Dads On Biological Behavior Of Sw480 Cells And Protein Expression Profiling Of

Part One Biological Effets of Diallyl Disulfide on SW480 Cell LineObjective: The biological effects of diallyl disulfide(DADS) was investigated in colonic cancer SW480 cell line.Methods: The SW480 cell cultured in vitro,proliferation inhibition of SW480 cell was tested by MTT assay. The change of cell morphologic was observed by HE staining and cell cycle was analyzed by flow cytometry(FCM).Results: MTT assay showed that DADS might significantly inhibited proliferation of SW480 cell in concentration-dependent manner. Morphologic observation presented that SW480 cell differentiation was induced by DADS. Flow cytometry analysis revealed that the percentage G₂/M phase and sub-duplation apex of SW480 cell,were increased by DADS.Conclusion: DADS significantly might inhibite proliferation of SW480 cells and arrest cell cycle in G₂/M. Part Two The Differential Proteomic Expression Analysis of SW480 Cells Induced by Diallyl DisulfideObjective: This study was designed to explore the differential proteomic expression induced by diallyl disulfide (DADS) in human colonic cancer SW480 cells and its related molecular mechanisms.Methods: A series of methods, including immobilized pH gradient-two dimensional polyacrylamide gel electrophoresis, peptide mass fingerpringting based on matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) ,database searching and so on, were used to separate and identify the differential proteomic expressions inducing effect of DADS on SW480 cells. Cells were processed for two-dimensional (2-D) electrophoresis and proteins were separated by isoelectric focusing and visualized by silver staining and Coomassie brilliant blue staining.After the pattern of polypeptide expression was evaluated by computer-assisted image analysis (PDQUEST), the different gel spots were excised and digested in situ and identified by MALDI-TOF-MS, and database searching.Results: The results showed that the good 2-DE pattern including high resolution and reproducibility was obtained. After silver staining, the 2-DE image analysis by PDQuest 2-DE software detected average (492±36) spots in SW480, and (523±24) spots in DADS treated SW480. And the average matching rate was 92% and 98% respectively. There were 167 spots that exhibited detectable quantitative changes between control and DADS-treated cells, 98 of them realized up-regulation while 69 of them realized down-regulation. Twenty of which were identified by mass spectrometry unambiguously. The expression of cytokeratin-18, IgE-dependent histamine-releasing factor, Cellular retinol-binding protein I, cytokeratin-1, myosin light chain, ubiquitin, H⁺-transporting ATPase, heat-shock protein-64, typothetical protein, guanylate cyclase-acting protein(GCAPs), oxytocinase/insulin-responsive aminopeptidase and sequence 9 from patent WO0164904 was increased in DADS-treated cells, whereas the expression of cytokeratin-19, actin-depolymerizing factor, V-1 protein, fructose bisphosphate aldolase, FK506-binding protein, capping protein, thioredoxin peroxidase, transformation-sensitive protein were down-regulation. These proteins were associated with cell differentiation, cell apoptosis, cell cycle, cell metastasis, transcription and cytoskeleton etc. DADS can inhibit the SW480 cells'ability of proliferation and induce the differentiation of SW480 cells by those protein.Conclusion:1. Up-regulated expression of guanylate cyclase-acting protein and myosin light chain or down-regulated of fructose bisphosphate aldolase and FK506-binding protein by DADS can inhibit the SW480 cells'ability of proliferation.Ubiquitin was involved in the effect of DADS on G2/M arrest in SW480 cells.2. DADS may induce the differentiation of SW480 cells via up-regulated expression of Cellular retinol-binding protein I, increase the expression of cytokeratin-18 and inhibit the expression of thioredoxin peroxidase induced SW480 cells apoptosis.3. DADS can inhibit the metastasis of colonic cnacer cells via down-regulated the expression of cytokeratin-19, capping protein and actin-depolymerizing factor.