大麻素受体1介导人类中性粒细胞dHL60的迁移功能

Cannabinoid receptor 1 mediated the migration of human neutrophil-like cell line dHL60

目的研究大麻素受体(cannabinoid receptors,CBs)对人类中性粒细胞(d HL60)迁移的影响。方法体外培养人早幼粒白血病细胞系HL60,使用二甲基亚砜(dimethylsulphoxide,DMSO)诱导为类中性粒细胞(d HL60),运用实时荧光定量聚合酶链反应检测其分化标志物CD11b mRNA的表达;应用琼脂糖凝胶电泳、Western blotting法及免疫荧光技术检测其大麻素受体1(CB1)及受体2(CB2)的表达;ACEA、AM281分别为CB1的药理学激动剂和拮抗剂,JWH133、AM630分别为CB2的药理学激动剂和拮抗剂,应用Boyden chamber法检测ACEA和JWH133对d HL60迁移的影响,并从药理学阻断CB1、CB2后,检测其迁移功能的变化;使用鬼笔环肽染细胞肌动蛋白纤维,并应用高内涵扫描分析的方法对肌动蛋白纤维的聚合进行分析。结果 d HL60在mRNA和蛋白质水平上均表达CB1、CB2;ACEA能够诱导d HL60的迁移及其细胞骨架的聚合,且其所诱导的迁移能够被CB1的药理学阻断剂AM281所阻断,而CB2的药理学阻断剂AM630对ACEA所诱导的迁移并无影响;给予CB2的激动剂JWH133对d HL60的迁移及细胞骨架的聚合无明显作用。结论激活CB1能够促进d HL60的迁移。

Objective To investigate the effect of cannabinoid receptors （CBs） on migration of human neutrophil-like cell line dHL60. Methods HL60 cell was cultivated in vitro and stimulated by dimethylsulphoxide （DMSO） to differentiate into neutrophil-like cells （dHL60）, RT-PCR was used to detect the CD11b mRNA expression as a differentiation marker. Western blotting and agarose gel electrophoresis were used to detect the expression of CB1 and CB2. Boyden chamber was used for cell migration assay. Filamentous actin （F-actin） stained with phalloidin was examined by high content analysis. Results In this study, we present evidence that dHL60 significantly expressed CB1 and CB2; the treatment of ACEA （agonist of CB1） caused an increase in dHL60 migration and F-actin remodeling, the pharmacological inhibition of CB1 markedly attenuated ACEA-induced migration; the treatment of JWH133 （agonist of CB2） has no effect on migration and F-actin remodeling. Conclusion CB1 mediated the migration of human neutrophil-like cell line dHL60.