Noncoding RNA Growth Research Articles

Long noncoding RNA growth arrest-specific transcript 5 alleviates renal fibrosis in diabetic nephropathy by downregulating matrix metalloproteinase 9 through recruitment of enhancer of zeste homolog 2.

Diabetic nephropathy (DN) is a frequently occurred microvascular complication associated with type I and type II diabetes mellitus. The participation of long noncoding RNAs (lncRNAs) in diabetes-related microvascular complications has been reported extensively. We attempted to unveil the possible regulatory mechanism of lncRNA growth arrest-specific transcript 5 (GAS5) and matrix metalloproteinase 9 (MMP9), an important inflammatory protein, in the progression of DN. A rat DN model was induced by streptozocin (STZ). The low expression of GAS5 and high expression of MMP9 in DN rats with DN was then determined by RT-qPCR and western blot analysis, and lentivirus-mediated GAS5 overexpression was shown to ameliorate STZ-induced renal interstitial fibrosis (RIF) and inflammatory reaction in the kidney of DN rats. Moreover, MMP9 was found to be upregulated in STZ-induced DN, while MMP9 silencing induced by lentivirus expressing shRNA against MMP9 reduced RIF and suppressed inflammation in the kidney of DN rats. RIP, RNA pull-down, and ChIP assays demonstrated that GAS5 downregulated MMP9 via recruiting enhancer of zeste homolog 2 (EZH2) in the promoter region of MMP9. Overall, our study reveals that GAS5 downregulates MMP9 expression through recruiting EZH2 to MMP9 promoter region and alleviates the progression of renal fibrosis in DN rats, which sheds new light on the therapeutic potential of GAS5-targeted therapies in combating that disease.

The Long Noncoding RNA, Growth Arrest-Specific 5, Suppresses Gastric Cancer by Downregulating miR-21 Expression

Background: Gastric cancer has become the second major cause of cancer death. The aim of the present study was to explore the relationship between miR-21 and the long noncoding RNA growth arrest-specific 5 (GAS5) in gastric cancer and the effect on gastric cancer cells. Methods: The expression of miR-21 and GAS5 mRNA was analyzed by quantitative real-time-PCR. Overexpression of GAS5 was used to investigate the biological functions of GAS5 in cells. The cell proliferation was detected by cell counting kit-8 assay and the cell migration and invasion were detected by Transwell. Cell apoptosis was evaluated by Annexin V-FITC/PI staining and apoptosis-related proteins were detected by western blot. The mechanism of GAS5 in vivo was evaluated by the tumorigenesis of nude mice, and dual luciferase reporter was used to determine if miR-21 is a GAS5 target. The inhibition of miR-21 and the simultaneous overexpression of GAS5 and miR-21 were further performed, and the above indicators were detected again. Results: GAS5 was low expression and miR-21 was high expression in gastric cancer tissues and cells. GAS5 overexpression reduced the proliferation, migration, and invasion of gastric cancer cells and increased the apoptosis of gastric cancer cells. The growth rate of GAS5 group slowed down and the volume of tumor decreased. miR-21 is a GAS5 target and GAS5 inhibits the proliferation of gastric cancer cells by targeting miR-21. Conclusion: Our research shown that overexpression of GAS5 can significantly inhibit the proliferation, migration, invasion and tumor formation of gastric cancer cells, and promote the apoptosis of gastric cancer cells, which may be related to the targeting inhibition of miR-21 expression by GAS5.

Long non-coding RNA GAS5 expression in patients with Down syndrome.

Trisomy 21, also known as Down Syndrome (DS), is the most common chromosome abnormality and causes intellectual disability. Long non-coding RNA (lncRNA) growth arrest-specific 5 (GAS5), whose differential expression has recently been reported in patients with Klinefelter syndrome, has been addressed to play a role in the development of inflammatory and autoimmune diseases, vascular endothelial cells apoptosis and atherosclerosis, all being common features in patients with DS. Therefore, the aim of this study was to assess the lncRNA GAS5 expression profile in DS patients and in controls. lncRNA GAS5 levels were evaluated by qRT-PCR assay in 23 patients with DS and 23 age-matched controls. A significant lncRNA GAS5 down-regulation was observed in patients with DS by RT-PCR analysis, The RNA sequencing experiments confirmed the qRT-PCR data. LncRNA GAS5 down-expression may play a role in the development of some typical features of the patients with DS and, particularly, in inflammatory and autoimmune diseases.

