| Objective: Human brain glioma is the most malignant primary intracranial tumor,which has poor prognosis and high mortality.It is important and necessary to elucidate the underlying mechanism of gliomagenesis as well as find novel targeted therapeutic strategy.Long noncoding RNAs(lncRNAs)implicated in tumorigenesis and progression of numerous tumors through participating in various important cellular biological behaviors,including proliferation,cell cycle,invasiveness and so on.LncRNAs could act as oncogenes or tumor suppressors in glioma,and be diagnostic or prognostic biomarkers for glioma.Small Nucleolar RNA Host Gene 5(SNHG5)gene belongs to lncRNA gene because it lacks the ability to encode protein.Recent researches suggested that SNHG5 showed abnormal expression and function in malignant tumors,such as bladder cancer,colorectal cancer and gastric cancer.Till now,there is no research on the expression level and functional roles of SNHG5 in glioma.It is well known that Wnt/?-catenin signaling pathway participates in the regulation of growth,development and metabolism,and plays important regulatory roles in almost all kinds of malignant tumors.Accumulating evidences showed Wnt/?-catenin signaling pathway was activated frequently during tumorigenesis and progress of glioma.Nevertheless,the interaction between Wnt/?-catenin signaling pathway and SNHG5 in glioma was poorly demonstrated.This study will investigate the impacts of SNHG5 gene on malignant cellular phenotypes of glioma cells and explore the possible mechanisms.Methods: 1.Cell culture of normal human astrocytes(NHA),U251 and U87 cells.2.Collection of glioma tissue and corresponding normal brain tissue(NBT)specimens.3.Arraystar LncRNA Array was used to screen differentially expressed lncRNAs between glioma and NBT specimens.4.Quantitative real time PCR(qRT-PCR)was used to detect the expression of SNHG5.5.The knockdown vectors of SNHG5 was transfected into U251 and U87 cells.6.Enhanced Cell Counting Kit-8 was used to examined the cell proliferation of U251 and U87 cells.7.Transwell assay was used to examined the cell invasiveness of U251 and U87 cells.8.Flow cytometry was used to examined the cell apoptosis of U251 and U87 cells.9.Luciferase reporter assay was used to quantify the activity of Wnt/?-catenin signaling pathway in U251 and U87 cells.10.Western blotting was applied to determine the expression of GSK3 B and CTNNB1 protein in U251 and U87 cells.Results: 1.The expression of SNHG5 in glioma specimens and cell lines(U251 and U87)were much higher than that in NBT specimens and NHA cell line(P<0.01).2.The increased expression of SNHG5 was correlated with high pathological grade of glioma(P<0.01),while was independent of other parameters,including age and gender of glioma patients(P>0.01).3.Transfection with sh-SNHG5 could knockdown the expression of SNHG5 in U251 and U87 cells(P<0.01).4.Knockdown of SNHG5 inhibited the cell proliferation ability of U251 and U87 cells(P<0.01).5.Knockdown of SNHG5 inhibited the cell invasiveness of U251 and U87 cells(P<0.01).6.Knockdown of SNHG5 advanced the cell apoptosis of U251 and U87 cells(P<0.01).7.The co-expression analysis in TCGA Pan-Cancer(PANCAN)database with 171 glioma samples displayed the expression of SNHG5 had significant negative correlation with GSK3B(r=-0.241,P<0.01)as well as positive correlation with CTNNB1(r=0.2071,P<0.01).8.Knockdown of SNHG5 inhibited the ratio of TOP/FOP luciferase values in U251 and U87 cells(P<0.01).9.Knockdown of SNHG5 increased the expression of GSK3 B protein and inhibited the expression of CTNNB1 protein in U251 and U87 cells(P<0.01).10.Compared with shSNHG5 + pE-NC groups,the ratio of TOP/FOP luciferase values sh-SNHG5 + pECTNNB1 groups U251 and U87 cells increased(P<0.01).11.Compared with sh-SNHG5 + pE-NC groups,the expression of CTNNB1 protein in sh-SNHG5 + pE-CTNNB1 groups U251 and U87 cells increased(P<0.01).12.Compared with sh-SNHG5 + pE-NC groups,the cell proliferation ability in sh-SNHG5 + pE-CTNNB1 groups U251 and U87 cells increased(P<0.01).13.Compared with sh-SNHG5 + pE-NC groups,the cell invasiveness in sh-SNHG5 + pE-CTNNB1 groups U251 and U87 cells increased(P<0.01).14.Compared with sh-SNHG5 + pE-NC groups,the cell apoptosis rate in sh-SNHG5 + pECTNNB1 groups U251 and U87 cells decreased(P<0.01).Conclusion: SNHG5 was highly expressed in gliomas and positively correlated with the pathological grade of gliomas.SNHG5 can positively regulate the biological behaviors of glioma cells through Wnt/beta-catenin signaling pathway. |
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