptosis rate was significantly increased in pcDNA3.1-RMRP-transfected MHCC97H and HuH7 cells. The migration rate and the number of invasive cells were significantly decreased in pcDNA3.1-RMRP-transfected MHCC97H and HuH7 cells. MiR-766 was a target of RMRP and eliminated the anti-tumor effect of RMRP on MHCC97H cells. The up-regulation of RMRP suppressed the growth of xenograft tumors in rats.ConclusionOverexpression of RMRP suppressed the tumorigenesis of HCC by targeting miR-766.

Long Noncoding RNA MALAT1 Promotes the Development of Colon Cancer by Regulating miR-101-3p/STC1 Axis.

PurposeColon cancer (CC) is a leading cause of cancer-related deaths worldwide. This study aimed to clarify the effect of long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) on CC progression and the potential mechanism.MethodsCC cell lines HCT116 and HT29 were selected for functional analysis. The expression of MALAT1, microRNA (miR)-101-3p, and stanniocalcin 1 (STC1) in CC tissues and cells were measured by quantitative reverse transcription PCR (qRT-PCR). Cell proliferation, apoptosis, migration and invasion were measured by Cell Counting Kit-8 (CCK-8), flow cytometry, wound scratch and transwell assay, respectively. The target relationships (MALAT1 and miR-101-3p, miR-101-3p and STC1) were validated by dual-luciferase reporter and RNA pull-down assay.ResultsThe expression of MALAT1 was elevated in CC tissues compared with adjacent normal tissues and was associated with lymph node metastasis, depth of invasion and tumor-node-metastasis (TNM) stage. Up-regulation of MALAT1 promoted the proliferation, migration, and invasion and inhibited the apoptosis of CC cells; while MALAT1 knockdown exhibited opposite results. MiR-101-3p was a target of MALAT1, which was negatively regulated by MALAT1. Silencing of miR-101-3p reverses the anti-tumor effect of MALAT1 knockdown on CC cells. STC1 was a target of miR-101-3p, which was negatively regulated by miR-101-3p. Silencing of STC1 reverses the tumor promoting effects of MALAT1 up-regulation and miR-101-3p down-regulation on CC cells.ConclusionMALAT1 may function as an oncogene in CC progression by affecting the miR-101-3p/STC1 axis, providing a hopeful therapeutic option for CC.

LncRNA-ANCR down-regulation suppresses invasion and migration of colorectal cancer cells by regulating EZH2 expression.

Our study aimed to explore the effects of long noncoding RNA (lncRNA)-ANCR on the invasion and migration of colorectal cancer (CRC) cells by regulating enhancer of zeste homolog 2 (EZH2) expression. CRC tissues and adjacent normal tissues were collected and CRC SW620 cells line and normal human intestinal epithelial cells (HIECs) were incubated. CRC SW620 cells line was transfected with ANCR-siRNA. The expressions of ANCR and EZH2 mRNA were measured by real-time quantitative polymerase chain reaction (RT-qPCR). EZH2 and trimethylation of H3K27 (H3K27me3) protein expressions were detected using Western blotting. The relationship between ANCR and EZH2 was determined through RNA pull-down and co-immunoprecipitation (co-IP) assays. Cell invasion and migration were determined by Trans-well and cell scratch assays. ANCR, EZH2 and H3K27me3 expressions were up-regulated in CRC tissues and SW620 cells (all P < 0.05). After transfected with ANCR-siRNA, SW620 cells showed decreased ANCR expression and EZH2 mRNA and protein expressions (all P < 0.05). According to the results of RNA pull-down and co-IP assays, ANCR could specifically bind to EZH2. The results of Trans-well and cell scratch tests showed that when ANCR expression was decreased, the invasion and migration abilities of SW620 cells significantly declined (both P < 0.05). In conclusion, these results suggest that lncRNA-ANCR could influence the invasion and migration of CRC cells by specifically binding to EZH2.

The Effect of Long Non-Coding RNA (lncRNA) HCP5 on Regulating Epithelial-Mesenchymal Transition (EMT)-Related Markers in Gastric Carcinoma Is Partially Reversed by miR-27b-3p.

