Total flavonoids of Salvia miltiorrhiza alleviate acetaminophen-induced acute liver injury in mice by suppressing hepatocyte ferroptosis via activating the Nrf2/HO-1 signaling pathway

doi:10.12122/j.issn.1673-4254.2024.11.17

Abstract

Abstract:

Objective To investigate the protective effect of total flavonoids of Salvia divinorum extract against acetaminophen (APAP)-induced acute liver injury (ALI) and its molecular mechanism. Methods The main chemical constituents of total flavonoids of Salvia divinorum were obtained through literature search, and their pharmacological mechanisms were predicted using bioinformatics analysis. In a mouse model of APAP-induced ALI, the protective effects of 100, 200 and 400 mg/kg total flavonoids of Salvia miltiorrhiza and 150 mg/kg bifidus were evaluated by observing changes in blood biochemistry and liver histopathology and detecting expressions of the key proteins in the Nrf2/HO-1 signaling pathway. Results Network pharmacology analysis suggested that the main active components in total flavonoids of Salvia divinorum for regulating APAP-induced liver injury included quercetin, lignocerol, caruric acid, and kaempferol, for which GO function enrichment analysis yielded 632 GO entries, including 472 involving biological processes, 42 involving cellular composition, and 118 involving molecular function. KEGG enrichment analysis showed that the total flavonoids of Salvia divinorum regulated APAP-induced liver injury mainly through ferroptosis-related signaling pathway. In mice with APAP-induced ALI, treatment with the total flavonoids significantly lowered ALT and AST levels, improved liver histopathology and inflammatory cell infiltration, reduced iron deposition in liver tissues, improved lipid peroxidation-related indexes, promoted the expressions of Nrf2, HO-1, SLC7A11, and GPX-4 proteins, and inhibited the expression of keap1 protein. Conclusion The total flavonoids of Salvia divinorum alleviate APAP-induced ALI in mice possibly by suppressing hepatocyte ferroptosis via activating the Nrf2/SLC7A11/GPX-4 signaling pathway.

Key words: total flavonoids of Salvia divinorum, Nrf2/HO-1, ferroptosis, network pharmacology, acute liver injury

Huajun CAI, Zhiqi CHEN, Wenting HU, Wei TAN, Hao WU, Chao WANG. Total flavonoids of Salvia miltiorrhiza alleviate acetaminophen-induced acute liver injury in mice by suppressing hepatocyte ferroptosis via activating the Nrf2/HO-1 signaling pathway[J]. Journal of Southern Medical University, 2024, 44(11): 2201-2208.

Figures/Tables 8

Fig.1 Result of network pharmacology analysis. A: Intersection target Wayne plot of the ethanol extract of Herba Desmodii Styracifolii and liver injury caused by acetaminophen. B: Protein-protein interaction (PPI) network. C: Drug-active ingredient-target-disease network.

Fig.2 GO analysis ( A) and KEGG enrichment analysis diagram ( B).

Fig.3 Effect of total flavonoids of Salvia divinorum extract (TFM) on serum ALT and AST levels in mice with APAP-induced ALI ( Mean±SD, n=6). ** P<0.01 vs control group; #P<0.05, ##P<0.01 vs APAP group.

Fig.4 Pathological changes in the liver tissues of mice with APAP-induced ALI treated with TFM (HE staining).

Fig.5 Effect of TFM on serum inflammatory factor TNF- α and IL-6 in the mice. ** P<0.01 vs control group; #P<0.05, ##P<0.01 vs APAP group.

Fig.6 Effects of TFM on the contents of 4-HNE ( A), MDA ( B), ROS ( C), and GSH( D) in the liver tissues of the mice. ** P<0.01 vs control group; #P<0.05, ##P<0.01 vs APAP group.

Fig.7 TFM reduces iron deposition in the liver tissues of the mice. A: Perls staining; B: Quantitative results ofPerls staining. ** P<0.01 vs control group; #P<0.05, ##P<0.01 vs APAP group.

