Effects Of Farnesyltransferase ? Silencing On The Proliferation,Migration And Invasion Of Tongue Squamous Cell Carcinoma Cells

Objective: Oral cancers are mostly squamous cell carcinomas,among which tongue squamous cell carcinoma(TSCC)is one of the most common malignancies of oral squamous cell carcinomas,with the characteristics of fast growth and strong infiltration.Local recurrence,lymph node metastasis and distant metastasis are the main factors for poor prognosis of tongue squamous cell carcinoma.Therefore,to elucidate the molecular mechanism of tongue squamous cell carcinoma proliferation,migration and invasion and determine its therapeutic target are crucial for clinical diagnosis and treatment.It is now found that Ras protein plays an important role in the process of tumor formation and development.HRAS mutation is more common in oral cancer.Post-translational modification of the RAS protein isopentenylation is essential for its cell membrane localization and biological activity.Farnesyl transferase(FTase)is a key enzyme involved in the post-translational modification of HRAS prenylation.In this study,RNA interference was used to silence FTase ?,and the effect and mechanism of silencing FTase ? on the proliferation,migration and invasion of TSCC cells were discussed.Methods: Based on the human FTase ? gene sequence,three FTase ?-siRNA sequences were designed and synthesized,and the three siRNAs were transfected into 3 groups of TSCC CAL27 cells by liposome-mediated method.At the same time,A negative control group(transfected with NC-siRNA)and a blank control group(only with cell transfection reagents,without any siRNA)were established.(1)Real-time quantitative polymerase chain reaction(PCR)was used to detect the mRNA expression of FTase ? and HRAS in each group of cells after silencing FTase.According to the expression of FTase ? mRNA,the group with the highest silencing efficiency was selected as the experimental group for further study.(2)Western blotting was used to detect the expression of FTase ?,HRAS,AKT,pAKT,p65,p-p65(S536),MMP-9,Cyclin D1,and survivin protein in each group after silencing FTase ?.(3)The effect of silencing FTase ? on CAL27 cell proliferation was detected by using CCK-8 method.(4)The effects at 48 hours post transfection on the migration and invasion of CAL27 cells were detected by the scratch test and Transwell test..The data was analyzed by SPSS 19.0.T test was used for the comparison among groups,and a single factor analysis of variance was used for the comparison among the various numbers.P<0.05 had statistical significance.Result:(1)The data showed that compared with the negative control group and the blank control group,the expression of FTase ? mRNA in the experimental group decreased(P<0.05).In the comparison of FTase ? mRNA expression among the experimental groups,the silencing efficiency of the FTase ?-siRNA-3 group was the highest,which was further studied as the experimental group.There was no statistical difference in HRAS mRNA expression among the groups(P>0.05).(2)The results showed that compared with the negative control group and the blank control group,the protein expression of FTase ?,pAKT,p-p65(S536),MMP-9,Cyclin D1,and survivin decreased in the experimental group(P<0.05).No significant difference of HRAS,AKT,and p65 protein expression expression was detected after FTase ? silencing(P>0.05).(3)cell proliferation activity of siRNA group was significantly decreased after 48 h transfection,and the difference was statistically significant(P<0.05).(4)Compared with the negative control group and the blank control group,the total number of invasive cells on the PET membrane of the FTase ?-siRNA group was significantly reduced(P<0.05).(5)Wound healing assay showed that wound migration rate was significantly reduced after FTase ? knockdown compared to the negative control group and the blank control group(P<0.05).Conclusion: In this study,the data suggested that silencing FTase ? in TSCC cells may reduce the protein expression of Cyclin D1 and survivin through the AKT pathway,thereby inhibiting cell proliferation;and may reduce the expression of MMP-9 protein through the NF-?B pathway,thereby inhibiting cell migration and invasion.This suggests that FTase plays a regulatory role in the proliferation,migration and invasion of TSCC cells,and further research on the specific target and mechanism of FTase may help provide important evidence for the targeted treatment of TSCC.