The Study Of Anti-tumor Effect Of Zinc Oxid Nanoparticles By Altering The Modification Status Of Histone

Objective:Zinc oxid nanoparticles(ZnO NPs)has attracted great interest because of its attractive properties and sufficiently used in a wide spectrum of fields.But the issue of the biosecurity is less than our expectation.The latest research demonstrated that high concentration of ZnO NPs have anticancer effect which mainly dependent on reactive oxygen formation.However,the effect and promote or inhibit mechanism to cancer cell which caused by low or sub-leghal concentrations of ZnO NPs that analogy to environmental exposure is still unclear.Besides,as far as we know,the urinary tract is the the mainly passage for ZnO NPs.We detected the altering of modified status of histone in three urinary system tumor under sub-lethal concentrations of ZnO NPs,and measured the changing expression of modified enzyme together with their targeted downstream gene.What's more,biological behavior was also detected belong to distinct concentrations of ZnO NPs.Methods:For this study,human kidney cancer cell line A498,prostate cancer cell line DU145,and bladder cancer cell line T24 were cultured in vitro.After 48 hr of exposure to concentration gradient of ZnO-NPs,the MTT Assay and Alamar Blue Assay were performed to measure cell viability and were used to choose specific concentrations to study.In order to demonstrate the epigenetics effect caused by ZnO particles,we extracted mRNA and protein from cells and used q-PCR and Western Bolt to detect the transcription and translation level of 4 histone related-enzyme,ultimately,we designated EZH2 protein which concentration-dependent upon ZnO exposure in T24 as a object of study.Then,we checked the expression of H3K27me3 which was deem to a specific modifier spot of EZH2.What's more,we detected the cellular ROS level and the degree of DNA damage so as to explore the effect of oxidative stress.Besides,we moved forward to demonstrate the effect which caused by histone status changed and found the downstream targeted gene RUNX3.What's more,we used CHIP-qPCR to verify the altering of histone status on the promoter ofRUNX3.In the end,wound healing,transwell and fluorescence activated cell sorting were prepared to detected the biological behaviour under different concentrations of ZnO.Results:The MTT And Alamar Blue Assay demonstrated that sub-lethal concentrations of ZnO particles have proliferation inhibition effect to urinary tumor cells(A498,DU145,T24)and ROS/DNA damage had no business of inhibition effect.Western Blot and q-PCR jointly demostrated that only EZH2 showed concentration-dependent upon ZnO exposure among four histone related enzyme(CBP,LSD1,EZH2,HDAC1),that with the increasing concentration of ZnO,the expression of EZH2 decreased.Besides,the changing expression of H3K27me3 in accord with EZH2.We further demonstrated that the expression of RUNX3 which is one of downstream target gene of EZH2,had inverse dynamic change compared with EZH2.CHIP-qPCR illustrated the negative correlation between H3K27me3 level and EZH2 expression on RUNX3 promoter.With the increasing concentration of ZnO in T24,the capacity of invasion and migration decreased,degree of apoptosis increased and cell cycle arrested in S phase.Conclusion:The proliferation of urinary tumor cells can be inhibited by low concentrations of ZnO particles and the effect wasn't caused by genotoxicity or cytotoxicity.The expression of EZH2 can be inhibited by ZnO particles resulted in H3K27me3 decreased on the promoter of RUNX3 in T24.As a result,the expression of RUNX3 increased.The elevated process of apoptosis and S phase arrest were found in T24 with the increased concentrations of ZnO particles.The ability of invasion and migration of T24 declined with the increased concentrations of ZnO particles.