BackgroundlncRNA HCP5 plays a cancer-promoting role in a variety of cancers. This study was the first to explore the mechanism of HCP5 in gastric carcinoma (GC).Material/MethodsThe differences in HCP5 between GC patients and healthy people were revealed in the TCGA database. The expression of HCP5 in GC tissues and adjacent tissues was compared by qRT-PCR. At the same time, the clinic pathological features of the patients were counted. Starbase and luciferase assay predicted and verified that miR-27b-3p is a targeted miRNA for HCP5. The expression of HCP5 and miR-27b-3p in various GC cells was detected by qRT-PCR. Cell viability and metastasis in different treatment groups were assessed by use of Cell Couting Kit-8 assay and clone formation assay, wound-healing assay, and transwell assay. Finally, expression of epithelial-mesenchymal transition (EMT)-associated markers was detected by Western blot.ResultsWe found that HCP5 was overexpressed in GC tissues. Patients with higher expression of HCP5 had larger tumors, were more likely to have lymph node metastasis, and had higher TNM stage. HCP5 was overexpressed in GC cells, but this was reversed by miR-27b-3p. Silencing HCP5 inhibited GC cell viability and metastasis by downregulating Vimentin and N-cadherin and up-regulating E-cadherin, but this effect was partially reversed by miR-27b-3p inhibitor.ConclusionsThe effect of silencing HCP5 on repressing GC cells viability and metastasis by regulating EMT-associated markers can be partially reversed by miR-27b-3p inhibitor.

Upregulation of lncRNA DANCR functions as an oncogenic role in non-small lung cancer by regulating miR-214-5p/CIZ1 axis.

Lung cancer has an unfavorable prognosis due to the lack of efficient diagnostic and therapeutic strategies. Therefore, this study sought to figure out the effect of long non-coding RNA (lncRNA) DANCR on lung cancer progression. LncRNA DANCR and miR-214-5p expressions in non-small cell lung cancer (NSCLC) were detected by Real Time-quantitative Polymerase Chain Reaction (RT-qPCR). Function assays, including Cell Counting Kit-8 (CCK-8) and flow cytometric analysis were conducted to clarify the role of DANCR and miR-214-5p in the progression of NSCLC. Western blot, Dual-Luciferase reporter assay, and RNA immunoprecipitation assay (RIP) were performed to elucidate the underlying mechanism. LncRNA DANCR was upregulated in NSCLC. The knockdown of lncRNA DANCR inhibited cell proliferation and accelerated cell apoptosis in NSCLC. LncRNA DANCR interacted with miR-214-5p. MiR-214-5p over-expression partially reversed the regulatory effects of DANCR on proliferation and apoptosis in NSCLC. In addition, CIZ1 was the downstream gene binding miR-214-5p. LncRNA DANCR could regulate the miR-214-5p/CIZ1 axis. Downregulation of lncRNA DANCR inhibited cell proliferation and induced cell apoptosis in NSCLC by regulating the miR-214-5p/CIZ1 axis. LncRNA DANCR may act as an oncogene and promote the progression of NSCLC.

LOC101060264 Silencing Suppresses Invasion and Metastasis of Human Colon Cancer.

BackgroundWe explored the regulatory effects of long noncoding RNA (lncRNA) LOC101060264 silencing mediated by shRNA on invasion and metastasis of human colon cancer.Material/MethodsInitially, 2 shRNA plasmids for LOC101060264 silencing – shRNA1 and shRNA2 – were introduced into LoVo cells. Following transfection, the expressions of LOC101060264, E-cadherin, and vimentin were determined. Next, to explore the regulatory effects of LOC101060264 silencing on cell growth, cell cycle, invasion, and migration abilities of LoVo cells, we performed MTT, flow cytometry, Transwell assay, and scratch assay, respectively. Furthermore, in nude mice with xenografted tumors, the tumor volume and weight were measured, and the expressions of PCNA, E-cadherin, vimentin, and MMP-9 in tumor tissues were determined by immunohistochemistry.ResultsThe level of E-cadherin increased and the level of vimentin decreased after LOC101060264 silencing mediated by shRNA1 and shRNA2 in LoVo cells. Silencing LOC101060264 repressed the migration, invasion, and proliferation of LoVo cells in vitro and inhibited tumor growth in nude mice in vivo. We also studied the expression of these proteins and found reduced expression of PCNA, vimentin, and MMP-9 protein, and found enhanced expression of E-cadherin protein. Moreover, the inhibitory effect of shRNA2 on the above cell behaviors was stronger than that of shRNA1.ConclusionsIn summary, LOC101060264 silencing decreased LoVo cell invasiveness via suppressing ETM and attenuated tumor metastasis, which provides a novel therapeutic target for patients with colon cancer.

