鸢尾素介导抗炎和抗氧化作用的机制

doi:10.3969/j.issn.1004-616x.2022.04.001

癌变·畸变·突变 ›› 2022, Vol. 34 ›› Issue (4): 247-254,261.doi: 10.3969/j.issn.1004-616x.2022.04.001

• 论著 •

鸢尾素介导抗炎和抗氧化作用的机制

涂永梅^1,2, 于卫华², 彭洁², 刘江正², 刘瑞², 吴昊², 孔德钦², 何改花², 李文丽², 王冲¹

1. 陕西中医药大学公共卫生学院, 陕西咸阳 712046;
2. 空军军医大学军事预防医学系军事毒理学与防化医学教研室, 特殊作业环境危害评估与防治教育部重点实验室, 陕西省自由基生物学与医学重点实验室, 陕西西安 710032

收稿日期:2021-12-24 修回日期:2022-04-28 发布日期:2022-08-05
通讯作者: 王冲
作者简介:涂永梅，E-mail：2296828447@qq.com;于卫华，E-mail：yuweihua1@fmmu.edu.cn。
基金资助:
国家自然科学基金(31800706，32171231)；陕西省自然科学基金(2020-JQ-467)

A mechanistic study on irisin-mediated anti-inflammatory and antioxidant effects in cells

TU Yongmei^1,2, YU Weihua², PENG Jie², LIU Jiangzheng², LIU Rui², WU Hao², KONG Deqin², HE Gaihua², LI Wenli², WANG Chong¹

1. School of Public Health, Shaanxi University of Chinese Medicine, Xianyang 712046;
2. Department of Toxicology, Key Lab of Hazard Assessment and Control in Special Operational Environment of Ministry of Education, Shaanxi Provincial Key Lab of Free Radical Biology and Medicine, School of Public Health, Air Force Medical University, Xi'an 710032, Shaanxi, China

Received:2021-12-24 Revised:2022-04-28 Published:2022-08-05

摘要/Abstract

摘要： 目的：探讨鸢尾素(irisin)介导的小鼠巨噬细胞RAW264.7抗炎和抗氧化作用的机制。方法：用200 ng/mL鸢尾素分别处理RAW264.7细胞0、24和48 h，采用实时荧光定量PCR (qPCR)检测抗炎因子(IL-10、Arg-1、CD206)，抗氧化基因(HO-1、SOD2)和PPAR-γ mRNA的表达；酶联免疫吸附实验(ELISA)检测细胞上清中抗炎因子含量；试剂盒测定抗氧化酶活性；Western blot检测PPAR-γ蛋白表达；免疫荧光法检测Nrf2、PPAR-γ的表达和定位。另用200 ng/mL鸢尾素预处理细胞4 h，100 ng/mL脂多糖(LPS)刺激巨噬细胞24 h，采用qPCR检测促炎因子表达，DCFH-DA和Mito-LX染色检测活性氧(ROS)水平。进一步用PPAR-γ抑制剂T0070907(30 μmol/mL)预处理细胞4 h，200 ng/mL鸢尾素培养24 h，采用qPCR检测抗炎因子表达。结果：与对照组相比，200ng/mL鸢尾素可促进IL-10、Arg-1和CD206等抗炎因子的表达(P<0.05)，并抑制LPS诱导的TNF-α和IL-6 mRNA表达升高(P<0.05)，表明鸢尾素具有抗炎作用。并且200 ng/mL鸢尾素处理RAW264.7细胞4 h后Nrf2发生核转位，HO-1和SOD2表达和酶活性显著增强，GSH含量也升高(P<0.05)。此外，鸢尾素刺激下细胞内PPAR-γ的mRNA和蛋白表达升高，并发生明显核转位，PPAR-γ抑制剂T0070907可阻断鸢尾素介导的抗炎因子表达升高(P<0.05)。结论：鸢尾素通过PPAR-γ启动巨噬细胞M2型极化发挥抗炎作用，并增强Nrf2及下游抗氧化酶谱表达，有望成为炎症疾病候选治疗药物。

关键词: 鸢尾素, 巨噬细胞, 抗炎, 抗氧化, 过氧化物酶体增殖物激活受体γ, 核因子E2相关因子2

Abstract: OBJECTIVE: To investigate mechanisms of antioxidant and anti-inflammatory effects mediated by Irisin in mouse macrophages RAW264.7. METHODS: RAW264.7 cells were treated with 200 ng/mL Irisin for 0, 24 and 48 h. Real-time PCR was used to detect mRNA expressions of anti-inflammatory factors (IL-10,Arg-1,and CD206),antioxidant genes (HO-1 and SOD2) and PPAR-γ. ELISA assay was used to detect IL-10, Arg-1 in cell supernatants. Commercial kits were used to determine antioxidant enzyme activities,and Western-blot and immunofluorescence to detect Nrf2 and PPAR-γ expression and localization. Furthermore, after 24 h with 100 ng/mL LPS stimulation of RAW264.7 and 4 h 200 ng/mL Irisin pretreatment, real-time PCR was used to detect pro-inflammatory cytokines. DCFH-DA and Mito-LX were used to detect ROS levels. PPAR-γ inhibitor T0070907 (30 μmol/mL) was pretreated for 4 h and 200 ng/mL Irisin for 24 h, real-time PCR was used to detect expressions of anti-inflammatory cytokines. RESULTS: Compared with the untreated cells,Irisin promoted the expression and release of anti-inflammatory factors such as IL-10, Arg-1 and CD206 (P<0.05). However, Irisin inhibited LPS-induced TNF-α and IL-6 mRNA expression (P<0.05), which indicated that Irisin has anti-inflammatory effects. In addition, Nrf2 nuclear translocation occurred after irisin treatment in RAW264.7 cells. Expressions and enzyme activities of HO-1 and SOD2 were significantly enhanced, and the contents of GSH increased (P<0.05). In response to Irisin stimulation, PPAR-γ expression was elevated and underwent nuclear translocation, while PPAR-γ inhibitor T0070907 blocked the Irisin-induced anti-inflammatory response (P<0.05). CONCLUSION: Irisin initiated macrophage M2-type polarization and anti-inflammatory responses through enhancement of PPAR-γ-related anti-inflammatory genes and Nrf2-related antioxidant enzymes. Therefore,Irisin may be a candidate drug for inflammation-related diseases.

Key words: irisin, macrophage, anti-inflammation, antioxidant, PPAR-γ, Nrf2

中图分类号:

R114

涂永梅, 于卫华, 彭洁, 刘江正, 刘瑞, 吴昊, 孔德钦, 何改花, 李文丽, 王冲. 鸢尾素介导抗炎和抗氧化作用的机制[J]. 癌变·畸变·突变, 2022, 34(4): 247-254,261.

TU Yongmei, YU Weihua, PENG Jie, LIU Jiangzheng, LIU Rui, WU Hao, KONG Deqin, HE Gaihua, LI Wenli, WANG Chong. A mechanistic study on irisin-mediated anti-inflammatory and antioxidant effects in cells[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2022, 34(4): 247-254,261.

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鸢尾素介导抗炎和抗氧化作用的机制

A mechanistic study on irisin-mediated anti-inflammatory and antioxidant effects in cells

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