The Study Of CXCR4Modulate Oral Squamous Cell Carcinoma EMT To Influence Tumor Cell Metastasis

Part oneObjectiveTo detect the expression level of CXCR4, β-catenin and EMT related proteins among OSCC samples with different LN metastasis status. To further detect the correlation between those proteins and relative clinicopathological features.MethodImmunohistochemistry stain was employed to detect the CXCR4, β-catenin and EMT related proteins expression in60cases OCSS, χ2test or Fish exact method was analysed the relationship between those proteins and clinicopathological features, Spearman was analysed the correlation between CXCR4and EMT related proteins.Result1.In lymph node metastasis group, the expression of CXCR4, N-cadherin, Twist, Snail were higher than those in non-lymph node metastasis group; however the expression of β-catenin and E-cadherin were lower than those in non-lymph node metastasis group. The difference was statistically significant (β<0.05).2.The expression of CXCR4and Twist were higher in moderate or poor differentiation, The expression of E-cadherin were higher in high differentiation (P <0.05).3.There was negatively correlated betweenβ-catenin and CXCR4, but positively orrelated with Twist, Snail, N-cadherin (P<0.05).ConclusionEMT plays an important role in the process of lymph node metastasis of oral squamous cell carcinoma, CXCL12/CXCR4biological axis may act on a particular aspect in the EMT, and regulate its occurrence.Part twoObjectiveWe observed migration, invasion, apoptosis, and the expression of Canonical Wnt pathway protein of the Tscca cell after silenced CXCR4by si RNA.Method1.We transfected CXCR4siRNA to Tscca cells by liposome transfection method, and detected the inhibition of siRNA by RT-PCR.2.We observed the ability of Tcssa cells migration after silenced CXCR4by wound 2.We observed the ability of Tcssa cells migration after silenced CXCR4by wound healing assay.3.We observed the ability of Tcssa cells invasion after silenced CXCR4by transwell chamber.4.We detect the apoptosis of Tcssa cells after silenced CXCR4by Annexin V.5.We detect the expression of Canonical Wnt pathway protein after silenced CXCR4by western blot.Result1.RT-PCR results showed that siRNA significantly inhibited the expression of CXCR4.2.The wound healing assay proved Tscca cell migration decrease after silenced CXCR4.3.Transwell invasion assay proved Tscca cell invasion decrease after silenced CXCR4, P<0.01.4.Flow cytometry was used to detect the early phage apoptosis of Tscca, the results showed that increase after silenced CXCR4, P<0.05.5.Western blot showed the expression level of MMP-2, MMP-9and N-cadherin was decrease, but E-cadherin was increase after cells were silenced CXCR4.Conclusion1. Silencing CXCR4gene can suppressed migration and invasion of tongue squamous cell carcinoma cell lines Tscca, CXCR4plays an important role in metastasis of oral squamous cell carcinoma.2.Silencing CXCR4gene can significantly inhibit the expression of MMP-2, MMP-9and-cadherin of Canonical Wnt pathway, but significantly increase the expression of E-cadherin. The CXCR4possible by regulating canonical Wnt pathway involved in the occurrence of EMT in oral squamous cell carcinoma.Part threeObjectThe explore the CXCR4siRNA transfected human tongue squamous cell carcinoma tumor model inhibit tumor growth and the change protein expression of Canonical Wnt pathway related factors.MethodThrough the establishment of the tongue squamous cell carcinoma tumor model, intratumorally inject mixture of lipofectamine2000and CXCR4siRNA, observe ResultSuccessfully established human tongue squamous cell carcinoma tumor model, measure tumor volume every three days. Compared to the control group and scramble group, the tumor of treatment group growed slowly after local multi-point injection lipofectamine2000and CXCR4siRNA mixture. IHC showed expression level of MMP-2, MMP-9, N-cadherin was decreased, but E-cadherin was increased after tumor specimens were knockdowned CXCR4.Conclusion1.Applying lipofectamine2000and CXCR4siRNA for human tongue squamous cell carcinoma tumor model, the results that the activity of the Canonical Wnt pathway was inhibited, and the expression of E-cadherin was increased after knockdown CXCR4, the same with in vitro.2.CXCR4may be effective target for inhibition of the metastasis to oral squamous cell carcinoma.