Inhibitory Effect Of Platycodin D On Human Multiple Myeloma Cells(RPMI8226) Proliferation And Its Mechanism

Objective: Research characteristics of Platycodin D(PD) on human multiple myeloma cells growth in vitro and the effect of whether to explore the mechanism of inhibition of cell proliferation by cycle arrest and induction of apoptosis.Multiple myeloma(multiple myeloma, MM) is a malignant tumor of the blood, incidence of a disease accounted for about 10% of the blood system of malignant tumor, which is the second largest blood system tumor.MM is occured in middle-aged and old, but in recent years, which has a tendency to incidence, early onset age.MM high molecular mechanism is complex and the heterogeneity of cytogenetics and have complex clonal evolution process.As proteasome inhibitors boron amine for meters and immunosuppressants Sally degrees, the drug lenalidomide, such as the emergence of new drugs, significantly improved the survival of young patients with MM, but still unable to overcome resistance and relapse of ending, and quite a number of patients often for economic reasons or severe toxicity reaction can’t tolerate the above treatment.MM is still considered incurable diseases, so look for high efficiency, low toxicity and cheap MM drug resistance is especially important.Rich resources of medicinal plants in our country, for thousands of years in practice has accumulated rich experience, which have unique structure compared with chemical synthetic drug resistance and wide biological activity,it is not easily and conveniently, and the advantages of low cost and little side effect, TCM active ingredients of anti-tumor research has become the modern medical research hot spot, that will open up new avenues for human anti-tumor treatment.Platycodin D is the main active ingredients of traditional Chinese herbal medicine platycodon grandiflorum, modern pharmacology study found that in addition to a cough expectorant, anti-inflammatory analgesic, protect liver, anti-oxidation, hypoglycemic lipid-lowering, a variety of action such as immune regulation, but also has extensive antitumor activity.In vitro experiments confirmed that PD can effectively inhibit liver cancer, gastric cancer, breast cancer, colon cancer, ovarian cancer, lung cancer, leukemia and other malignant tumor cell growth, and has no obvious influence on the normal tissue cells, its mechanism mainly includes the induction of apoptosis, block the cell cycle, cut telomerase, inhibit cancer metastasis and start to produce synergistic antitumor effect, etc., are likely to develop, become the new anti-cancer drugs.However, PD has inhibitory effect on whether human multiple myeloma cells,which has yet to see literature reports.This experiment by observing the PD on the influence of multiple myeloma RPMI8226 cell growth, and from the perspectives of inducing apoptosis and cycle arrest, discuss the mechanism of inhibition of cell proliferation, for PD is used to provide theoretical experimental basis for clinical treatment of multiple myeloma.Methods: In vitro human multiple myeloma RPMI8226 cells, using different concentrations determined by MTT colorimetric method PD function cells after 24 h, 48 h, 72 h, the impact on the cell proliferation activity;Annexin V- FITC/PI double staining flow cytometry to detect cell apoptosis rate after 24 h PD action;C organism iodide(PI) single staining flow cytometry to detect PD role within 24 h after the cell cycle distribution;Western blot detecting PD on cell cycle dependent protein kinase 1(CDK1) expression level and the effects of phosphorylation.Results:1 Determined by MTT colorimetry to detect RPMI 8226 cell survival rate: PD can obviously inhibit RPMI8226 cell proliferation, within the scope of 10-60μM and its inhibition in time and dose dependence, 24 h, 48 h and 72 h half inhibitory concentration(IC50 alue concentration), respectively 20.75μM、16.03μM and 14.54μM.2 Flow cytometry to detect cell apoptosis(Annexin V-FITC/PI double dye) : 10, 15, 20 microns RPMI8226 cells after 24 h PD treatment, compared with control group, cell apoptosis rate increased significantly, respectively, of 19.50%±5.1%, 35.80%±5.3% and 43.80%±4.0%(P< 0.05, the control group 8.9%±5.7%), with the increase of drug concentration, live cells reduce gradually, gradually increased apoptotic cells and cell death.3 Flow cytometry to detect the cell cycle(PI): 10, 15, 20 microns cells after 24 h PD, compared with the control group, the proportion of G2/M phase cells increased significantly, 72.83%±5.26% and 76.82%±5.26% respectively(P<0.05, the control group 56.780%±4.92%), with the increase of drug concentration, proportion of G2/M phase cells gradually increased.4 CDK1 and Western blot detection protein phosphorylation CDK1-Thr(14) : by different concentration of PD after the intervention, and G2/M phase of RPMI8226 cell block CDK1 related protein expression level has no obvious change, and CDK1 phosphorylation level increased.Conclusion:1 PD can inhibit the effect of multiple myeloma cell proliferation, within a certain range, the time- dose dependent2 PD for multiple myeloma proliferation inhibition and induction of apoptosis and the cell G2/M phase retardation.3 PD by inhibition of CDK1 phosphorylation, thus inhibiting the activation, blocked the cells in the G2/M phase.