Studies On Mechanism Of Phycoerythrin Inducing MCF-7 Cells Senescence Through P53-p21-pRB Pathways

This study explores the actions of the inhibition of human breast cancer cells MCF-7 cultured in vitro by Phycoerythrin (PE) and described its regulation of p53-p21-pRB pathways to induce human breast cancer cells MCF-7 of the molecular mechanism of senescence.In vitro studies using mononuclear cell direct cytotoxicity assay(the MTT method) and found PE inhibit human breast cancer cells MCF-7 growth inhibition effect obviously,with IC50 being 134.87μg·mL-1.By morphological identification of PE senescence effect on the tumor cells, usingβ-galactosidase staining kit, DAPI staining of tumor cells stained, using ordinary optical microscope and fluorescence microscope observations, tumor cells have the typical senescence characteristics, found that great changes of tumor cells become flattened, cell shrinkage, cytoplasmic vacuolization and begin to sprout, blistering. The P-galactosidase (SA-P-gal) staining, inverted microscope and found that treatment group characteristic of senescence cells to secrete P-galactosidase was increased by dyeing to produce blue material. DAPI staining observed by fluorescence microscopy, the nucleus of drug-treated DNA showed punctate aggregated distribution, and with PE concentration increased, the phenomenon of more and more obviously that increasing the proportion of cells senescence.Subsequently, the use of Fluo-3/AM dyed, using confocal laser scanning microscope detection of PE on the tumor cells, calcium ion concentration. The study found that with increasing concentration of PE, PE tumor cells can significantly increase the concentration of calcium ions. And showed a definite dose-dependent relationship. Carried out using TMRE staining, using confocal laser scanning microscope phycoerythrin on the tumor cells, mitochondrial membrane potential impact. The study found that with the PE concentration increasing, PE can significantly reduce the mitochondrial membrane potential in tumor cells. The use of 2',7'-dichloro fluorescein hydrogenated fat (DCFH-DA) staining of tumor cells, using confocal laser scanning microscope of tumor cells, PE content of reactive oxygen species. The study found, with the PE concentration increased, the tumor cells of reactive oxygen species levels are also increasing. On this basis, we initially examined the PE inducing tumor cells senescence way. In the confocal laser scanning microscope observation, different concentrations of PE effect on human breast cancer cells MCF-7 48h after the tumor cells intracellular p53, p21, and pRB proteins have increased fluorescence intensity and concentration with the PE increase, p53, p21, and pRB protein in the fluorescence intensity also increasesd. This effect shows PE can induce human breast cancer cells MCF-7 expressed senescence-associated protein, p53, p21, and pRB. From the above, it can be concluded that PE can inhibit in vitro the growth of human breast cancer cells MCF-7. By morphological observation, tumor cells can be induced by PE of the typical senescence characteristics. Role in the mechanism may be related to tumor cells by increased intracellular calcium concentration and raised the ROS level, which activate cells senescence start-protein-p53 protein, thereby inducing cells senescence occurred. In a word, PE has the anti-tumor effect obviously in vitro and PE can induce the human breast cancer cells MCF-7 senescence via p53-p21-pRB pathways.