Abstract
BackgroundThe prognosis of patients with hepatocellular carcinoma (HCC) still remains very dismal, which is mainly due to metastasis. In our previous studies, we found that chromosome 8p deletions might contribute to metastasis of HCC. In this study, we aimed to identify the candidate metastatic suppressor gene on chromosome 8p.MethodsOligo-nucleotide microarrays which included 322 genes on human chromosome 8p were constructed to analyze the difference in gene expression profiles between HCC tissues with and without metastasis. The leading differentially expressed genes were identified and selected for further analysis by real-time PCR and Western blotting. Recombinant expression plasmid vectors for each target gene were constructed and transfected into HCC cells and its in vitro effects on proliferation and invasion of HCC cells were also investigated.ResultsSixteen leading differentially expressed genes were identified from the HCC tissues with metastasis compared with those without metastasis (p < 0.01, q < 16 %). Among of the 10 significantly down-regulated genes in HCC with metastasis, methionine sulfoxide reductase A (MSRA) had the lowest p value and false discovery rate (FDR), and was considered as a potential candidate for metastasis suppressor gene. Real-time PCR and Western blotting confirmed that the mRNA and protein expression levels of MSRA were significantly decreased in HCC with metastasis compared with those without metastasis (p < 0.001), and MSRA mRNA level in HCCLM6 cells (with high metastatic potential) was also much lower than that of other HCC cell lines. Transfection of a recombinant expression plasmid vector and overexpression of MSRA gene could obviously inhibit cell colony formation (4.33 ± 2.92 vs. 9.17 ± 3.38, p = 0.008) and invasion (7.40 ± 1.67 vs. 17.20 ± 2.59, p= 0.0001) of HCCLM6 cell line.ConclusionMSRA gene on chromosome 8p might possess metastasis suppressor activity in HCC.
Highlights
The prognosis of patients with hepatocellular carcinoma (HCC) still remains very dismal, which is mainly due to metastasis
Of the 322 genes on the microarrays, 16 genes were found differentially expressed at a level of p < 0.01 between metastatic HCC and non-metastatic HCC, and the corresponding estimated false discovery rate (FDR) for q value thresholds was 16 % (Table 1)
RT-PCR (A, upper) and Western blotting (A, low) showed that the expression level was significantly higher in HCCLM6 transfected with pIRES2-EGFP-methionine sulfoxide reductase A (MSRA) (M) compared with the control that transfected with pIRES2-EGFP (P) and HCCLM6 cell line (LM6)
Summary
The prognosis of patients with hepatocellular carcinoma (HCC) still remains very dismal, which is mainly due to metastasis. Many advances have been made as a result of HCC clinical studies during the past decades that include early detection, surgical resection and liver transplantation, the general prognosis of the patients with HCC still remains dismal [3]. This outcome has been attributed to the high possibility of intra-hepatic metastases and recurrence after curative treatment [3,4,5,6]. The traditional metastasis paradigm has been challenged by the observations that most of the genetic and epigenetic changes necessary for metastasis appear to be the hallmarks of cancer [8,9,10]
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