Department of Anesthesiology, Chengde Central Hospital, China.
Department of Endocrinology, Chengde Central Hospital, China.
J Toxicol Sci. 2023;48(7):387-398. doi: 10.2131/jts.48.387.
Morphine-induced microglia activation and neuroinflammation have been considered as the contributors of morphine tolerance. Corilagin (Cori) has been reported to exhibit strong anti-inflammatory property. The present study aims to investigate whether and how Cori alleviates morphine-induced neuroinflammation and microglia activation. Mouse BV-2 cells were exposed to different concentrations of Cori (0.1, 1 and 10 μM) prior to morphine stimulation (200 μM). Minocycline (10 μM) acted as the positive control. Cell viability was determined by CCK-8 assay and trypan blue assay. The levels of inflammatory cytokines were determined using ELISA. IBA-1 level was examined via immunofluorescence. TLR2 expression level was examined by quantitative real-time PCR and western blot. The expression levels of corresponding proteins were measured by western blot. It was found that Cori was non-toxic to BV-2 cells but greatly inhibited morphine-induced IBA-1 expression, overproduction of pro-inflammatory cytokines, activation of NLRP3 inflammasome and endoplasmic reticulum stress (ERS), and upregulation of COX-2 and iNOS. TLR2 was negatively regulated by Cori, and could promote the activation of ERS. A high affinity between Cori and TLR2 protein was confirmed via Molecular docking investigation. Moreover, TLR2 overexpression or tunicamycin (TM), an agonist of ERS, partly abolished the inhibitory effects of Cori on morphine-induced alternations on neuroinflammation and microglial activation in BV-2 cells as above. In summary, our study suggested that Cori effectively alleviated morphine-induced neuroinflammation and microglia activation through inhibiting TLR2-mediated ERS in BV-2 cells, providing a novel potential drug to overcome morphine tolerance.
吗啡诱导的小胶质细胞激活和神经炎症被认为是吗啡耐受的原因。鞣花酸（Cori）已被报道具有很强的抗炎作用。本研究旨在探讨 Cori 是否以及如何减轻吗啡诱导的神经炎症和小胶质细胞激活。将小鼠 BV-2 细胞暴露于不同浓度的 Cori（0.1、1 和 10 μM）之前用吗啡刺激（200 μM）。米诺环素（10 μM）作为阳性对照。通过 CCK-8 测定法和台盼蓝测定法测定细胞活力。通过 ELISA 测定炎症细胞因子水平。通过免疫荧光法检查 IBA-1 水平。通过定量实时 PCR 和 Western blot 检查 TLR2 表达水平。通过 Western blot 测量相应蛋白的表达水平。结果发现 Cori 对 BV-2 细胞无毒，但能显著抑制吗啡诱导的 IBA-1 表达、促炎细胞因子的过度产生、NLRP3 炎性体和内质网应激（ERS）的激活以及 COX-2 和 iNOS 的上调。Cori 负调节 TLR2，并能促进 ERS 的激活。通过分子对接研究证实了 Cori 与 TLR2 蛋白之间的高亲和力。此外，TLR2 过表达或衣霉素（TM），ERS 的激动剂，部分消除了 Cori 对吗啡诱导的 BV-2 细胞神经炎症和小胶质细胞激活改变的抑制作用，如前所述。综上所述，我们的研究表明，Cori 通过抑制 TLR2 介导的 ERS，有效缓解吗啡诱导的神经炎症和小胶质细胞激活，为克服吗啡耐受提供了一种新的潜在药物。