Fig.8 　Expression levels of Nrf2, HO-1, keap1, GPX-4 and SLC7A11 proteins in the liver tissues of the mice detected by Western blotting. **P<0.01vs control group; #P<0.05, ##P<0.01vs APAP group.

References 31

1	Zhang Y, Wang C, Yu B, et al. Gastrodin protects against ethanol-induced liver injury and apoptosis in HepG2 cells and animal models of alcoholic liver disease[J]. Biol Pharm Bull, 2018, 41( 5): 670- 9.
2	Kaplowitz N, Than TA, Win S. Acetaminophen hepatotoxicity: Strong offense and weakened defens[J]. Hepatology, 2020, 71( 5): 1530- 2.
3	Yamada N, Karasawa T, Kimura H, et al. Ferroptosis driven by radical oxidation of n-6 polyunsaturated fatty acids mediates acetaminophen-induced acute liver failure[J]. Cell Death Dis, 2020, 11( 2): 144.
4	Teschke R. Treatment of drug-induced liver injury[J]. Biomedicines, 2022, 11( 1): 15.
5	高俊斌, 王璇, 陈燕红, 等. HPLC法同时测定鼠曲草中7种成分[J]. 中成药, 2018, 40( 5): 1116- 9.
6	Morimoto M, Kumeda S, Komai K. Insect antifeedant flavonoids from Gnaphalium affine D. don[J]. J Agric Food Chem, 2000, 48( 5): 1888- 91.
7	刘婉婷, 林婉琪, 卓辰茜, 等. 鼠曲草抗COPD有效成分及作用机制研究：基于UPLC-QE-MS结合网络药理学[J]. 亚太传统医药, 2023, 19( 08): 168- 75.
8	Zeng WC, Zhang WC, Zhang WH, et al. The antioxidant activity and active component of Gnaphalium affine extract[J]. Food Chem Toxicol, 2013, 58: 311- 7.
9	王海东, 林飞, 林伟青. 鼠曲草对大鼠膀胱癌形成的抑制作用研究[J]. 世界中医药, 2021, 16( 11): 1708- 11.
10	姜丽君, 朴锦花, 刘宇, 等. 鼠曲草提取物对四氯化碳所致小鼠急性肝损伤保护作用[J]. 时珍国医国药, 2008, 19( 8): 1901- 2.
11	张德胜, 康照金, 尤双梅, 等. 鼠曲草花中黄酮的提取及抗氧化活性研究[J]. 现代农业科技, 2017( 24): 237- 40, 244.
12	石青浩, 李荣, 姜子涛. 大孔树脂纯化鼠曲草总黄酮的研究[J]. 中国食品添加剂, 2012( 3): 93- 8.
13	陶艳艳, 陈高峰, 刘成海. 常用急性肝损伤动物模型评价及其在中药药理研究中的应用[J]. 上海中医药杂志, 2019, 53( 11): 11- 9.
14	田彩云. 药食同源植物鼠曲草中6种黄酮类成分的含量测定[J]. 人参研究, 2018, 30( 6): 30- 2.
15	覃萍, 张薇, 陈紫莹, 等. 槲皮素对对乙酰氨基酚诱导的小鼠急性肝损伤的保护作用[J]. 广东化工, 2022, 49( 12): 63- 6.
16	宋金玥, 任锋, 张向颖, 等. 山奈酚干预对D-氨基半乳糖/脂多糖诱导小鼠急性肝衰竭的保护作用[J]. 中国中西医结合杂志, 2017, 37( 5): 569- 74.
