Comparative proteomics of side population cells derived from human hepatocellular carcinoma cell lines with varying metastatic potentials

Abstract

Metastasis and recurrence following surgery are major reasons for the high mortality rate and poor prognosis associated with hepatocellular carcinoma (HCC). Cancer stem cells (CSCs) are thought to be able to cause cancer, and to be the primary cause of tumor recurrence and metastasis. The underlying mechanisms of the metastatic potential of CSCs is poorly understood. In the present study, side population (SP) cells were isolated from 4 HCC cell lines, and their self-renewal and migratory abilities were compared. The results demonstrate that SP cells from different cell lines exhibited similar self-renewal abilities but different metastatic potentials. Furthermore, the overall proteomes of the SP cells were systematically quantified. This revealed 11 and 19 differentially expressed proteins (DEPs), upregulated and downregulated, respectively, associated with increased metastatic potential. These proteins were involved in the ‘regulation of mRNA processing’ and ‘cytoskeleton organization’ biological processes. The majority of the proteins were involved in ‘cell proliferation’, ‘migration’ and ‘invasion of cancer’, and may promote HCC metastasis in a synergistic manner. The AKT and nuclear factor-κB signaling pathways may contribute to the regulation of HCC metastasis through regulating the DEPs in SP cells. To the best of our knowledge, the present study is the first to demonstrate the overall proteome difference among SP cells from the different HCC cell lines with different metastatic potentials. The present study provides novel information regarding the metastatic potential of CSCs, which will facilitate further investigation of the topic.

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“…The data analysis procedure was similar to our previous reports with certain adaption. 18 Briefly, MS data acquiring were processed using Proteome Discoverer (Thermo Fisher Scientific, Bremen, Germany; version 2.1) against the uniprot human_database (released at 2014-04-10, 20,264 entries). The enzyme specificity of trypsin was set and maximally up to two missed cleavages were allowed.…”

Quantitative Analysis of Tissue Secretome Reveals the Diagnostic and Prognostic Value of Carbonic Anhydrase II in Hepatocellular Carcinoma

“…The proteomic studies and data analyses were modified from our previously reported protocols [44]. Briefly, the thiol groups of the above extracted protein samples from the culture supernatant of 3 groups were alkylated with 50 mM iodoacetamide (cat No.…”

“…The data processing strategy was also carried out according to our previous publications, with some modification [44]. The LC-MS/MS data acquisition was processed using Proteome Discoverer (version 1.4, Thermo Fisher Scientific, Germany) and searched using the Sequest HT (version 2.5.1, Matrix Science, United Kingdom) search algorithms against the human database (Uniprot, 20,264 entries, released at April 10, 2014).…”

“…Western blotting was performed according to a previous publication [44]. Briefly, proteins were separated on a 12% SDS-PAGE, and the gel was transferred to a nitrocellulose membrane (cat No.…”

“…The cell migration and invasion assays were carried out according to our previously published protocols [44]. The cell migration ability was investigated by using transwell units with 8 μm pores (cat No.…”