Ooi Bee Kee, Phang Su Wen, Yong Phelim Voon Chen, Chellappan Dinesh Kumar, Dua Kamal, Khaw Kooi-Yeong, Goh Bey Hing, Pusparajah Priyia, Yap Wei Hsum
School of Biosciences, Taylor's University, Subang Jaya, Selangor Darul Ehsan 47500, Malaysia.
Department of Life Sciences, School of Pharmacy, International Medical University (IMU), Bukit Jalil, 57000, Kuala Lumpur, Malaysia.
Life Sci. 2021 Aug 1;278:119658. doi: 10.1016/j.lfs.2021.119658. Epub 2021 May 25.
Maslinic acid (MA) is a naturally occurring pentacyclic triterpene known to exert cardioprotective effects. This study aims to investigate the involvement of nuclear factor erythroid 2-related factor 2 (Nrf2) for MA-mediated anti-inflammatory effects in atheroma pathogenesis in vitro, including evaluation of tumor necrosis factor-alpha (TNF-α)-induced monocyte recruitment, oxidized low-density lipoprotein (oxLDL)-induced scavenger receptors expression, and nuclear factor-kappa B (NF-ĸB) activity in human umbilical vein endothelial cells (HUVECS) and human acute monocytic leukemia cell line (THP-1) macrophages.
An in vitro monocyte recruitment model utilizing THP-1 and HUVECs was developed to evaluate TNF-α-induced monocyte adhesion and trans-endothelial migration. To study the role of Nrf2 for MA-mediated anti-inflammatory effects, Nrf2 inhibitor ML385 was used as the pharmacological inhibitor. The expression of Nrf2, monocyte chemoattractant protein-1 (MCP-1), vascular cell adhesion molecule 1 (VCAM-1), cluster of differentiation 36 (CD36), and scavenger receptor type A (SR-A) in HUVECs and THP-1 macrophages were investigated using RT-qPCR and Western blotting. The NF-κB activity was determined using NF-κB (p65) Transcription Factor Assay Kit.
The results showed opposing effects of MA on Nrf2 expression in HUVECs and THP-1 macrophages. MA suppressed TNF-α-induced Nrf2 expression in HUVECs, but enhanced its expression in THP-1 macrophages. Combined effects of MA and ML385 suppressed MCP-1, VCAM-1, and SR-A expressions. Intriguingly, at the protein level, ML385 selectively inhibited SR-A but enhanced CD36 expression. Meanwhile, ML385 further enhanced MA-mediated inhibition of NF-κB activity in HUVECs. This effect, however, was not observed in THP-1 macrophages.
MA attenuated foam cell formation by suppressing VCAM-1, MCP-1, and SR-A expression, as well as NF-κB activity, possibly through Nrf2 inhibition. The involvement of Nrf2 for MA-mediated anti-inflammatory effects however differs between HUVECs and macrophages. Future investigations are warranted for a detailed evaluation of the contributing roles of Nrf2 in foam cells formation.
山楂酸（MA）是一种天然存在的五环三萜，已知具有心脏保护作用。本研究旨在探讨核因子红细胞2相关因子2（Nrf2）在MA介导的体外动脉粥样硬化发病机制抗炎作用中的参与情况，包括评估肿瘤坏死因子-α（TNF-α）诱导的单核细胞募集、氧化型低密度脂蛋白（oxLDL）诱导的清道夫受体表达，以及人脐静脉内皮细胞（HUVECS）和人急性单核细胞白血病细胞系（THP-1）巨噬细胞中的核因子-κB（NF-κB）活性。
建立了一种利用THP-1和HUVECs的体外单核细胞募集模型，以评估TNF-α诱导的单核细胞黏附和跨内皮迁移。为研究Nrf2在MA介导的抗炎作用中的作用，使用Nrf2抑制剂ML385作为药理抑制剂。采用RT-qPCR和蛋白质印迹法研究HUVECs和THP-1巨噬细胞中Nrf2、单核细胞趋化蛋白-1（MCP-1）、血管细胞黏附分子1（VCAM-1）、分化簇36（CD36）和A类清道夫受体（SR-A）的表达。使用NF-κB（p65）转录因子检测试剂盒测定NF-κB活性。
结果显示MA对HUVECs和THP-1巨噬细胞中Nrf2表达有相反的作用。MA抑制TNF-α诱导的HUVECs中Nrf2的表达，但增强其在THP-1巨噬细胞中的表达。MA和ML385的联合作用抑制了MCP-1、VCAM-1和SR-A的表达。有趣的是，在蛋白质水平上，ML385选择性抑制SR-A但增强CD36的表达。同时，ML385进一步增强了MA介导的HUVECs中NF-κB活性的抑制。然而，在THP-1巨噬细胞中未观察到这种作用。
MA可能通过抑制Nrf2，抑制VCAM-1、MCP-
