佛波酯对人单核白血病细胞THP-1中颗粒溶素表达的激活作用

Activation of expression of granulysin in human monocytic cell line THP-1 by phorbol 12-myristate 13-acetatefor

目的探讨佛波酯(phorbol 12-myristate 13-acetatefor,PMA)对人单核白血病细胞THP-1中颗粒溶素(granulysin,GLS)表达的激活作用。方法分别用不同浓度的PMA(130、160、180 nmol/L)诱导THP-1细胞24 h,RT-PCR法检测细胞中GLS基因mRNA的转录,筛选PMA最佳诱导浓度;用PMA最佳诱导浓度分别诱导THP-1细胞12、24和48 h,RT-PCR法检测GLS基因mRNA的转录,筛选最佳诱导时间;用PMA最佳诱导浓度分别诱导THP-1细胞12、24、48和72 h,Western blot法检测GLS蛋白的表达,免疫细胞化学法检测48 h时GLS蛋白的表达。结果以160 nmol/L的PMA诱导THP-1细胞24 h,细胞中GLS基因mRNA的转录水平最高;诱导48 h后可检测到GLS蛋白大量表达。结论 PMA可激活THP-1细胞中GLS的表达,本实验为后续进一步研究GLS表达的调控机制奠定了基础。

Objective To investigate the activation effect of phorbol 12-myristate 13-acetatefor（PMA） on expression of granulysin（GLS）in human monocytic cell line THP-1. Methods THP-1 cells were induced by PMA at various concentrations（130,160 and 180 nmol / L）for 24 h,and determined for transcription level of GLS mRNA by RT-PCR,based on which the PMA concentration for induction was optimized. THP-1 cells were induced with PMA at the optimal concentration for 12,24 and 48 h respectively and determined for transcription level of GLS mRNA by RT-PCR,based on which the time for induction was optimized. THP-1 cells were induced by PMA at the optimal concentration for 12, 24, 48 and 72 h,and determined for expression of GLS protein by Western blot. Meanwhile,the expression of GLS protein 48 h after induction was also determined by immunocytochemical assay. Results The transcription level of GLS mRNA reached the maximum in THP-1 cells 42 h after induction with 160 nmol / L PMA. However,GLS protein was expressed in a large quantity 48 h after induction. Conclusion PMA activated the expression of GLS in THP-1 cells,which laid a foundation of further study on regulatory mechanism of GLS expression.