| Background:Stomach cancer is the fourth most common cancer worldwide. It is a disease with a high death rate making it the second most common cause of cancer death worldwide after lung cancer. The mechanism of gastric cancer incidence,invasion and metastasis is still unclear. There is study reported that nuclear matrix protein SATB1 was abnormal expression in some tumor. RNA-interference-mediated knockdown of SATB1 in highly aggressive (MDA-MB-231) cancer cells reversed tumorigenesis by restoring breast-like acinar polarity and inhibited tumor growth and metastasis in vivo. Conversely, ectopic SATB1 expression in non-aggressive (SK-BR-3) cells led to gene expression patterns consistent with aggressive-tumour phenotypes, acquiring metastatic activity in vivo. SATB1 functions as a genome organizer during tumorigenesis to reprogramme expression and promote metastasis. Our studies have found that SATB1 was also abnormal expression in gastric cancer, and its expression was consistent with the expression of heparanase. We used transcription factor binding site analysis Software Mat-inspector and found that there are 8 sites in the upstream promoter region of heparanase may combine with SATB1 protein, indicating possible link between them.Objective:investigate the expression of SATB1 in gastric cancer and Gastric cancer cell lines. Regulate the expression of SATB1 to know the changes in the biological behavior and the heparanase expression of gastric cancer cell line. It may provide new idea for the diagnosis and treatment of gastric cancer.Method:Immunohistochemistry and RT-PCR detect the expression of SATB1 protein and mRNA in gastric cancer and adjacent tissues; RT-PCR and Western blot detect the expression of SATB1 mRNA and protein in MDA-MB-435S,AGS,SGC-7901 and GES-1 cell line. Construction the eukaryotic expression vector pcDNA3.0-SATB1 and DNA double restriction enzyme digestion identification of plasmid. After transient transfection of SGC-7901 cells 48 h, RT-PCR and Western blot detection the expression of SATB1 mRNA and protein; MTT assay detection the cell proliferation; Flow cytometry analysis of cell-cycle arrest and apoptosis; transwell chamber assay detection the cell invasion; RT-PCR and Western blot detection the expression of heparanase. Design and synthesis of SATB1 siRNA oligonucleotides, After transient transfection of AGS cells 48 h, RT-PCR and Western blot detection the expression of SATB1 mRNA and protein; MTT assay detection the cell proliferation; Flow cytometry analysis of cell-cycle arrest and apoptosis; transwell chamber assay detection the cell invasion; RT-PCR and Western blot detection the expression of heparanase.Results:Immunohistochemical analysis of 48 gastric cancer samples showed that 41 samples were positively expressed and the positive rate was 85.42%(41/48), however, the positive rate was only 4.17%(2/48) in adjacent tissues. RT-PCR showed in cancer samples and adjacent tissues the positive expression of SATB1 mRNA were 87.50%(42/48) and 25 %(3/48), respectively (p<0.01). The SATB1/β-actin mRNA rate in MDA-MB-435S, AGS, SGC-7901 and GES-1 cell lines were 0.437±0.021; 0.269±0.017; 0.019±0.006 and 0.011±0.003, respectively. Western Blot showed that the expression of SATB1 protein in AGS cell was higher than that in SGC-7901 cell (p<0.01). After transient transfection of SGC-7901 cells 48 h, RT-PCR showed that the SATB1/β-actin mRNA was 0.537±0.028 in pcDNA3.0-SATB1 transfection group and 0.098±0.007 in blank control group(p<0.05). Western Blot showed the SATB1/β-actin protein was 0.196±0.013 in pcDNA3.0-SATB1 transfection group and 0.067±0.008 in blank control group (p<0.05). MTT assay showed the proliferation ability of the SGC-7901 cell line in pcDNA3.0-SATB1 transfection group was enhanced (p<0.05).Flow cytometry showed that cycle distribution (G0/G1, S, G2/M) of blank control group, pcDNA3.0 transfection group and pcDNA3.0-SATB1 transfection group were G0/G1:65.27±5.38,78.36±4.64,52.28±4.13; S:24.52±1.68,10.25±1.13, 27.95±0.31, G2/M:10.21±1.13,11.39±0.96,19.77±1.02, respectively; the rate of G2/M in pcDNA3.0-SATB1 group increased (p<0.05). The apoptosis rates of three group were similar (p>0.05). Transwell chamber assay showed that the cell in blank control group, pcDNA3.0 transfection group and pcDNA3.0-SATBl transfection group were 47.26±3.39, 44.65±3.11 and 128.67±9.13, respectively. The cell number in pcDNA3.0-SATB1 transfection group was higher than those of other two group (p<0.05). RT-PCR and Western Blot showed that the expression of heparanase was unregulated in pcDNA3.0-SATB1 transfection group (p<0.05). aiRNA transfection could effectively silence gene expression. When AGS cell was transfected with SATB1-siRNA or SATB1-aiRNA, the expression of SATB1 was down-regulated in both group compared with control group (p<0.05), and SATB1-aiRNA could more effectively silence SATB1 mRNA expression (p<0.05). MTT assay showed the proliferation of the AGS cell line in SATBl-siRNA and SATB1-aiRNA transfection group was inhibited (p<0.05). Flow cytometry showed that percentage of G0/G1 was increased in SATBl-siRNA and SATBl-aiRNA transfection group (p<0.05), indicating of G0/G1 arrest. The apoptosis rate in control, SATB1-siRNA and SATBl-aiRNA transfection group was 2.69±0.09,6.37±0.42,10.12±0.61, respectively (p<0.05). Transwell chamber assay showed that the cells in control group, SATB1-siRNA and SATB1-aiRNA transfection group were 137.67±12.36,73.54±10.61 and 31.55±4.27, respectively (p<0.05). RT-PCR and Western Blot showed that the expression of heparanase were both down-regulated in SATB1-siRNA and SATB1-aiRNA transfection group (p<0.05).Conclusion:The SATB1 gene was high positive expression in gastric cancer, and the expression of SATB1 is higher in AGS cell line than in SGC-7901 cell line. Up-regulation the expression of SATB1 in SGC-7901 cell could induce the cell proliferation, accelerate cell cycle progression, enhance invasive potential, and increase the expression of heparanase; however, down-regulation the expression of SATB1 in AGS cell could inhibit the cell proliferation, induce G0/G1 cell cycle arrest and cell apoptosis, reduce invasive potential and decrease the expression of heparanase. SATB1 could be involved in the progression of gastric cancer. It may be a new important prognostic indicators and therapeutic target of gastric cancer.
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