过表达IL-18抑制人结直肠癌细胞HCT-116的增殖

IL-18 over-expression inhibits proliferation of human colorectal cancer HCT-116 cells

目的:研究过表达IL-18基因对人结直肠癌(colorectal cancer,CRC)HCT-116细胞体内外增殖的影响及其可能的机制。方法:构建含人IL-18基因片段的载体,采用慢病毒转染法获得稳定过表达人IL-18的CRC HCT-116细胞株HCT-116/IL-18,CCK-8法检测HCT-116/IL-18细胞与野生型HCT-116细胞的增殖,Western blotting检测两组细胞内IL-18、Cyclin D1、增殖细胞核抗原(PCNA)、DNA损伤修复酶(PARP)蛋白的表达。将HCT-116、HCT-116/IL-18细胞分别接种于裸鼠左右两侧腋下,观察成瘤性及移植瘤的生长情况,免疫组化法检测移植瘤组织中IL-18及PCNA的表达。结果:IL-18基因在HCT-116细胞内过表达,可延缓HCT-116的增殖(P<0.05或P<0.01);与HCT-116细胞相比,HCT-116/IL-18细胞内PARP表达明显增强,PCNA、Cyclin D1表达减弱(P<0.01)。HCT-116/IL-18细胞系在裸鼠体内成瘤率明显降低,成瘤率为43%,与对照组比较其移植瘤成瘤时间晚、生长慢、肿块小,且HCT-116/IL-18异种移植瘤PCNA蛋白表达下调(P<0.01)。结论:过表达IL-18基因对HCT-116细胞生长增殖具有抑制作用,其机制可能与IL-18调控细胞周期和促进DNA损伤有关。

Objective： To investigate the effects of interleukin-18 over-expression on the in vitro and in vivo proliferation of human colorectal cancer（CRC） HCT-116 cells. Methods： A recombinant lentivirus vector containing human IL-18 gene fragment was constructed. Then the CRC HCT-116 cell line stably expressing human IL-18（HCT-116/IL-18） was obtained by recombinant lentivirus transfection. In vitro proliferation of HCT-116/IL-18 cells and wild-type HCT-116 cells was determined by CCK-8 method. The expressions of IL-18, Cyclin D1, proliferating cell nuclear antigen（PCNA） and DNA damage repair enzyme（PARP） were detected by Western blotting. HCT-116 and HCT-116/IL-18 cells were inoculated into left and right axillas of Balb/c nude mice, respectively. Then the tumorigenicity and the growth of transplanted tumor were observed. The expressions of IL-18 and PCNA in xenograft tissues were detected by immunohistochemistry analysis. Results： IL-18 gene over-expression in HCT-116 cells could delay the proliferation of HCT-116 cells（P〈0.05 or P〈0.01）. PARP expression was increased significantly and PCNA, Cyclin D1 expression were decreased in HCT-116/IL-18 cells as compared to that of HCT-116 cells（P〈0.01）.The tumorigenicity of HCT-116/IL-18 cell was significantly decreased in nude mice with a tumor-formation rate of 43%; Compared with control group, HCT-116/IL-18 cell line had a longer tumorigensis time,slower growth and smaller tumor volume; moreover, PCNA protein expression was down-regulated in HCT-116/IL-18 xenograft tissuesas shown by immunohistochemistry analysis（P〈0.01）. Conclusion： IL-18 over-expression inhibited the growth and proliferation of HCT-116 cells both in vitro andin vivo, and the mechanism might be related with IL-18 regulating cell cycle and promoting DNA damage.