ABSTRACT
Objective: To investigate the effects of Chinese herbal formula Qinghuofu (QHF) on the migration and invasion of breast cancer MCF-7 cells and its possible molecular mechanisms, thereby providing a theoretical basis to find effective anti-cancer medicine and therapeutic targets for the treatment of anti-migration and anti-invasion of breast cancer. Methods: Breast cancer MCF-7 cells were treated with different QHF and other different reagents, CCK8 assay was used to detect the influence of the reagents on the proliferation of MCF-7 cells; Scrape migration and Transwell assay were used to quantitatively determine the migration and invasion effects of QHF and hepatocyte growth factor (HGF) on the MCF-7 cells. Subsequently, the c-Met inhibitor and its downstream ERK and PI3K inhibitors were used to investigate the relationship between the migration and invasion of MCF-7 cells, as well as its downstream MAPK/ERK and PI3K/Akt signaling pathways. The expression levels of HGF, c-Met, ERK, p-Akt, p-c-Met, p-ERK, p-Akt, MMP2, MMP9, and VEGF in breast cancer MCF-7 cells treated with QHF and other reagents were also examined. Results: The result indicated that formula QHF not only significantly inhibited the proliferation of MCF-7 cells, but also significantly suppressed the effects of HGF (40 ng/mL) on the proliferation and movement of MCF-7 cells, reducing the ability of the cells to invade and migrate. Western blot analysis indicated that QHF and c-Met inhibitor significantly decreased the expression of p-c-Met, p-ERK1, p-ERK2, p-Akt, MMP-2, MMP-9, and VEGF, while HGF significantly increased the expression of p-c-Met in MCF-7 cells; c-Met downstream ERK and PI3K inhibitors also significantly decreased the expression of MMP-2, MMP-9, and VEGF in MCF-7 cells; But the difference among c-Met, PI3K, ERK, and QHF group were not statistically significant. Conclusion: QHF can prevent the proliferation, migration, and invasion of MCF-7 cells by inhibiting the HGF/c-Met and its downstream PI3K/Akt and MAPK/ERK signaling pathways; Thereby down-regulating the expression of HGF, p-Met, p-ERK1, p-ERK2, p-Akt, MMP-2, MMP-9, and VEGF.
ABSTRACT
AIM:To investigate the effect of β-elemene on reversing hepatocyte growth factor(HGF)-induced resistance to gefitinib in PC-9 cells, and to explore its possible mechanisms.METHODS: The gefitinib-resistant PC-9 cells induced by HGF were treated with β-elemene or/and gefitinib.The cell activity was measured by MTT assay.The effect of β-elemene on the invasion ability in HGF-induced resistance to gefitinib in PC-9 cells was detected by Transwell migration assay.The protein levels of p-Met, c-Met, p-AKT and AKT in PC-9 cells of each group were determined by Western blot.RESULTS:The results of MTT assay showed that the cell activity of PC-9 cells was significantly inhibited by β-elemene(P<0.05).IC50of β-elemene for PC-9 cells was 169.31 mg/L.IC50of gefitinib for PC-9 cells was 0.30 μmol/L.Exogenously adding recombinant HGF induced significantly resistance to gefitinib in PC -9 cells.Moreover, SU11274(an inhibitor of c-Met)significantly decreased the viability of the cells exposed to HGF and gefitinib(P <0.05).Combined treatment with β-elemene and gefitinib in the presence of HGF(50 μg/L)significantly decreased the viability of PC-9 cells as compared with the PC-9 cells treated with gefitinib alone in the presence of HGF(P<0.05),so did the result of the cell migration.The protein levels of p-Met and p-AKT were significantly up-regulated by HGF,while the protein levels of p-Met and p-AKT were markedly down-regulated in the cells treated with β-elemene and gefitinib com-pared with gefitinib alone in the presence of HGF.CONCLUSION: β-elemene reverses HGF-induced resistance to ge-fitinib in lung cancer PC-9 cells,likely due to the inhibition of HGF-induced activation of c-Met and its down streams sig-naling pathways(P<0.01).
ABSTRACT
AIM: To investigate the role of HGF/c-Met signaling pathway in crizotinib-induced apoptosis of different lung carcinoma cell lines and to analyze its potential regulatory mechanisms .METHODS: EML4-ALK positive cell line H2228, c-Met proliferation cell line H1993 and control cell line A549 were treated with crizotinib at different doses for different time periods .The viability of the cell lines was measured by MTT assay .The apoptosis was analyzed by flow cytometry with PI staining.The protein levels of MET and phosphorylated MET (p-MET) of HGF/c-Met signaling pathway as well as its down-stream key proteins AKT , ERK, p-AKT and p-ERK in the cell lines before and after crizotinib treatment were examined by Western blot .RESULTS:The growth of H1993, H2228 and A549 cell lines was inhibited after crizoti-nib treatment for 72 h in a dose-dependent manner .Apoptotic rates of H1993 cells and H2228 cells were increased with the crizotinib concentration and exposure time .Down-regulation of p-MET, p-AKT and p-ERK at protein levels in H1993 cells and H2228 cells after exposure to crizotinib for 72 h was confirmed by Western blot .No obvious change of the related-pro-teins of HGF/c-Met signaling pathway was found in A 549 cell line.CONCLUSION: HGF/c-Met signaling pathway may contribute to crizotinib-induced apoptosis of H1993 cells and H2228 cells, which provides the experimental basis for MET-targeting treatment of lung cancer .