携Jagged 2基因siRNA慢病毒表达载体的构建及其转染对胰腺癌细胞影响

目的:构建携带Jagged 2(JAG2)基因siRNA的慢病毒表达载体,并观察其转染人胰腺癌细胞生物学特性的影响。方法:采用基因重组技术构建若干携JAG2基因siRNA片段的慢病毒表达载体,将其转染经酶解法从胰腺癌组织标本中原代分离的胰腺癌细胞;选择对JAG2基因抑制效率最高的siRNA片段,通过MTT法、流式细胞术、Transwell小室实验观察其转染对原代胰腺癌细胞生长、凋亡、细胞周期及侵袭转移能力的影响。结果:所构建的携JAG2基因siRNA片段的慢病毒表达载体均能不同程度降低原代胰腺癌细胞JAG2 mRNA与蛋白的表达。JAG2 siRNA转染后,胰腺癌细胞的增殖明显降低、凋亡与S期阻滞明显增强、侵袭转移能力明显减弱(均P0.05)。结论:成功构建携JAG2基因siRNA慢病毒表达载体,其转染能有效削弱人胰腺癌细胞的恶性生物学特征。

Construction of lentiviral vector bearing siRNA targeting Jagged 2 gene and effects of its transfection on pancreatic cancer cells

Objective: To construct lentiviral vector carrying siRNA targeting Jagged 2 (JAG2) gene and observe the effects of its transfection on biological characteristics in human pancreatic cancer cells. Methods: Several lentiviral expression vectors carrying different siRNA fragments targeting JAG2 gene were constructed using gene recombination technology, and were transfected into pancreatic cancer cells primarily isolated from the human pancreatic cancer tissue specimens through enzymolysis approach. Then, the siRNA fragment showing strongest inhibitory effect on JAG2 gene was selected for testing the influences of its transfection on proliferation, apoptosis, cell cycle and invasion and metastasis abilities in human pancreatic cancer cells by MTT assay, flow cytometry and transwell chamber assay, respectively. Results: All the lentiviral expression vectors carrying different siRNA fragments showed inhibitory effects, to different extents, on mRNA and protein expressions of JAG2. In pancreatic cancer cells after transfection with the JAG2 siRNA, the proliferation was decreased, apoptosis and S-phase arrest were enhanced, and invasion and metastasis abilities were reduced significantly (all P<0.05). Conclusion: lentiviral vector bearing JAG2 siRNA are successfully constructed and their transfection can effectively moderate the malignant biological characteristics of human pancreatic cancer cells.