| Objective:Training the CNE2side population cells (CNE2-SP cells) and which is identified; Research Yiqijiedu side of CNE2side population cells proliferative activity of nasopharyngeal CNE2-SP and induce apoptosis and its mechanism.Methods:water extraction Yiqijiedu side and freeze-dried; serum-free suspension culture method for culturing CNE2side population cells (CNE2-SP); by CNE2-SP self-renewal, differentiation potential, resistance and CD44expression in four identify aspects of their stem cell properties; CCK-8by the method of detection within72hours Yiqijiedu side of CNE2-SP proliferation inhibition activity, and calculate the relative rate of living cells under different concentrations of liquid IC50value and role; with fluorescent probe JC-1staining CNE2-SP apoptosis and apoptotic rate; Western-blot assay with P53, Survivin, Cleaved PARP, cleaved caspase-8, cleaved caspase-9, cleaved caspase-3, cleaved caspase-7protein.Results:1, serum-free culture of CNE2side population cells (CNE2-SP). Self-renewal capacity:CEN2into a ball rate (19±2.6)%o, CNE2-SP first generation into a ball rate (37±1.9)‰, the second generation into a ball rate (39±2.2)‰, the third generation of a ball rate (39±1.0)‰o. CNE2-SP was significantly higher than the ball into CNE2cells, and P<0.01, significant statistical significance. That group of stem cell self-renewal ability to CNE2cells, consistent with the characteristics of stem cells; differentiation potential:CNE2-SP cells can be cultured in serum-containing fluid and serum-free medium which converts alternating with pluripotency. Resistance:drug effects at low concentrations, CNE2cell proliferation that began to appear, and CNE2-SP cells were not affected, as the drug concentration increased, CNE2-SP cells appear only partially inhibited cells, but this time CNE2cells have all died, CNE2-SP showed a greater resistance. CD44expression:protein CNE2-SP cells was significantly higher than that of CD44CNE2cells. CNE2-SP relative expression of CD44protein was higher than CNE2cells (8.1±0.92) times, identified as CNE2side population cells with stem cell properties. 2, Yiqijiedu side effects after72hours, the cells under different concentrations of drugs subject to varying degrees of inhibition. Omg/living cell rate (100±9.2)%,0.5mg/ml of live cell rate (87.2±6.8)%ml after the drug effect,1mg/ml of live cell rate (66.5±5.7)%,2mg/ml of live cell ratio (48.9±5.2)%,4mg/ml living cell rate (11.8±1.2)%. Yiqijiedu party for CNE2-SP cells significantly inhibited the proliferation of, and directly proportional to the concentration. IC50at2mg/ml or so.3, Yiqijiedu party intervention CNE2-SP cells24h,36h,48h after staining with JC-1, under a fluorescence microscope concept, early apoptotic cells appear obvious, over time, the cell vacuoles, cytoplasm shrinkage, nuclear chromatin margination, late apoptotic bodies.48h control group apoptosis rate of (7.6±1.1)%, drug24h, apoptosis rate (33.4±4.6)%, while the rate of apoptosis36h (48.5±8.1)%, while the rate of apoptosis48h (85.9±10.2)%. Three time points apoptosis rate compared with the control group P<0.01, statistically significant. Apoptotic cells in apoptotic cells and the sum of the proportion of viable cells (i.e., apoptosis rate) proportional to the time of drug action.4, Yiqijiedu party acting on CNE2-SP cells24h,36h,48h after, Western-blot detection of apoptosis-related proteins showed that, P53protein increased, an early marker of apoptosis Cleaved PARP protein expression increased apoptosis Survivin inhibitor gene downregulated, Cleaved caspase-9upregulation activate Cleaved caspase-3, Cleaved caspase-7these apoptotic effects caspase, significantly upregulated, indicating Yiqijiedu order to promote apoptosis. Cleaved caspase-8protein did not express the extrinsic pathway, suggesting Yiqijiedu square through the intrinsic pathway CNE2-SP-induced apoptosis. Apoptotic protein expression levels in the drug has reached the maximum36hours,48hours have declined.Conclusion: Yiqijiedu order effectively inhibit proliferation CNE2-SP cells and induce apoptosis mechanism Or cause DNA damage,(mitochondrial) pathway within the start endogenous...
|
PDF Full Text Request