X C Chen1, Y G Zhu, X Z Wang, L A Zhu, C Huang. 1. Fujian Institute of Geriatrics, Union Hospital, Fujian Medical Univisity, Fuzhou 350001, China. ZJCXCH@Public5.FZ.FJ.cn
Abstract
AIM: To explore the possible molecular mechanism of exogenous dopamine-induced apoptosis in PC12 cells and the protective effect of ginsenoside Rg1. METHODS: Flow cytometric assay was used to quantify the apoptotic cells and measure the percentage of cells with positive Bcl-2 and Bax proteins. The morphology of apoptotic cells was evaluated by transmission electron microscope. DNA fragmentation was observed by gel electrophoresis. Caspase-3 activity was determined by fluorescent spectrofluorometer and the expressive bcl-2 and bax mRNA by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Dopamine 0.15, 0.30, 0.45, and 0.60 mmol/L induced PC12 cell apoptosis from 1.1 % +/- 0.4 % (control) to 41 % +/- 3 %, 46.4 % +/ -2.7 %, 53 % +/ -3 %, and 64.5 % +/- 2.7 %, respectively. After treatment with dopamine 0.45 mmol/L following pretreatment with Rg1 10 micromol/L for 24 h, the percentage of apoptotic cells and caspase-3 activity decreased from 53 % +/- 3 % and 683 +/- 8 (mean fluorescence intensity, MFI) to 1.9 % +/- 0.6 % and 325 +/- 5, and the percentage of cells with positive Bcl-2 protein increased from 14.3 % +/- 1.1 % to 25.9 % +/- 1.6 %, however, the percentage of cells with positive Bax protein decreased from 48 % +/- 3 % to 35 % +/- 3 %, compared with group treated with DA 0.45 mmol/L alone. CONCLUSION: Ginsenoside Rg1 protected PC12 cells against apoptosis by inhibiting the activation of caspase-3 and regulating the ratio of Bcl-2 to Bax protein.
AIM: To explore the possible molecular mechanism of exogenous dopamine-induced apoptosis in PC12 cells and the protective effect of ginsenoside Rg1. METHODS: Flow cytometric assay was used to quantify the apoptotic cells and measure the percentage of cells with positive Bcl-2 and Bax proteins. The morphology of apoptotic cells was evaluated by transmission electron microscope. DNA fragmentation was observed by gel electrophoresis. Caspase-3 activity was determined by fluorescent spectrofluorometer and the expressive bcl-2 and bax mRNA by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). RESULTS:Dopamine 0.15, 0.30, 0.45, and 0.60 mmol/L induced PC12 cell apoptosis from 1.1 % +/- 0.4 % (control) to 41 % +/- 3 %, 46.4 % +/ -2.7 %, 53 % +/ -3 %, and 64.5 % +/- 2.7 %, respectively. After treatment with dopamine 0.45 mmol/L following pretreatment with Rg1 10 micromol/L for 24 h, the percentage of apoptotic cells and caspase-3 activity decreased from 53 % +/- 3 % and 683 +/- 8 (mean fluorescence intensity, MFI) to 1.9 % +/- 0.6 % and 325 +/- 5, and the percentage of cells with positive Bcl-2 protein increased from 14.3 % +/- 1.1 % to 25.9 % +/- 1.6 %, however, the percentage of cells with positive Bax protein decreased from 48 % +/- 3 % to 35 % +/- 3 %, compared with group treated with DA 0.45 mmol/L alone. CONCLUSION:Ginsenoside Rg1 protected PC12 cells against apoptosis by inhibiting the activation of caspase-3 and regulating the ratio of Bcl-2 to Bax protein.