【摘要】 目的　了解三氧化二砷 (As2 O3 )诱导人T淋巴细胞的差异表达基因 ,探讨砷的免疫毒性和免疫抑制的机制。方法　在体外用As2 O3 处理人JurkatT淋巴细胞 (5 μmol/L ,2 4h)后 ,应用抑制性消减杂交技术 (SSH)和克隆技术构建差异表达的cDNA文库 ,用聚合酶链反应技术和测序技术鉴定阳性克隆。结果　用SSH技术成功地构建了由As2 O3 诱导的JurkatT淋巴细胞的差异表达基因的正向消减cDNA文库。经测序分析该文库含有 30个基因片段 ,其中仅有 1个是重复基因片段。与基因库的序列比较显示 ,这些基因序列的同源性在 95 %～ 10 0 %。该文库的基因有 :氧化代谢基因 (包括磷酸丙糖脱氢酶、酰胺腺嘌呤二核苷酸 4亚单位、焦磷酸合成酶、16S核糖体、琥珀酰CoA合成酶和ATP合成酶 6 ) ,转录和翻译基因 (包括多聚腺苷结合蛋白、泛醌素特异性蛋白酶 14、核糖体蛋白L2 3、核糖体蛋白S15a、真核翻译起始因子 3、Rab5相互作用蛋白、剪切因子和ADP 核糖体因子 6样相互作用蛋白 ) ,与氧化应激相关的基因 (铁蛋白重链和高泳动类蛋白 2 ) ,与细胞分化或凋亡相关基因 (髓细胞分化初期应答蛋白和凋亡相关蛋白 ) ,参与蛋白活化或信号传导的基因 (酪氨酸激酶、丝氨酸激酶和磷脂酰 4磷接头蛋白 1相关蛋白 ) ;5个功能不详的基因是KIAA0 0 92更多还原

【Abstract】 Objective To understand the differentially expressed genes in human T lymphocytes induced by arsenic trioxide (As 2O 3) and to explore mechanism of its immunotoxicity and immune suppression. Methods Human Jurkat T cell line was treated by arsenic trioxide ( 5 μmol/L, 24 h) in vitro, as a sample model.Then, the differentially expressed genes were cloned and the subtractive cDNA library from Jurkat T cell line was constructed by suppression subtractive hybridization (SSH).Polymerase chain reaction (PCR) and sequencing techniques were applied to identify positive clones. Results The forward subtracted cDNA library contained differentially expressed genes from Jurkat T cell line induced by arsenic trioxide was constructed, including 29 different gene fragments and only replicated one in the subtracted cDNA library identified by PCR and sequencing analysis.These gene sequences were 95%—100% analogous to the genes in public database (GenBank/EMBL).The cDNA library contained oxidative metabolic genes in mitochondria(triose phosphate dehydrogenase, NADH4, pyrophosphate synthase, 16S rRNA ribosome, succinate-CoA ligase and ATP synthase 6); transcriptional and tranlation genes poly (A) binding protein, t-RNA-guanine transglycoslase, ribosomal protein L23, ribosomal protein S15A, eukaryotic translation initiation factor 3, Rab interaction protein 5, splicing factor-arginine serine rich 5, and ADP-ribosylation factor-like 6 interacting protein), oxide stress related genes (ferritin high chain and high-mobility group protein 2); protein activating and signaling pathway related genes (casein kinase, serine kinase 2 and phosphatidylinositol-four-phosphate adaptor protein-1-associated protein); cell diffenrentiation and apoptosis associated genes (NB4 cell apoptosis related protein and myeloid differentiation primary response protein) and five genes with unknown function (KIAA0092, CGI-147protein, GCI-35, nucleolar phosphoprotein Nopp34 and Mus muscular partial mRNA for hypothetical protein), as well as a novel gene unmatched to the sequence in GenBank. Conclusions The forward subtracted cDNA library contained differentially expressed genes from Jurkat T cell line induced by arsenic trioxide was successfully constructed.And, genes not involved in previous research on arsenic were found.Results of analysis for these genetic function suggested that there should be many genes involved in process of T lymphocytes apoptosis or injury induced by arsenic trioxide and that there should still be many genes associated with arsenic that were not reported in the past.更多还原