The relationships of genetic polymorphisms of the long noncoding RNA growth arrest-specific transcript 5 with uterine cervical cancer.

The purposes of the investigation were to examine the implications of long noncoding RNA growth arrest-specific transcript 5 (GAS5) in progression and clinicopathological factors of uterine cervical cancer, and patient survival in Taiwan. Genotypic distributions of two GAS5 genetic variants rs145204276 and rs55829688 were detected in 208 patients including 111 patients with invasive cancer, 97 with precancerous lesions as well as 307 control women using real-time polymerase chain reaction. It explored that patients with cervical precancerous lesion had lower rate of AGGCA deletion (Del) in both alleles (Del/Del) of GAS5 rs145204276 as compared with control women. Patients with invasive cancer did not exhibit higher rate of Del/Del. Meanwhile, there were no different genotypic distributions in rs55829688 among patients with cervical invasive cancer and those with precancerous lesions as well as control women. Moreover, cervical cancer patients with Ins (insertion, AGGCA)/Del and Del/Del (-/-) in GAS5 rs55829688 tended to have poorer hazard ratio (HR) of 5 years survival. In addition, lymph node metastasis status exerted the most significantly predictive of 5 years survival rate. Conclusively, GAS5 polymorphism rs145204276 is probably applicable to predict 5 years survival HR of cervical cancer patients. However, the mechanism elucidating the methylation status and transcription function of rs145204276 in uterine cervical cancer needs to be delineated for its unique implication in uterine cervical cancer.

Long noncoding RNA-GAS5 attenuates progression of glioma by eliminating microRNA-10b and Sirtuin 1 in U251 and A172 cells.

Long noncoding RNA (lncRNA) growth arrest-specific 5 (GAS5) is implicated in several cancers via modulating microRNAs (miRs). However, little information is available about the correlation between GAS5 and miR-10b. Therefore, we sought out to investigate the biological role of GAS5-miR-10b node mainly in glioma cells. We artificially modulated GAS5 to explore its roles in viability assayed by cell counting kit-8 (CCK-8), motile activities by 24-Transwell assay, as well as apoptosis by a flow cytometer and Western blot assay. miR-10b and Sirtuin 1 (Sirt1) were quantified by qRT-PCR. After co-transfection, we analyzed the viability, migration, invasion, apoptosis, and Sirt1 expression. Western blot was implemented to detect the phosphorylated forms of PTEN, PI3K, AKT, MEK, and ERK. GAS5 inhibited proliferation and motile behaviors, and fortified apoptosis. As for the viability and motile activities, the property of GAS5 was reversed in miR-10b-replenished U251 and A172 cells, while maintained in miR-10b-deficient cells. Additionally, GAS5-induced apoptosis was abolished by miR-10b overexpression while fortified by miR-10b silence. Besides, GAS5 negatively modulated Sirt1 via miR-10b. Moreover, Sirt1 negatively modulated PTEN and positively mediated the abovementioned regulators. GAS5 represses the process of glioma cells by decreasing miR-10b, which as accompanied by Sirt1 silence-induced inactivation of PTEN/PI3K/AKT and MEK/ERK cascades.

LncRNA GAS5 suppresses inflammatory responses and apoptosis of alveolar epithelial cells by targeting miR-429/DUSP1

