State Key Laboratory of Traditional Chinese Medicine Syndrom，The second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, 510120, China.
School of Biomedical and Pharmaceutical Sciences, Guangdong University of Technology, Guangzhou, 510006, China.
Inflamm Res. 2024 Jan;73(1):47-63. doi: 10.1007/s00011-023-01819-8. Epub 2023 Dec 26.
NLRP3 inflammasome-mediated pyroptosis of macrophage acts essential roles in the progression of sepsis-induced acute lung injury (ALI). Tangeretin (TAN), enriched in citrus fruit peel, presents anti-oxidative and anti-inflammatory effects. Here, we aimed to explore the potentially protective effect of TAN on sepsis-induced ALI, and the underlying mechanism of TAN in regulating NLRP3 inflammasome.
The effect of TAN on sepsis-induced ALI and NLRP3 inflammasome-mediated pyroptosis of macrophage were examined in vivo and in vitro using a LPS-treated mice model and LPS-induced murine macrophages, respectively. The mechanism of TAN regulating the activation of NLRP3 inflammasome in sepsis-induced ALI was investigated with HE staining, Masson staining, immunofluorescent staining, ELISA, molecular docking, transmission electron microscope detection, qRT-PCR, and western blot.
TAN could evidently attenuate sepsis-induced ALI in mice, evidenced by reducing pulmonary edema, pulmonary congestion and lung interstitial fibrosis, and inhibiting macrophage infiltration in the lung tissue. Besides, TAN significantly suppressed inflammatory cytokine IL-1β and IL-18 expression in the serum or bronchoalveolar lavage fluid (BALF) samples of mice with LPS-induced ALI, and inhibited NLRP3 inflammasome-mediated pyroptosis of macrophages. Furthermore, we found TAN inhibited ROS production, preserved mitochondrial morphology, and alleviated excessive mitochondrial fission in LPS-induced ALI in mice. Through bioinformatic analysis and molecular docking, Polo-like kinase 1 (PLK1) was identified as a potential target of TAN for treating sepsis-induced ALI. Moreover, TAN significantly inhibited the reduction of PLK1 expression, AMP-activated protein kinase (AMPK) phosphorylation, and Dynamin related protein 1 (Drp1) phosphorylation (S637) in LPS-induced ALI in mice. In addition, Volasertib, a specific inhibitor of PLK1, abolished the protective effects of TAN against NLRP3 inflammasome-mediated pyroptosis of macrophage and lung injury in the cell and mice septic models.
TAN attenuates sepsis-induced ALI by inhibiting ROS-mediated NLRP3 inflammasome activation via regulating PLK1/AMPK/DRP1 signaling axis, and TAN is a potentially therapeutic candidate against ALI through inhibiting pyroptosis.
NLRP3 炎性小体介导体细胞焦亡在脓毒症诱导的急性肺损伤（ALI）的进展中起关键作用。橙皮素（TAN）富含于柑橘类水果皮，具有抗氧化和抗炎作用。本研究旨在探讨 TAN 对脓毒症诱导的 ALI 的潜在保护作用，以及 TAN 调节 NLRP3 炎性小体的作用机制。
通过 LPS 处理的小鼠模型和 LPS 诱导的鼠巨噬细胞，分别在体内和体外研究 TAN 对脓毒症诱导的 ALI 和 NLRP3 炎性小体介导的巨噬细胞焦亡的影响。采用 HE 染色、Masson 染色、免疫荧光染色、ELISA、分子对接、透射电镜检测、qRT-PCR 和 Western blot 等方法研究 TAN 调节脓毒症诱导的 ALI 中 NLRP3 炎性小体激活的机制。
TAN 可明显减轻 LPS 诱导的 ALI 小鼠的肺损伤，表现为肺组织水肿、淤血和肺间质纤维化减轻，以及肺组织中巨噬细胞浸润减少。此外，TAN 明显抑制 LPS 诱导的 ALI 小鼠血清或支气管肺泡灌洗液（BALF）中炎症细胞因子 IL-1β和 IL-18 的表达，并抑制 NLRP3 炎性小体介导的巨噬细胞焦亡。进一步研究发现，TAN 抑制 LPS 诱导的 ALI 小鼠 ROS 产生，维持线粒体形态，减轻线粒体过度分裂。通过生物信息学分析和分子对接，发现 Polo 样激酶 1（PLK1）是 TAN 治疗脓毒症诱导的 ALI 的潜在靶点。此外，TAN 明显抑制 LPS 诱导的 ALI 小鼠中 PLK1 表达、AMP 激活蛋白激酶（AMPK）磷酸化和动力相关蛋白 1（Drp1）磷酸化（S637）的减少。此外，PLK1 的特异性抑制剂 Volasertib 消除了 TAN 在细胞和小鼠脓毒症模型中对 NLRP3 炎性小体介导的巨噬细胞焦亡和肺损伤的保护作用。
TAN 通过调节 PLK1/AMPK/DRP1 信号通路抑制 ROS 介导的 NLRP3 炎性小体激活，减轻脓毒症诱导的 ALI，TAN 通过抑制细胞焦亡可能成为治疗 ALI 的潜在治疗靶点。
