Daaka Y, Friedman H, Klein T W
Department of Medical Microbiology and Immunology, University of South Florida, College of Medicine, Tampa, USA.
J Pharmacol Exp Ther. 1996 Feb;276(2):776-83.
Brain-type cannabinoid receptor (CB1) mRNA has been demonstrated in several peripheral tissues; however, the function of this message is not clear. In the present study, we examined the levels of CB1 mRNA and receptor protein in stimulated immune cells to link message and protein expression with cell activation. Consistent with previous reports, immune cell lines from human and mouse were positive by reverse transcription-polymerase chain reaction for CB1 mRNA; however, one cell line, Jurkat, was only weakly positive. Mitogen activation of Jurkat cells, however, increased CB1 mRNA within 2 hr after stimulation and equilibrium binding studies, using Jurkat membranes, showed [3H]CP55,940 specific binding was negative early after mitogen stimulation but positive at 40 hr poststimulation. To investigate whether this observed increase in CB1 mRNA and specific binding activity was associated with expression of the CB1 protein, polyclonal antibodies were produced to a fusion protein consisting of glutathione S-transferase and a 342 amino acid portion (residues 33 through 374) of the CB1 protein. Western blotting analysis showed expression of several immunoreactive proteins on membranes from mitogen-activated Jurkat cells, but not on membranes from unstimulated cells. These results demonstrate a link between the level of CB1 mRNA and surface protein in activated immune cells, suggesting the possibility of a functional role of CB1 in immune cell activation.
脑型大麻素受体（CB1）mRNA已在多种外周组织中得到证实；然而，这种信息的功能尚不清楚。在本研究中，我们检测了受刺激免疫细胞中CB1 mRNA和受体蛋白的水平，以将信息和蛋白表达与细胞活化联系起来。与先前的报道一致，人和小鼠的免疫细胞系通过逆转录-聚合酶链反应检测CB1 mRNA呈阳性；然而，一种细胞系Jurkat仅呈弱阳性。然而，Jurkat细胞经丝裂原激活后，在刺激后2小时内CB1 mRNA增加，使用Jurkat细胞膜进行的平衡结合研究表明，[3H]CP55,940特异性结合在丝裂原刺激后早期为阴性，但在刺激后40小时为阳性。为了研究观察到的CB1 mRNA增加和特异性结合活性是否与CB1蛋白的表达相关，制备了针对由谷胱甘肽S-转移酶和CB1蛋白的342个氨基酸部分（第33至374位残基）组成的融合蛋白的多克隆抗体。蛋白质印迹分析显示，丝裂原激活的Jurkat细胞膜上有几种免疫反应性蛋白表达，但未刺激细胞的膜上没有。这些结果表明活化免疫细胞中CB1 mRNA水平与表面蛋白之间存在联系，提示CB1在免疫细胞活化中可能具有功能作用。
