Figure 3 The effect of c-Myb on OPN mTOR activation expression of HCCLM6 cells. (A) OPN mRNA expression in HCCLM6 cells transfected with c-Myb siRNA was significantly decreased. (* P < 0.05, vs control). The mRNA expression of OPN in cells transfect with scramble siRNA was used as control. (B) OPN protein expression in HCCLM6 cells transfected with c-Myb siRNA was significantly reduced compared
with cells transfected with sramble siRNA. Blot was representative of three experiments. 3.4 Migration and invasion of HCCLM6 cells in vitro were inhibited by c-Myb siRNA As migratory and invasive behaviors are the indicators HMPL-504 in vitro of the metastatic potential, we examined migration and invasion of HCCLM6 cells in vitro using the transwell assay after c-Myb expression was inhibited by c-Myb siRNA. The average numbers of HCCLM6 cells transfected with c-Myb siRNA migrating toward the conditioned medium
or invading through the Matrigel were significantly fewer than those transfected with scramble siRNA (Migration assay: 17.60 ± 4.04 vs 33.60 ± 4.67, P < 0.05; Invasion assay: 8.00 ± 2.55 vs 18.8 ± 4.15, P < 0.05, Figure 4), This result showed that the capability of migration and invasion in HCCLM6 cells was significantly PLX3397 purchase decreased after inhibition of c-Myb, suggesting that c-Myb is an important contributor to the migration and invasion of HCC cells. Figure 4 Migration and invasion of HCCLM6 cells in response to transfection of c-Myb siRNA. The c-Myb siRNA could significantly inhibit the migration and invasion of HCCLM6 cells compared with cells treated with scramble siRNA (* P < 0.05). The migration and invasion assays were assessed by transwell chambers. Data were expressed as means ± SD of three experiments. Molecular motor Discussion Metastasis remains one of the major challenges for HCC patients undergoing various therapies including liver resection, local ablation
and chemoembolization [2, 3]. Previous work at our institute has shown that OPN gene is over-expressed in the metastatic HCC . In this study, we searched for transcription factors that were correlated with OPN expression in HCC cells and revealed that transcription factor c-Myb was positively associated with OPN expression in HCC cells, which can bind the OPN promoter and increase its transcription activity. Inhibition of c-Myb by siRNA decreased the transcription activity of the OPN promoter, reduced the expression of OPN, and compromised the ability for migration and invasion of HCC cells. Therefore, our results demonstrate that c-Myb plays an important role in regulating OPN expression in HCC cells, suggesting c-Myb might be a novel target for therapeutic intervention. OPN is known to mediate correlates of metastatic biology in a variety of cancers including HCC. Thus, modulating OPN expression might be a novel approach of suppressing tumor metastasis [17–19].