挥发性麻醉药七氟烷抑制Toll样受体1/2激活
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挥发性麻醉药七氟烷抑制Toll样受体1/2激活
翻译：任文鑫 编辑：冯玉蓉 审校：曹莹
背景：虽然麻醉药物的免疫调节作用已经被越来越多的人认识到，但其潜在的分子机制尚未完全清楚。Toll样受体（Toll-like receptors，TLRs）是识别入侵病原体和受损宿主组织发出的危险信号并启动免疫反应的主要受体之一。在TLR家族中，TLR2和TLR4可识别广泛的配体，在围术期病理生理学中具有重要作用。基于我们新近的研究发现——挥发性麻醉药可调节TLR4功能，我们假设并验证挥发性麻醉药也可调节TLR2功能。
方法：采用报告分析法检测麻醉药异氟烷、七氟烷、异丙酚和右美托咪定对TLR2活性的影响。对影响TLR2激活的麻醉药进行计算机精确对接模拟。为了验证我们的假设，七氟烷和TLR1/TLR2配体Pam3CSK4会竞争TLR2的同一位点，我们使用稳定转染了TLR2 的含七氟烷或不含七氟烷的HEK细胞(HEK-TLR2)，进行Pam3CSK4竞争结合试验。我们研究了不同麻醉药对TLR2配体刺激的人中性粒细胞功能的影响。采用Kruskal-Wallis检验和Mann-WhitneyU检验进行统计分析。
结果：我们观察到，七氟烷暴露时可减弱TLR1/TLR2活性，而异氟烷、异丙酚或右美托咪定暴露时则没有。TLR2/TLR6活性在所有受试麻醉药中均未出现衰减。计算机精确对接模拟预测七氟烷和Pam3CSK4结合在TLR1/TLR2复合物的同一位点中。七氟烷存在的情况下，Pam3CSK4与HEK-TLR2细胞的结合受损，这表明七氟烷和Pam3CSK4竞争位点，正如计算机模拟预测的那样。Pam3CSK4刺激中性粒细胞可诱导L-选择素脱落，但不影响吞噬功能和活性氧的产生。中性粒细胞分泌的L-选择素仅被七氟烷减弱，这与我们报告分析结果一致。
结论：我们发现七氟烷可减弱TLR1/TLR2活性，但在临床相关浓度下，我们未发现异氟烷、异丙酚或右美托咪定可减弱TLR1/TLR2活性的证据。我们的结构性分析和竞争性试验支持在TLR1/TLR2复合物的中间相时，七氟烷直接与TLR2结合。七氟烷抑制中性粒细胞L-选择素脱落，是中性粒细胞迁移的重要步骤。
文献来源：Mitsui Y, Hou L, Huang X, et al. Volatile Anesthetic Sevoflurane Attenuates Toll-Like Receptor 1/2 Activation.[J].Anesth Analg 2020;131:631–9.
Volatile Anesthetic Sevoflurane Attenuates Toll-Like Receptor 1/2 Activation
Abstract
BACKGROUND: Although immunomodulatory effects of anesthetics have been increasingly recognized, their underlying molecular mechanisms are not completely understood. Toll-like receptors (TLRs) are one of the major receptors to recognize invading pathogens and danger signals from damaged host tissues to initiate immune responses. Among the TLR family, TLR2 and TLR4 recognize a wide range of ligands and are considered to be important players in perioperative pathophysiology. Based on our recent finding that volatile anesthetics modulate TLR4 function, we tested our hypothesis that they would also modulate TLR2 function.
METHODS: The effect of anesthetics isoflurane, sevoflurane, propofol, and dexmedetomidine on TLR2 activation was examined by reporter assays. An anesthetic that affected the activation was subjected to in silico rigid docking simulation on TLR2. To test our prediction that sevoflurane and a TLR1/TLR2 ligand Pam3CSK4 would compete for the same pocket of TLR2, we performed Pam3CSK4 competitive binding assay to TLR2 using HEK cells stably transfected with TLR2 (HEKTLR2) with or without sevoflurane. We examined the effect of different anesthetics on the functions of human neutrophils stimulated with TLR2 ligands. Kruskal–Wallis test and Mann–Whitney U test were used for statistical analysis.
RESULTS: We observed that the attenuation of TLR1/TLR2 activation was seen on sevoflurane exposure but not on isoflurane, propofol, or dexmedetomidine exposure. The attenuation of TLR2/TLR6 activation was not seen in any of the anesthetics tested. The rigid docking simulation predicted that sevoflurane and Pam3CSK4 bound to the same pocket of TLR1/TLR2 complex. The binding of Pam3CSK4 to HEK-TLR2 cells was impaired in the presence of sevoflurane, indicating that sevoflurane and Pam3CSK4 competed for the pocket, as predicted in silico. The stimulation of neutrophils with Pam3CSK4 induced L-selection shedding but did not affect phagocytosis and reactive oxygen species production. L-selectin shedding from neutrophils was attenuated only by sevoflurane, consistent with the result of our reporter assays.
CONCLUSIONS: We found that TLR1/TLR2 activation was attenuated by sevoflurane, but we found no evidence for attenuation by isoflurane, propofol, or dexmedetomidine at clinically relevant concentrations. Our structural analysis and competition assay supported that sevoflurane directly bound to TLR2 at the interphase of the TLR1/TLR2 complex. Sevoflurane attenuated neutrophil L-selectin shedding, an important step for neutrophil migration.
贵州医科大学高鸿教授课题组
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