重组腺病毒介导的野生型p53对人胰腺癌细胞凋亡的诱导作用

目的研究恢复外源性野生型（ｗｔｐ）ｐ５３的表达对人胰腺癌细胞生长和凋亡的作用。方法构建了一个复制缺陷型５型腺病毒ｗｔｐ５３表达载体Ａｄ５ＣＭＶｗｔｐ５３，含有人ＣＭＶ启动子，人野生型ｐ５３和ＳＶ４０ｐｏｌｙＡ信号，经腺病毒包装细胞２９３细胞包装、扩增后，转染胰腺癌ＰＣ２细胞。ＰＣＲ和免疫沉淀技术证实转染的细胞有外源ｗｔｐ５３基因的存在和表达。结果转染的ＰＣ２细胞的生长率和３Ｈ掺入率降低。原位凋亡检测、流式细胞术和ＤＮＡ凝胶电泳显示转染细胞凋亡明显增多。而ＰＣ２细胞和用Ａｄ５ｐＸＪ转染的细胞没有这些改变。结论复制缺陷型腺病毒是一种安全高效的基因转移载体，恢复ｗｔｐ５３的表达可以有效地诱导凋亡和抑制胰腺癌细胞生长

Apoptosis induced by adenovirus-mediated wild-type p53 expression in human pancreatic cancer cells

OBJECTIVE: To investigate the biological effects of wtp53 expression on the growth rate and apoptosis of human pancreatic carcinoma. METHODS: The Ad5CMVwtp53 a recombinant replication-deficient adenoviral vector containing a human CMV promoter, a human wild-type p53 and the SV40 polyadenylation signal was amplified in 293 packaging cells. The pancreatic carcinoma cell line (PC-2), carrying a mutation of p53 gene at codon 240, was transfected using Ad5CMV wtp53 and the control adenoviral vector Ad5pXJ. RESULTS: The cells transfected with Ad5CMV wtp53 showed that presence and expression of wtp53 gene were demonstrated using PCR and immunoprecipitation, that growth rate and 3H-TdR incorporation rate were decreased. The restoration of wtp53 encoded protein in PC-2 cells induced apoptosis assessed by in situ TUNEL apoptosis, flow cytometry, DNA agarose gel electrophoresis analyses, whereas noninfected cells or the cells infected with control virus didn't show these changes. CONCLUSION: Replication-deficient adenoviral vector is an efficient vector in transferring wtp53 gene and antitumor therapy using the p53 gene is a valuable method in inhibiting pancreatic cancer cells growth by inducing apoptosis.