| Background:Glioma is a typical intracranial malignant tumor.it is often unclear from the surrounding normal tissues because of high invasive growth characteristics,making it difficult to achieve total resection.The recurrence rate is also high and the survival time of patients is short because of its high degree of malignancy and resistant to chemotherapeutic drugs.At present,surgery and chemoradiotherapy are the most mainstream treatment options in clinical applications,but there are still some limitations and poor treatment effects.Temozolomide is currently the most widely used chemotherapy agent for glioma.It can penetrate the blood-brain barrier and induce DNA alkylation of tumor cells,promoting tumor cell apoptosis.It is the only chemotherapy drug that can significantly improve the quality of life of glioma patients.How to reduce the resistance of glioma to temozolomide and strengthen the efficacy of temozolomide is the focus of current research on glioma treatment.MGMT is an important mechanism of TMZ resistance in glioma cells.It reverses TMZ alkylation and repairs damaged DNA by transferring the methyl group on TMZ-induced O6-methylguanine,making tumor cells resistant to TMZ.Studies have shown that the energy source of tumor cells is mainly through glycolysis,and the increase of aerobic glycolysis level is an important reason for tumor resistance.Therefore,current scientific research is trying to explore how to promote the apoptosis of tumor cells by inducing glycolysis dysfunction of tumor cells in order to obtain better therapeutic effects.The effect of TMZ on the glycolysis function of tumor cells and the relationship between glycolysis and MGMT need to be further studied to provide theoretical value for improving the prognosis of glioma patients.Objective:Randomly selected glioma pathology samples and human glioma cell lines were examined for the expression of MGMT,glycolytic regulatory proteins HK Ⅱ,PFKP,and PKM2,and the role of glycolytic dysfunction in the killing of glioma cells by temozolomide and the expression of MGMT in tumor cells was investigated.Methods:1.Western blotting detect the expression levels of glycolytic regulatory proteins HK Ⅱ,PFKP and PKM2 in 4 groups of primary gliomas and their peritumoral tissues,as well as 6 primary gliomas of different pathological grades and 2 peritumoral tissues.2.Western blotting detect the expression levels of MGMT and glycolytic regulatory proteins HKⅡ,PFKP and PKM2 in U87 and U251 glioma cell lines after TMZ treatment at different time points.The content of ATP and pyruvate in the final products of two glycolytic products was detected.3.Different concentrations of TMZ were applied to U87 and U251 cells for grouping.Different concentrations of sodium pyruvate were added and a blank control group was set up to detect the cell death rate of each group.Different concentrations of TMZ were applied to U87 and U251 cells for grouping,PAS was added respectively and a blank control group was set up to detect the expression level of MGMT in each group of cells.4.Different concentrations of TMZ were applied to U87 and U251 glioma cell lines,and MGMT inhibitor lomeguatrib was added to each group and a blank control group was set up.The expression levels of MGMT and glycolytic regulatory proteins HK Ⅱ,PFKP,PKM2,cell death rate,ATP and pyruvate content were detected in each group.5.U87 and U251 cells were divided into administration group and non-administration group according to whether TMZ was administered.Each group was further divided into positive transfection group,negative transfection group and blank control group according to whether si RNA was transfected.The expression levels of MGMT and glycolysis regulatory proteins HK Ⅱ,PFKP and PKM2 in cytoplasm and nucleus of tumor cells,cell death rate and ATP and pyruvate content were detected in groups.Results:1.The expression levels of HK Ⅱ,PFKP and PKM2 in the cytoplasm of tumor tissues in the four groups were significantly higher than those in the peritumoral tissues.The higher the pathological grade of glioma,the higher the expression level of glycolytic mediation proteins HK Ⅱ,PFKP and PKM2.2.With the prolongation of TMZ action time,the expression level of MGMT increased,and the expression levels of HKⅡ,PFKP and PKM2 decreased.The intracellular ATP and pyruvate contents were significantly decreased after TMZ treatment.3.Under the same concentration of TMZ,the cell death rate and MGMT expression level decreased after PAS was added.4.Under the same concentration of TMZ,after adding MGMT inhibitor lomeguatrib,the expression level of MGMT decreased,the cell death rate further increased,the expression levels of glycolytic regulatory proteins HK Ⅱ,PFKP,PKM2 and the contents of ATP and pyruvic acid further decreased.5.The expression levels of MGMT,HK Ⅱ,PFKP,PKM2,ATP and pyruvate in the positive transfection group were lower than those in the negative transfection group or the blank control group,while the cell death rate in the positive transfection group was higher than that in the negative transfection group or the blank control group.Conclusion:1.The glycolytic regulatory proteins HK Ⅱ,PFKP and PKM2 are highly expressed in glioma tissues,and the expression level is up-regulated with the increase of pathological grade of glioma tissues.2.TMZ-induced glycolytic dysfunction promotes MGMT upregulation3.MGMT inhibits TMZ-induced glycolysis dysfunction... |
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