中文摘要：

目的：用TGF—-β1shRNA稳定转染涎腺粘液表皮样癌Ms细胞，观察shRNA的基因沉默效应及其对细胞增殖的影响．方法：用Liperfetamine2000将TGF—β1shRNA转染Ms细胞．以G418（600mg／L）筛选21d，挑出单克隆获得稳定转染细胞系．RT—PCR法和免疫组化方法检测细胞中TGF—B1mRNA和蛋白的表达利用MTT、软琼脂克隆形成实验、流式细胞仪、透射电镜等检测转染前后Ms细胞的增殖和形态特点，以空载体转染和未转染细胞为对照：结果：获得稳定转染TGF—β1shRNA的细胞系：TGF—β1shRNA阻断了Ms细胞中TGF—β1mRNA和蛋白的表达，未转染组，转染空载体组和转染shRNA组．细胞倍增时间分别为393h，40．1h和45．3h，G1期细胞所占比例分别为54．2％，56．8％和61．7％，增殖指数P1分别为0．46，0．43和0，38；软琼脂克隆形成率分别为34．7％，33，3％和15．8％：超微结构观察见转染细胞细胞器扩张和肿胀，核浆比例变小，核仁减小，数目减少，核分裂像减少。结论：应用shRNA沉默TGF—β1基因可抑制Ms细胞的增殖。

英文摘要：

Objective： To study the effect of TGF-β1 gene silencing with shRNA on the proliferation of human salivary gland mucoepidermoid carcinoma Ms cells. Methods： Ms cells were transfected with TGF-β1 shRNA vector or the empty vector, and the stably transfected cells were selected after exposure to 600 mg/L of G418 for 21 days. The positive clones were identified by RT- PCR and immunohistochemical staining. MTT assay, soft agar clone formation assay, flow cytometry （FCM） and transmission electron microscopy （TEM） were used to observe cell proliferation and morphology. Results： TGF-β1 shRNA blocked the expression of TGF-β1 mRNA and protein in Ms cells. The population doubling time （h） of untransfected cells, empty vector transfected cells and shRNA vector transfected cells was 39.2, 40.1, and 45.3, while the percentage of cells in phase G1 was 54.2%, 56.8% and 61.7%, respectively. The proliferation index （PI） 0.46, 0.43 and 0.38, and the clonogenecity （%） 34.7, 33.3 and 15.8, respectively, for untransfected cells, empty vector transfected cells and shRNA vector transfected cells. TEM observation revealed that microvilli on the surface of shRNA vector transfected cells were less abundant, the endoplasm web was obviously expanded and the ratio of nucleus to cytoplasm was reduced. Conclusions： Transfection of TGF-β1 shRNA may silence the gene and inhibit cell proliferation.