Effect of long non-coding RNA maternally expressed gene 3 on proliferation and invasion capacity of glioma cells through the Wnt/human-catenin signal pathway

Objective To investigate the effect of long non-coding RNA maternally expressed gene 3 (MEG3) on the proliferation and invasion capacity of glioma cells through the Wnt/human-catenin signal pathway. Methods Glioma cells were divided into 3 groups: the blank control group (CON), the gene transfection group (MEG3) and the gene inhibition group (siMEG3). The cells in blank group were left untreated and cultured normally. MEG3 gene was transfected in the gene transfection group, and gene interference was conducted in the gene inhibition group. Protein expression was determined by Western blotting, and gene expression was detected by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Results After transfection of MEG3 gene, the apoptosis rate increased to 61.05±2.77 with the prolongation of culture time, which was higher than that of the control group and the gene inhibition group (F=23.543, P<0.05). After MEG3 gene knockout, the apoptosis rate of glioma cells was higher than that of the control group (P<0.05). After 48 hours of transfection, the number of cell migration was 105.36±15.27 (F=33.350, P<0.05). When MEG3 gene was inhibited, the number of cell migration was 989.41±11.06, higher than that of the control group (F=40.667, P<0.05). After MEG3 gene transfection, the number of invasion cells was 251.25±35.85, which was lower than that of the control group and gene knockout group (F=31.167, P<0.05). After MEG3 gene was knocked out, the number of invasion cells was 1 500.00± 84.76, which was higher than that of the blank control group (F=63.762, P<0.05). Compared with the gene inhibition group, the c-Jun gene of the control group and the gene transfection group decreased, and the gene transfection group was lower than the blank control group (F=15.426, P<0.05). Conclusion Long non-coding RNA MEG3 can inhibit the proliferation and invasion of glioma cells through the Wnt/human-catenin signal pathway. Key words: Glioma; Protein; Gene

Expression and Effects of Long Non-Coding RNA, LINC01124, in Non-Small Cell Lung Cancer.

ObjectiveTo investigate the expression and evaluate the clinical significance of long non-coding RNA, LINC01124, in non-small cell lung cancer (NSCLC) and to study its influence in this tumor.MethodsHundred specimens of NSCLC tissues and normal lung tissues after surgery were collected. The qRT-PCR for LINC01124 expression was performed on cancerous and normal lung tissues. The correlations between the expression of LINC01124 and pathological characteristics were analyzed. PcDNA-LINC01124 was transfected to upregulate LINC01124 expression in NSCLC cells, and the transfection efficiency was evaluated by the qRT-PCR. CCK8 assay, wound-healing assay, and the Transwell assay were performed to evaluate the effect of ectopic LINC01124 expression on proliferation, migration, and invasive of NSCLC cells.ResultsThe expression level of LINC01124 was downregulated in tumor tissues when compared with the paired normal lung tissues (P<0.05). The expression of LINC01124 was associated with patients’ age and distant metastasis (P<0.05). Enforced expression of LINC01124 significantly inhibited the proliferation, migration, and invasive ability of NSCLC cells.ConclusionThe expression of LINC01124 was decreased in patients with NSCLC of older age and with those having distant metastasis. LINC01124 may inhibit cell proliferation, migration, and invasive ability.

LncRNA KCNQ1OT1 contributes to the cisplatin resistance of tongue cancer through the KCNQ1OT1/miR-124-3p/TRIM14 axis.