Acute lung injury (ALI) is a life-threatening syndrome characterized by excessive inflammation and apoptosis of alveolar epithelial cells. This study firstly investigated the role and mechanism of long non-coding RNA (lncRNA) growth arrest-specific 5 (GAS5) in regulating lipopolysaccharide (LPS)-induced inflammatory response and apoptosis of murine alveolar epithelial cell line MLE-12. The expression of GAS5, miR-429, and dual-specificity phosphatase 1 (DUSP1) were examined using quantitative Real-Time PCR (qRT-PCR) and western blot. The inflammatory responses were evaluated by detecting the levels of pro-inflammatory cytokines using ELISA. Cell apoptosis was assessed by TUNEL assay. The interactions among GAS5, miR-429, and DUSP1 were examined using luciferase reporter assay. The results showed that GAS5 and DUSP1 expression were decreased, whereas miR-429 was increased in lung tissues from LPS-induced ALI mice and LPS-treated MLE-12 cells. Furthermore, GAS5 overexpression decreased cell inflammatory responses and apoptosis in LPS-treated MLE-12 cells, which was reversed by miR-429 mimic and DUSP1 knockdown. Mechanistically, GAS5 acted as a competitive endogenous RNA by sponging miR-429 to facilitate DUSP1 expression. Our findings suggest that GAS5 suppresses inflammatory responses and apoptosis of alveolar epithelial cell MLE-12 by targeting miR-429/DUSP1 axis.

Abstract B17: Long noncoding RNA growth arrest specific 5 (GAS5) as a proliferation «brake» in aggressive population of CD133+ cells responsible for recurrence in PDAC

Abstract The presence of quiescent, therapy-resistant population of tumor cells is often attributed to extreme metastasis and tumor recurrence. This population, often described as cancer stem cells (CSCs), or tumor-initiating cells (TICs), is enriched in a tumor as a result of microenvironmental (hypoxia) or chemotherapeutic stress. In pancreatic cancer, CD133+ cells have been described as a representation of this aggressive TIC population and are most responsible for tumor relapse in patients. This population adapts to stress by turning on mechanisms to halt cell cycle progression. Upon removal of the stress, these cells restart their cell cycle and regain their proliferative nature. Growth Arrest Specific 5 (GAS5) is a long noncoding RNA critical for this process and is overexpressed in PDAC. 13C-Glucose labeling was used for glucose flux assay to track the use of glucose and other metabolites in CD133+ and CD133- populations of pancreatic cancer cells. CD133+/- cell populations were sorted with magnetic sorting or flow cytometry sorting techniques. Lentiviral stable transfection of human CD133 cDNA in MIA PaCa-2 cell line was used to obtain a CD133 overexpressing cell line. siGAS5 and siSOX2 were used to knock down genes in CD133Hi cell lines. Glucocorticoid receptor (GR) activity was assessed using GRE dual luciferase assay system (Qiagen). MTT-based assay was used for proliferation and BrDU incorporation assay was used for measure of nucleic acid synthesis. Fluorescent in situ hybridization using human GAS5 RNA probe (Stellaris) was used to visualize GAS5 localization in human PDAC vs. normal adjacent FFPE samples as well as xenograft FFPE samples. Our results show for the first time that the emergence of CD133+ population coincides with upregulation of GAS5, which reprograms cell cycle to slow proliferation by inhibiting GR-mediated cell cycle control. The CD133+ population further routed glucose through the pentose phosphate pathway, a predominantly biosynthetic pathway, in spite of being quiescent in nature. However, this did not result in immediate nucleic acid synthesis. Upon inhibiting GAS5, these cells were released from growth arrest and restarted nucleic acid synthesis and proliferation. Additionally, this study shows that GAS5 regulates cell cycle through GR, which it is known to bind and inhibit. CD133Hi cells with high GAS5 expression had low GR activity, and stimulation of these cells with dexamethasone was able to increase cell proliferation. Our study thus shows that GAS5 acts as a molecular switch for regulating quiescence and growth arrest in CD133+ population, allowing these cells to overcome chemical and environmental stressors and leading to the aggressive nature of pancreatic tumors. Citation Format: Nikita S. Sharma, Brittany C. Durden, Prisca Gnamlin, Vineet K. Gupta, Kousik Kesh, Vanessa T. Garrido, Ashok Saluja, Sulagna Banerjee. Long noncoding RNA growth arrest specific 5 (GAS5) as a proliferation «brake» in aggressive population of CD133+ cells responsible for recurrence in PDAC [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr B17.

