| BackgroundGlioblastoma(GBM)is the most common malignant tumor of the central nervoussystem.GBM patients have a poor prognosis due to the rapid proliferative capacity ofGBM cells,the invasive growth pattern and the high tolerance to radiotherapy andchemotherapy.Studies have found that the proliferation of GBM cells and theoccurrence and development of GBM are regulated by a variety of genes,so it is apolygenic lesion in nature.Therefore,it has become a new breakthrough point in gliomatreatment to study the relationship between related factors and seek new molecular genetargets at the level of gene regulation.Micro RNAs(miRNAs)are endogenous,single-stranded non-coding smallmolecule RNAs with highly conserved sequences.They can participate in generegulation through different signaling pathways and they can bind with target genes tochange the expression of target proteins.Mi RNAs play an important role in the normalphysiological function of glial cells,in the regulation of gene expression in glioma cells,and in the signal transduction caused by them.The role of miRNA-16 in the malignant progression of glioma has been found.Ithas been reported that miRNA-16 is overexpressed in GBM U87 and U251 cells,it caninhibit the adhesion and invasion of tumor cells,down-regulate gene expression,and itmay play a role in glioma invasion through the mechanism of epithelium-mesenchymaltransformation(EMT).These results suggest that miRNA-16 may be a potentialtherapeutic target and prognostic indicator for glioblastoma.Target Scan(http://www.targetscan.org)online tool was used to predict the Targetgenes of miRNA-16-1.These genes included the proliferation related genes Cyclin D1(CCND1),Cyclin E1(CCNE1),CDK6 and SOX5.It is inferred that miRNA-16-1 mayaffect the proliferation of GBM by regulating the expression levels of these target geneproteins,and thus affect the prognosis of GBM patients.ObjectiveTo investigate the possible regulatory role of miRNA-16-1 and some of its target genes in the proliferation of GBM cells and their relationship with the prognosis of patients.Method1.Paraffin-embedded specimens were collected from 132 cases of pathological diagnosis of GBM.And clinical pathological data of the patients were collected.2.The Target genes of miRNA-16-1 were verified by the Dual Luciferase Report Assay.3.The expression of miRNA-16-1 in 132 cases of GBM tissues was detected by RTPCR.4.The m RNA expression levels of CCND1,CCNE1,CDK6 and SOX5 in 132 GBM tissues were detected by RT-PCR.5.The expressions of CCND1,CCNE1,CDK6,SOX5 and Ki67 protein in 132 GBM tissues were detected by immunohistochemistry.6.Kaplan-Meier method was used to analyze the relationship between miRNA-16-1expression and OS of GBM patients,and the relationship between the level of each target gene and overall survival(OS)of GBM patients.Result1.In 132 cases of GBM paraffin tissues,miRNA-16-1 was low expressed in 77cases(58.33%)(1.81±0.72)and high in 55 cases(41.67%)(16.61±0.65),the difference was statistically significant.2.Dual luciferase assay confirmed that CCND1,CCNE1,CDK6 and SOX5 in GBM cells were the target genes of miRNA-16-1.3.The expression of CCND1,CCNE1,CDK6,and SOX5 decreased in the cases with high miRNA-16-1 expression,and increased in the cases with low miRNA-16-1 expression.The difference was statistically significant.4.The GBM cell proliferation index Ki67 of miRNA-16-1 high expression group was significantly decreased,while the GBM cell proliferation index Ki67 of miRNA-16-1 low expression group was significantly increased,the difference was statistically significant(p< 0.001).5.The median OS of patients in the miRNA-16-1 high expression group and low expression group were 19.00±2.61 months and 10.00±3.36 months.The survival time of the former was significantly higher than that of the latter,and the difference was statistically significant(p<0.05).The survival time of GBM patients in the low expression group of CCND1 and SOX5 were longer than that in the high expression group,and the differences were statistically significant(p<0.05).The levels of CDK6 and CCNE1 were not significantly correlated with OS in GMB patients.Conclusion1.Mi RNA-16-1 inhibits the proliferation of GBM by regulating the m RNA of target genes CCND1,CCNE1,CDK6 and SOX5.2.GBM patients with low miRNA-16-1 expression have poor prognosis,which can be used as a potential molecular marker for the prognosis of GBM. |
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