Elevated levels of 2-arachidonoylglycerol promote atherogenesis in ApoE-/- mice.
Elevated levels of 2-arachidonoylglycerol promote atherogenesis in ApoE-/- mice.
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DOI：
10.1371/journal.pone.0197751
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发表时间：
2018
期刊：
影响因子：
3.7
通讯作者：
Nickenig G
中科院分区：
文献类型：
--
作者：
Jehle J;Schöne B;Bagheri S;Avraamidou E;Danisch M;Frank I;Pfeifer P;Bindila L;Lutz B;Lütjohann D;Zimmer A;Nickenig G
内源性大麻素 (eCB) 2-花生四烯酰甘油 (2-AG) 是一种已知的炎症调节剂，也是促炎性大麻素受体 1 (CB1) 和抗炎性 CB2 的配体。虽然这两种受体在动脉粥样硬化形成中的作用已被广泛研究，但 2-AG 对动脉粥样硬化形成的重要性尚不清楚。在 ApoE-/- 小鼠的两个治疗组中研究了 2-AG 对动脉粥样硬化形成的影响。一组接受单酰基甘油脂肪酶 (MAGL) 抑制剂 JZL184 [5 mg/kg 腹腔注射]，它会损害 2-AG 降解，从而导致 2-AG 水平升高，另一组接受载体治疗 4 周。同时，两组均接受高胆固醇饮食。油红 O 染色后，通过主动脉窦冰冻切片评估动脉粥样硬化斑块负荷，并通过靶向 CD68 的免疫荧光检测浸润巨噬细胞。在体外，通过博伊登室实验评估了 2-AG 对 B6MCL 巨噬细胞迁移的影响。通过 qPCR 评估巨噬细胞中粘附分子和趋化因子受体的转录。正如预期的那样，MAGL 抑制剂 JZL184 的应用导致血管组织中 2-AG 水平显着增加（98.2 ± 16.1 nmol/g 对比 27.3 ± 4.5 nmol/g；n = 14–16；p < 0.001）。与载体处理的对照相比，2-AG 水平升高的 ApoE-/- 小鼠表现出显着增加的斑块负荷（0.44 ± 0.03 对比 0.31 ± 0.04；n = 14；p = 0.0117）。与此同时，动脉粥样硬化血管壁内浸润巨噬细胞显着增加（0.33 ± 0.02 vs. 0.27 ± 0.01；n = 13–14；p = 0.0076）。虽然 JZL184 治疗动物的白细胞计数没有变化，但与媒介物相比，2-AG 在体外增强巨噬细胞迁移能力 1.8 ± 0.2 倍（n = 4-6；p = 0.0393），而联合给予 CB1 或 CB2 受体拮抗剂则完全消除了这一现象。 2-AG 刺激的巨噬细胞的 qPCR 分析显示，趋化因子 CCL5（1.59 ± 0.23 倍；n = 5-6；p = 0.0589）及其相应受体 CCR1（2.04 ± 0.46 倍；n = 10-11；p = 0.0472）和 CCR5（2.45 ± 0.62 倍；p = 0.0472）的转录增强。 n = 5–6；p = 0.0554)。总而言之，升高的 2-AG 水平似乎会促进体内动脉粥样硬化形成。我们的数据表明，2-AG 可能通过 CCL5-CCR5/CCR1 轴促进巨噬细胞迁移，从而导致血管炎症。因此，降低血管 2-AG 水平可能是动脉粥样硬化和冠心病患者的一种有前景的治疗策略。
The endocannabinoid (eCB) 2-arachidonoylglycerol (2-AG) is a known modulator of inflammation and ligand to both, pro-inflammatory cannabinoid receptor 1 (CB1) and anti-inflammatory CB2. While the role of both receptors in atherogenesis has been studied extensively, the significance of 2-AG for atherogenesis is less well characterized. The impact of 2-AG on atherogenesis was studied in two treatment groups of ApoE-/- mice. One group received the monoacylglycerol lipase (MAGL)-inhibitor JZL184 [5 mg/kg i.p.], which impairs 2-AG degradation and thus causes elevated 2-AG levels, the other group received vehicle for four weeks. Simultaneously, both groups were fed a high-cholesterol diet. The atherosclerotic plaque burden was assessed in frozen sections through the aortic sinus following oil red O staining and infiltrating macrophages were detected by immunofluorescence targeting CD68. In vitro, the effect of 2-AG on B6MCL macrophage migration was assessed by Boyden chamber experiments. Transcription of adhesion molecules and chemokine receptors in macrophages was assessed by qPCR. As expected, application of the MAGL-inhibitor JZL184 resulted in a significant increase in 2-AG levels in vascular tissue (98.2 ± 16.1 nmol/g vs. 27.3 ± 4.5 nmol/g; n = 14–16; p < 0.001). ApoE-/- mice with elevated 2-AG levels displayed a significantly increased plaque burden compared to vehicle treated controls (0.44 ± 0.03 vs. 0.31 ± 0.04; n = 14; p = 0.0117). This was accompanied by a significant increase in infiltrating macrophages within the atherosclerotic vessel wall (0.33 ± 0.02 vs. 0.27 ± 0.01; n = 13–14; p = 0.0076). While there was no alteration to the white blood counts of JZL184-treated animals, 2-AG enhanced macrophage migration in vitro by 1.8 ± 0.2 -fold (n = 4–6; p = 0.0393) compared to vehicle, which was completely abolished by co-administration of either CB1- or CB2-receptor-antagonists. qPCR analyses of 2-AG-stimulated macrophages showed an enhanced transcription of the chemokine CCL5 (1.59 ± 0.23 –fold; n = 5–6; p = 0.0589) and its corresponding receptors CCR1 (2.04 ± 0.46 -fold; n = 10–11; p = 0.0472) and CCR5 (2.45 ± 0.62 –fold; n = 5–6; p = 0.0554). Taken together, elevated 2-AG levels appear to promote atherogenesis in vivo. Our data suggest that 2-AG promotes macrophage migration, possibly by the CCL5-CCR5/CCR1 axis, and thereby contributes to vascular inflammation. Thus, decreasing vascular 2-AG levels might represent a promising therapeutic strategy in patients suffering from atherosclerosis and coronary heart disease.
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通讯作者：
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