Role of NR3C1 and GAS5 genes polymorphisms in multiple sclerosis

Introduction: Multiple sclerosis (MS) as a progressive chronic disease of the central nervous system (CNS) is characterized by demyelination and axonal loss. Results of genetic studies and clinical trials have proved a key role for the immune system in the pathogenesis of MS. Glucocorticoids (GR) are regarded as potent therapeutic compounds for autoimmune and inflammatory diseases which act through their receptors encoded by Nuclear Receptor Subfamily 3 Group C Member 1 (NR3C1) gene. Meanwhile, the long non-coding RNA (lncRNA) growth arrest specific 5 (GAS5) interacts with GR through binding to the DNA-binding domain (DBD) region and reduces GR transcriptional activity.Methods: The purpose of our study was to evaluate the association between MS and polymorphisms within NR3C1 (rs6189/6190, rs56149945, rs41423247) and GAS5 (rs55829688) genes in 300 relapsing-remitting MS patients and 300 healthy subjects.Results: We demonstrated significant differences in distribution of genotype, allele and haplotype frequencies of rs6189, rs41423247 and rs55829688 between the study groups.Conclusion: Our data may suggest that rs6189, rs41423247 and rs55829688 are associated with the increased risk of MS development. Future studies are needed to verify our results in larger sample sizes and elaborate the underlying mechanisms for contribution of these variants in MS disease.

Silencing of GAS5 Alleviates Glaucoma in Rat Models by Reducing Retinal Ganglion Cell Apoptosis.

Retinal ganglion cells (RGCs) play a key role in the pathogenesis and development of glaucoma. The present study aims to investigate the underlying mechanism of long noncoding RNA growth arrest-specific transcript 5 (GAS5) in glaucoma development through regulating the apoptosis of RGCs. Rat models of chronic glaucoma were successfully established by translimbal laser photocoagulation. Retinal tissues were collected to determine the density of RGCs through Toluidine blue staining. The overexpression vector or short hairpin RNA for GAS5 or enhancer of zeste homolog 2 (EZH2) was transfected into RGCs after in vitro pressurization culture to examine the function of GAS5 in RGC apoptosis. The involvement of EZH2 and ATP-binding cassette transporter A1 (ABCA1) was further identified. Cell apoptosis after laser treatment and transfection was assessed by flow cytometry. We found abundant GAS5 expression and a reduction in RGC density in the retinal tissues of glaucoma rats. Silencing of GAS5 led to increased EZH2 expression and decreased ABCA1 expression in RGCs. In addition, upregulation of EZH2 promoted trimethylation of lysine 27 on histone H3, thereby suppressing ABCA1 expression and eventually leading to the inhibition of RGC apoptosis. These findings provide further understanding of the function of GAS5 in RGC apoptosis. We conclude that downregulation of GAS5 could help relieve glaucoma symptoms. GAS5 is therefore a promising target for developing novel therapeutic approaches for treating patients with glaucoma.

Long Non-Coding RNA GAS5 and Intestinal MMP2 and MMP9 Expression: A Translational Study in Pediatric Patients with IBD.

Background: The long non-coding RNA (lncRNA) growth arrest–specific transcript 5 (GAS5) seems to be involved in the regulation of mediators of tissue injury, in particular matrix metalloproteinases (MMPs), implicated in the pathogenesis of inflammatory bowel disease (IBD). We investigated the role of GAS5 in regulating MMP2 and MMP9 expression in pediatric patients with IBD and in vitro. Methods: In total, 25 IBD patients were enrolled: For each patient paired inflamed and non-inflamed biopsies were collected. RNA was extracted and GAS5, MMP2, and MMP9 were quantified by TaqMan assay. The expression of GAS5 and MMPs was also determined in the human monocytic THP1 cells differentiated into macrophages and stimulated with lipopolysaccharide (LPS). The function of GAS5 was assessed by overexpressing the lncRNA and evaluating the MMPs levels. Results: Real-time PCR results demonstrated a downregulation of GAS5 and an upregulation of both MMPs in inflamed tissues. In vitro data confirmed the trend observed in patients for the three genes: The stimulation with LPS promoted a downregulation of GAS5 while an increase of MMPs was observed. Overexpression experiments showed that higher levels of GAS5 lead to a decrease of both enzymes. Conclusion: These results provide new information about the role of GAS5 in IBD: The lncRNA could mediate tissue damage by modulating the expression of MMPs.

