ABSTRACT
OBJECTIVE: To investigate the effect of saikosaponins-b2 (SS-b2) on the migration of HepG2 cells, and to explore its potential molecular mechanism. METHODS: The effect of SS-b2 on the viability of HepG2 cells was detected by MTT method before the cells were divided into normal control group, SS-b2 (15, 30 and 60 mg·L-1) groups and positive control [doxorubicin (DOX) 2 mg·L-1] group according to the results of MTT. Wound-healing assay was performed to observe the influence of SS-b2 on the migration of HepG2 cells. The protein expressions of vascular endothelial growth factor (VEGF), matrix metalloproteinase 2 (MMP-2), MMP-9, hypoxia inducible factor-lot (HIF-1α) and the phosphorylation of extracellular regulated protein kinase 1/2 (ERK1/2) in HepG2 cells were measured by Western blotting. RESULTS: The wound-healing rate was 74.71%, 72.50% and 66.82% after treatment with SS-b2 15, 30 and 60 mg·L-1for 48 h, respectively. The wound-healing rate decreased significantly with the increae of SS-b2 doses. Compared with the normal control group, the wound-healing rate of SS-b2 30 and 60 mg·L-1was significantly reduced (P<0.05). The expressions of VEGF and HIF-1α in HepG2 cells were significantly decreased in SS-b2 groups (P<0.05, P<0.01). SS-b2 60 mg·L-1could reduce the protein expressions of MMP-2 and MMP-9 and the phosphorylation of ERK1/2 significantly (P< 0.05, P<0.01). The positive control group could significantly reduce the expressions of the above four proteins and the phosphorylation of ERK1/2 (P<0.05, P<0.01). Compared with the positive control group, the protein expressions of VEGF and MMP-2 were significantly down-regulated in 60 mg·L-1SS-b2 group (PO.05, P<0.01), but there was no significant difference in the expression of HIF-1α between SS-b2 groups and the positive control group. However, the down-regulation effect of SS-b2 on MMP-9 protein expression was not so significant as in the positive control group (P<0.01). The effect of SS-b2 60 mg·L-1on the phosphorylation of ERK1/2 was similar to that of the positive control group. CONCLUSION: SS-b2 can inhibit the migration of HepG2 cells, which may be related to the inhibition of angiogenesis-related proteins.
ABSTRACT
OBJECTIVE Angiogenesis therapy has attracted interest as a potential treatment for hepatocellular carcinoma (HCC). In this study, we investigated the anti-proliferative activities and anti-angiogenesis effects of saikosaponins (SS)-b on hepatocellular carcinoma (HCC) and its regulation on VEGF/ERK/HIF-1α signal pathway. METHODS H22 hepatoma-bearing mice model and HepG-2 cells were used to study the anti-tumor and anti-angiogenesis effects of SS-b in vivo and in vitro. Pathological change of tumor tissue was observed by HE staining, the microvascular changes were detected by immunohistochemical method. The effects of SS-b on angiogenesis were examined by using the chick embryo chorioallantoic membrane (CAM) model. The effects of SS- b on proliferation, migration and invasion were investigated by MTT assay, scratch wound healing assay and transwell assay inhuman umbilical vein endothelial cell (HUVEC) and HepG2 cells in vitro. Vascular endothelial growth factor (VEGF), matrix metalloproteinase-2/9(MMP-2/9), hypoxia-inducible factor-1α (HIF-1α) expression and the phosphorylation of extracellular regulated kinase(ERK) were analyzed using RT-PCR and Western-blot. RESULTS SS-b effectively inhibited the tumor growth of H22 mice in vivo. The inhibitory rate of tumor was 49.1%, 50.7%, 66.1% in SS-b 5, 10 and 20 mg·kg-1 group respectively. HE staining results showed that SS-b induced tumor necrosis and nuclear dissolution in H22 mice. Moreover, SS-b also reduced the number of microvessels of tumor tissue in H22 mice significantly and suppressed the angiogenesis of CAM induced by b-FGF. SS-b had an obvious inhibitory effect on cell proliferation, migration and invasion of HUVEC cells and HepG-2 cells. These effects were associated with down-regulation of the expression of MMP2/9 and suppression of VEGF/ERK/HIF-1α signaling in H22 mice and Hep-G2 cells. CONCLUSION Our findings showed that SS-b exerts anti-tumor effects by inhibit?ing tumor angiogenesis via regulating VEGF/ERK/HIF-1α signal pathway in vivo and in vitro.