Chunjing Zhang1, Haitao Yu, Jincai Hou. 1. Department of Biology Genetics, Department of Biochemistry and Molecular Biology, Qiqihar Medical College, Qiqihar 161042, China.
Abstract
OBJECTIVE: To evaluate and explore the effects of 20(S)-ginsenoside Rh2 and 20(R)-ginsenoside Rh2 on the cytotoxicity, proliferation and the apoptosis of human lung adenocarcinoma A549 cells, and to illustrate the structure-activity relationship and possible mechanisms of anti-tumor active ingredients of ginseng. METHOD: A549 cells were treated with different concentration gradient of ginsenoside Rh2 (S and R structure) and incubated for different time. Cell proliferation and cytotoxicity studies were detected by methyl thiazolyl tetrazolium (MTT) colorimetric assay, cell cycle and apoptotic was analyzed by PI stains and combination of Annexin V/Prop idium iodide double staining with flow cytometric analysis. The influences of activation on Caspase-3 were also detected by the immunofluorescence staining with fluorescence microscope. RESULT: MTT test indicated that ginsenoside Rh2 had a strong cytotoxicity activity to A549 cells. Ginsenoside Rh2 could obviously inhibit the cell proliferation in human lung adenocarcinoma cell line A549 at the effective doses of 25 mg x L(-1) treated with 48 h. The inhibition ratio and the value of IC50 for48 h of 20(R)-Rh2 and 20(S)-Rh2 were respectively 28.5%, 33.6% and 33.4, 28.5 mg x L(-1). The inhibition of ginsenoside Rh2 to A549 showed structure relationship significantly, time-dependent and concentration-dependent. Flow cytometric analysis (FACS) with PI stains analysis results showed that the proportion of A549 cells in G1 phase increased, while the number of cells in S phase decreased significantly and those in G2 phase reduced slightly. This result indicated structure relationship significantly, especially in the 20(S) -ginsenoside Rh2 inhibited the proliferation of A549 cell dramatically and retarded A549 cell cycle at G0/G1 phase. The immunofluorescent of combination with Annexin VFITC/PI by flow cytometric suggested ginsenoside Rh2 can induce inchoate apoptsis rate and late apoptosis rate of A549 cell significantly. All the results showed structure relationship significantly, especially in the 20(S)-ginsenoside Rh2. The immunofluorescent with fluorescence microscope suggested the activity of Caspase-3 were enhanced after ginsenoside Rh2 treated. CONCLUSION: 20 (R) and 20(S)-ginsenoside Rh2 had a significant inhibitory effect on the proliferation. Compared with 20(S)-ginsenoside Rh2, 20 (S)-ginsenoside Rh2 has been shown to have significant anticancer effects and to be capable of blocking cell proliferation and causing G1 phase arrest in human lung adenocarcinoma A549 cells. 20 (R) and 20(S)-ginsenoside Rh2 have been shown to have anticancer effects and to be capable of increasing inchoate apoptotic rate, reducing apoptotic rate significantly, enhancing the activity of Caspase-3 and inducing apoptosis in human lung adenocarcinoma A549 cells.
OBJECTIVE: To evaluate and explore the effects of 20(S)-ginsenoside Rh2 and 20(R)-ginsenoside Rh2 on the cytotoxicity, proliferation and the apoptosis of humanlung adenocarcinoma A549 cells, and to illustrate the structure-activity relationship and possible mechanisms of anti-tumor active ingredients of ginseng. METHOD: A549 cells were treated with different concentration gradient of ginsenoside Rh2 (S and R structure) and incubated for different time. Cell proliferation and cytotoxicity studies were detected by methyl thiazolyl tetrazolium (MTT) colorimetric assay, cell cycle and apoptotic was analyzed by PI stains and combination of Annexin V/Prop idium iodide double staining with flow cytometric analysis. The influences of activation on Caspase-3 were also detected by the immunofluorescence staining with fluorescence microscope. RESULT: MTT test indicated that ginsenoside Rh2 had a strong cytotoxicity activity to A549 cells. Ginsenoside Rh2 could obviously inhibit the cell proliferation in humanlung adenocarcinoma cell line A549 at the effective doses of 25 mg x L(-1) treated with 48 h. The inhibition ratio and the value of IC50 for48 h of 20(R)-Rh2 and 20(S)-Rh2 were respectively 28.5%, 33.6% and 33.4, 28.5 mg x L(-1). The inhibition of ginsenoside Rh2 to A549 showed structure relationship significantly, time-dependent and concentration-dependent. Flow cytometric analysis (FACS) with PI stains analysis results showed that the proportion of A549 cells in G1 phase increased, while the number of cells in S phase decreased significantly and those in G2 phase reduced slightly. This result indicated structure relationship significantly, especially in the 20(S) -ginsenoside Rh2 inhibited the proliferation of A549 cell dramatically and retarded A549 cell cycle at G0/G1 phase. The immunofluorescent of combination with Annexin VFITC/PI by flow cytometric suggested ginsenoside Rh2 can induce inchoate apoptsis rate and late apoptosis rate of A549 cell significantly. All the results showed structure relationship significantly, especially in the 20(S)-ginsenoside Rh2. The immunofluorescent with fluorescence microscope suggested the activity of Caspase-3 were enhanced after ginsenoside Rh2 treated. CONCLUSION:20 (R) and 20(S)-ginsenoside Rh2 had a significant inhibitory effect on the proliferation. Compared with 20(S)-ginsenoside Rh2, 20 (S)-ginsenoside Rh2 has been shown to have significant anticancer effects and to be capable of blocking cell proliferation and causing G1 phase arrest in humanlung adenocarcinoma A549 cells. 20 (R) and 20(S)-ginsenoside Rh2 have been shown to have anticancer effects and to be capable of increasing inchoate apoptotic rate, reducing apoptotic rate significantly, enhancing the activity of Caspase-3 and inducing apoptosis in humanlung adenocarcinoma A549 cells.