The Study On Anti-Bladder Cancer Of Honkiol

Objective:For NMIBC,whether postoperative immunotherapy or intravascular chemotherapy,the recurrence and progress rate is still quite high,and most of the toxic side effects of drugs were very obvious.For MIBC,the radical cystectomy is alwalys the firist choice,adjuvant chemotherapy was alwalys applied postoperative.The platinum-based chemotherapy drugs is widely used in the clinical,but wchich emerged much toxic side effects to patients,and multidrug resistance alwalys generated after the used of adjuvant chemotherapy.So a drug with small side effects,and the good effect is immediately for the treatment of bladder cancer.Honkiol,which has been confirmed has multanti-tumor effort in vivo and vitro with the low side efforts.The efforts of anti-bladder cancer has been reported about honkiol,but the study was shallow.The aim of this study is to investigate the effect of honkiol on proliferation and apoptosis of bladder cancer T24 and EJ cell lines in vitro.And try to explore the possible mechanism,which will provide basic research for improvement of bladder cancer treatment.Methods: T24 and EJ cells were cultured in vitro,and the cells were treated with different concentrations of Honkiol after the logarithmic growth phase.First of all,to confirm the effect before the test,observe the cell proliferation condition after treated with drugs is needed.MTT and monoclonal assay were used to observe the cell proliferation.Second,using the flow cytometry to observe the cell cycle and cell apoptosis.Thirdly,the m RNA expression levels of the proteins were screened by PCR,Finally,Western Blot technique was used to check the expression of protein related to apoptosis and cyclin,the possible pathways were predicted.Each experiment in the test was repeated at least three times.Results: 1.MTT results: Both T24 and EJ cell proliferation was significantly inhibited when honkiol was used.They showed time-effect and concentration-effect relationships within a certain range.Monoclonal formation assay showed that Honkiol had a significant inhibitory effect on both tumor cells,and showed dose-dependent.2.Flow cytometry results: 48 h after treated with different concentration drugs,tumor cells are mostly inhibited in the G1 / S phase,indicating that Honkiol can prevent tumor cells G1 phase to S phase transformation,inhibition of tumor cell mitosis.The cells were significantly apoptotic and showed a dose-dependent effect 48 h after treated with different concentration drugs.3.PCR results: The changes of the related genes such as apoptosis,proliferation,and angiogenesis were observed at the level of m RNA.The m RNA levels of PCNA,CDK-4,CDK-6,BCL-2,MCL-1 and VEGF were significantly decreased,and the m RNA expression of Bad,Bax and Bak was significantly increased after treated with honkiol.The results showed that Honkiol inhibited the proliferation of bladder cancer cell lines,induced apoptosis of bladder cancer cells and inhibited the angiogenesis of tumor cells.4.Caspase-3,8,9 results: With the increase of concentration of Honkiol,the expression of Caspase increased gradually,showing a dose-dependent relationship,indicating that Honkiol-induced apoptosis showed a Caspase-dependent pathway.5.Western-Blot results: The expression of CDK-4,CDK-6 and BCL-2 protein could be inhibited,the expression of Bax protein is enhanced after treated with honkiol and showed in a dose-dependent manner.Conclusion: Honkiol can inhibit the proliferation of bladder cancer cells T24 and EJ cells,the main mechanism of its inhibition is preventing tumor cells transformation from G1 phase to S phase,inhibition of tumor cell mitosis.,which showed time-effect and concentration-effect relationships;Honkiol can induce apoptosis of bladder cancer cells T24 and EJ cells.There are two ways to produce apoptosis,namely Caspase-dependent pathway and Caspase-independent pathway.In addition,Honkiol not only affects the apoptosis of bladder cancer cells T24 and EJ cells through endogenous pathways,but also affects the apoptosis of tumor cells through exogenous pathways;Honkiol can inhibit tumor angiogenesis,but the specific mechanism remains to be explored.In conclusion,Honkiol has a good effect on bladder cancer cells,which can inhibit the effect of bladder cancer cells in a variety of ways,but still need to explore the relevant path in order to clinical application.