白细胞介素18在人白血病细胞系J6-1的表达及其意义

Expression of interleukin-18 in human leukemia cell line J6-1 and its significance

目的 阐明白细胞介素 18(IL 18)在人白血病细胞系J6 1的表达与调控 ,探讨IL 18在白血病发生中的意义。方法 应用逆转录浆合酶链反应检测IL 18mRNA的表达 ,酶切鉴定其特异性 ;构建IL 18的重组质粒 ,测序分析其cDNA序列的同源性 ;应用反义核酸技术 ,观察不同浓度的IL 18反义寡脱氧核苷酸 (ASODN)对J6 1细胞增殖的影响。结果 J6 1细胞组成性高表达IL 18mRNA ,测序结果表明 ,其cDNA序列与文献报道IL 18的同源性为 99% ,仅有一处同义点突变 (35UCA→UCC)。正常人外周血单个核细胞 (PBMC)IL 18mRNA的表达水平很低 (0 13± 0 0 5 ) ,CpG寡脱氧核苷酸 (CpG ODN)显著上调IL 18在PBMC的表达 (0 82± 0 2 4) ,但对J6 1细胞无类似上调作用。IL 18在J6 1细胞的表达水平 (0 98± 0 2 9)明显高于在K5 6 2、HL 6 0、U937和LCL细胞系的表达。IL 18ASODN能显著抑制J6 1细胞的增殖 (抑制率 43 3% ) ,而对HL 6 0的抑制作用不明显 (抑制率16 7% )。结论 J6 1细胞组成性高表达IL 18;IL 18基因的编码区在J6 1细胞不存在有意义的突变 ;IL 18可能通过自分泌途径正向调节J6 1细胞的增殖。

Objective To elucidate the expression and regulation of interleukin 18 (IL 18) in human leukemia cell line J6 1, and investigate its possible role in leukemogenesis. Methods IL 18 mRNA expression was detected using reverse transcriptase ploymerase chain reaction (RT PCR), and the specificity of IL 18 cDNA was verified by restriction endonucleases analysis. To sequence and analyze its cDNA alignment, a recombinant plasmid of IL 18 was constructed. Employing the antisense technique, we ionvestigated the effects of IL 18 antisense oligodeoxynucleiotide (ASODN) on the proliferation of J6 1 cells. Results J6 1 cells constitutively expresses IL 18 mRNA, and its cDNA sequence shared 99% homology with the published IL 18 sequence,except that a synonymous point mutation (Codon35 UCA to UCC) was confirmed by sequencing and alignment analysis. Peripheral blood mononuclear cells (PBMC) of healthy volunteers weakly expresses IL 18 mRNA (0.13±0 05), while CpG ODN significantly up modulated IL 18 expression in PBMC (0 82±0 24), but not in J6 1 cells. Compared with K562?HL 60?U937 and LCL cells, IL 18 mRNA level expression of in J6 1 cells was the highest(0 98±0 29). ASODN of IL 18 markedly inhibited the proliferation of J6 1 cells (inhibition rate 43.3%), but did not show significant effects on HL 60 cells(inhibition rate 16.7%). Conclusions J6 1 cells constitutively expresses IL 18 mRNA, and no significant mutation was detected coding region of IL 18 cDNA. IL 18 might be a positive regulation factor for the proliferation of J6 1 cells after in an autocrine model.