Tongue cancer is a common malignant tumor in the oral and maxillofacial region, most of which is squamous cell carcinoma. Cisplatin (DDP) is one of the chemotherapy drugs for patients with tongue squamous cell carcinoma (TSCC). However, DDP resistance has become a major obstacle to its clinical application. Our study aimed to investigate the effects of long non-coding RNA (lncRNA) KCNQ1 overlapping transcript 1 (KCNQ1OT1) on DDP resistance of tongue cancer and the underlying mechanism. The levels of KCNQ1OT1, miR-124-3p, and tripartite motif containing 14 (TRIM14) were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The maximum size of tumor (MTS) assay was used to detect the cell survival rates. Furthermore, the cell proliferation was detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Transwell assay was performed to detect the cell migration and invasion. Western blot assay was used to detect the protein levels of Vimentin, N-cadherin, E-cadherin, and TRIM14. The functional targets of KCNQ1OT1 and miR-124-3p, miR-124-3p and TRIM14 were predicted by starBase 3.0 and TargetScan. The relationship between KCNQ1OT1 and miR-124-3p was confirmed by Dual-Luciferase reporter assay, RNA immunoprecipitation (RIP) and RNA pull-down. Further, the relationship between miR-124-3p and TRIM14 was verified by Dual-Luciferase reporter assay. Animal experiment revealed the effect of KCNQ1OT1 on DDP resistance of tongue cancer cells in vivo. KCNQ1OT1 was upregulated in DDP-resistant tongue cancer tissues and cells, and mainly expressed in cytoplasm. Functionally, the knockdown of KCNQ1OT1 inhibited the survival rate, proliferation, migration, invasion, and EMT of the DDP-resistant tongue cancer cells. Of note, miR-124-3p acted as a target of KCNQ1OT1 and KCNQ1OT1 could reduce the expression of miR-124-3p. Moreover, miR-124-3p targeted TRIM14 and the downregulation of TRIM14 reduced the DDP resistance of tongue cancer cells. Importantly, KCNQ1OT1 regulated the TRIM14 expression by targeting miR-124-3p. Furthermore, KCNQ1OT1 knockdown reduced the DDP-resistant tumor growth and weight through the miR-124-3p/TRIM14 axis in vivo. LncRNA KCNQ1OT1 promotes the DDP resistance of tongue cancer by sponging miR-124-3p to regulate TRIM14 expression.

LncRNA PAPAS aggravates the progression of gastric cancer through regulating miRNA-188-5p.

To uncover the biological effect of long non-coding RNA (lncRNA) PAPAS on the progression of gastric cancer (GC) by mediating microRNA-188-5p (miRNA-188-5p) level. The relative level of PAPAS was determined in GC tissues and cell lines by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The Kaplan-Meier method was introduced to assess the prognostic potential of PAPAS in the overall survival of GC patients. Regulatory effects of PAPAS on proliferative, migratory, and invasive abilities of HGC-27 and AGS cells were detected by cell counting kit-8 (CCK-8), transwell, and wound closure assay, respectively. Subsequently, the binding relation between PAPAS and miRNA-188-5p was verified by the Dual-luciferase reporter gene assay. Correlation between expression levels of PAPAS and miRNA-188-5p in GC tissues was explored. Finally, rescue experiments were conducted to uncover the role of PAPAS/miRNA-188-5p axis in the progression of GC. PAPAS was upregulated in GC tissues and cell lines compared to controls. GC patients expressing a high level of PAPAS suffered worse prognosis relative to those with low level. The silence of PAPAS remarkably attenuated proliferative, migratory, and invasive abilities of HGC-27 cells. Overexpression of PAPAS in AGS cells obtained the opposite trends. MiRNA-188-5p was the direct target of PAPAS, which was negatively regulated by PAPAS. MiRNA-188-5p was able to reverse the regulatory effects of PA-PAS on proliferative, migratory, and invasive abilities of GC cells. LncRNA PAPAS is upregulated in GC and closely related to lymphatic metastasis, distant metastasis, and poor prognosis of GC patients. PAPAS aggravate the malignant progression of GC by negatively regulating the miRNA-188-5p level.

Downregulation of DLGAP1-Antisense RNA 1 Alleviates Vascular Endothelial Cell Injury Via Activation of the Phosphoinositide 3-kinase/Akt Pathway Results from an Acute Limb Ischemia Rat Model

