Effects of down-regulation of stormal interaction molecule 1 on cell cycle and apoptosis of pancreatic cancer cell line SW1990

Abstract

Objective To investigate effects of down-regulation of STIM1 on cell cycle and apoptosis of pancreatic cancer cell line SW1990 and explore its possible molecular mechanism. Methods Lentiviral vectors, which carrying siRNA of STIM1 gene, were contructed. Then lentiviral vectors were used to transfect the SW1990 cells and were divided into three groups:an untreated group, an empty vector group and an STIM1 group. The effects of down-regulation of STIM1 expression on cell proliferation, cell cycle and apoptosis were assessed by MTT assay and flow cytometry, respectively. Further expression changes of genes related to cell cycle and apoptosis were detected by semi-quantitative RT-PCR and Western blot. All values present the mean±SD.ttest was used between the two groups. Results The STIM1 expression of STIM1 group mRNA(0.261 ± 0.029), protein(0.120 ± 0.032)was down-regulated compared with the empty vector group[mRNA(1.002 ± 0.091) t= 20.74,P< 0.01],[protein(0.996 ± 0.053) t = 26.89,P< 0.01]in the SW1990 cells. Cell proliferation of SW1990 in the STIM1 group in 24 h, 48 h, 72 h(0.122 ± 0.008),(0.252 ± 0.031),(0.373±0.028)was significantly inhibited compared with the empty vector group[(0.223± 0.035) t= 6.48,P< 0.01],[(0.618±0.017) t= 16.90,P< 0.01],[(0.924 ± 0.140) t= 23.99,P < 0.01]. The results of flow cytometry revealed that the percentage of cells at G2/M phase in the STIM1 group was(41.47±0.66)﹪, significantly higher than that in the empty vector group[(10.30±2.24)﹪,t= 23.14,P< 0.01]. In addition, the percentage of apoptotic cells in the STIM1 group was(25.21±1.96)﹪, significantly higher than that in the empty vector group[(3.71 ±1.23)﹪,t= 16.03,P< 0.01]. Moreover, the results of semi-quantitative RT-PCR and Western blot showed that Cyclin B1 expression of the STIM1 group[mRNA(0.344±0.031)],[protein(0.776±0.042)]was down regulated compared with the empty vector group[mRNA(1.011±0.060)t= 40.06,P= 0.000][protein(1.034±0.036)t= 8.51,P< 0.01], P21 expression of STIM1 group[mRNA(1.970±0.107)],[protein(1.315±0.093)]was regulated compared with the empty vector group[mRNA(1.025 ±0.044)t= 17.10,P< 0.01],[protein(0.998±0.036)t = 9.52,P< 0.01], Bcl-2 expression of STIM1 group[mRNA(0.156±0.025)],[protein(0.381±0.028)]was down regulated compared with the empty vector group[mRNA(1.010±0.072)t= 15.46,P< 0.01],[protein(0.980±0.057) t= 14.63,P< 0.01]. Survivin expression of STIM1 group[mRNA(0.188±0.022)],[protein(0.022±0.019)]was down regulated compared with the empty vector group[mRNA(1.016±0.090)t= 58.08,P < 0.01],[protein(0.994±0.047)t= 442.58,P< 0.01]. Procaspase-3 expression of STIM1 group[protein(0.389±0.030)]was down regulated compared with the empty vector group[protein(1.008± 0.040)t= 19.22,P< 0.01]. Conclusion STIM1 may play a pivotal role in the proliferation and apoptosis of pancreatic cancer cell line SW1990, which could become a molecular target for therapy of pancreatic carcinoma. Key words: Pancreatic cancer; Stormal interaction molecule 1; Cell cycle; Cell apoptosis