摘要
目的 探讨反义寡核苷酸（ＡＳＯＤＮ） 对ｂｃｌ２ 基因的表达调控及诱导胃癌细胞凋亡的作用。方法 应用ＭＴＴ方法比较两条人工合成的ＡＳＯＤＮ 直接作用及其以脂质体（ＤＯＴＡＰ） 为载体对胃癌细胞的抑制效果。应用流式细胞术和ＲＴＰＣＲ 方法检测ＡＳＯＤＮ 对ｂｃｌ２ 蛋白和ｍＲＮＡ 表达量的调控。应用相差显微镜、电子显微镜、流式细胞术等方法，观察ｂｃｌ２ ＡＳＯＤＮ 诱导ＢＧＣ８２３ 胃癌细胞凋亡的效果。结果 ＭＴＴ实验显示两条ＡＳＯＤＮ 抑制胃癌细胞增殖的作用呈浓度和时间依赖性，且靶位点于ｍＲＮＡ蛋白编码区（ＡＳＯＤＮ２） 和以ＤＯＴＡＰ 为载体的ＡＳＯＤＮ 对胃癌细胞的增殖抑制效果为佳。ＡＳＯＤＮ作用于胃癌细胞，降低ｂｃｌ２ 蛋白和ｍＲＮＡ 表达。ｂｃｌ２ ＡＳＯＤＮ处理胃癌细胞，观察到肿瘤细胞缩小、凋亡小体出现、染色质浓缩、凋亡峰等凋亡特征性改变。结论 ｂｃｌ２ ＡＳＯＤＮ可降低ｂｃｌ２ ｍＲＮＡ和蛋白表达。
Objective To study the regulation of bcl 2 gene expression and induction of apoptosis by bcl 2 antisense oligodeoxynucleotides (AS ODN) on human gastric cancer cell line BGC 823 in vitro.Methods Two bcl 2 AS ODNs were synthesized, one covering the initiation sequence of translation of bcl 2 mRNA (AS ODN1) and the other covering the protein coding region (AS ODN2). BGC 823 cells in logarithmic phase of growth were cultured in the presence of free or liposome (DOTAP) encapsulated AS ODN. Cell growth was assessed by MTT method. The expression of bcl 2 at mRNA and protein levels was examined by RT PCR and flow cytometry, respectively. Electron microscopy and flow cytometry were used to demonstrate apoptotic changes in AS ODN treated cells.Results Both AS ODNs inhibited proliferation of BGC 823 cells. The inhibitory activity of AS ODN2 was stronger than that of AS ODN1. AS ODNs encapsulated in liposome led to more marked inhibition of cell growth than free AS ODNs. Both AS ODNs reduced bcl 2 expression of BGC 823 cells at mRNA and protein levels. Apoptosis of BGC 823 cells were demonstrated by the appearance of apoptotic bodies, chromatin condensation and pre G 1 peak on flow cytometric analysis.Conclusion Antisense oligodeoxynucleotide of bcl 2 decreases bcl 2 gene expression and induces apoptosis of human gastric cancer cells in vitro.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
1999年第4期253-255,共3页
Chinese Journal of Oncology
基金
辽宁省自然科学基金
