Differential Proteomic Analysis of Cancer Stem Cell Properties in Hepatocellular Carcinomas by Isobaric Tag Labeling and Mass Spectrometry

Abstract

Malignant tumors are relatively resistant to treatment due to their heterogeneous nature, drug resistance, and tendency for metastasis. Recent studies suggest that a subpopulation of cancer cells is responsible for the malignant outcomes. These cells are considered as cancer stem cells (CSC). Although a number of molecules have been identified in different cancer cells as markers for cancer stem cells, no promising markers are currently available for hepatocellular carcinoma cells. In this study, two clones of Hep3B cell lines were functionally characterized as control or CSC-like cells, based on properties including spheroid formation, drug resistance, and tumor initiation. Furthermore, their protein expression profiles were investigated by isobaric tags for relative and absolute quantitation (iTRAQ), and a total of 1,127 proteins were identified and quantified from the combined fractions; 50 proteins exhibited at least 2-fold differences between these two clones. These 50 proteins were analyzed by GeneGo and were found to be associated with liver neoplasms, hepatocellular carcinoma (HCC), and liver diseases. They were also components of metabolic pathways, immune responses, and cytoskeleton remodeling. Among these proteins, the expressions of S100P, S100A14, and vimentin were verified in several HCC cell lines, and their expressions were correlated with tumorigenicity in HCC cell lines. The functional significance of vimentin and S100A14 were also investigated and verified.

Comparative analysis of primary hepatocellular carcinoma with single and multiple lesions by iTRAQ-based quantitative proteomics

“…For instance, Huang et al have reported the iTRAQ-based quantitative analysis of the recurrence/metastasis of Huge HCC (diameter is larger than 10 cm) at different stages, and identified two biomarkers for distinguishing and predicting the early recurrence/metastasis behaviors of Huge HCC [26]; Wang et al have reported the serum proteome of metastatic HCC using the iTRAQ-based quantitative analysis and identified a metastasis-associated biomarker [27]. Ko et al have reported the iTRAQ-based quantitative analysis of HCC cancer stem cell proteome [28]. Meanwhile, the iTRAQ-based quantitative study of the proteome changes and the secreted proteome changes during HBV infection have also been reported [29,30].…”

Quantitative proteomics analysis of early recurrence/metastasis of huge hepatocellular carcinoma following radical resection

“…So far, several groups have reported the applying of iTRAQ based quantitative proteomics approach in the study of hepatic cancer, especially for the screening of diagnostic or prognostic protein biomarkers. He et al have reported the serum biomarker screening of AFP negative HBV related HCC through the iTRAQ based approach [35]; Ko et al have reported the iTRAQ based quantitative analysis of HCC cancer stem cell proteome [36]; Huang et al have reported the iTRAQ based serum biomarker screening of the HCC micro-vascular invasion [37]; Qin et al, Yu et al and Wang et al have reported the screening of metastatic related proteins of HCC through iTRAQ based approach [33,38,39]; the iTRAQ based quantitative study of proteome change during HBV infection has also been reported [40,41]. However, the application of iTRAQ labeling in studying the molecular mechanisms and screening for biomarkers associated with the early recurrence/metastasis of H-HCC has not yet been reported to our knowledge.…”

Differential proteomics of monosodium urate crystals‐induced inflammatory response in dissected murine air pouch membranes by iTRAQ technology

“…The samples were loaded on 24 OFFGEL wells. The separation was carried out for 48 h using the 3100 OFFGEL fractionator (Agilent Technologies, Wilmington, DE, USA) with a limiting current of 50 μA and a limit of 50 kV· h before holding the voltage at 500 V. The samples from 24 fractions were cleaned‐up using C18 Spin columns for further analysis .…”