Ginsenoside Rb1’s Protective Effects On Amyloid Peptide-Induced PC12Cell Cytotoxicity

Senile dementia diseases such as Alzheimer’s disease (AD) have a serious impacton human health with the intensification of social aging. In this study, the protectiveeffects of ginsenoside Rb1on the model of AD cells were analyzed by atomic forcemicroscope (AFM), laser scanning confocal microscope (LSCM), flow cytometry,immunofluorescence labeling technologies and other methods. The innovative resultsare as follows:1. After cells were treated with25μmol/L β-amyloid peptide (Aβ)25-35, theCCK-8results showed that the OD value reduced from1.68±0.13to1.45±0.15. Itmeans that cell activity was significantly decreased. Through the flow cytometry, itwas found that the apoptosis rate was increased from (3.53±0.37)%to (16.17±0.79)%, the intracellular ROS expression was increased, and the intracellular calciumlevels was also increased. Cell topography and cellular ultra-structure were analyzedby AFM, the results showed that cell surface became rough and uneven, there weremany ostioles on membrane, and the size of particle on cell surface was changed. Inthis work, it was the first time that the role of β-amyloid peptide in the ADpathological mechanisms of cell damage was analyzed by AFM combined withbiological methods.2. The cell viability of AD model cells were increased from50.42±5.51%to102.72±4.34%after cells were treated with50μM ginsenoside Rb1. Compared withAD model group, the DCF fluorescence intensity of cells in ginsenoside Rb1treatment group was decreased, which means that the ROS level of AD model cellswas suppressed by ginsenoside Rb1. And the MDA level was decreased to2.16±0.20μmol mg-1protein from4.41±0.43μmol mg-1protein of AD model group. Aftertreated with ginsenoside Rb1, cell morphology tended to be plump and intact, andmembrane ultra-structure was smooth. Through cytoskeleton analyzed by AFM andLSCM, it was found that ginsenoside Rb1could protect cytoskeleton from destructionwhich was induced by Aβ. And these results provided a new insight on prevention and treatment of AD.3. Quantitative and qualitative analysis results of cholesterol and PPARγ moleculesin the cells of different groups were obtained by LSCM and flow cytometry. Theresults showed that the cholesterol was distributed in whole cell, and the level ofcholesterol was decreased with addition of ginsenoside Rb1. The FILIPINfluorescence intensity of AD model group was2543±539, while the fluorescenceintensity of ginsenoside Rb1treatment group was reduced to1674±262. Theexpression of PPARγ in ginsenoside Rb1treatment group was significantly higherthan AD model group and control group. LSCM and Rhodamine-labeled rabbitanti-PPARγ antibody analysis results showed that PPARγ was mainly distributed inthe cytoplasm, a small part of the PPARγ in the nucleus in the control group and theAD model group cells. While in ginsenoside Rb1treatment group, the PPARγexpression of nuclei was significantly increased. These results indicated that PPARγmolecules could be activated by ginsenoside Rb1. It was the first time to analysis therole of ginsenoside Rb1as a PPARγ agonist to protect cells against amyloid damageon the molecular level.In this study, AFM and other biological methods were used to analysis theprotective mechanism of ginsenoside Rb1on the model of AD cells. And this workhas an important significance for AD prevention and treatment research.