This study aimed to investigate the effect of long non-coding RNA (lncRNA) DLGAP1 antisense RNA 1 (DLGAP1-AS1) on vascular endothelial cell (VEC) injury via the phosphoinositide 3-kinase (PI3K)/Akt pathway in rat models of acute lower limb ischaemia-reperfusion (I/R). Differentially expressed lncRNAs related to I/R were screened using the gene expression omnibus database. Acute lower limb I/R models were induced in male Wistar rats, in which the regulatory mechanisms of DLGAP1-AS1 silencing were analysed after the treatment of small interfering RNA (siRNA) against DLGAP1-AS1 or an inhibitor of the PI3K/Akt pathway. The relationship between DLGAP1-AS1 and the PI3K/Akt pathway was analysed. The levels of tumour necrosis factor (TNF)-α and vascular cell adhesion molecule-1 (VCAM-1), as well as malondialdehyde (MDA) concentration and creatine kinase (CK) activity, were measured. The number of circulating endothelial cells (CECs) and apoptosis of VECs were identified. Microarray based analysis indicated that DLGAP1-AS1 was highly expressed in I/R, which was further confirmed by detection of expression in rat models of acute lower limb I/R. Notably, the treatment of siRNA against DLGAP1-AS1 led to the activation of the PI3K/Akt pathway. In response to siRNA against DLGAP1-AS1, the levels of TNF-α and VCAM-1 were decreased, and MDA concentration and CK activity was downregulated. Reduced CEC numbers and suppressed VEC apoptosis were also observed. DLGAP1-AS1 silencing could further suppress the oxidative stress, exert an anti-apoptosis effect, and reduce inflammatory reaction, whereby VEC injury is alleviated by activation of the PI3K/Akt pathway in rats with acute lower limb I/R.

LncRNA SOX2OT Mediates Mitochondrial Dysfunction in Septic Cardiomyopathy.

Researches establish an indispensable role of mitochondrial dysfunction in septic cardiomyopathy. We aimed to investigate the effects of long noncoding RNA (LncRNA) SOX2 overlapping transcript (SOX2OT) on mitochondrial dysfunction in septic cardiomyopathy. We observed an obvious overexpression of SOX2OT in septic hearts and cardiomyocytes. Knockdown of SOX2OT in mice recovered the reduced cardiac function, and improved the mitochondrial membrane potential impaired by lipopolysaccharide (LPS). SOX2OT overexpressed mice showed the opposite situation. In parallel, knockdown of SOX2OT in cardiomyocytes restored the mitochondrial membrane potential, along with reduced mitochondrial reactive oxygen species production induced by LPS, while overexpression of SOX2OT reversed these effects. Mechanistically, SOX2OT could regulate mitochondrial dysfunction in septic cardiomyopathy via SOX2. In general, SOX2OT contributed to mitochondrial dysfunction progression via inhibiting SOX2 expression in septic cardiomyopathy, which may provide a new insight for treatment of septic cardiomyopathy.

Long Noncoding RNA NEAT1 Promotes Cell Proliferation And Invasion And Suppresses Apoptosis In Hepatocellular Carcinoma By Regulating miRNA-22-3p/akt2 In Vitro And In Vivo.

BackgroundHepatocellular carcinoma (HCC) is one of the most aggressive cancers that is associated with cirrhosis and other chronic liver diseases. Although remarkable progress has been made in past decades, it is still necessary to continue exploring the pathology and development of HCC.ObjectiveIn this study, we elucidated the effect of long noncoding RNA (lncRNA) NEAT1 on HCC development and underlying mechanisms.MethodsClinicopathological features of HCC patients were collected and the correlations with NEAT1 expression were assessed. To determine cell activities, CCK-8, flow cytometry, invasion assays, and TUNEL assays were performed. Real-time PCR, Western blot, and luciferase reporter assays were performed to investigate the related mechanism of HCC.ResultsThe results revealed that NEAT1 expression was associated with tumor size and differentiation where NEAT1 was upregulated in both HCC tissues and cell lines. Overexpression of NEAT1 promoted proliferation and invasion while inhibited apoptosis in HCC cells, which was opposite to the effect of NEAT1 knockdown. Also, AKT2 was increased in HCC tissues. Downregulation of AKT2 was associated with reduced cell proliferation and invasion while increased apoptosis, while overexpression of AKT2 exerted opposite roles. In addition, the expression of miRNA-22-3p displayed an inverse association with NEAT1. miRNA-22-3p mimic and inhibitor suppressed and promoted HCC development, respectively. The luciferase assay revealed that both NEAT1 and AKT2 were direct target genes of miRNA-22-3p. Furthermore, knockdown and overexpression of NEAT1 suppressed and promoted tumor growth in the HCC mouse model, which were abolished by the miRNA-22-3p inhibitor and mimic, respectively.ConclusionIn conclusion, the results demonstrate that NEAT1 promotes the development of HCC, both in vitro and in vivo, through regulating miRNA-22-3p/AKT2, and provides insight into developing a new strategy for HCC treatment.