Down-regulation of ANXA7 decreases metastatic potential of human hepatocellular carcinoma cells in vitro.

We report for the first time the influence of ANXA7 gene on human hepatocellular carcinoma cells (HCC). We down-regulated ANXA7 in human HCC cell line (HepG2) using siRNA method. By Western Blot analysis, we confirmed about 70% down-regulation of the gene in the shRNA-ANXA7 transfected cells (shRNA-ANXA7-HepG2) compared to the non-specific sequence shRNA transfected cells (control-shRNA-HepG2) and the un-manipulated-HepG2 cells. We used CCK-8 cell proliferation kit and observed about 65% reduction in the shRNA-ANXA7-HepG2 cells where the two controls exhibited comparable cell proliferation rates. Also, by using PI staining followed by flow cytometry, we noticed a cell cycle arrest at G0/G1 with more than one fold reduction of shRNA-ANXA7-HepG2 cell population in the S-phase of the cell cycle. Also of particular note was a significant aneuploidy in the controls compared to zero aneuploidy in the ANXA7 down-regulated cells. Migration of the cells was detected using Boyden's transwell chamber and scratch wound healing assay which showed 50% and 30% respective reductions in shRNA-ANXA7-HepG2 cells migration. Furthermore, the control-shRNA-HepG2 cells and the un-manipulated-HepG2 cells invaded through the ECM-coated transwell plates two times more than the shRNA-ANXA7-HepG2 cells. We have found ANXA7 to be functioning like a tumour promoter in HepG2 human hepatocellular carcinoma cells and could have a potential as a therapeutic window into the management of liver cancer.