论文部分内容阅读
目的检测人硫酸酯酶1在胰腺癌细胞株中的表达水平,及转染后对胰腺癌细胞膜的结构及生长增殖能力的影响。方法应用逆转录-聚合酶链反应(RT-PCR)检测7株胰腺株硫酸酯酶1 mRNA的表达水平。采用脂质体Lipofectamine方法对Panc-1细胞进行硫酸酯酶1基因转染。Western blot检测转染前后Panc-1细胞硫酸酯酶1蛋白表达。共聚焦显微镜检测细胞膜表面硫酸肝素蛋白多糖(HSPG)结构的变化,应用噻唑蓝(MTT)比色分析法测定胰腺癌细胞的生长增殖能力,并检测转染前后的胰腺癌细胞对3种化疗药物(5-氟尿嘧啶、吉西他滨及放线菌素D)敏感性的影响。结果7株人胰腺癌细胞株中3株硫酸酯酶1 mRNA呈高表达,其余4株低表达或无表达。硫酸酯酶1稳定转染后,Panc-1细胞稳定表达硫酸酯酶1蛋白成分。转染后细胞膜表面HSPG结构明显改变,并且硫酸酯酶1表达细胞增殖速度减慢,转染组细胞倍增时间为(68.2±4.3)h,明显慢于对照组(50.3±3.2)h,P<0.05。转染后的胰腺癌细胞对化疗药物的敏感性并无明显改变。结论人硫酸酯酶1的表达通过影响细胞膜表面HSPG的结构,影响胰腺癌细胞的生长增殖能力,可能作为胰腺癌基因治疗的靶点。
Objective To detect the expression of human sulfatase 1 in pancreatic cancer cell lines and the effect of transfection on the cell membrane structure and growth and proliferation of pancreatic cancer cells. Methods The expression of sulfatase-1 mRNA in 7 pancreas strains was detected by reverse transcription-polymerase chain reaction (RT-PCR). Sulfatase 1 gene was transfected into Panc-1 cells by Lipofectamine method. Western blot was used to detect the expression of sulfatase-1 protein in Panc-1 cells before and after transfection. Confocal microscopy was used to detect the changes of the structure of heparin sulfate proteoglycan (HSPG) on the surface of the cell membrane. The growth and proliferation of pancreatic cancer cells were determined by MTT colorimetric assay. The effects of three chemotherapeutic drugs (5-fluorouracil, gemcitabine and actinomycin D) sensitivity. Results Three of the seven human pancreatic cancer cell lines showed high expression of sulfatase-1 mRNA, and the remaining four were either low or no expression. After stable transfection of Sulfatase 1, the Panc-1 cells stably expressed the sulfatase 1 protein component. After transfection, the structure of HSPG on the surface of cell membrane changed obviously, and the proliferation rate of S-1 expression cells slowed down. The doubling time of transfection group was (68.2 ± 4.3) h, which was significantly slower than that of control group (50.3 ± 3.2) h, P < 0.05. Transfection of pancreatic cancer cells on the sensitivity of chemotherapeutic drugs did not change significantly. Conclusions The expression of human sulfatase 1 may play a role in the gene therapy of pancreatic cancer by affecting the structure of HSPG on the cell surface and affecting the growth and proliferation of pancreatic cancer cells.