Saroz Yurii, Kho Dan T, Glass Michelle, Graham Euan Scott, Grimsey Natasha Lillia
Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, Centre for Brain Research, Faculty of Medical and Health Sciences, and Department of Molecular Medicine and Pathology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, 85 Park Road, Grafton, Auckland, 1023, New Zealand.
Department of Pharmacology and Toxicology, School of Biomedical Sciences, Division of Health Sciences, University of Otago, Dunedin, 9016, New Zealand.
ACS Pharmacol Transl Sci. 2019 Oct 1;2(6):414-428. doi: 10.1021/acsptsci.9b00049. eCollection 2019 Dec 13.
Cannabinoid receptor 2 (CB) is a promising therapeutic target for immunological modulation. There is, however, a deficit of knowledge regarding CB signaling and function in human primary immunocompetent cells. We applied an experimental paradigm which closely models the state of human primary leukocytes (PBMC; peripheral blood mononuclear cells) to characterize activation of a number of signaling pathways in response to a CB-selective ligand (HU308). We observed a "lag" phase of unchanged cAMP concentration prior to development of classically expected Gα-mediated inhibition of cAMP synthesis. Application of G protein inhibitors revealed that this apparent lag was a result of counteraction of Gα effects by concurrent Gα activation. Monitoring downstream signaling events showed that activation of p38 was mediated by Gα, whereas ERK1/2 and Akt phosphorylation were mediated by Gα-coupled βγ. Activation of CREB integrated multiple components; Gα and βγ mediated ∼85% of the response, while ∼15% was attributed to Gα. Responses to HU308 had an important functional outcome-secretion of interleukins 6 (IL-6) and 10 (IL-10). IL-2, IL-4, IL-12, IL-13, IL-17A, MIP-1α, and TNF-α were unaffected. IL-6/IL-10 induction had a similar G protein coupling profile to CREB activation. All response potencies were consistent with that expected for HU308 acting via CB. Additionally, signaling and functional effects were completely blocked by a CB-selective inverse agonist, giving additional evidence for CB involvement. This work expands the current paradigm regarding cannabinoid immunomodulation and reinforces the potential utility of CB ligands as immunomodulatory therapeutics.
大麻素受体2（CB）是免疫调节中一个很有前景的治疗靶点。然而，关于CB信号传导和在人类原代免疫活性细胞中的功能，我们还知之甚少。我们应用了一种实验范式，该范式紧密模拟人类原代白细胞（外周血单个核细胞，PBMC）的状态，以表征一系列信号通路在响应CB选择性配体（HU308）时的激活情况。我们观察到，在经典预期的Gα介导的cAMP合成抑制出现之前，cAMP浓度有一个不变的“滞后”阶段。应用G蛋白抑制剂显示，这种明显的滞后是由同时发生的Gα激活对Gα效应的抵消作用导致的。监测下游信号事件表明，p38的激活由Gα介导，而ERK1/2和Akt磷酸化由Gα偶联的βγ介导。CREB的激活整合了多个成分；Gα和βγ介导了约85%的反应，而约15%归因于Gα。对HU308的反应有一个重要的功能结果——白细胞介素6（IL-6）和10（IL-10）的分泌。IL-2、IL-4、IL-12、IL-13、IL-17A、MIP-1α和TNF-α不受影响。IL-6/IL-10的诱导与CREB激活具有相似的G蛋白偶联模式。所有反应效力均与HU308通过CB起作用的预期一致。此外，CB选择性反向激动剂完全阻断了信号传导和功能效应，为CB的参与提供了额外证据。这项工作扩展了当前关于大麻素免疫调节的范式，并强化了CB配体作为免疫调节治疗药物的潜在效用。
