RhoC overexpression-induced migration and invasion of human normal hepatocytes

Abstract

Objective To investigate the role of Ras-homologous GTPase C (RhoC) overexpres-sion in inducing migration and invasion for malignant transformation in normal human hepatocytes.Methods pcDNA3-RhoC vector was transfected into human normal liver cell HL7702.Scratch-healing test combined with Transwell was used to examine cell migration and invasion,and matrix metalloproteinases (MMPs)gelatinase activities were assayed through zymography.Reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting were used to detect the migration and invasion associated genes.RhoC transfected HL7702 cells were inoculated into the nude mice to test tumor formation.Results Cells transfected with pcDNA3-RhoC acquired migration capacity,and the migration distance was notably increased as compared with parental cells and negative control groups [(63.33 ± 10.07)％ vs.(28.67 ± 7.02)％ and (28.33 ±6.66)％,P ＜0.01].The number of pcDNA3-RhoC cells through the Transwell was increased significantly (65.33 ± 6.80 vs.24.33 ± 5.79 and 23.33 ± 5.73,P ＜ 0.01),and the invasion capacity was significantly enhanced in pcDNA3-RhoC-transfected group as compared with control groups (43.67 ±7.59 vs.13.33 ± 3.40 and 15.33 ± 4.11,P ＜ 0.01).MMPs gelatinase activities were increased,the expression of migration and invasion associated genes and proteins such as MMP-2,MMP-9,vascular endothelial growth factor (VEGF),P27RF-Rho and Survivin was significantly up-regulated,and the expression of tissue inhibitor of tissue inhibitorof metalloproteinase 1 (TIMP1),TIMP2,p53 and phosphatase and tensin homolog deleted on chromosome ten (PTEN) was down-regulated in pcDNA3-RhoC-transfected group as compared with control group.The tumor formation ratio in nude mice was 40％.Conclusion RhoC overexpression can induce migration and invasion for malignant transformation in human normal hepatocytes. Key words: RhoC ; Hepatocyte ; Vicious transformation