The Synergy Of Specific-Recombinant Oncolytic Adenovirus Gene Target Therapy Combined With Radiation Therapy In Bladder Cancer Cells

Objective Current treatments for bladder cancer include surgery, radiotherapy and chemotherapeutics. However, current treatments are not effective for recurrent and advanced disease. Gene therapy is one of the most important therapeutic straegy for improving the antitumor efficacy. Rdiosensitiveness which is the key point for radio therapy determines the antitumor efficacy.This study was designed to clarify the synergy of Specific-Recombinant oncolytic adenovirus gene target therapy combined with radiation therapy in bladder Cancer Cells and approach the underlying mechanisms as the observation system that bladder cancer cells were transfected with recombinant oncolytic adenovirus.Methods The bladder cancer cells BIU-87and EJ were cultured. We detected the tite and the purity of the oncolytic adenovirus by TC150method and determined plaque forming unit(PFU);by the electronmicroscope, we can observe that bladder cancer cells changed into a rounded shape and condensation of the cytoplasm which is the cytopathic effect (CPE);We detected the survival rate of the bladder cancers infected with oncolytic adenovirus or irradiation alone or combination treatment by the MTT assay method;Caspase-3apoptosis assay was applied to show whether the apoptosis of the bladder cancer cells after irrdadiation increased; By the flow cytometry,we study the revolution and apoptosis of the bladder cancer cells after intervention; electron lens was used to observe adenovirus particle that infected into cells and the changes of cellular structure and shapes; Western blot was aimed to study the path way of the DNA break repair which involved the protein of the MREll,γ-H2AX-Ser139, chk2-thr68and research whether the adenovirus receptor up-regulation and the expression of both the hexon protein and the El A multed;A TCID50assay showed the difference in the virus yield between the bladder cancer cells irrdadiated and the unirrdadiated.Result Specific recombinant oncolytic adenovirus has high purity and titer, we definited the optimization quantity of the virus.we detected that adenoviral replication in the bladder cancer cells infected by the virus results a rounded shape and condensation of the cytoplasm that is the cytopathic effect (CPE).The study determined the timing of the oncolytic adenovirus intervention with regard to radiation was that virus infection was scheduled24hr after irradiation and proved that oncolytic adenovirus combined with radiotherapy improved antitumor efficacy compared to either alone agent.meanwhile the survival rate of the bladder cancer cells interfered by combination treatment was timedependent and dosage-dependent. We can see the apoptosis in the bladder cancer cells by combination treatment was increased with the increasing of the dosage of the ray which did not have an effect on the tite of the virus.Autophagic vacuoles were seen in both the bladder cancer cells infected by virus and irradiated, which was the greatest in combination treated cells, and then virus particle that existed scatterly and gatherly observation by electron lens proved that oncolytic adenovirus could enter into the cellule and replicate.By flow cytometry, we analyzed that cell which was effected72hours by combination treatment cycles was inhibited in the G2/M stage.The protein of the DNA break repair MRE11, chk2-thr68were decreased while γ-H2AX-Ser139increased, at the some time,the E1A gene and the hexon protein of the the oncolytic adenovirus expression were increased following with the adenovirus receptor increasing.Conclusion The paper proved the synergy of specific-recombinant oncolytic adenovirus gene target therapy combined with radiation therapy in bladder cancer cells and investigated the synergy cellular mechanisms,what was more,the study provided the combination treatment therapeutic strategy for the bladder cancer.