| Glioma is the most common and invasive primary malignant tumor of the central nervous system(CNS),with high recurrence rate,high mortality and low cure rate.The treatment of glioma remains a major challenge in the field of neurosurgery.Although some progress has been made in surgery,radiotherapy,chemotherapy,molecular targeted therapy and tumor electric field therapy(TTFields)in recent years,the prognosis of glioma patients is still unsatisfactory.Especially for patients with glioblastoma,the median survival time is only about 14 months.The unclear pathogenesis of glioma is an important reason restricting its diagnosis and treatment.Therefore,it is the focus of research in the field of neurosurgery to further explore the molecular biological mechanism in the tumorigenesis and progression of glioma and find specific therapeutic targets.Long non-coding RNA(lnc RNA)is a group of RNAs with a length over 200 nucleotides and no protein coding function.Lnc RNA can participate in the regulation of target genes at multiple levels,including transcription,post-transcription,translation and post-translation,and thus play important biological functions.The aberrant expression of Lnc RNA can affect the tumorigenesis and progression,and act as potential molecular marker and therapeutic target for the diagnosis and prognosis in glioma.Although several lnc RNAs that are differentially expressed in gliomas have been identified and studied in depth,the biological functions and molecular mechanisms of most lnc RNAs in glioma remain unclear.By screening the microarray data completed by our research team earlier,we found that LINC00909 was differentially expressed in gliomas.LINC00909 is a novel long non-coding RNA molecule,and there is no report on tumor research.In this study,we confirmed by q RT-PCR that LINC00909 was overexpressed in glioma tissues,and the expression level of LINC00909 was positively correlated with the WHO grade of glioma.Kaplan-Meier survival analysis showed that increased expression of LINC00909 was associated with poor prognosis in glioma patients.Function assays in vitro showed that interfering the expression of LINC00909 inhibited proliferation,clone formation,migration and invasion,and affected the epithelial-mesenchymal transition(EMT)process of glioma cells.Study on the mechanism of lnc RNAs as competing endogenous RNAs(ce RNAs)in glioma is one of the current hotspots.Lnc RNAs can regulate the expression of target genes by sponging mi RNAs,thus participating in the regulation of biological functions of tumor cells.Therefore,we speculated that LINC00909 may alsofunction as ce RNA in glioma.Bioinformatics analysis and luciferase reporter assay revealed that both LINC00909 and the 3'UTR of MUC1 have the region of mi R-194 binding site,and could adsorb mi R-194.q RT-PCR and Western blot showed that interference with LINC00909 down-regulated the expression level of MUC1 in glioma cells,while mi R-194 inhibitor reversed this effect.LINC00909 may promote the progression of glioma by competitively binding mi R-194 to regulate the expression of MUC1.Our results indicate that LINC00909 has the potential to be a novel diagnostic marker and therapeutic target for glioma.Part ? Study on the expression and clinical significance ofLINC00909 in gliomaObjective: Identifying the expression of LINC00909 in glioma tissues and normal brain tissues,and analyzing its relationship with the prognosis and clinical indicators of patients,in order to provide theoretical basis for proving whether LINC00909 could be used as a molecular marker for glioma diagnosis and prognosis evaluation in patients.Methods: A total of 112 cases tissue specimens(102 glioma tissues and 10 normal brain tissues)were collected in Shanghai Changzheng Hospital from 2014 to 2016.The RNA expression level of LINC00909 in gliomas and normal brain tissues was detected by q RT-PCR.The clinical information of the corresponding patients was collected,and the correlation between the expression of LINC00909 and prognosis of the patients and other clinical observation indicators was analyzed by statistics.Results: The results of q RT-PCR showed that,compared with normal brain tissues,the expression of LINC00909 was significantly up-regulated in glioma tissues,and its expression in high-grade glioma tissues was significantly higher than that in the low-grade gliomas tissues.Based