Effect Of Grp78 On Invasion And Metastasis Of Human Gastric Carcinoma SGC7901 Cells In Vitro And Theirs Mechanism

Objective: To study the effect of BAPTA-AM and A23187 on GRP78 expression of gastric carcinoma SGC-7901 cells in vitro, examine the role of GRP78 in invasion and metastasis of these tumor cells, and investigate their mechanism initially.Methods: The SGC7901 cells were cultured in medium in vitro and divided into three groups: BAPTA-AM-treated group, control group and A23187-treated group. RT-PCR and Western blot were used to assess the expression of GRP78 at both mRNA and protein levels. The invasion and metastasis levels of SGC7901 cells were evaluated by matrigel invasion assay and a scratch wound assay. Immuno-cytochemistry and Western blot were performed to test the expression of E-cadherin and N-cadherin.Results:1. The ratios of gray values in A23187 group, control group, 20μmol/LBAPTA-AM group, 40μmol/LBAPTA-AM group, 60μmol/LBA -PTA-AM group were as follow: 1.070±0.005,1.011±0.012,0.701±0.009, 0.543±0.009,0.348±0.013, the expression of GRP78-mRNA was elevated in A23187 group and reduced in BAPTA-AM groups compared with that in control group(P<0.01); The ratios of GRP78 protein gray values andβ-actin gray values in A23187 group, control group, 40μmol/L BAPTA- -AM group were as follow: 0.911±0.004,0.823±0.007,0.647±0.015, the expression of GRP78 protein was elevated in A23187 group and reduced in BAPTA-AM groups compared with that in control group(P<0.01).2. In A23187 group, the degree of healing was higher than that in control group(79.68%±0.91% vs 57.32%±0.80%,P<0.01) and the mean value of the invasive cells in one filed was more than that in control group(89±2 vs 59±2,P<0.01). In BAPTA-AM group, the degree of healing was lower than that in control group (39.92%±0.79% vs 57.32%±0.80%, P<0.01)and the mean value of the invasive cells in one filed was less than that in control group(43±1 vs 59±2,P<0.01). The invasion and migration levels were increased in A23187 group and decreased in BAPTA-AM group compared with that in control group.3. Immuno-cytochemistry, Western blot showed the expression level of E-cadherin protein was decreased in A23187 group(0.106±0.003 vs 0.122±0.003,P<0.01; 0.495±0.010 vs 0.709±0.008,P<0.01)and increased in BAPTA-AM group compared with that in control group(0.133±0.003 vs 0.122±0.003,P<0.01; 0.915±0.017 vs 0.709±0.008,P<0.01); The expression level of N-cadherin protein was elevated in A23187 group(0.143±0.003 vs 0.131±0.003,P<0.01;0.541±0.009 vs 0.344±0.006,P<0.01)and reduced in BAPTA-AM group compared with that in control group(0.104±0.004 vs 0.131±0.003,P<0.01;0.210±0.004 vs 0.344±0.006,P<0.01).Conclusion: BAPTA-AM was an inhibitor of GRP78 expression in gastric carcinoma SGC7901 cells, and A23187 could induce expression of GRP78. Moreover, the regulation of GRP78 could affect the invasion and metastasis levels of tumor cells in vitro and it may be resulted from the change of N-cadherin expression and E-cadherin expression, which play an important role in EMT.