The Study Of VHL Modulates Epithelial-mesenchymal Transition In Oral Squamous Cell Carcinoma

ObjectiveLoss of Von Hippel-Lindau (VHL) gene expression has been implicated in the development of human cancers. However, its function in oral squamous cell carcinoma (OSCC) remains undefined. We previously reported that the epithelial mesenchymal transition (EMT) process is linked with OSCC metastasis and invasion. The aim of this study was to clarify the VHL expression in OSCC,to analyse correlation of VHL with the EMT and to explore the underlying mechanisms of VHL in modulating the EMT of OSCC.Methodl.The expression of VHL,HIF-1α and EMT related proteins were evaluated by immunohistochemistry (IHC) in80OSCC tissues. The interrelationships of VHL expression with clinicopathologic features were analyzed using x2or Fisher exact test. Spearman’s correlation test was used to analyze the correlation of VHL with the EMT related proteins. Kaplan-Meier and time series test (log-rank test) was used for univariate survival analysis. Cox regression analysis was used for multivariate analysis.2. The VHL plasmid was transfected respectively into Tscca and Tca8113P160human tongue squamous cell carcinoma cell lines by Iipofectamine2000. Western blot was used to detect upregulation effect.Morphology of cells were observed.MTT assay was used to determine cells proliferation.The migratory and invasive abilities were compared by scratch and transwell assay. EMT related proteins were measured by western blot and immunofluorescence(IF).Result1.The rate of VHL high expression in OSCC(31.3%)was significant lower than that in peritumoral tissue(69.2%,P=0.013).Loss of VHL expression was associated with tumor grade,lymphatic metastasis. The expression of VHL,β-catenin,N-cadherin, E-cadherin, MMP-2/9were significantly associated with lymph node metastasis. An increase in HIF-la was noted from T1&T2stage to T3&T4stage.VHL was negatively correlated with (β-catenin,N-cadherin,snail,MMP-9, and HIF-la.The5-year survival rate of patients with VHL-positive(56.0%)was significantly higher than that of patient with VHL-negative(23.6%,β=0.002).COX model analysis shows that VHL expression, tumor differentiation, lymph node metastasis were independent prognosis factor for OSCC.2.Tscca and Tca8113P160showed a VHL low expression level,VHL expression was elevated by transfecting a pCDNA3plasmid earring a VHL gene.The morphology of cells changed, acquired the phenotype of epithelial after reexpression of VHL.MTT assay,cell invasion assay and scratch assay indicated that VHL inhibited the cell proliferation,motivation and invasion. Western blot shown that P-catenin, snail, N-cadherin and MMP-2, MMP-9, HIF-1α, VEGF, were decreased in both of the cell lines after transfected with VHL plasmid,while E-cadherin was upregulated.IF staining and western blot indicated P-catenin expression in nucleus was inhibited significantly.ConclusionThe expression of VHL is low in OSCC. Aberrant expression of VHL was related to EMT process of OSCC and may affect invasion,metastasis and prognosis by modulating EMT. VHL modulates OSCC morphology, proliferation, invasion and migration and EMT relative protein. Loss of VHL expression contributes to EMT in OSCC by regulating β-catenin and its downstream fectors.