Medroxyprogesterone Acetate-induced Colon Cancer SW480 Cell Apoptosis And Differential Expression Protein Analysis

ObjectivesTo establish the apoptosis model that SW480 are treated with medroxyprogesterone acetate(MPA). In addition, to investigate changes in the protein spectrum of SW480 cells MPA-treated with comparative proteome research and provide new experimental evidence to further elucidate the mechanism of MPA-mediated cell apoptosis from protein molecular level.Methods1.SW480 cells treated with MPA were induced to be apoptosis.MTT and flow cytometry were used to detect the SW480 cells inhibition rate in order to select the appropriate drug concentration and induction time. To observe the changes on subcellular structure in SW480 by transmission electron microscope before and after treatment with MPA.Cell cycle and apoptosis detection were used to verify the apoptosis of treated SW480 cells, to establish the apoptosis model.2.We separated proteins of SW480 cells in MPA-treated and control group by two dimensional gel electrophoresis(2-DE) and the 2-DE images were analyzed with PDQuest8.0 software. Trypsin digested differentia protein spots of SW480 cells treated and untreated. HPLC-Chip/MS was adopted to on-line analyze and indentify differentially expressed protein. 3.Mascot software was used to search for the differentially expressed proteins in UniProtKB/SWISS-PORT, bioinformatics analysis can gain MS/MS spectrum of peptide from differentially expressed proteins.Results1.The proliferation of SW480 cells significantly was inhibited, and SW480 cell apoptosis was induced by different concentrations of MPA.The IC50(50%inhibitory concentration)dose was about 50μmol/L. Cell cycle analysis indicated that the MPA-induced apoptosis in SW480 cell may be related to cell cycle arrest-related, which was in a concentration - dependent. After SW480 cells treated by 50μmol/L for 48h, nuclear condensation,fragmentation,chromatin condensation,aggregation and a lot of apoptotic bodies was observed under in transmission electronic microscope.2.Screening 5 differentia protein spots in analysis of 2-DE images used by PDQUEST software.Compared with control, 2 proteins were up-regulated while 5 were down-regulated. The 7 kinds of SW480 cell apoptosis-related proteins have been identified by HPLC-Chip/MS.They were Ubiquitin,LMO3,HSP10,Solute carrier family 22 member 11, UCHL 2,NDPK-A,Laminin receptor 1.ConclusionsMPA can induce homo SW480 cell apoptosis which is possible related to arrest of cell cycle.The 7 differentialy expressed proteins identified in this study may serve as potential MPA anti-colon cancer biomarkers and play very important roles in MPA-mediated cell apoptosis.