The Proteomics Study Of Hepatocellular Carcinoma Serum

Background and Objective:Hepatocellular carcinoma(HCC)is a leading cause in cancer related death.Novel serum biomarkers are required to increase the sensitivity and specificity of serum screening for the early diagnosis of HCC.In this study,a quantitative proteomics strategy was employed to study the differential expression of serum glycoproteins between HCC and normal control serum samples.Meanwhile,in order to improve the comprehensiveness and convenience of serum protein detection,we established an 2D LC-MS/MS system to analysis the proteome of the whole serum without depleting abundant protein.Methods:Lectin affinity chromatography was used to enrich glycoproteins from the serum samples.Quantitative mass spectrometric analysis combing dimethyl stable isotopic labeling and two dimensional liquid chromatography(2D LC)separations was used to examine the differential levels of the detected proteins between HCC and control serum samples.The differential expression levels of three serum proteins were examined using Western blotting.And an SCX-RP column integrated with a LTQ linear ion trap mass spectrometer was applied to study the the proteome of the whole serum without depleting abundant protein.Results:Using the 2D LC-MS/MS method,2280 protein groups were identified in the serum samples from HCC patients.Thirty-six proteins were found to be upregulated in the HCC serum,and 19 proteins were downregulated.Three differential glycoproteins were validated using Western blotting,including fibrinogen gamma chain(FGG),FOS-like antigen 2(FOSL2)and alpha-1,6-mannosylglycoprotein 6-beta-N-acetylglucosaminyltransferase B(MGAT5B).All these three proteins were upregulated in the HCC serum samples.An integrated HPLC column consist of a SCX and a C18 reverse-phase section was applied to separate online 2D LC,which was connected to a linear ion trap mass spectrometer.We identified over a thousand protein groups from a single serum sample.Conclusion:Quantitative glycoproteomics method was established and proved to be useful for mining potential novel biomarkers for HCC.And 2D LC-MS/MS system was established and this method enabled 2D separation of tryptic peptides of complex biological samples with a conventional one-dimensional chromatography system.