The effects of Gentiana dahurica Fisch on alcoholic liver disease revealed by RNA sequencing
The effects of Gentiana dahurica Fisch on alcoholic liver disease revealed by RNA sequencing
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RNA测序揭示白花龙胆对酒精性肝病的影响
DOI：
10.1016/j.jep.2020.113422
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发表时间：
2021-07-15
影响因子：
5.4
通讯作者：
Jin, Ling
中科院分区：
文献类型：
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作者：
Cao, Houkang;Xi, Shaoyang;Jin, Ling
民族药理学相关性：达乌里秦艽（在中国称为秦艽）的根是一种传统中药，在中国用于治疗酒精性肝病（ALD），但目前尚无关于其治疗ALD的科学报道。
研究目的：研究达乌里秦艽乙醇提取物（GDEE）对ALD的治疗效果，并利用RNA测序揭示其可能的作用机制。
材料和方法：通过对小鼠连续灌胃酒精建立ALD模型，并用GDEE治疗ALD。进行病理观察（HE染色、油红O染色）和生化指标检测以评估肝脏组织损伤和GDEE的疗效。对肝脏组织进行RNA测序分析以阐明ALD的发病机制以及GDEE的保肝作用机制。通过实时定量聚合酶链反应（PCR）和蛋白质印迹法检测乙酰辅酶A羧化酶α（Acacα）、脂肪酸合酶（Fasn）和肉碱棕榈酰转移酶1A（Cpt1a）的mRNA和蛋白质表达来验证RNA测序结果。
结果：生化参数测量表明，GDEE能够抑制酒精引起的血清中转氨酶活性升高和肝脏中脂质水平升高。观察到GDEE能够减轻酒精在肝脏组织中引起的脂肪变性、水肿和细胞坏死。肝脏组织的RNA测序分析发现，酒精和GDEE分别使719个基因和1137个基因发生显著变化。GDEE逆转了酒精上调的大多数甘油三酯合成相关基因的变化。GDEE上调了ALD小鼠中大多数参与脂肪酸降解的基因，而酒精对它们影响较小。此外，GDEE抑制了酒精上调的大多数参与胆固醇合成的基因。GDEE上调了ALD小鼠中与胆汁酸合成相关的基因，下调了与胆汁酸重吸收相关的基因，而酒精对胆汁酸代谢相关基因无显著影响。在验证实验中，通过实时PCR和蛋白质印迹法定量的Acacα、Fasn和Cpt1a表达与RNA测序结果一致。
结论：GDEE能够减轻ALD小鼠的肝脏损伤和脂肪变性，其作用机制可能与调节甘油三酯和胆固醇的过程有关。
Ethnopharmacological relevance: The root of Gentiana dahurica Fisch (called Qin-Jiao in China), a traditional Chinese medicine, is used in China to treat alcoholic liver disease (ALD), but there has been no scientific report on the treatment of ALD. Aim of the study: To investigate the therapeutic effects of Gentiana dahurica Fisch ethanol extract (GDEE) on ALD and to reveal its possible mechanism of action using RNA sequencing. Materials and methods: The model of ALD was established by continuous gavage with alcohol in mice, and GDEE was used to treat ALD. Pathological observation (HE staining, oil red O staining) and biochemical indicators were performed to evaluate liver tissue lesions and efficacy of GDEE. RNA sequencing analysis of liver tissues was carried out to elucidate the pathogenesis of ALD and the mechanism of hepatoprotective effect by GDEE. The RNA sequencing results were verified by detecting mRNA and protein expressions of acetyl coenzyme A carboxylase alpha (Acac alpha), fatty acid synthase (Fasn) and carnitine palmitoyltransferase 1A (Cpt1a) by quantitative real-time polymerase chain reaction (PCR) and Western blot. Results: Measurements of biochemical parameters showed that GDEE could inhibit the increased transaminase activities in the serum and lipid levels in the liver caused by alcohol. It was observed that GDEE could alleviate fatty degeneration, edema and cell necrosis caused by alcohol in the liver tissue. RNA sequencing analysis of liver tissues found that 719 genes and 1137 genes were significantly changed by alcohol and GDEE, respectively. GDEE reversed most of the changes in triglycerides synthesis-related genes up-regulated by alcohol. GDEE upregulated most of the genes involved in the fatty acid degradation in ALD mice, while alcohol had little effect on them. In addition, GDEE suppressed most of the genes involved in cholesterol synthesis that were up-regulated by alcohol. GDEE up-regulated genes related to bile acid synthesis in ALD mice, and down-regulated genes related to bile acid reabsorption, while alcohol had no significant effect on genes related to bile acid metabolism. In the validation experiments, the Acac alpha, Fasn and Cpt1a expressions quantified by real-time PCR and Western blot were consistent with the RNA sequencing results. Conclusions: GDEE can alleviate liver damage and steatosis in ALD mice, and its mechanism of action may be related to the process of regulating triglycerides and cholesterol.