艾纳香油对NF-κB及Nrf2/HO-1信号通路的作用研究

doi:10.11843/j.issn.0366-6964.2021.04.013

摘要/Abstract

摘要： 旨在通过研究NF-κB、Nrf2/HO-1通路来揭示艾纳香油（Blumea balsamifera（L.）DC Oil，BBO）发挥抗炎效果的作用机制。本研究利用LPS诱导的RAW264.7巨噬细胞炎症模型，试验分为空白组、LPS模型组、BBO组（低、中、高剂量）、BBO阴性对照组，每组3个重复，采用Hoechst 33342和PI双染检测BBO对细胞凋亡的影响；采用ELISA和分光光度法检测BBO对LPS诱导巨噬细胞后分泌IL-1β、TNF-α、PGE₂、LTB₄、NO含量及iNOS活力的影响；采用RT-PCR检测BBO对TNF-α、IL-1β的mRNA表达水平的影响；采用Western blotting检测BBO对NF-κB及Nrf2/HO-1信号通路关键蛋白水平的影响。结果显示，BBO在80 μg·mL^-1剂量范围内可以扭转LPS导致的巨噬细胞形态变化及细胞凋亡发生；与LPS模型组相比，BBO 60~80 μg·mL^-1剂量下可以极显著抑制LPS诱导的细胞炎性因子和炎症介质IL-1β、TNF-α、PGE₂、LTB₄、NO的分泌及iNOS活力（P<0.01），并极显著抑制LPS导致的细胞IL-1β、TNF-α mRNA表达（P<0.01）；同时，BBO 60~80 μg·mL^-1剂量下极显著下调LPS导致的COX-2、5-LOX、p-IKKα、p-P65、p-IκB-α、胞质Nrf2蛋白表达（P<0.01），极显著促进HO-1、核Nrf2蛋白表达（P<0.01），并呈现剂量依赖性关系。结果提示，艾纳香油有良好的抑制细胞炎性和抗凋亡效果，其可能通过抑制NF-p-IκBB通路中关键蛋白磷酸化和炎症因子的产生从而促进Nrf2/HO-1通路中主要抗炎基因表达最终发挥抗炎作用。

关键词: 艾纳香油, 脂多糖, RAW264.7细胞, NF-κB, Nrf2/HO-1通路

Abstract: The purpose of this study was to reveal the mechanism of anti-inflammatory effect of BBO by studying NF-κB and Nrf2/HO-1 pathway. RAW264.7 macrophage inflammation model induced by LPS was used. The control group, LPS model group, BBO group (low, middle and high dose) and BBO negative control group were set up in this experiment, with 3 repeats in each group. The effect of BBO on apoptosis was detected by Hoechst 33342 and PI double staining. The contents of IL-1β, TNF-α, PGE₂, LTB₄, NO and the activity of iNOS in LPS-induced macro-phage were detected by ELISA and spectrophotometry. The effect of BBO on the mRNA expression of TNF-α and IL-1β was detected by RT-PCR. The effect of BBO on the key proteins expression in NF-κB and Nrf2/HO-1 signal pathway was detected by Western blotting. The results showed that BBO could reverse the morphological changes and apoptosis of macrophages induced by LPS within the dose range of 80 μg·mL^-1. Compared with LPS model group, BBO of 60-80 μg·mL^-1 could extremely significantly inhibit the secretion of LPS-induced cellular inflammatory factors and inflammatory mediators IL-1β, TNF-α, PGE₂, LTB₄, NO and iNOS activity(P<0.01); BBO could also extremely significantly inhibit the expression of IL-1β and TNF-α mRNA(P<0.01) induced by LPS. At the same time, BBO of 60-80 μg·mL^-1 could extremely significantly down-regulate the protein expression of COX-2, 5-LOX, p-IKKα, p-P65, p-IκB-α and cytoplasmic Nrf2 induced by LPS(P<0.01), and extremely significantly promote the protein expression of HO-1 and nuclear Nrf2 in a dose-dependent manner(P<0.01). The results suggest that BBO has good effects on anti-inflammatory and anti-apoptotic. It may play an anti-inflammatory role by inhibiting the phosphorylation of key proteins and the production of inflammatory factors in NF-κB pathway and promote the expression of major anti-inflammatory genes in Nrf2/HO-1 pathway.

Key words: Blumea balsamifera(L.) DC Oil, LPS, RAW264.7 cell, NF-κB, Nrf2/HO-1 pathway

中图分类号:

S852.2

王万林, 高月, 廖加美, 彭俊超, 蔡亚玲, 易琼, 王鲁. 艾纳香油对NF-κB及Nrf2/HO-1信号通路的作用研究[J]. 畜牧兽医学报, 2021, 52(4): 976-986.

WANG Wanlin, GAO Yue, LIAO Jiamei, PENG Junchao, CAI Yaling, YI Qiong, WANG Lu. Effect of Blumea balsamifera (L.) DC Oil on NF-κB and Nrf2/HO-1 Signal Pathway[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(4): 976-986.

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