The LncRNA MIR155HG Promotes Temozolomide Resistance By Activating The Wnt/β-catenin Pathway Via Binding To PTBP1 In Glioma

Objectives: TMZ is widely used for glioma therapy in the clinic.The development of TMZ resistance has largely led to poor prognosis.Very little is understood about the role of MIR155 HG,as a long noncoding RNA,in TMZ resistance.Materials and Methods: qRT-PCR,western blotting and immunofluorescence were used to measure the levels of MIR155 HG and proteins.RNA-binding protein immunoprecipitation assay was used to analyze the interaction between MIR155 HG and PTBP1.Flow cytometry apoptosis assay,CCK-8 assay,colony formation assay and animal experiments were used to detect the effects of MIR155 HG on TMZ resistance in glioma.Results: Bioinformatics analysis showed that MIR155 HG expression was higher in glioma specimens than in normal brain specimens,and the survival rate was significantly lower in patients with high expression of MIR155 HG than in patients with low expression of MIR155 HG.CCK-8 and apoptosis experiments showed that the cell viability was significantly higher in pMSCV-MIR155 HG group than in pMSCV group and the apoptosis rate was lower in pMSCV-MIR155 HG group than in pMSCV group after TMZ treatment.In contrast,the cell viability was decreased after transfection with siMIR155 HG,compared with the cell viability in the NC group.A higher percentage of apoptotic cells was observed in the siMIR155 HG groups than in the NC groups.After TMZ treatment,the average final weight and volume of tumors was higher in sMSCV-MIR155 HG group than in sMSCV group.After TMZ treatment,the cell viability was lower in siPTBP1 group than in NC group,and the apoptosis rate was higher in siPTBP1 group than in NC group.RIP assay revealed that MIR155 HG was specifically enriched using the PTBP1 antibody in U251 cells.Changes in the mRNA level of PTBP1 were statistically insignificant in U251 cells with MIR155 HG overexpression or knockdown.However,there was obviously a positive correlation between the MIR155 HG expression and the protein level of PTBP1 by western blotting and immunofluorescence assay.Western blotting and immunofluorescence assay demonstrated a dramatic increase in the expression of the proteins related to the Wnt/β-Catenin pathway in pMSCVMIR155 HG group compared with pMSCV group.Furthermore,a significant decrease in the expression of the proteins related to the Wnt/β-Catenin pathway could be seen in the siMIR155 HG group and in the siPTBP1 group compared with the control group.As determined by CCK-8 assays and flow cytometry,the increase of the cell viability and the decrease of apoptosis rate caused by MIR155 HG overexpression could be reversed by reducing the expression of PTBP1.LiCl significantly weakened the decrease of viability and the increase of apoptosis rate caused by PTBP1 knockdown.Conclusions: MIR155 HG level is markedly higher in glioma patients than in normal controls and that the survival rate is positively correlated with MIR155 HG expression.It is apparent that TMZ sensitivity is promoted by downregulation of MIR155 HG,and this can be reversed by MIR155 HG overexpression in vivo and in vitro.Furthermore,PTBP1 is proven to bind with MIR155 HG and regulate MIR155HG-related TMZ resistance.Further investigation indicates that the expression levels of both MIR155 HG and PTBP1 influence the expression of relevant proteins in the Wnt /β-catenin pathway.Above all,the study demonstrates that the knockdown of MIR155 HG increases glioma sensitivity to TMZ by inhibiting Wnt /β-catenin pathway activation via potently downregulating PTBP1.