背景:酒精性肝病(ALD)显著导致全球肝脏相关的发病率和死亡率。天然产物在ALD的预防和治疗中起着至关重要的作用。羟基红花黄色素A(HSYA),红花的独特和主要成分(红花),表现出不同的药理活性。然而,HSYA对ALD的影响和机制尚未完全阐明。
目的:本研究的目的是采用综合药理学方法来评估HSYA抗ALD的多靶向机制。
方法:网络药理学和分子对接技术用于分析HSYA抗ALD的潜在治疗信号通路和靶点。建立了斑马鱼幼虫ALD模型。幼虫用HSYA预处理,然后暴露于乙醇。通过在肝脏特异性转基因斑马鱼品系Tg(fabp10a:DsRed)和肝脏组织H&E染色中的荧光表达分析测量肝脏损伤。肝脏脂肪变性通过整体油红O染色和TG水平确定。此外,在体外建立乙醇诱导的肝细胞损伤模型,以观察肝细胞损伤(细胞活力,ALT水平),脂质积累(油红O染色,TC和TG),和氧化应激(ROS,MDA,用或不用HSYA处理的HepG2细胞中的GPx和SOD)。最后,qRT-PCR结合网络药理学和分子对接来验证HSYA对靶标的影响。
结果:HSYA表现出显著的,乙醇诱导的斑马鱼幼虫和HepG2细胞肝损伤的剂量依赖性改善。网络药理学分析显示,HSYA可能通过341个潜在靶标对ALD发挥药理作用。这些靶标参与各种信号通路,包括脂质代谢和动脉粥样硬化,PI3K-Akt信号通路,MAPK信号通路,和ALD本身。分子对接研究表明,HSYA对IL1B的结构域具有很强的结合亲和力,IL6,TNF,PPARA,PPARG,HMGCR和ADH5。qRT-PCR实验证明HSYA能有效逆转乙醇诱导的SREBF1、FASN、ACACA,CPT1A,PPARA,IL1B,IL6,TNFα,ADH5和ALDH2在体内和体外。
结论:本研究使用综合药理学方法对HSYA的抗ALD机制进行了全面研究。HSYA的潜在靶标可能与增强乙醇分解代谢有关,减少脂质积累,缓解氧化应激,抑制炎症反应。
BACKGROUND: Alcoholic liver disease (ALD) significantly contributes to global liver-related morbidity and mortality. Natural products play a crucial role in the prevention and treatment of ALD. Hydroxysafflor yellow A (HSYA), a unique and primary component of Safflower (Carthamus tinctorius l.), exhibits diverse pharmacological activities. However, the impact and mechanism of HSYA on ALD have not been fully elucidated.
OBJECTIVE: The purpose of this study was to employ an integrative pharmacology approach to assess the multi-targeted mechanism of HSYA against ALD.
METHODS: Network pharmacology and molecular docking techniques were used to analyze the potential therapeutic signaling pathways and targets of HSYA against ALD. An ALD model in zebrafish larvae was established. Larvae were pretreated with HSYA and then exposed to ethanol. Liver injury was measured by fluorescence expression analysis in the liver-specific transgenic zebrafish line Tg (fabp10a:DsRed) and liver tissue H&E staining. Liver steatosis was determined by whole-mount oil red O staining and TG level. Additionally, an ethanol-induced hepatocyte injury model was established in vitro to observe hepatocyte damage (cell viability, ALT level), lipid accumulation (oil red O staining, TC and TG), and oxidative stress (ROS, MDA, GPx and SOD) in HepG2 cells treated with or without HSYA. Finally, qRT-PCR combined with network pharmacology and molecular docking was employed to validate the effects of HSYA on targets.
RESULTS: HSYA exhibited a significant, dose-dependent improvement in ethanol-induced liver injury in zebrafish larvae and HepG2 cells. Network pharmacology analysis revealed that HSYA may exert pharmacological effects against ALD through 341 potential targets. These targets are involved in various signaling pathways, including lipid metabolism and atherosclerosis, PI3K-Akt signaling pathway, MAPK signaling pathway, and ALD itself. Molecular docking studies displayed that HSYA had a strong binding affinity toward the domains of IL1B, IL6, TNF, PPARA, PPARG, HMGCR and ADH5. qRT-PCR assays demonstrated that HSYA effectively reversed the ethanol-induced aberrant gene expression of SREBF1, FASN, ACACA, CPT1A, PPARA, IL1B, IL6, TNFα, ADH5, and ALDH2 in vivo and in vitro.
CONCLUSIONS: This study offers a comprehensive investigation into the anti-ALD mechanisms of HSYA using an integrative pharmacology approach. The potential targets of HSYA may be implicated in enhancing ethanol catabolism, reducing lipid accumulation, mitigating oxidative stress, and inhibiting inflammatory response.