17	Xiao C, Xia ML, Wang J, et al. Luteolin attenuates cardiac ischemia/reperfusion injury in diabetic rats by modulating Nrf2 antioxidative function[J]. Oxid Med Cell Longev, 2019, 2019: 2719252.
18	Shojaie L, Iorga A, Dara L. Cell death in liver diseases: a review[J]. Int J Mol Sci, 2020, 21( 24): 9682.
19	Li P, Yu J, Huang F, Zhu YY, et al. SLC7A11-associated ferroptosis in acute injury diseases: mechanisms and strategies[J]. Eur Rev Med Pharmacol Sci, 2023, 27( 10): 4386- 98.
20	Zeng L, Jin XL, Xiao QA, et al. Ferroptosis: action and mechanism of chemical/drug-induced liver injury[J]. Drug Chem Toxicol, 2023: 1- 12.
21	Li HY, Lin YJ, Zhang L, et al. Ferroptosis and its emerging roles in acute pancreatitis[J]. Chin Med J, 2022, 135( 17): 2026- 34.
22	Li XB. Ferroptosis and drug-induced liver injury. Chin J Hepatol, 2023, 31( 4): 345- 8.
23	Jaeschke H, Adelusi OB, Ramachandran A. Ferroptosis and acetaminophen hepatotoxicity: are we going down another rabbit hole[J] ? Gene Expr, 2021, 20( 3): 169- 78.
24	Sun XF, Ou ZH, Chen RC, et al. Activation of the p62-Keap1-NRF2 pathway protects against ferroptosis in hepatocellular carcinoma cells[J]. Hepatology, 2016, 63( 1): 173- 84.
25	Fan Z, Wirth AK, Chen D, et al. Nrf2-Keap1 pathway promotes cell proliferation and diminishes ferroptosis[J]. Oncogenesis, 2017, 6( 8): e371.
26	Bellezza I, Giambanco I, Minelli A, et al. Nrf2-Keap1 signaling in oxidative and reductive stress[J]. Biochim Biophys Acta Mol Cell Res, 2018, 1865( 5): 721- 33.
27	Dodson M, Castro-Portuguez R, Zhang DD. NRF2 plays a critical role in mitigating lipid peroxidation and ferroptosis[J]. Redox Biol, 2019, 23: 101107.
28	张北雪, 孙建之, 姜宇珺, 等. 酒龙胆中总多糖和总环烯醚萜苷保肝作用的研究[J]. 中南药学, 2019, 17( 7): 1001- 5.
29	Wei YY, Gao C, Wang HR, et al. Mori fructus aqueous extracts attenuates liver injury by inhibiting ferroptosis via the Nrf2 pathway[J]. J Anim Sci Biotechnol, 2023, 14( 1): 56.
30	黄晓佳, 李永金, 李静, 等. 鼠曲草总黄酮抑制疼痛模型小鼠炎症因子产生而致镇痛作用[J]. 食品科学, 2014, 35( 21): 240- 3.
31	黄燕, 覃璐璐, 管少兴, 等. 金缕半枫荷的水提取物抑制胰腺癌的作用机制:活性成分、关键靶点和信号通路[J]. 南方医科大学学报, 2024, 44( 7): 1336- 44.

