An Experimental Study Of Resveratrol On The Apoptosis In Human Esophageal Cancer Eca109 Cells And The Expression Of Apoptosis-related Genes

Objective: To explore the mechanism of resveratrol as an potential antineoplastic for the treatment of esophageal cancer through inducing apoptosis of esophageal cancer Eca109 cells and the effect of resveratrol on the expression of apoptosis-related genes including survivin, bcl-2 and bax in esophageal cancer Eca109 cells. The study may provide experimental foundation for the therapy of esophageal cancer.Methods: Human esophageal cancer Eca109 cells were cultured with resveratrol in different time and various concentrations in the environment of 37℃, 5%CO2.1 MTT assay was used to determine the cell growth inhibitory rate.2 The change of morphology was observed by inverted phase contrast microscope, transmission electron microscope and HE staining method.3 Flow Cytometry was used to detect the apoptosis and cell cycle of Eca109 cells treated by resveratrol.4 Reverse transcription polymerase chain reaction (RT-PCR) manner was used to observe the expression of survivin, bcl-2 and bax mRNA.5 Flow Cytometry was used to detect the expression of survivin, bcl-2 and bax protein.Results:1 The results of the inhibitory of cell growth by MTT assay: After human esophageal cancer Eca109 cells were treated with resveratrol at various concentrations including 0μg/ml, 10μg/ml, 20μg/ml, 40μg/ml, 80μg/ml and 100μg/ml for 24 hours, 48 hours and 72 hours respectively, resveratrol inhibited the growth of esophageal cancer Eca109 cells in a dose- and time-dependent manner. The biggest cell growth inhibitory rate arrived at 76.42%±0.04% (p<0.01).2 The change of morphology: The growth of drug-treated cells was inhibited and cell granulations were increased and thicken some cells became round and suspension in the medium observed by inverted phase contrast microscope. Transmission electron microscope and HE staining manner showed that resveratrol induced Eca109 cells to undergo apoptosis with typically apoptotic characteristics, including morphological change of nuclear chromatin condensation and the bulks of nuclear lessening.3 The analysis of apoptosis and cell cycle by flow Cytometry: There was a sub-G0/G1 peak in the graph, after Eca109 cells were treated by resveratrol (20μg/ml, 40μg/ml) for 72 hours. The peak was a typical apoptotic peak which was not shown in the graph of control cells (Eca109 cells with no drug-treated). And the apoptosis rates were 8.54%±0.52% and 18.79%±0.73%(p<0.01)respectively. The quantity of drug-treated cells in G0/G1 phase increased, while in S and G2/M phase decreased. Most Eca109 cells were blocked at G0/G1 phase, and the percent of drug-treated cells arrived at 51.5%±1.70% and 62.4%±1.82% respectively, which were significantly higher than control cells (38.0%±1.53%) (p<0.01).4 The results of the mRNA expression by RT-PCR manner: after Eca109 cells were treated by resveratrol (20μg/ml, 40μg/ml) for 72 hours,with the concentrations increasing, the expression of survivin and bcl-2 mRNA were down-regulated while bax mRNA was up-regulated.5 The analysis of the protein expression by flow cytometry: The expression of survivin and bcl-2 protein was decreased (Fluorescence index were 0.82±0.04,0.69±0.06 and 0.91±0.03, 0.74±0.04 respectively) (p<0.01) while the expression of bax protein was increased (Fluorescence index were 1.05±0.03,1.17±0.04) (p<0.05).Conclusion:1 Resveratrol had significantly inhibitory effect on the proliferation of human esophageal cancer Eca109 cells.2 A certain range of resveratrol's concentrations could effectively induce apoptosis of human esophageal cancer Eca109 cells and block the cell cycle at G0/G1 phase.3 Resveratrol could modulate the expression of apoptosis -related genes including survivin, bcl-2 and bax. Its molecular mechanisms were possibly related to up regulate bax and down regulate survivin, bcl-2.The inducing apoptosis of human esophageal cancer Eca109 cells demonstrated that resveratrol as a potential antineoplastic agent for the treatment of esophageal cancer had a great applied foreground. The study would provide experimental foundation for the therapy of esophageal cancer in the future.