砷对20个基因表达水平影响的研究

Effect of sodium arsenite to the level of twenty genes expression

目的 :研究砷 (亚砷酸钠 )对 2 0个相关基因表达的影响。 方法 :依据已报道的砷中毒机理 ,选择与其相关的 2 0个基因 ,并设内对照和阴性对照 ,然后制备这些基因的寡核苷酸探针 (长度在 2 3～ 30 m er之间 ) ,并制备寡核苷酸芯片。选取人正常肝细胞 ( L- 0 2 )培养并用不同浓度的砷染毒 ,提取染毒细胞的总 RNA,纯化 m RNA,反转录获第一链 c DNA,并用 Cy3、Cy5生物标记 ,切断后同已制备好的寡核苷酸的基因探针杂交 ,最后读片 ,根据信号强度并进行一定计算来判定砷对每个基因表达水平的影响。 结果 :0 .5 μM和 2 .0 μM染砷组与对照组相比 ,C- myc癌基因、抗砷蛋白基因 ( ARS)和 CAAT结合转录因子基因 ( CTF)的表达水平均出现下调 ,染砷 2 .0 μM还可抑制肝细胞生长因子基因 ( HGF)表达 ,同时诱导 p5 3抑癌基因表达。 结论 :砷可抑制 CTF基因的表达 ,使基因转录的起始受阻 ,从而导致大部分基因表达水平下调 。

Objective: Study the level of 20 genes expression related with arsenic (sodium arsenite). Methods: First, 20 related genes were selected according to the mechanism of arsenism having reported and two control groups were set up. Then, the probs (23～30 mer) and arrays of oligonucleotide were made. Second, human hepatic cells (L 02) were cultured for two weeks exposing different arsenic concentrations, and total RNA was extracted, mRNA purificated, first strand cDNA synthesized, cDNA labelled using Cy\+5 and Cy\+3, cDNA broken using DNA enzyme, then cDNA and the probs hybridized and fluorescence signal scanned. Last, the values of Cy\+5: Cy\+3 were calculated to decide the level of those genes expression. Results: As compared with control, in 0.5 μM and 2.0 μM sodium arsenite groups, expression of C myc, ARS and CTF genes were inhibited, in 2.0 μM,expression of HGF gene was inhibited, but the expression of p53 gene was induced. Conclusion: Arsenic can inhibit the expression of CTF gene, starting of gene transcript was blocked, so it can cause the expression of many of genes to be declined. This result may be ones of the mechanisms of arsenical genetic toxicity.