Research On The Growth Inhibition Effects Of Pristimerin On Human Nasopharyngeal Carcinoma Cells

Objective:Observe the growth inhibition effects of pristimerin on human nasopharyngeal carcinoma cell,and the possible mechanism of inducing tumor cell apoptosis.Methods:(1)Methyl thiazolyl tetrazolium(MTT)assay was used for cell proliferation determination in vitro by pristimerin in human nasopharyngeal carcinoma cells HNE-2、CNE-2、HONE-1;(2)Using flow cytometry to investigate pristimerin effect on apoptosis in HNE-2 cells;(3)MTT assay was used to evaluate the influence of the caspase inhibitor to pristimerin on cell apoptosis effects of HNE2 cells;(4)Western blot Analysis was used to detect the influence of pristimerin on the expression of cleved Caspase3、Caspase9、PARP 、 Bcl-2 family,and HSP90 protein expression.;(5)Co immunoprecipitation assay was used to evaluate the influence of pristimerin on the interaction between HSP90 and CDC37;(6)MTT assay was used for cell proliferation determination in vitro by pristimerin in human nasopharyngeal carcinoma cell which is restrained by 5-fluorouracil;(7)Western blot Analysis was used to detect the influence of pristimerin alone and the combination pristimerin with5-fluorouracil in thymine synthase protein of human nasopharyngeal carcinoma Cell HNE2;(8)The study of the possible mechanism that pristimerin reduce thymine synthetase by using the Co-Immunoprecipitation.Results:(1)Pristimerin was dose-dependently inhibition in human nasopharyngeal carcinoma cells HNE-2、CNE-2、HONE-1;(2)Pristimerin can enhance HNE-2 cells apoptosis on dose-dependently mode;(3)Pristimerin can use the down-regulation Bim and up-regulated Bcl-x L in Bcl-2 family proteins,also canreduce the expression of endoplasmic reticulum stress signaling related proteins such as Bi P,PEAK,etc.to induced apoptosis.;(4)Pristimerin induced customer proteins of HSP90,such as EGFR,C-MET,with the increase of pristimerin concentration and showed a decreasing trend.It also can inhibited the interaction between HSP90 and its chaperone CDC37;(5)Pristimerin can enhance synergistically the inhibitory effect of5-fluorouracil on human nasopharyngeal carcinoma HNE-2 cell proliferation;(6)Pristimerin can reduce TS in a concentration dependent manner by itself,and it can also reduce TS content combined with 5-fluorouracil;(7)Pristimerin can obviously inhibited the interaction between HSP90 and TS.Conclusion:Pristimerin has the effect of inhibiting proliferation and inducing apoptosis of nasopharyngeal carcinoma cells.Through the study found that pristimerin induced apoptosis in nasopharyngeal carcinoma cells and the activation of the intrinsic apoptotic pathway and endoplasmic reticulum stress pathway is closely related to.Pristimerin as a new type of HSP90 inhibitor,can lead to customer due to loss of HSP90 protein protection and degradation,but also through the interference of HSP90 and its auxiliary interacting partner CDC37,thereby inducing cell apoptosis. In addition,pristimerin can enhance synergistically the inhibitory effect of 5-fluorouracil on human nasopharyngeal carcinoma HNE-2 cell proliferation,can reduce the synergy 5-fluorouracil thymidylate synthase protein.The mechanism may be related to pristimerin by inhibiting HSP90 activity and promote the degradation of thymidylate synthase.All of these provide a new way to solve the problem of how to improve the sensitivity of 5-fluorouracil chemotherapy and how to reverse the drug resistance problem that has been solved in clinic.For the future will provide a theoretical basis for the development of pristimerin high selectivity low adverse natural antitumor drugs,indicates a new direction for the optimization of chemotherapy in patients with nasopharyngeal carcinoma.