Wnt5a Cene Affects Cell Biological Behavior Of HCC

ObjectivePrevious experimental studies indicated that Wnt5a had a tumor-suppressing role in the development of hepatocellular carcinoma, so we desigened Wnt5a expression vecter to explore the effects on the cytobiological characteristics of the transfected HCC cell, and investigated the interaction between Wnt5a and the downstream gene of Wnt5a.Mathods:1. Human liver cell lines of L02 and MHCC97L HCC were transfected with the pcDNA3.1-Wnt5a or the empty vector controls.2. Using RT-PCR and Western Blot methods, the expression of Wnt5a before and after transfection were detected, and transfection systerm was established.3. Flow cytometry used to detect the change of cell cycle, scratch assays to examine cell migration.4. RT-PCR and Wsetern Blot were employed to detect the expression chang of Ror2,β-catenin mRNA and protein before and after transfection.Results:1. The S phase fraction in L02 liver cell line and MHCC97L HCC line with Wnt5a expression vector transfected were 20.09% and 21.69%, compared with emoty vector transfected cells in S phase fraction decreased(respectively 29.63% and 29.03%), suggested Wnt5a lead to decrease of cell proliferation.2. Cells transfected with Wnt5a expression vectors display lower motility than vectors control transfected cells. Blocking this pathway using antibodies to Wnt5a, there was an increase in cell migration.3. The Ror2 mRNA and protein expression was higher in the Wnt5a transfectants, which suggested Ror2 to be a receptor for Wnt5a in the development of HCC.4. In Wnt5a-transfected cells, P-β-catenin protein levels were increased andβ-catenin were decreased as compared with vectors control transfected cells, showing Wnt5a inhibits the canonical Wnt/β-catenin pathway by promotingβ-catenin degradation.Conclusion:1. Our studies in vitro showed that cell proliferation and motility decreased with expression Wnt5a in cells of L02 and MHCC97L.2. Cells were transfected with Wnt5a expression vectors constitutively increased expression of Ror2 and P-β-catenin protein and decrease expression ofβ-catenin protein, suggested Ror2 act as a receptor for Wnt5a to activate non-canonical Wnt signaling pathway to antagonize the canonical Wnt/β-catenin signaling pathway.3. Wnt5a expression resulted in suppression cell proliferation and invation through antagonize Canonical Wnt signaling pathway, suggested Wnt5a plays the role of a tumor suppressor gene in HCC. Wnt5a maybe a new target of gene therapy for hepatocellular carcinoma.