Silencing of GAS5 represses the malignant progression of atherosclerosis through upregulation of miR-135a

Long non-coding RNA growth arrest-specific 5 (GAS5) has been demonstrated to be involved in the pathogenesis of atherosclerosis (AS). The purpose of the present study was to investigate the underlying mechanisms of GAS5 on the inflammation and lipid metabolic disorders of AS. ApoE−/− mice were fed on a high fat diet (HFD) and THP-1 macrophages were treated with ox-LDL to construct AS model in vivo and in vitro, respectively. The detections of blood lipids and inflammatory cytokines were performed using corresponding assay kits. qRT-PCR was used to assess the expression of GAS5 and miR-135a. Western blot was performed to detect PPARα and CPT1 levels. The targeted interaction between GAS5 and miR-135a was determined by dual-luciferase reporter assay and RNA immunoprecipitation assay. Our data revealed that GAS5 was upregulated in AS mice model and ox-LDL-treated macrophages. GAS5 silencing alleviated lipid metabolic disorders and inflammation in AS mice and ox-LDL-treated macrophages. Moreover, GAS5 directly targeted miR-135a and repressed miR-135a expression. MiR-135a expression restoration abrogated the alleviative effects of GAS5 silencing on inflammation and lipid metabolic disorders in ox-LDL-treated macrophages. In conclusion, our study suggested that GAS5 silencing repressed the malignant progression of AS at least partly through upregulation of miR-135a. Targeting GAS5 might be a promising treatment strategy for AS management.

LncRNA GAS5 overexpression downregulates IL-18 and induces the apoptosis of fibroblast-like synoviocytes.

Long non-coding RNA (lncRNA) growth arrest specific transcript 5 (GAS5) negatively regulates interleukin-18 (IL-18) in ovarian cancer, while IL-18 contributes to the development of rheumatoid arthritis (RA). Therefore, GAS5 may also participate in RA. GAS5 and IL-18 in plasma of RA patients (n = 60) and healthy controls (n = 60) were measured by RT-qPCR and ELISA, respectively. Linear regression was performed to analyze the correlations between plasma levels of IL-18 and GAS5 in both RA patients and healthy controls. In the present study, we found that plasma GAS5 was downregulated, while IL-18 was upregulated in RA patients than in healthy controls. A significant and inverse correlation between GAS5 and IL-18 was found in RA patients but not in healthy controls. IL-18 treatment did not significantly alter the expression of GAS5 in fibroblast-like synoviocytes, while GAS5 overexpression led to the inhibited expression of IL-18. GAS5 overexpression also resulted in the promoted apoptosis of fibroblast-like synoviocytes. Therefore, GAS5 overexpression may improve RA by downregulating IL-18 and inducing the apoptosis of fibroblast-like synoviocytes. Key points • The present study mainly showed that overexpression of GAS5 may assist the treatment of RA. • The mechanism of GAS5 for the treatment of RA involves the downregulating inflammatory IL-18 and mediating the apoptosis of fibroblast-like synoviocytes. • GAS5 and IL-8 were correlated in RA patients but not in healthy controls.

Indole-3-acetic acid has long-term effects on long non-coding RNA gene methylation and growth in Populus tomentosa.

DNA methylation and long non-coding RNAs (lncRNAs) regulate plant growth and development, but their relationship and effect on responses to the auxin phytohormone indole-3-acetic acid (IAA) remain largely unknown, particularly in woody plants such as poplar (Populus tomentosa). Following treatment of 1-year-old clonal plants with 100µM IAA, key poplar lncRNA genes showed changes in methylation, but whole-genome methylation levels showed no significant change. Moreover, 100µM IAA inhibited growth of the 1-year-old poplar clones, possibly through the suppression of photosynthesis. This inhibition had a long-term effect, persisting at 1month after removal of the exogenous IAA. Transcriptome analysis identified two candidate lncRNA genes that show changes in expression following IAA treatment, TCONS_00003480 and TCONS_00004832. TCONS_00003480 contains the same microRNA target sites of ptc-miR6464 as the 4-coumarate: CoA ligase 2 transcript, which encodes a lignin biosynthesis enzyme. And TCONS_00004832shares the same target sites of ptc-miR6437a with the Photosystem II reaction center protein D and Cytochrome C Oxidase 17 transcripts, which are related to photosynthesis. The two lncRNAs as the mimics to corresponding target genes of miRNAs to prevent them from degrading. Examination of lncRNA gene expression and methylation revealed a negative relationship (r = - 0.29, P < 0.05); moreover, hypermethylation of the two candidate lncRNA genes remained 1month after IAA treatment, suggesting that changes in methylation might be involved in the long-term effects of plant hormones. Therefore, our study reveals a long-term effect of IAA on the growth of P. tomentosa, possibly via methylation-mediated epigenetic changes in lncRNA gene expression and the interaction with corresponding miRNAs, leading to regulation of genes related to photosynthesis and growth.

