School of Pharmaceutical, Guangzhou University of Chinese Medicine, Guangzhou University Town, No. 232 Waihuan Dong Rd., Panyu District, Guangzhou Higher Education Mega Center, Guangzhou 510000, China.
Department of Pharmacy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, No. 1095 Jiefang Avenue, Wuhan 430030, China.
Phytomedicine. 2019 Mar 1;55:310-319. doi: 10.1016/j.phymed.2018.10.013. Epub 2018 Oct 11.
Recent studies indicate that vascular complications are closely related to diabetes mellitus; in particular, inflammatory-mediated endothelial dysfunction plays a crucial role in diabetes-induced cardiovascular diseases. Therefore, exploring effective methods to suppress endothelial dysfunction via inhibition of inflammatory responses is imperative. Puerarin (Pu), a flavonoid common in Pueraria, has been widely and successfully used to treat cardiovascular diseases in China for many years. However, information on its protective properties in hyperglycemia-induced vascular complications is insufficient. Hypothesis/Purpose: In this study, we investigate the protective effects of puerarin against high glucose-induced endothelial dysfunction and the underlying mechanism of the flavonoid.
we investigated the protective effects of Pu against hyperglycemia-induced inter-endothelial junction by permeability and transendothelial electrical resistance (TEER) assay. In addition, changes in the Nlrp3 inflammasome activation via reactive oxygen species (ROS)-dependent oxidative pathway were investigated using western blot, immunofluorescence microscopy analyses and flow cytometry. ROS scavenger and Nlrp3 gene silencing were used to determine the roles of the ROS-Nlrp3 pathway involved in the molecular mechanism of Pu.
Our findings demonstrate that puerarin inhibits high glucose-induced Nlrp3 inflammasome formation and activation, as shown by fluorescence confocal microscopy and Western blot. Puerarin decreases Nlrp3 protein, which is a critical factor necessary to form an inflammasome complex. We demonstrate that puerarin exerts anti-oxidation and ROS scavenged effects, similar to apocynin (APO). Interestingly, thioredoxin-interacting protein (TXNIP) protein and TXNIP binding to Nlrp3 markedly decreased with puerarin treatment. Together with these changes, puerarin could decrease high mobility group box 1 (HMGB1) release from mouse vascular endothelial cell (mMVECs). We also demonstrate the decreased expression of the tight junction proteins ZO-1/ZO-2, which are related to endothelial permeability after stimulation by high glucose in endothelial cells. Puerarin could recover the gap junction protein and decrease monolayer cell permeability in endothelial cells. In conclusion, we reveal a new protection mechanism of puerarin that inhibits Nlrp3 inflammasome activation and decreases subsequent caspase-1 activation, triggering the release of HMGB1 by reducing ROS generation.
Our findings indicate that puerarin exhibits immense potential and specific therapeutic value in hyperglycemia-related cardiovascular disease and the development of innovative drugs.
最近的研究表明，血管并发症与糖尿病密切相关；特别是，炎症介导的内皮功能障碍在糖尿病引起的心血管疾病中起着至关重要的作用。因此，探索通过抑制炎症反应来有效抑制内皮功能障碍的方法是当务之急。葛根素（Pu）是葛根中的一种黄酮类化合物，多年来在中国被广泛用于治疗心血管疾病。然而，关于其在高血糖诱导的血管并发症中的保护作用的信息还不够充分。假设/目的：在这项研究中，我们研究了葛根素对高葡萄糖诱导的内皮功能障碍的保护作用及其黄酮类化合物的潜在机制。
我们通过通透性和跨内皮电阻（TEER）测定研究了 Pu 对高糖诱导的内皮细胞间连接的保护作用。此外，通过 Western blot、免疫荧光显微镜分析和流式细胞术研究了 Nlrp3 炎性小体激活的变化，通过活性氧（ROS）依赖性氧化途径。使用 ROS 清除剂和 Nlrp3 基因沉默来确定 ROS-Nlrp3 途径在 Pu 的分子机制中所涉及的作用。
我们的研究结果表明，葛根素抑制高葡萄糖诱导的 Nlrp3 炎性小体的形成和激活，如荧光共聚焦显微镜和 Western blot 所示。葛根素降低了 Nlrp3 蛋白，这是形成炎性小体复合物所必需的关键因素。我们证明，葛根素有抗氧化和清除 ROS 的作用，类似于 apocynin（APO）。有趣的是，随着葛根素的治疗，硫氧还蛋白相互作用蛋白（TXNIP）蛋白和 TXNIP 与 Nlrp3 的结合明显减少。与这些变化一起，葛根素可以减少高迁移率族蛋白 1（HMGB1）从小鼠血管内皮细胞（mMVECs）中的释放。我们还证明了在高糖刺激后内皮细胞中紧密连接蛋白 ZO-1/ZO-2 的表达减少，这与内皮通透性有关。葛根素可以恢复间隙连接蛋白并降低内皮细胞单层细胞通透性。总之，我们揭示了葛根素抑制 Nlrp3 炎性小体激活并减少随后 caspase-1 激活，从而通过减少 ROS 生成来触发 HMGB1 释放的新保护机制。
我们的研究结果表明，葛根素在高血糖相关心血管疾病和创新药物的发展中具有巨大的潜力和特定的治疗价值。
