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  • Title: [Construction of a eukaryotic vector expressing MASPIN gene and its effect on cell apoptosis in gastric cancer cell line SGC7901].
    Author: Zheng AH, Yi YF, Zhou WW.
    Journal: Ai Zheng; 2008 Nov; 27(11):1161-5. PubMed ID: 19000446.
    Abstract:
    BACKGROUND & OBJECTIVE: MASPIN gene is closely associated with carcinogengsis and plays a key role in cell proliferation, adhesion, migration and apoptosis. This study was to construct a recombinant eukaryotic vector expressing MASPIN, and explore the effect of MASPIN overexpression on the apoptosis in human gastric carcinoma cell line SGC7901. METHODS: A eukaryotic expression vector MASIPIN/PCR2.1 was constructed and transfected into SGC7901 cells. RT-PCR and Western blot were used to detect the expression changes of MASPIN and Bax/Bcl-2. TNF-related apoptosis inducing ligand (TRAIL) (50ng/ml) was used to induce apoptosis at different time courses. DNA apoptotic ladders were determined using agarose gel electrophoresis (AGE). Cell apoptosis was measured by flow cytometry (FCM). RESULTS: Recombinant plasmid MASIPIN/PCR2.1 was successfully constructed and transfected into SGC7901 cells. The mRNA and protein levels of MASPIN were significantly higher in the MASPIN/PCR2.1 group (33.6+/-1.2, 23.4+/-1.6) than in the PCR2.1(15.0+/-1.5, 12.3+/-1.5)and the untreated group (13.7+/-2.0, 12.0+/-1.3) (P<0.05). After transfection of MASIPIN/PCR2.1, DNA apoptotic ladders appeared in SGC7901 cells and the induction of apoptosis was in a time-dependent manner. The apoptosis rates were 8.0%, 16.3% and 25.8%in the MASPIN/PCR2.1 plus TRAIL group, 3.0%, 8.2%, 14.4% in the MASPIN/PCR2.1 group, and 4.1%, 9.8%,15.9% in the TRAIL group at 12, 24, and 48 h(P<0.05). The expression levels of Bax mRNA and protein at 48 h after MASIPIN/PCR2.1 transfection were significantly higher in MASPIN/PCR2.1 plus TRAIL group(55.3+/-2.1, 75.4+/-1.3) than in the PCR2.1 group (34.3+/-1.2, 40.7+/-1.8) and the TRAIL group (43.2 +/-1.8,36.2+/-1.3)(P<0.05). The expression of Bcl-2 mRNA in the MASPIN/PCR2.1 plus TRAIL group, PCR2.1 group and TRAIL group were 28.3+/-2.5, 34.3+/-1.2, 32.8+/-2.1, respectively (P<0.05), and those of Bcl-2 protein were 17.4+/-1.5, 45.1+/-2.1, 42.8+/-1.5 in the three groups, respectively (P<0.05). CONCLUSIONS: upregulation of MASPIN/PCR2.1 can significantly enhance the sensibility of gastric cancer cell line SGC7901 to the apoptosis inducer. This maybe related to the upregulation of Bax and downregulation of Bcl-2.
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