摘要
目的:探讨全反式维甲酸(ATRA)诱导人急性早幼粒细胞白血病细胞株NB4向粒细胞分化过程中肽酰基精氨酸脱亚胺酶4(PADI4)的表达变化及其是否参与炎性因子的表达。方法:建立ATRA诱导的NB4细胞分化模型,采用Wright-Giemsa染色观察细胞形态学变化;采用流式细胞术检测细胞表面分化抗原CD11b的表达;RT-PCR方法检测PADI4基因转录水平的表达;Western bolt方法检测PADI4翻译水平的表达;ELISA检测培养液中肿瘤坏死因子(TNF)-α和白细胞介素(IL)-1β的表达;siRNA技术干扰PADI4表达后ELISA检测TNF-α和IL-1β的表达。结果:ATRA诱导后NB4细胞胞浆增多,核浆比例降低,核有凹陷,杆状和分叶现象增多。流式细胞术检测结果显示细胞表面分子CD11b表达明显升高(P <0.01)。在分化模型中,RT-PCR及Western blot检测结果显示PADI4转录水平和翻译水平表达均明显升高(P <0.05)。ELISA方法检测培养液中TNF-α和IL-1β的表达水平明显升高(P<0.05),而siRNA技术干扰PADI4表达后TNF-α和IL-1β的表达水平明显下降(P <0.05)。结论:ATRA诱导NB4细胞向粒细胞分化过程中PADI4的表达升高,同时PADI4参与了NB4细胞分化过程中炎性因子的表达。
Objective: To investigate the expression of peptidylarginine deiminase 4 PADI4) during the process of differentiation into granulocyte of NB4 cells induced by all-trans-retinoic acid( ATRA) and whether PADI4 is involved in the inflammatory cytokines expression. Methods: Granulocyte differentiation model of NB4 cells induced by ATRA was established. The cell morphology changes were observed by Wright-Giemsa staining. The expression of cell differentiation marker CD11 b was analyzed by flow cytometry. The mRNA and protein expression of PADI4 was detected by RT-PCR and Western blot,respectively. The expression of tumor necrosis factor( TNF) α and interleukin( IL) 1β was analyzed by ELISA,and also examined with the knockdown of PADI4 expression by siRNA. Results:After NB4 cells induced by ATRA,the cytoplasm increased and the ratio of nuclear to cytoplasmic was reduced. Nuclear dented,and rod-shaped nucleus,lobulated phenomenon increased( P < 0. 05). Flow cytometry analysis results showed that the cell surface molecule CD11 b expression increased( P < 0. 01). RT-PCR and Western blot showed the expression of PADI4 increased at both transcriptional and translational levels during the process of the differentiation. ELISA showed TNF-α and IL-1β secretion increased in differentiated macrophages,while they could be inhibited by PADI4-specific siRNA. Conclusion: During the differentiation into granulocyte of NB4 cells induced by ATRA,PADI4 expression increased. Furthermore,PADI4 appeared to play a critical role in inflammatory cytokines secretion.
作者
郭庆伟
李府
宋丽
王亚屏
杨晓梅
GUO Qing-Wei;LI Fu;SONG Li;WANG Ya-Ping;YANG Xiao-Mei(Department of Oncology&Hematology,Qilu Children's Hospital of Shandong University,Jinan 250022,Shandong Province,China)
出处
《中国实验血液学杂志》
CAS
CSCD
北大核心
2021年第4期1065-1070,共6页
Journal of Experimental Hematology
基金
山东省医药卫生科技发展计划项目(2016WS0107)
济南市科技创新发展计划(202019072)。