on statistical analysis of various clinical indicators of patients,the results suggested that the expression of LINC00909 was positively correlated with the WHO grade of glioma.Survival analysis results showed that the expression of LINC00909 was negatively correlated with the overall survival time of patients.Univariate analysis showed that the expression of LINC00909 was significantly correlated with the prognosis of glioma patients.Multivariate COX regression analysis showed that the expression of LINC00909 was an independent risk factor affecting the prognosis of patients.CONCLUSION: The expression of LINC00909 in glioma tissues is significantly up-regulated.The expression level of LINC00909 is positively related to the WHO gradeof glioma and negatively related to the overall survival time of patients.The expression of LINC00909 is an independent risk factor for the prognosis of patients.LINC00909 can be used as a potential biomarker for diagnosis and prognosis evaluating of glioma.Part ? Study on the biological function of LINC00909 in gliomacellsObjective: To investigate the biological function of LINC00909 in the progression of glioma.Methods: Small interfering RNAs targeting LINC00909 were designed and synthesized,and then transfected into glioma cell lines U87 MG and U251 using Lipofectamine 3000.MTT assay,clone formation assay,Transwell migration and invasion assay,and Western blot assay were performed to observe the effect of LINC00909 interference on proliferation,clone formation,migration and invasion,and epithelial-mesenchymal transition(EMT)process of glioma cells.Results: LNC00909 was highly expressed in glioma cell lines U87 MG and U251,and the expression of LINC00909 was significantly down-regulated after transfection with si RNA.After interference with LINC00909 expression,the proliferation,clone formation,migration and invasion capabilities of glioma cell lines U87 MG and U251 were significantly decreased,and the epithelial-mesenchymal transition process was inhibited.Conclusion: LINC00909 can affect the proliferation,clone formation,migration,invasion,and epithelial-mesenchymal transition of glioma cells,and may play the role of oncogenes in glioma.LINC00909 can be used as a potential therapeutic target of glioma.Part ? Study on the molecular mechanism of LINC00909 ingliomaObjective: To explore the molecular mechanism of LINC00909 in gliomas,and to provide new targets for clinical treatment of gliomas.Methods: The mi RNA and its downstream target genes that may interact with LINC00909 were predicted by bioinformatics analysis.The regulatory mechanism of LINC00909 as ce RNA in glioma was studied by q RT-PCR,luciferase reporter assay,and Western blot.Results: Bioinformatics analysis showed that mi R-194 was differentially expressed inglioma and may have a binding site with LINC00909.The results of q RT-PCR revealed that the expression of mi R-194 in glioma cell lines U87 MG and U251 was up-regulated after interference with LINC00909;the expression of mi R-194 was significantly down-regulated in glioma tissue and negatively correlated with the expression of LINC00909.The luciferase reporter assay confirmed that LINC00909 has the binding site of mi R-194,and could adsorb it.MUC1 was predicted by bioinformatics analysis to be one of the possible target genes of mi R-194 in glioma.The results of q RT-PCR showed that the expression of MUC1 in glioma cell lines U87 MG and U251 was significantly decreased after overexpression of mi R-194.Analysis results from TCGA database showed that the expression of MUC1 was up-regulated in glioma tissues,and the increased expression of MUC1 was correlated with the poor prognosis of glioma patients.The luciferase reporter assay confirmed that the 3'UTR region of MUC1 also has the binding site of mi R-194,and could adsorb it as well.The results of q RT-PCR and Western blot showed that interference with LINC00909 down-regulated the m RNA and protein expression levels of MUC1 in glioma cell lines U87 MG and U251,while mi R-194 inhibitor reversed this effect.Conclusion: LINC00909 can adsorb mi R-194,and MUC1 is the target gene of mi R-194.LINC00909 can regulate the expression level of MUC1 by competitively binding mir-194.As an oncogenic lnc RNA,LINC00909 may promote the progression of glioma by regulating the mi R-194 /MUC1 axis. |
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