Effects Of Eukaryotic Elongation Factor 2 Kinase (eEF2K) On Proliferation, Migration And Invasion Of Glioma Cells

Objective: Glioma is one of the most common tumors in the central nervous system.Due to its strong invasion ability,it is difficult to remove completely by surgery,and the existing treatment methods can only moderately prolong the survival of patients.Finding new treatment methods has become the main goal of current research.In recent years,many studies have shown that eEF2 k is closely related to the occurrence and development of various tumors.Therefore,exploring the effect of eEF2 k on the proliferation,migration and invasion of glioma cells can provide new targets and ideas for treatment.Methods:(1)Select normal astrocytes HA1800 as the control group,and glioma cell lines U87 and T98 as the experimental group.The expression of eEF2 k m RNA in normal astrocytes and glioma cells was detected by real-time fluorescence quantitative PCR;The protein expression of eEF2 k in normal astrocytes and glioma cells was detected by Western blot.(2)The glioma cell lines U87 and T98 were transfected with lentivirus,and the transfection efficiency was verified by green fluorescence test and western blot test.(3)The proliferation of U87 and T98 cells at different time periods(0h,24 h,48h,72 h,96h)was detected by CCK8 proliferation test.(4)The effect of eEF2 k on the migration and invasion ability of glioma U87 and T98 cells was detected by cell Migration test and invasion test.(5)To further verify the effect of eEF2 K on the tumorigenicity of U87 glioma cells through in vivo experiments: the 4-week-old nude mice were fed for one week to adapt to the environment and were randomly divided into two groups,U87 group and U87-sh RNA group,which were injected into the right armpit of the nude mice,and the tumor size was measured every 5 days,and the tumor body was killed and weighed after 35 days.Results:(1)Compared with normal astrocyte HA1800,the expression of m RNA and protein in glioma cell lines U87 and T98 was significantly higher(p<0.05).(2)After 72 hours of lentivirus transfection,the expression rate of green fluorescence was more than 80% under the fluorescence microscope.Western blot results showed that compared with sh-con group,the expression of eEF2 k protein in U87 cells in sh-1,sh-2 and sh-3 groups decreased significantly(P<0.001);The expression of eEF2 k in T98 cells in sh-3 group decreased(P<0.001),while there was no significant difference in protein expression between sh-1 group and sh-2 group and sh-con group(P>0.05).(3)The results of CCK8 showed that compared with sh-con group,the proliferation ability of U87 and T98 cells decreased significantly after48 hours(**P<0.01,***P<0.001).(4)The results of cell scratch test showed that compared with sh-con group,the scratch healing rate of sh-eEF2 K glioma cell group decreased significantly(**P<0.01,***P<0.001).(5)Transwell invasion test results showed that compared with sh-con group,the number of cells passing through the compartment in sh-eEF2 K glioma cell group was significantly reduced,with a statistically significant difference(**P<0.01,***P<0.001).(6)Compared with U87 group,the volume and weight of tumor in U87-sh RNA group were significantly reduced after knocking down eEF2 K in nude mice tumorigenesis experiment,and the difference was statistically significant(***P<0.001).Conclusion: eEF2 K is highly expressed in glioma cells.Knocking down eEF2 K can inhibit the proliferation,migration and invasion of glioma U87 and T98 cells.