[1]	Mingyuan LI, Wei ZHANG, Mengqing HUA. Bardoxolone methyl alleviates acute liver injury in mice by inhibiting NLRP3 inflammasome activation [J]. Journal of Southern Medical University, 2024, 44(9): 1662-1669.
[2]	Kai CHEN, Zhaofei MENG, Jingting MIN, Jiahui WANG, Zhenghong LI, Qin GAO, Junfeng HU. Curcumin alleviates septic lung injury in mice by inhibiting TXNIP/TRX-1/GPX4-mediated ferroptosis [J]. Journal of Southern Medical University, 2024, 44(9): 1805-1813.
[3]	Mingzi OUYANG, Jiaqi CUI, Hui WANG, Zheng LIANG, Dajin PI, Liguo CHEN, Qianjun CHEN, Yingchao WU. Kaixinsan alleviates adriamycin-induced depression-like behaviors in mice by reducing ferroptosis in the prefrontal cortex [J]. Journal of Southern Medical University, 2024, 44(8): 1441-1449.
[4]	Xingmei CHEN, Qinwen LIU, Yi LI, Xiaoyu ZHONG, Qiling FAN, Ke MA, Liuting LUO, Daogang GUAN, Zhibo ZHU. Analysis of core functional components in Yinchenhao Decoction and their pathways for treating liver fibrosis [J]. Journal of Southern Medical University, 2024, 44(8): 1508-1517.
[5]	Shanyuan ZHANG, Qiaoyan CAI, Jianghan QI, Kaixin YIN, Chenchen HE, Zhuye GAO, Ling ZHANG, Jianfeng CHU. Pharmacodynamics of Qingxin Jieyu Granules for treatment of atherosclerosis and its regulatory mechanism for lipid metabolism [J]. Journal of Southern Medical University, 2024, 44(8): 1518-1528.
[6]	Yinliang ZHANG, Zetan LUO, Rui ZHAO, Na ZHAO, Zhidong XU, Di AO, Guyi CONG, Xinyu LIU, Hailun ZHENG. Sanguinarine induces ferroptosis of colorectal cancer cells by upregulating STUB1 and downregulating GPX4 [J]. Journal of Southern Medical University, 2024, 44(8): 1537-1544.
[7]	Yuming ZHANG, Shicheng XIA, Linlin ZHANG, Mengxi CHEN, Xiaojing LIU, Qin GAO, Hongwei YE. Protective effect of Lonicerae japonicae flos extract against doxorubicin-induced liver injury in mice [J]. Journal of Southern Medical University, 2024, 44(8): 1571-1581.
[8]	Jinjin WANG, Wenfei CUI, Xuewei DOU, Binglei YIN, Yuqi NIU, Ling NIU, Guoli YAN. Euonymus alatus delays progression of diabetic kidney disease in mice by regulating EGFR tyrosine kinase inhibitor resistance signaling pathway [J]. Journal of Southern Medical University, 2024, 44(7): 1243-1255.
[9]	Linyue WANG, Wenyue QI, Jihua GAO, Maosheng TIAN, Jiancheng XU. Tongyangxiao Lotion promotes postoperative wound healing in a rat model of anal fistula by downregulating inflammatory factors and suppressing inflammation [J]. Journal of Southern Medical University, 2024, 44(7): 1256-1265.
[10]	Wenxiang ZHANG, Huixian GU, Pengde CHEN, Siyu WU, Hongyan MA, Lan YAO. Compound Yuye Decoction protects diabetic rats against cardiomyopathy by inhibiting myocardial apoptosis and inflammation via regulating the PI3K/Akt signaling pathway [J]. Journal of Southern Medical University, 2024, 44(7): 1306-1314.
[11]	Yan HUANG, Lulu QIN, Shaoxing GUAN, Yanping GUANG, Yuru WEI, Ailing CAO, Dongmei LI, Guining WEI, Qibiao SU. Therapeutic mechanism of aqueous extract of Semiliquidambar cathayensis Chang root for pancreatic cancer: the active components, therapeutic targets and pathways [J]. Journal of Southern Medical University, 2024, 44(7): 1336-1344.
[12]	Yuanguo WANG, Peng ZHANG. Ferroptosis suppressor genes are highly expressed in esophageal cancer to inhibit tumor cell ferroptosis [J]. Journal of Southern Medical University, 2024, 44(7): 1389-1396.
[13]	Zhijun REN, Jianxin DIAO, Yiting WANG. Xionggui Decoction alleviates heart failure in mice with myocardial infarction by inhibiting oxidative stress-induced cardiomyocyte apoptosis [J]. Journal of Southern Medical University, 2024, 44(7): 1416-1424.
[14]	Huaxing HE, Lulin LIU, Yingyin LIU, Nachuan CHEN, Suxia SUN. Sodium butyrate and sorafenib synergistically inhibit hepatocellular carcinoma cells possibly by inducing ferroptosis through inhibiting YAP [J]. Journal of Southern Medical University, 2024, 44(7): 1425-1430.
[15]	Zhixian REN, Beixian ZHOU, Linxin WANG, Jing LI, Rongping ZHANG, Xiping PAN. Inhibitory effect of 5-hydroxy-6,7-dimethoxyflavone on H1N1 influenza virus-induced ferroptosis and inflammation in A549 cells and its possible mechanisms [J]. Journal of Southern Medical University, 2024, 44(6): 1070-1078.