Article

Dynamic alteration of protein expression profiles during neoplastic transformation of rat hepatic oval-like cells

Liver Cancer Institute, Affiliated Zhongshan Hospital of Fudan University, Shanghai, China.
Cancer Science (Impact Factor: 3.52). 03/2010; 101(5):1099-107. DOI: 10.1111/j.1349-7006.2010.01513.x
Source: PubMed

ABSTRACT

To explore the molecular basis of neoplastic transformation of hepatic oval cells, a proteomic strategy was utilized to examine the global protein expression alterations during neoplastic transformation of rat hepatic oval-like cells. N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-initiated WB-F344 cells were treated with H(2)O(2) for neoplastic transformation. The transformed cells were identified by soft agar assay and MTT assay. The subsequent proteomic separation and identification were performed with 2-DE followed by MALDI-TOF-MS/MS analysis. Of the 148 differentially displayed protein spots analyzed, 121 spots representing 79 distinct proteins were finally identified. The expression levels of interested proteins were validated by western blotting including 40 S ribosomal protein A (RPSA) and cytokeratin 8. Bioinformatics annotations indicated that these identified proteins were enriched with oxidoreduction and stress response; transcription, translation, and protein processing; and energy/metabolism functions. Interestingly, 17 of the identified proteins were also found to be involved in early hepatic differentiation of mouse embryonic stem (ES) cells in our previous study. Twenty-six proteins had been reported as being dysregulated in hepatocellular carcinoma and other cancers. It suggested that these changed proteins may be implicated in neoplastic transformation of WB-F344 cells. The results may provide some clues for understanding the molecular mechanisms of hepatocarcinogenesis as viewed from dysregulation of differentiation.

Download full-text

Full-text

Available from: Xuefei Li, Feb 10, 2015
  • Source
    • "The rat hepatic oval cell line WB-F344 (abbreviated WB cells) used in this study is structurally and phenotypically simple epithelial cells that were isolated from the liver of an adult male Fischer 344 rat [27]. Their morphological and biological properties have been previously characterized as mostly resembling the oval cells [28]. Furthermore, these cells could not form tumors when injected into nude mice [9]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatocellular carcinoma (HCC) can be derived from malignant transformed adult hepatic progenitor cells. However, the regulatory factors and molecular mechanisms underlying the process are not well defined. Our previous microRNA (miRNA) microarray analysis revealed a significant decrease of miR-200a level in F344 rat HCC side population (SP) fraction cells versus their normal counterparts. In the present study, we further investigated the effect of miR-200a on hepatic oval cell (HOC) phenotypes. We first confirmed downregulated miR-200a levels in rat hepatoma cells compared with WB-F344 cells. Next, by lentivirus-mediated loss-of-function studies, we showed that stable knockdown of miR-200a confers a mesenchymal phenotype to WB-F344 cells, including an elongated cell morphology, enhanced cell migration ability and expression of epithelial mesenchymal transition (EMT)-representative markers. Concomitantly, several cancer stem cell (CSC)-like traits appeared in these cells, which exhibit enhanced spheroid-forming capacity, express putative hepatic CSC markers and display superior resistance to chemotherapeutic drugs in vitro. Furthermore, bioinformatics analysis, luciferase assays and western blot analysis identified β-catenin (CTNNB1) as a direct and functional target of miR-200a. Knockdown of miR-200a partially activated Wnt/β-catenin signaling, and silencing of β-catenin functionally attenuated anti-miR-200a effects in vitro in WB-F344 cells. At length, in vivo xenograft assay demonstrated the acquisition of tumorigenicity of WB-F344 cells after miR-200a siliencing. Collectively, our findings indicate that miR-200a may function as an important regulatory factor in neoplastic transition of HOCs by targeting the β-catenin pathway.
    Full-text · Article · Nov 2013 · PLoS ONE
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatocellular carcinoma (HCC) is the third most common cause of cancer mortality. Significantly downregulated histidine-rich glycoprotein (HRG) during the dynamic stages (WB, WB7, and WB11) of neoplastic transformation of WB F344 hepatic oval-like cells was screened out by iTRAQ labeling followed by 2DLC-ESI-MS/MS analysis. HRG expression was significantly lower in HCC tissues. HRG overexpression in Huh7 and MHCC-97H hepatoma cell lines led to decreased cell proliferation, colony-forming ability, and tumor growth, and increased cell apoptosis. HRG could inhibit cell proliferation via the FGF-Erk1/2 signaling pathway by reducing Erk1/2 phosphorylation. On the other hand, the functional expression of HRG was also dependent on the glycosylation status at its N-terminal, especially at the glycosylation site Asn 125. The glycosylation of HRG may play a key competitive role in the interaction between HRG and heparin sulfate for binding bFGF and activating the FGF receptor. These findings provide novel insights into the molecular mechanism of HRG in HCC.
    Preview · Article · Aug 2015 · Oncotarget
  • [Show abstract] [Hide abstract]
    ABSTRACT: Histone deacetylase (HDAC) can blockDNA replication and transcription and altered HDAC expressionwas associated with tumorigenesis. This study investigated the effects of HDAC inhibitors on hepatic oval cells and aimed to delineate the underlying molecularevents. Hepatic oval cells were treated with two different HDAC inhibitors, suberoylanilidehydroxamic acid (SAHA) and trichostatin-A (TSA). Cells were subjected tocell morphology, cell viability, cell cycle, and wound healing assays.The expression of proteins related to both apoptosis and the cell cycle, and proteins of the AKT/mammalian target of rapamycin (mTOR) signaling pathway were analyzed by Western blot. The data showed that HDAC inhibitors reduced oval cell viability and migration capability, and arrested oval cells at the G0/G1 and S phases of the cell cycle, in a dose- and time-dependent manner. HDAC inhibitors altered cell morphology and reduced oval cell viability, and downregulated the expression of PCNA, cyclinD1, c-Myc and Bmi1 proteins, while also suppressing AKT/mTOR and its downstream target activity. In conclusion, this study demonstrates that HDAC inhibitors affect oval cells by suppressing AKT/mTOR signaling.
    No preview · Article · Nov 2015 · Archives of Biochemistry and Biophysics