Effect Of Ginsenoside Rg3 On The Epidermal Growth Factor Receptor Signaling Pathways In Human Breast Cancer Cell Line MCF-7

Objective: Ginsenoside Rg3 is the monomer extracted from red ginsenoside with anti-tumor drugs. Studies have shown that, It exerts the inhibition of tumor growth, invasion and metastasis, and inhibition of tumor angiogenesis and improving immunity and supplying synergy role, when combined with chemotherapy, Its anti-tumor action and signal transduction mechanism is unclear.EGFR is one of the epidermal growth factor receptor family members. It is usually highly expressed in a variety of malignant tumors, and abnormal activation is closely correlated with tumor cell biology, as well as the malignant tumor in patients with poor prognosis is closely related. EGFR is a tyrosine-protein kinase receptor; the key steps of EGFR activation are the formation of dimers and their tyrosine phosphorylation. Tyrosine phosphorylation may raise signaling molecules containing SH2 domain, which will leads to downstream signal transduction molecules, resulting in a series of downstream signaling pathway activation.This study investigates the effect of ginsenoside Rg3 on the growth human breast cancer cell line MCF-7 of EGFR signaling. Proposed ginsenosides Rg3 may serve as a signal molecule provide experimental basis of EGFR-mediated signal-regulated network view for clinical applications ginsenoside Rg3 in anti-tumor targeted therapy. It will also explore the molecular of Rg3 in anti-tumor.Methods:1. Human breast cancer cell line MCF-7 was exposed to ginsenoside Rg3 at the same time with the differential concentration respectively. The cell proliferation inhibition was measured by 3-〔4,5-dimethylthiazo-2-yl〕-2, 5 diphenyl tetrazolium bromide (MTT) assay. 2. Different concentrations of ginsenoside Rg3 exposed to human breast cancer cell line MCF-7 at the same time, to test MCF-7 cell surface receptor expression level of EGFR, phosphorylated EGFR, phosphorylation of tyrosine kinase receptor by Flow cytometry.3. To test the expression of EGFR,pEGFR and PY20,different concentrations of ginsenosides Rg3 exposed to Human breast cancer cell line MCF-7 at the same time, to test fluorescence intensity of MCF-7 cell surface receptors, such as EGFR, phosphorylation EGFR, Phosphorylated of tyrosine kinase receptor by cell immunofluorescence method.Results:1.After a certain period of time, the action of different concentrations of Rg3 to MCF-7 cell proliferation inhibition. Human breast cancer cell line MCF-7 was exposed to ginsenoside Rg3 at a concentration of 12.5μg/ml, 25μg/ml, 50μg/ml, 100μg/ml, 200μg/ml, accessory (50μg/ml), respectively. Under the microscope, we can observe the cells adherence, fullness, polygon in control group, the cell amount sparse, suspended, quasi-round, and no refraction nature in experiment groups; the higher the concentration, the greater the morphological change of cells became. The inhibition ratio was 10.3%, 15.2%, 20.7%, 26.3%, 50.4%, 92.7%, respectively and that of accessory group was 22.6% after 24 hours, statistical significance could be found from 100μg/ml to 200μg/ml compared with the control group (p﹤0.05). The inhibition ratio was 6.0%, 12.4%, 30.3%, 60.6%, 69.1%, 74.0%, respectively. Accessory group was 14.8% after 48 hours. Statistical significance can be found from 25μg/ml to 200μg/ml compared with the control group (p﹤0.05).2. MCF-7 cell surface receptor expression level of EGFR by Flow cytometry was 25μg/ml (40.32±0.84)%,50μg/ml (32.09±1.63)%,100μg/ml (25.22±2.15)%,200μg/ml (15.17±0.95)%. Significantly lower than the control group (71.54±1.89)% and accessories group (68.49±2.87)% (p﹤0.05); MCF-7 Cell surface receptors expression rates of phosphorylation EGFR by Flow cytometry was 25μg/ml (38.16±0.79)%, 50μg/ml (27.94±0.59)%, 100μg/ml (19.22±1.41)%, 200μg/ml (10.53±0.99)%. Significantly lower than the control group (57.10±0.35)% and Accessories Group (54.49±0.43)% (p﹤0.05); MCF-7 Cell surface receptors expression rates of phosphorylation of tyrosine kinase receptor by Flow cytometry was 25μg/ml (38.90±1.01)%, 50μg/ml (25.73±1.19)%, 100μg/ml (17.64±1.29)%, 200μg/ml (9.73±0.75)%. Significantly lower than the control group (53.98±1.16)% and Accessories Group (58.04±0.96)% (p﹤0.05); expression rates of pEGFR, and phosphorylation of tyrosine kinase receptor was lower than expression rates of EGFR.3. With the increasing of the Rg3 concentration, the fluorescence intensity of membrane surface gradually weakened, in the control group that the fluorescence intensity of membrane surface was higher than the drug group.Conclusion:1. Ginsenoside Rg3 has obvious anti-tumor effect on human breast cancer cell line MCF-7, the higher the concentration, and the greater the inhibitory effect on tumor cells proliferation.2. In a certain range (25μg/ml, 50μg/ml, 100μg/ml, 200μg/ml), ginsenosides Rg3 reduced the EGFR expression of the MCF-7 in the cell surface, as well as the expression of pEGFR and PY20. With the increasing of drug concentration, the expression of EGFR, pEGFR, PY20 gradually decreased in a dose-dependent manner, and there was a statistically significant difference.3. After the action of ginsenosides Rg3, the expression of EGFR, pEGFR and PY20 were positively correlated.4. Ginsenoside Rg3 inhibited the growth of tumor cells by blocking MCF-7 cell surface EGFR and pEGFR, PY20, suggesting that ginsenoside Rg3 inhibited the growth of tumor cell by targets EGFR and its downstream signal transduction pathways.