Kouam Arnaud F, Yuan Fei, Njayou Frédéric N, He Hongtao, Tsayem Roméo F, Oladejo Babayemi O, Song Fuhang, Moundipa Paul F, Gao George F
Laboratory of Pharmacology and Toxicology, Department of Biochemistry, Faculty of Science, University of Yaoundé 1Yaoundé, Cameroon.
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of SciencesBeijing, China.
Front Pharmacol. 2017 Sep 19;8:653. doi: 10.3389/fphar.2017.00653. eCollection 2017.
Drug-induced liver injury (DILI) is a major clinical problem where natural compounds hold promise for its abrogation. (Meliaceae) is used in Cameroonian traditional medicine for the treatment of liver related diseases and has been studied for its hepatoprotective properties. Till date, reports showing the hepatoprotective molecular mechanism of the plant are lacking. The aim of this study was therefore to identify compounds from the plant bearing hepatoprotective activity and the related molecular mechanism by assessing their effects against acetaminophen (APAP)-induced hepatotoxicity in normal human liver L-02 cells line. The cells were exposed to APAP (10 mM) or co-treated with phytochemical compounds (40 μM) over a period of 36 h and, biochemical and molecular parameters assessed. Three known limonoids namely 17-epi-methyl-6-hydroxylangolensate, 7-deacetoxy-7-oxogedunin and deacetoxy-7R-hydroxygedunin were identified. The results of cells viability and membrane integrity, reactive oxygen species generation and lipid membrane peroxidation assays, cellular glutathione content determination as well as expression of cytochrome P450 2E1 demonstrated the protective action of the limonoids. Immunoblotting analysis revealed that limonoids inhibited APAP-induced c-Jun N-terminal Kinase phosphorylation (p-JNK), mitochondrial translocation of p-JNK and Bcl-associated X Protein, and the release of Apoptosis-inducing Factor into the cytosol. Interestingly, limonoids increased the expression of Mitogen-activated Protein Kinase Phosphatase (Mkp)-1, an endogenous inhibitor of JNK phosphorylation and, induced the nuclear translocation of Nuclear Factor Erythroid 2-related Factor-2 (Nrf2) and decreased the expression of Kelch-like ECH-associated Protein-1. The limonoids also reversed the APAP-induced decreased mRNA levels of Catalase, Superoxide Dismutase-1, Glutathione--Transferase and Methionine Adenosyltransferase-1A. The obtained results suggest that the isolated limonoids protect L-02 hepatocytes against APAP-induced hepatotoxicity mainly through increase expression of Mkp-1 and nuclear translocation of Nrf2. Thus, these compounds are in part responsible of the hepatoprotective activity of and further analysis including and toxicological studies are needed to select the most potent compound that may be useful as therapeutic agents against DILI.
药物性肝损伤（DILI）是一个重大临床问题，天然化合物有望消除这一问题。楝科植物在喀麦隆传统医学中用于治疗肝脏相关疾病，并已对其保肝特性进行了研究。迄今为止，缺乏显示该植物保肝分子机制的报告。因此，本研究的目的是通过评估其对正常人类肝脏L-02细胞系中对乙酰氨基酚（APAP）诱导的肝毒性的影响，从该植物中鉴定具有保肝活性的化合物及其相关分子机制。将细胞暴露于APAP（10 mM）或与植物化学化合物（40 μM）共同处理36小时，然后评估生化和分子参数。鉴定出三种已知的柠檬苦素，即17-表甲基-6-羟基兰戈伦酸酯、7-脱乙酰氧基-7-氧代格杜宁和脱乙酰氧基-7R-羟基格杜宁。细胞活力和膜完整性、活性氧生成和脂质膜过氧化测定、细胞内谷胱甘肽含量测定以及细胞色素P450 2E1表达的结果证明了柠檬苦素的保护作用。免疫印迹分析显示，柠檬苦素抑制APAP诱导的c-Jun N端激酶磷酸化（p-JNK）、p-JNK的线粒体转位和Bcl相关X蛋白，以及凋亡诱导因子释放到细胞质中。有趣的是，柠檬苦素增加了丝裂原活化蛋白激酶磷酸酶（Mkp）-1的表达，Mkp-1是JNK磷酸化的内源性抑制剂，并诱导核因子红系2相关因子-2（Nrf2）的核转位，降低了类Kelch样ECH相关蛋白-1的表达。柠檬苦素还逆转了APAP诱导的过氧化氢酶、超氧化物歧化酶-1、谷胱甘肽-S-转移酶和甲硫氨酸腺苷转移酶-1A的mRNA水平降低。所得结果表明，分离出的柠檬苦素主要通过增加Mkp-1的表达和Nrf2的核转位来保护L-02肝细胞免受APAP诱导的肝毒性。因此，这些化合物部分负责该植物的保肝活性，需要进一步分析，包括体内研究和毒理学研究，以选择最有效的化合物，其可能用作抗DILI的治疗剂。
