Study On ITRAQ Quantitative Proteomics In Human Lung Adenocarcinoma Of A549Cell

Objective:Using a relative quantitative method, the protein expression profile of a spectrumof human lung adenocarcinoma of A549cells after treatment with elemene and/orX-irradiation, was analyzed. HSP27and COX-3were proved to be expressedsignificantly different between elemene and/or X-irradiation treated group and controlgroup. To explore the potential value of HSP27and COX-3as the sensitive protein forthe diagnosis and treatment of lung adenocarcinoma, the difference was further verifiedby biological validation strategies.Methods:1. In vitro iTRAQ labeling combined with liquid chromatography-massspectrometry, the relative change in protein expression after treatment with elemenesingle-agent, X-ray irradiation, elemene combined with X-ray irradiation, was analyzedand compared with normal cultured human lung adenocarcinoma A549cells.2. The protein expression of HSP27and COX-3in cells of elemene monotherapygroup, X-ray irradiation group, elemene combined with X-ray irradiation group andnormal cultured human lung adenocarcinoma A549cells was profiled by ELISAdouble-antibody sandwich methods and a statistical analysis was performed on theseresults.Results:1. A quantitative proteomic approach with isobaric labeling (iTRAQ) was appliedto evaluate the protein expression differences between each experimental groups and thecontrol group,514differentially expressed proteins were identified. And33proteinsexpressed significantly different, when a differential expression filter (1.5fold) wasapplied to the protein expression values. Among them,15proteins deemed statistically significant by the P<0.05filter. The function of the15proteins involved in cellularenergy metabolism, RNA processing and transit, signal transduction, cell differentiationand apoptosis.2. The expression of HSP27protein in each experimental group: the concentrationof protein in the irradiation group was obviously higher than that of control group (P <0.05), the drug group was significantly lower than the control group (P <0.05), jointgroup was higher than the control group (P <0.05), the expression of protein in the druggroup was significantly lower than the combination group (P <0.05), the irradiationgroup was higher than the combined group (P <0.05).The expression of COX-3protein in each group was the same to HSP27protein.Conclusion:For the first time, iTRAQ coupled with LC-LTQ-orbitrap mass spectrometry wasapplied to explore a spectrum of human lung adenocarcinoma of A549cells. Based onthe isotope labeling proteomics relatively quantitative techniques, the differentiallyexpressed proteins after treatment by different methods were obtained. The function ofthese proteins mainly involved in cellular energy metabolism, DNA damage and repair,regulation of gene expression, signal transduction, and cell proliferation related to theprocess of differentiation and apoptosis. These findings were expected to provideexperimental evidence for revealing the molecular mechanisms of elemene andradiation therapy or finding the sensitive protein of related treatment.The results obtained from the verification of HSP27and COX-3showed that theirexpression was increased to different levels after radiation or elemene and X-rayirradiation therapy. However, the expression of HSP27and COX-3were decreasedsignificantly when treated with elemene alone. The results confirm that HSP27andCOX-3play a different role in the lung adenocarcinoma cells when treated by elemeneand/or X-ray irradiation, and their expression may closely related to the developmentof lung adenocarcinoma. Therefore, HSP27and COX-3proteins may become thesensitive protein of related treatment to evaluate the elemene and/or radiation therapyof lung adenocarcinoma.