A Single Nucleotide Polymorphism in GAS5 lncRNA is Associated with Risk of Bladder Cancer in Iranian Population

Down-regulation of the long non-coding RNA (lncRNA) growth arrest-specific 5 (GAS5) has a pathogenic role in bladder cancer. Moreover, genomic variants of this lncRNA have been associated with risk of diverse cancers. In the present project, we genotyped two putative functional SNPs (rs2067079 and rs6790) in 122 bladder cancer patients and 150 age- and sex-matched healthy subjects. The rs2067079 was associated risk of bladder cancer in recessive inheritance model (TT vs.CC + CT: OR (95% Confidence interval (CI)) = 2.67 (1.27-5.62), adjusted P value = 0.02). The T G haplotype (rs2067079 and rs6790) increased the risk of bladder cancer in the assessed population (OR (95% CI) = 1.73 (1.18-2.56), adjusted P value = 0.02). Consequently, in the current project we introduced a novel risk locus for bladder cancer in Iranian population.

Expression Pattern of Long Non-coding RNA Growth Arrest-specific 5 in the Remission Induction Therapy in Childhood Acute Lymphoblastic Leukemia.

SummaryBackgroundLong non-coding RNA growth arrest-specific 5 (GAS5) is deregulated in many cancers because of its role in cell growth arrest and apoptosis. Additionally, GAS5 interacts with glucocorticoid receptor, making it a potential pharmacotranscription marker of glucocorticoid (GC) therapy. In this study, we aimed at analysing GAS5 expression in the remission induction therapy phase of childhood acute lymphoblastic leukemia (ALL), in which GCs are mandatorily used, and to correlate it with therapy response.MethodsGAS5 expression was measured in peripheral blood mononuclear cells taken from 29 childhood ALL patients at diagnosis, on day 15 and day 33 of remission induction therapy using RT-qPCR methodology.ResultsOur results have shown interindividual differences in GAS5 expression at all time points. For each ALL patient, GAS5 expression was higher on day 15 in comparison to its level at diagnosis (p<0.0005). On day 33, the level of GAS5 expression decreased in comparison with day 15 (p<0.0005), but it was still significantly higher than at diagnosis for the majority of patients (p=0.001). Patients whose number of blasts on day 8 was below 100 per μL of peripheral blood had a higher GAS5 expression at diagnosis (p=0.016), and lower ratio day 15/diagnosis (p=0.009).ConclusionsOur results suggest that the expression level of GAS5 could be a potential marker of therapy response in remission induction therapy of childhood ALL.

LncRNA GAS5 acts as a ceRNA for miR-21 in suppressing PDGF-bb-induced proliferation and migration in vascular smooth muscle cells.

Aberrant proliferation and migration of vascular smooth muscle cells (VSMCs) contribute significantly to the development of many human cardiovascular diseases, including hypertension. The present study aimed to evaluate the in vitro functional roles of long noncoding RNA growth arrest-specific transcript 5 (GAS5) in VSMCs and explored the underlying molecular mechanisms. We found that GAS5 was significantly downregulated in PDGF-bb-treated VSMCs, and overexpression of GAS5 remarkably attenuated PDGF-bb-induced VSMC proliferation and migration. In addition, miR-21 was observed to potentially bind to GAS5, and we also identified that PDCD4 might be a direct target of miR-21 in VSMCs. Cotransfection of miR-21 mimics remarkably reduced the PDCD4 protein expression in GAS5 overexpressing VSMCs. Further, rescue experiments showed that enforced expression of miR-21 attenuated the inhibitory effects of GAS5 on VSMC proliferation and migration. Collectively, our results demonstrated that GAS5 inhibits PDGF-bb-induced VSMC proliferation and migration, partly through acting as a competing endogenous RNA of miR-21, and provided new evidence that GAS5 may serve as a potential therapeutic target for hypertension.

