Cardiovascular disease is one of the main causes of global mortality, and there is an urgent need for effective treatment strategies. Gut microbiota-dependent metabolite trimethylamine-N-oxide (TMAO) promotes the development of cardiovascular diseases, and shizukaol C, a natural sesquiterpene isolated from with various biological activities, might exhibit beneficial role in preventing TMAO-induced vascular inflammation. The purpose of this study was to investigate the anti-inflammatory effects and the underlying mechanisms of shizukaol C on TMAO-induced vascular inflammation. The effect and underlying mechanism of shizukaol C on TMAO-induced adhesion molecules expression, bone marrow-derived macrophages (BMDM) adhesion to VSMC were evaluated by western blot, cell adhesion assay, co-immunoprecipitation, immunofluorescence assay, and quantitative Real-Time PCR, respectively. To verify the role of shizukaol C , TMAO-induced vascular inflammation model were established using guidewire-induced injury on mice carotid artery. Changes in the intima area and the expression of GSTpi, VCAM-1, CD68 were examined using haematoxylin-eosin staining, and immunofluorescence assay. Our data demonstrated that shizukaol C significantly suppressed TMAO-induced adhesion molecule expression and the bone marrow-derived macrophages (BMDM) adhesion in vascular smooth muscle cells (VSMC). Mechanically, shizukaol C inhibited TMAO-induced c-Jun N-terminal kinase (JNK)-nuclear factor-kappa B (NF-κB)/p65 activation, and the JNK inhibition was dependent on the shizukaol C-mediated glutathione-S-transferase pi (GSTpi) expression. By further molecular docking and protein-binding analysis, we demonstrated that shizukaol C directly binds to Keap1 to induce Nrf2 nuclear translocation and upregulated GSTpi expression. Consistently, our experiment showed that shizukaol C elevated the expression level of GSTpi in carotid arteries and alleviates TMAO-induced vascular inflammation. : Shizukaol C exerts anti-inflammatory effects in TMAO-treated VSMC by targeting Keap1 and activating Nrf2-GSTpi signaling and resultantly inhibits the downstream JNK-NF-κB/p65 activation and VSMC adhesion, and alleviates TMAO-induced vascular inflammation , suggesting that shizukaol C may be a potential drug for treating TMAO-induced vascular diseases.

中文翻译：

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Shizukaol C 通过激活血管平滑肌细胞 Keap1-Nrf2-GSTpi 通路减轻三甲胺氧化物诱导的炎症

心血管疾病是全球死亡的主要原因之一，迫切需要有效的治疗策略。肠道微生物依赖的代谢物三甲胺-N-氧化物 (TMAO) 会促进心血管疾病的发生，而 Shizukaol C 是一种从具有多种生物活性的天然倍半萜烯中分离出来的物质，可能在预防 TMAO 引起的血管炎症方面发挥有益作用。本研究的目的是探讨 Shizukaol C 对 TMAO 诱导的血管炎症的抗炎作用及其潜在机制。通过蛋白质印迹、细胞粘附测定、免疫共沉淀、免疫荧光测定和定量实时 PCR 评估静香酚 C 对 TMAO 诱导的粘附分子表达、骨髓源性巨噬细胞 (BMDM) 粘附 VSMC 的影响和潜在机制， 分别。为了验证shizukaol C的作用，采用导丝诱导小鼠颈动脉损伤建立TMAO诱导的血管炎症模型。采用苏木精-伊红染色和免疫荧光法检测内膜面积的变化以及GSTpi、VCAM-1、CD68的表达。我们的数据表明，shizukaol C 显着抑制 TMAO 诱导的粘附分子表达和血管平滑肌细胞 (VSMC) 中骨髓源性巨噬细胞 (BMDM) 的粘附。机械上，shizukaol C 抑制 TMAO 诱导的 c-Jun N 末端激酶 (JNK)-核因子-κ B (NF-κB)/p65 激活，并且 JNK 抑制依赖于 shizukaol C 介导的谷胱甘肽-S-转移酶pi (GSTpi) 表达式。通过进一步的分子对接和蛋白结合分析，我们证明shizukaol C直接与Keap1结合诱导Nrf2核转位并上调GSTpi表达。我们的实验一致表明，shizukaol C 提高了颈动脉中 GSTpi 的表达水平，并减轻了 TMAO 诱导的血管炎症。 ：Shizukaol C 通过靶向 Keap1 并激活 Nrf2-GSTpi 信号传导，在 TMAO 处理的 VSMC 中发挥抗炎作用，从而抑制下游 JNK-NF-κB/p65 激活和 VSMC 粘附，并减轻 TMAO 诱导的血管炎症，表明 Shizukaol C C可能是治疗TMAO引起的血管疾病的潜在药物。