摘要
目的探讨载体介导的转化生长因子(TGF)-β1小干扰RNA(siRNA)对人胃癌细胞株SGC-7901生物学行为的影响。方法构建TGF-β1特异性siRNA真核表达载体。转染胃癌细胞72 h后,以半定量逆转录一聚合酶链反应(RT-PCR)、Western blot检测细胞中TGF-β1的mRNA和蛋白表达的变化；以噻唑蓝(MTT)比色法、流式细胞术、细胞附壁试验和Miliicell小室测定细胞增殖、周期、体外黏附和侵袭力,并观察裸鼠体内成瘤性。结果与未处理组比较,TGF-β1-siRNA载体能明显下调TGF-β1 mRNA[(0．910±0．066)与(0．617±0．025)比较,P＜0．01]和蛋白[(120．00±8．72)与(41．00±5．57)比较,P＜0．01]。在胃癌细胞中的表达；细胞抑制率达30．18%,体外黏附、侵袭力下降,裸鼠体内成瘤性受到抑制。结论载体介导的TGF-β1 siRNA可显著下调TGF-β1在胃癌细胞中的表达,在一定程度上抑制其恶性生物学行为。
Objective To investigate the effects of transforming growth factor （TGF）β1 on biological behaviors of human gastric cancer cell line SGC-7901. Methods A vector used to transcribe functional short interfering RNA （siRNA） targeting TGF-β1 was constructed and was transiently transfected into human gastric cancer cell line SGC-7901. Seventy-two h after transfection, changes in expression of TGF-β1 rnRNA and protein were determined by semi-quantitative RT-PCR and Western blot. The effect of growth suppression and cell cycle was observed by MTT assay and flow cytometry. The adhesion and invasion to ECM were measured by cell attachment assay and Millicell chamber method, and nude mice inoculation test was carried. Results Electrophoresis and DNA sequencing confirmed that the TGF-β1- specific siRNA was synthesized and cloned into the expression vector pcPUR βcassette successfully. Semiquantitative RT-PCR and Western blot revealed that the expression of TGF-β1 mRNA （0.910 ± 0.066 vs 0.617±0.025, P〈0.01） and protein （120.00 ±8.72 vs 41.00 ± 5.57, P〈0.01） in human gastric cancer cell line SGC-7901 could be remarkably decreased at 72 h after transfection of TGF-β1-siRNA in comparison with the untreated group, respectively. TGF-β1-siRNA had great inhibitory effect on the growth of SGC-7901 cell with the inhibition ratio （IR） being 30.18%. Adhesion and invasion assay at the same time presented a decrease. The tumorigenic rate was also reduced in mice. Conclusion Vectorbased TGF-β1- siRNA could down-regnlate significantly TGF-β1 expression in human gastric cancer cell line SGC-7901 ahd suppress its malignant phenotype to some extent.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2006年第6期651-653,共3页
Chinese Journal of Experimental Surgery
基金
国家重点基础研究发展规划973项目(G1998051203)
辽宁省自然科学基金(2001225002-3)