Tumor-derived exosomal lncRNA GAS5 as a biomarker for early-stage non-small-cell lung cancer diagnosis.

Diagnosis and treatment at an early stage may improve survival of non-small-cell lung cancer (NSCLC). Previous studies have found that long noncoding RNA growth arrest-specific transcript 5 (GAS5) is essential to cancer progression. However, the expression and diagnostic value of GAS5 in exosomes (Exo-GAS5) remain unclear. One hundred and four participants were enrolled, including subjects with NSCLC (n = 64) and healthy subjects ( n = 40). The total Exosome Isolation Kit was applied to isolate exosomes from serum. Total RNA was extracted and the AS5 expression was analyzed using quantitative reverse transcription polymerase chain reaction. Receiver operating characteristic (ROC) curve analysis was applied to evaluate the diagnostic value of Exo-GAS5 in NSCLC. Our data indicated that the Exo-GAS5 was downregulated in patients with NSCLC compared with healthy controls ( p < 0.001). Furthermore, patients with NSCLC with larger tumor size ( p = 0.025) and advanced TNM (T: extent of the primary tumor; N: lymph node involvement; M: metastatic disease) classification ( p = 0.047) showed lower Exo-GAS5 expression. ROC curve analysis using Exo-GAS5 combined with carcinoembryonic antigen showed an area under curve (AUC) of 0.929. Exo-GAS5 could be used to distinguish patients with Stage I NSCLC with an AUC of 0.822. In conclusion, Exo-GAS5 may function as an ideal noninvasive serum-based marker for identifying patients with early NSCLC.

LncRNA GAS5 restrains CCl4-induced hepatic fibrosis by targeting miR-23a through the PTEN/PI3K/Akt signaling pathway.

Hepatic fibrosis is chronic liver damage with many causes that has a relatively high death rate. The current study showed that long noncoding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5), microRNA-23a (miR-23a), and phosphatase and tensin homolog (PTEN) play important roles in the pathological process of hepatic fibrosis but have a relatively unclear regulatory mechanism. This study aimed to investigate the roles of lncRNA GAS5, miR-23a, and PTEN in the pathological process of hepatic fibrosis and hepatic stellate cell (HSC) activation. We used carbon tetrachloride (CCl4) intraperitoneal injections to establish a rat hepatic fibrosis model and exogenous transforming growth factor-β1 to establish an HSC activation model. Quantitative RT-PCR, Western blot, dual-luciferase reporter system, and RNA pull-down assays were used to investigate which microRNAs and lncRNAs participate in the process of hepatic fibrosis and HSC activation. miR-23a expression increased significantly in hepatic fibrosis tissues and activated HSCs. miR-23a interaction with and degradation of PTEN further influenced the downstream signaling pathway phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin/Snail (PI3K/Akt/mTOR/Snail), causing E-cadherin expression levels to decrease and α-smooth muscle actin and collagen I expression levels to increase. lncRNA GAS5 can be used as a sponge platform for miR-23a to decrease miR-23a expression levels competitively. We revealed the role of the lncRNA GAS5/miR-23a/PTEN/PI3K/Akt/mTOR/Snail signaling pathway in hepatic fibrosis, providing molecular targets for the treatment of hepatic fibrosis. NEW & NOTEWORTHY This is the first study revealing that microRNA-23a (miR-23a) promotes hepatic fibrosis through the phosphatase and tensin homolog/phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin/Snail signaling pathway, and long noncoding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5) can act as a sponge platform for miR-23a. Therefore, lncRNA GAS5/miR-23a may bring molecular targets for hepatic fibrosis therapy.

