摘要
目的 GC蛋白在大部分恶性肿瘤中呈高表达,且对肿瘤的形成、发生及发展起着重要的作用,然而GC蛋白对结肠癌的形成进展影响并不清楚,本文探讨结肠癌中的GC蛋白表达及其对细胞生物学行为的影响。方法应用RT-PCR检测结肠癌细胞中GC蛋白的表达情况,选取GC蛋白表达量最高的SW1116细胞株为研究对象,分为干扰组(GC-siRNA组),阴性对照组(Negative control siRNA组),空白对照组(Blank组),运用siRNA干扰技术,沉默SW1116中GC蛋白的表达,分别对各组细胞运用MTT法检测细胞增殖的情况,运用Transwell试验检测细胞迁移能力的变化,运用Annexin.V.PI双染法流式细胞术检测细胞的凋亡状况,观察沉默GC基因对SW1116细胞生物特性的影响。结果 GC在大肠癌细胞中呈高表达,显著高于正常结肠细胞株CCD18Co,差异有统计学意义(SW480 vs.CCD18Co,P=0.003;SW620 vs.CCD18Co,P<0.01;SW116 vs.CCD18Co,P<0.01),其中SW1116细胞表达最高(RQ值4.3±3.5);沉默SW1116细胞GC基因后,相比两对照组,GC-siRNA组细胞活性率在转染后48 h和72 h显著下降,差异有统计学意义(24 h,P=0.034;48 h,P=0.009);Transwell实验结果显示GC-siRNA组迁移过纤维膜细胞数为(51.6±9.4)个,阴性对照组(182.1±16.1)个,空白对照组(174.3±22.8)个,迁移能力减低(P<0.01);Annexin V-FITC细胞凋亡实验结果显示干扰组凋亡率为(28.5±5.9)%,明显高于阴性对照组(5.6±2.1)%和空白对照组(6.1±2.3)%,细胞诱导凋亡增加(P<0.01);两对照组比较差异无统计学意义(P>0.05)。结论沉默结肠癌细胞中的GC蛋白表达后,细胞的增殖减慢,迁移能力减低,凋亡诱导增加,GC基因可能与结肠癌的发生与进展相关,或可作为结肠癌基因治疗的靶点。
Objective GC protein( group-specific component,i, e. Vitamin D-binding protein) is highly expressed in most malignant tumor, and plays an important role in the tumor formation, occurrence and development;but the influence of GC protein on the formation and development of colon cancer remains unclear. The aim of this paper is to investigate the expression of GC protein in colon cancer cells and its influence on cell and biological behavior. Methods RT-PCR was used to detect the expression of GC protein in colon cancer cell lines. SWl116 cell lines with the highest amount of GC protein expression was selected and divided into 3 groups, the interference group ( GC-siRNA group), negative control group( Negative control group siRNA), blank control group (Blank). siRNA interference technology was conducted to si- lence GC gene in SW1116 cells. The MTr assay was performed to assess cell viability ;the Transwell migration assay was used to altered migratory behavior; a double staining flow cytometric assay was employed to detect the apoptotic leuco- cytes. The effects of GC gene silence on biological characteristics of SW1116 cells were evaluated. Results GC expressed actively in colorectal cancer cells, that was significantly higher than in normal colon cell lines CCD18Co, the difference was statistically significant[ (SW480 vs. CCD18Co,P 〈 0.05 ; SW620 vs. CCD18Co, P 〈 0.01 ; SW116 vs. CCD18Co, P 〈 O. O1 ) ]. After GC gene silence in SWI 116 cells, the cell activity rate in GC-siRNA group decreased significantly in the 48 h and 72 h after transfection as compared with the negative control group and blank control group, the difference was statistically significant (24 h, P = 0. 034 ;48 h, P = O. 009). The Transwell migration assay showed a declined numbers of migrated cells in the three groups (51.6 ± 9.4 in GC-siRNA group, 182.1 ± 16. 1 in negative control group and 174.3 ± 22.8 blank control group), P 〈 0.01;Annexin V-FITC assay showed that the apoptosis rate in the interference group was (28.5±5.9 )% , which was significantly higher than the negative control group( 5.6 ± 2.1 )% and blank control group(6.1 ±2.3 )%, P 〈 0.01 ;there was no statistical significance difference between the two control group (P 〉 0.05). Conclusion To silence the expression of GC protein in colon cancer cells can slow cell proliferation,reduce the migration ability,increase apoptosis induction. GC gene may be associated with the occurrence and progress of colon cancer,it can be used as a colon cancer gene therapy targets.
出处
《中华全科医学》
2015年第3期393-395,497,F0003,共5页
Chinese Journal of General Practice
关键词
结肠癌
GC蛋白
SIRNA
Colan cancer
Vitamin D-binding protein
Small interfering RNA(siRNA)