干扰素诱导跨膜蛋白 1 对结直肠癌细胞 SW480 增殖、侵袭和转移的影响。

Influences of the interferon induced transmembrane protein 1 on the proliferation, invasion, and metastasis of the colorectal cancer SW480 cell lines.

机构信息

Department of Oncology, Huaian First People's Hospital Affiliated to Nanjing Medical University, Huaian, Jiangsu 223300, China.

出版信息

Chin Med J (Engl). 2012 Feb;125(3):517-22.

PMID:22490413

Abstract

BACKGROUND

Interferon-induced transmembrane protein 1 (IFITM1) has been identified as a molecular marker of the colorectal tumors; however its influences on the biological behaviors of the colorectal cancer cells are currently unknown. We aimed to study the influences of IFITM1 on the proliferation, invasion, and metastasis of the colorectal cancer SW480 cell lines.

METHODS

We constructed IFITM1/pEGFP-C3 recombinant plasmids and transfected them into the colorectal cancer SW480 cell lines. IFITM1/pEGFP-C3 recombinant plasmids were identified by means of immunofluorescence, laser confocal scanning microscopy, and reverse transcription polymerase chain reaction. IFITM1/SW480 cells with stable over-expression of IFITM1 were confirmed by G418 screening. The influences of IFITM1 on the proliferation of the SW480 cell lines were investigated by MTT assay and tumor transplantation experiments in nude mice. Cell invasion experiments were performed to determine the invasion capacity of the IFITM1/SW480 cells. Matrix metalloproteinase 2 (MMP-2) and MMP-9 activities were detected by the gelatin zymographic analysis, and MMP-9 expression by the Western blotting analysis.

RESULTS

IFITM1/pEGFP-C3 recombinant plasmids were successfully constructed in this study, and the IFITM1/SW480 cells with stable IFITM1 gene over-expression were confirmed by G418 screening. MTT results showed that the proliferation of the IFITM1/SW480 cells was significantly enhanced (P < 0.01). Tumors were harvested from four weeks old mice. Tumor volumes were (1347.00 ± 60.94) mm(3), (1032.40 ± 111.38) mm(3) and (1018.78 ± 28.83) mm(3); and tumor weights were (1522.34 ± 62.76) mg, (1137.78 ± 97.22) mg and (1155.76 ± 133.31) mg for mice inoculated with the IFITM1/SW480 cells, pEGFP-C3/SW480 cells and SW480 cells, respectively. Tumor volumes and weights from mice inoculated with the IFITM1/SW480 cells were significantly increased (P < 0.01). In addition, the numbers of the SW480 cells and IFITM1/SW480 cells that migrated through Matrigel were 448.64 ± 38.09 and 540.45 ± 44.61, respectively; so the invasive ability of the SW480 cells transfected with IFITM1 gene was significantly greater than that of the SW480 cells (P < 0.01). Gelatin zymographic analysis showed that MMP-9 and MMP-2 protein activities in the IFITM1/SW480 cells were significantly enhanced, and Western blotting analysis showed that MMP-9 expression in the IFITM1/SW480 cells was also increased.

CONCLUSION

IFITM1 can enhance the proliferation, invasion, and metastasis of the colorectal cancer SW480 cell lines.

摘要

背景

干扰素诱导跨膜蛋白 1（IFITM1）已被鉴定为结直肠肿瘤的分子标志物；然而，其对结直肠癌细胞的生物学行为的影响目前尚不清楚。我们旨在研究 IFITM1 对结直肠癌细胞 SW480 系增殖、侵袭和转移的影响。

方法

我们构建了 IFITM1/pEGFP-C3 重组质粒，并将其转染至结直肠癌细胞 SW480 系中。通过免疫荧光、激光共聚焦扫描显微镜和逆转录聚合酶链反应鉴定 IFITM1/pEGFP-C3 重组质粒。通过 G418 筛选鉴定 IFITM1 稳定过表达的 IFITM1/SW480 细胞。通过 MTT 检测和裸鼠肿瘤移植实验研究 IFITM1 对 SW480 细胞系增殖的影响。通过基质金属蛋白酶 2（MMP-2）和 MMP-9 活性的明胶酶谱分析和 MMP-9 表达的 Western 印迹分析检测 IFITM1/SW480 细胞的侵袭能力。

结果

本研究成功构建了 IFITM1/pEGFP-C3 重组质粒，并通过 G418 筛选证实了 IFITM1 基因稳定过表达的 IFITM1/SW480 细胞。MTT 结果表明，IFITM1/SW480 细胞的增殖明显增强（P<0.01）。四周龄小鼠采集肿瘤。肿瘤体积分别为（1347.00±60.94）mm3、（1032.40±111.38）mm3和（1018.78±28.83）mm3；肿瘤重量分别为（1522.34±62.76）mg、（1137.78±97.22）mg和（1155.76±133.31）mg，接种 IFITM1/SW480 细胞、pEGFP-C3/SW480 细胞和 SW480 细胞的小鼠。接种 IFITM1/SW480 细胞的小鼠的肿瘤体积和重量明显增加（P<0.01）。此外，穿过 Matrigel 的 SW480 细胞和 IFITM1/SW480 细胞的数量分别为 448.64±38.09 和 540.45±44.61，因此转染 IFITM1 基因的 SW480 细胞的侵袭能力明显强于 SW480 细胞（P<0.01）。明胶酶谱分析显示 IFITM1/SW480 细胞中 MMP-9 和 MMP-2 蛋白活性明显增强，Western 印迹分析显示 IFITM1/SW480 细胞中 MMP-9 表达也增加。