目的研究异硫氰酸苯己酯(PHI)对骨髓瘤u266细胞株P16基因的去甲基化作用并探讨其作用机制。方法应用半定量逆转录-聚合酶链反应检测u266细胞经过PHI处理后dnA甲基转移酶dnMT1、dnMT3A和dnMT3b基因的MrnA的表达变化,用甲基化特异性聚合酶链反应检测PHI作用前后u266细胞株P16基因甲基化状态的变化。结果使用不同浓度的PHI处理u266细胞72H后,u266细胞表达dnMT1、dnMT3A、dnMT3bMrnA的水平明显降低并呈浓度依赖性。以不同浓度的PHI处理u266细胞10d后,P16基因的甲基化状态被逐渐逆转。结论 PHI可能通过抑制dnA甲基转移酶的表达水平诱导P16基因产生去甲基化,使失活的抑癌基因重新激活,诱导瘤细胞凋亡。Objective To study the effect of demethylation of P16 and discuss the mechanisms by phenylhexyl isothiocyanate(PHI) on Myeloma U266 cells.Methods The mRNA expression of DNA methyltransferase 1,DNA methyltransferase 3a and DNA methyltransferase 3b were measured by RT-PCR,Modified methylation specific PCR(MSP) was used to screen p16-M and p15-U mRNA on U266 cells treated with PHI.Results After 72 hours treatment with different concentration PHI,the mRNA expression of DNMT1,DNMT3a and DNMT3b was decreased and related to PHI concentration,After 10 days PHI treatment,P16 gene hypermethylation was reversed at 10 μmol/L concentration.Conclusion By decreasing the level of DNA methyltransferase,PHI can reverse hypermethylation of gene P16 and re-actived silenced P16 gene and induce myeloma cells apoptosis in U266 cells.厦门市科技计划项目(3502Z20104034
