The Preparation Of Realgar Nano Particle And The Studies Of Inducing Apoptosis In MCF-7 Cells

Realgar comes from the mineral sulfide realgar family,which is a traditional chinese medicine.In this study,Realgar Nanoparticles?NRP?were prepared by high-energy ball milling technology,and the influence of particle size and the content of As₂O₃ in different ball milling time were investigated.To investigate the effect of water,acid water and alkali water processing methods on NRP for preparing realgar nanoparticles of high purity and low toxicity,with the indicators of processed products yield,weight of the residue,particle size distribution and the content of As₂S₂ and As₂O₃.At the same time,the inhibition of Crude Realgar?CR?,NRP and Realgar Nanoparticles Processed?NRPP?were determined by MTT method in human breast cancer MCF-7 cells.And Hoechst33258 dye,PI staining and AnnexinV-FITC/PI double marking methods were applied to investigate the difference of inducing apoptosis on MCF-7 cells by CR and NRPP.To preliminary research the mechanism of NRPP induced apoptosis in MCF-7 cells,the mitochondrial membrane potential was detectd by Rh-123 staining method,and the expression of apoptosis related proteins such as bcl-2,caspase8,bax,caspase9,caspase3 and p53 protein were investigated by western blotting.The results showed that with the extension of milling time,the size of particle showed a tendency of increase with the decrease of the first,and the minimum average size was 157.3nm when the Realgar were milled for 16 h,while the content of As₂O₃ increased.The acid water processed has the minimum particle size,the highest content of As₂S₂ and lowest of As₂O₃,thus acid water processing was the best processing method in three methods.CR,NRP and NRPP could inhabited the proliferation of MCF-7 cells in dose-and time-dependent manners by MTT method.CR andNRPP could induce MCF-7 cell apoptosis in vitro through Hoechst33258 dye and Annexin V-FITC/PI double marking method with two aspects of qualitative and quantitative.CR could make MCF-7 cells cycle arrested in G2/M phase by PI staining,while NRPP arrested in S phase.The mitochondrial membrane potential of MCF-7 cells was markedly decreased in a dose-and time-dependent by NRPP on Rh-123 staining method.The expression of p53,caspase8,bax,cyt-c,caspase9 and caspase3 proteins dramatically increased in dose-and time-dependent manners,while the expression of bcl-2 decreased,then the ratio of the bcl-2 / bax declined.In general,the present results demonstrated that CR,NRP and NRPP could both inhibit tumor growth in MCF-7 cells,and the inhibition of CR and NRPP was through the cell cycle arrested and apoptosis induced;the mechanism of NRPP induced apoptosis in MCF-7 cell may be related to the activation of p53-bcl-2-caspase9,caspase8-mitochondria-caspase9 and caspase8-caspase3 pathways.