激活大麻素受体1诱导的单核巨噬细胞J774A.1的迁移依赖RNA结合蛋白HuR

发布时间：2018-05-22 13:51

本文选题：人抗原R + 大麻素受体　；参考：《首都医科大学学报》2016年01期

【摘要】：目的研究大麻素受体1(cannabinoid receptor 1,CB1)在单核巨噬细胞迁移中的重要作用以及RNA结合蛋白人抗原R(human antigen R,HuR)参与其中的可能机制。方法选用单核巨噬细胞系J774A.1,应用琼脂糖凝胶电泳和免疫荧光染色技术鉴定J774A.1中CB1以及HuR的表达;ACEA和AM281分别为CB1的药理学激动剂和拮抗剂,应用Boyden chamber法检测ACEA和AM281对J774A.1迁移活性的影响。HuR的基因干扰用于确定激活CB1诱导的J774A.1迁移功能是否依赖HuR;胞质蛋白的分离用于探究激活CB1是否能引起J774A.1胞质中HuR的富集;RT-qPCR和Western blotting法检测CB1和HuR mRNA和蛋白质的变化情况。结果该研究证明J774A.1在基因和蛋白质水平上均表达CB1和HuR;激活CB1能够促进J774A.1的迁移(P0.01)并且能够被其药理学拮抗剂AM281所抑制;激活CB1诱导的J774A.1的迁移依赖HuR;激活CB1促进了J774A.1胞质中HuR的富集进一步影响了CB1的表达,由此HuR参与了激活CB1诱导的J774A.1的迁移。结论激活CB1能够诱导单核巨噬细胞系J774A.1的迁移,且此过程依赖RNA结合蛋白HuR。
[Abstract]:Objective to study the important role of cannabinoid receptor 1(cannabinoid receptor 1 (CB1) in the migration of mononuclear macrophages and the possible mechanism of the involvement of RNA binding protein human antigen R(human antigen Ru Hur in the migration of mononuclear macrophages. Methods Mononuclear macrophage cell line J774A.1was used to identify the expression of CB1 and HuR in J774A.1 by agarose gel electrophoresis and immunofluorescence staining. ACEA and AM281 were pharmacological agonists and antagonists of CB1, respectively. The effect of ACEA and AM281 on J774A.1 migration activity was detected by Boyden chamber assay. The gene interference of Hur was used to determine whether the J774A.1 migration function induced by CB1 was dependent on HuR.The separation of cytoplasmic proteins was used to investigate whether activating CB1 could induce HuR in J774A.1 cytoplasm. The changes of CB1, HuR mRNA and protein were detected by RT-qPCR and Western blotting. Results this study demonstrated that J774A.1 expressed CB1 and Hur at both gene and protein levels, activation of CB1 promoted the migration of J774A.1 (P0.01) and was inhibited by its pharmacological antagonist AM281. Activation of CB1 induced migration of J774A.1 was dependent on Hur, and activation of CB1 promoted the accumulation of HuR in the cytoplasm of J774A.1 and further affected the expression of CB1, thus HuR was involved in CB1 induced J774A.1 migration. Conclusion Activation of CB1 can induce the migration of monocyte macrophage J774A.1, and this process is dependent on the RNA binding protein Hur.
【作者单位】：首都医科大学基础医学院细胞生物学系‘肝脏保护与再生调节’北京市重点实验室;
【基金】：国家自然科学基金(81170407,31301154) 北京市属高等学校创新团队建设与教师职业发展计划项目(IDHT20150502)~~
【分类号】：R392

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