Study On Glycoprotein Proteomics Of Hepatocellular Carcinoma Based On Liquid Chromatography - Mass Spectrometry

Part Ⅰ Collection of the serum from hepatocellular carcinoma (HCC) patients and healthy controlsObjective: Serum samples of HCC patients and healthy controls were collected for the following proteomic analysis.Methods:Between2012.3and2012.8, in the department of gastroenterology and invasive technology, peripheral blood was collected from patients in ward and outpatient. Serum samples were prepared and preserved. Corresponding clinical information was also collected.Results:104samples were collected in total, including52samples from HCC patients and52samples form healthy controls;54samples from men and50samples from women. Serum samples were prepared and preserved. Personal essential and clinical information tables were designed and completed.Conclusion:Serum samples and corrresponding information were standardly collected. Information of high quanlity can contribute to the following validation.Part Ⅱ Serum proteins’ separation, enrichment and analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS)Objective:Serum glycoproteins from the HCC patients and healthy controls were analysed through proteomic technique.Methods:After removal of the high abundant proteins, enrichment of the glycoproteins and digestion by trypsin and peptide-N-glycosidase F (PNGase F), the samples were analysed using LC-MS/MS.Results:The removal of the high abundant proteins and enrichment of the glycoproteins were both efficient and reliable. The spectrogram was clear and showed no contamination.Conclusion:Our work provided a systematic and quantitative method for glycoprotein and glycosylation sites identification from serum samples. The spectrum accessed can be used for the following bioinformatics analysis.Part Ⅲ Bioinformatics analysis of serum glycoprotomics of HCC patients and healthy controlsObjective:Differentially expressed serum glycoproteins and glycosylation sites were explored between HCC patients and healthy controlsMethods:Protein searches were performed with MaxQuant1.3.0.5against the IPI databases. Glycopeptides were submitted to the NetNGlyc1.0Server, CBS information tools to predicte the N-glycosylation sites. Results derived were compared with the experiment results. Gene ontology (GO) analysis were performed on all the proteins identified.Results:152glycosylation sites and54glycoproteins were identified. Glycosylation sites derived from the experiment are in general accord with those predicted by the database, which suggested the dependability of the methodology of the experiment.10of the54proteins were identified differently expressed between HCC patients and healthy controls, which had been reported to be related with several malignant tumors, and may be involved in several biological process, cellular component and molecular function of HCC.Conclusion:Glycoroteins differently expressed between HCC patients and healthy controls may be promising HCC biomarkers and may be helpful to unveil the underlying mechanisms of hepatocarcinogenesis and development.