Low GAS5 Levels as a Predictor of Poor Survival in Patients with Lower-Grade Gliomas

Introduction Gliomas are infiltrative neoplasms of a highly invasive nature. Different stages of gliomas feature distinct genomic, genetic, and epigenetic changes. The long noncoding RNA Growth Arrest Specific Transcript 5 (GAS5) is an identified tumour suppressor involved in several cancers. However, the underlying roles of the GAS5 gene in lower-grade glioma (LGG) patients are not clear. Methods Via bioinformatic analysis based on TCGA-LGG and TCGA-GBM data, we explored the mechanisms of GAS5 expression in LGG (grades II and III) and high-grade glioma (glioblastoma multiforme, grade IV). The log-rank test and multivariate Cox analysis were performed to find the association between GAS5 and overall survival (OS) in LGG patients. Weighted gene coexpression network analysis (WGCNA) and RNA-Seq analysis were applied to find the key gene network associated with GAS5. Results We found that GAS5 expression was downregulated in both LGG and glioblastoma multiforme (GBM) compared with normal brain tissue. Low methylation in the GAS5 promoter region was detected in both LGG and GBM tissues. The amplification type was the predominant type of GAS5 gene alteration in both LGG and GBM. High GAS5 expression was more associated with long overall survival (OS) in LGG patients than in GBM patients. The multivariate survival analysis of GAS5 and clinical and molecular characteristics in LGG patients further confirmed the association between GAS5 and OS in LGG patients. We then developed a nomogram for clinical use. WGCNA and RNA-Seq analysis indicated that ribosomal biogenesis and translation initiation were the predominant events regulated by GAS5 in LGG patients. Conclusion Taken together, these results demonstrate that GAS5 expression is associated with OS in LGG patients and that its underlying roles involve the regulation of ribosomal biogenesis and translation initiation, which may aid in identifying a new target for the treatment of LGG.

Inhibition of long noncoding RNA growth arrest–specific 5 attenuates cerebral injury induced by deep hypothermic circulatory arrest in rats

ObjectiveWe sought to investigate cerebroprotection by targeting long noncoding RNA growth arrest–specific 5 in a rat model of prolonged deep hypothermic circulatory arrest. MethodsDeep hypothermic circulatory arrest was conducted for 60 minutes when the pericranial temperature was cooled to 18°C in rats. Dual luciferase assay was used to detect the binding relationship between growth arrest–specific 5 and putative target microRNAs. Adeno-associated viral vectors containing growth arrest–specific 5 small interfering RNA or negative control small interfering RNA were administered by intracerebroventricular injection 14 days before deep hypothermic circulatory arrest. Expressions of growth arrest–specific 5, microRNA-23a, phosphate and tension homology, Bcl-2–associated X protein, Bcl-2, phospho-protein kinase B, protein kinase B, and cleaved caspase-3 in the hippocampus were measured by quantitative reverse transcription polymerase chain reaction and Western blot. Spatial learning and memory functions were evaluated by the Morris water maze test. The hippocampus was harvested for histologic examinations and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining. ResultsLuciferase assay showed that growth arrest–specific 5 targeted and inhibited microRNA-23a expression. After deep hypothermic circulatory arrest, hippocampal growth arrest–specific 5 expression was significantly enhanced with a robust decrease of hippocampal microRNA-23a expression. Small interfering RNA growth arrest–specific 5 significantly inhibited growth arrest–specific 5 expression and enhanced microRNA-23a expression in the hippocampus, accompanied with decreases of phosphate and tension homology and Bcl-2–associated X protein expression, and increases of Bcl-2 expression and phospho-protein kinase B/protein kinase B ratio. Growth arrest–specific 5 knockdown inhibited neuronal apoptosis, attenuated histologic damages, and increased the number of surviving neurons in the hippocampus. Spatial learning and memory functions after deep hypothermic circulatory arrest were also markedly improved by growth arrest–specific 5 inhibition. ConclusionsInhibition of large noncoding RNA growth arrest–specific 5 can provide a powerful cerebroprotection against deep hypothermic circulatory arrest, which may be mediated through microRNA-23a/phosphate and tension homology pathway.