背景:随着人们对中药和食品安全性的认识不断提高,以及对花椒DC的药理活性和毒性的深入研究。(ZADC),已经发现ZADC具有肝毒性。然而,毒性物质的基础和作用机制尚未完全阐明。羟基-α-sanshool(HAS)属于ZADC果实中的酰胺化合物,可能有肝毒性.然而,HAS的具体影响,包括肝脏毒性,不清楚。
目的:本研究的目的是确定HAS如何影响肝脏脂质代谢,确定HAS积累肝脏脂质的潜在机制,并为HAS的安全管理提供保证。
方法:通过用各种剂量的HAS(5、10和20mg/kg)给C57BL/6J小鼠进行体内实验。生化指标测定,并进行组织学分析以评估HAS肝毒性。使用脂质指数和油红O(ORO)染色确定肝脏脂质水平。在体外用不同浓度的HAS处理HepG2细胞后,通过生化分析和ORO染色确定细胞内脂质含量。线粒体膜电位,呼吸链复合酶,和ATP水平通过线粒体的荧光标记来评估。使用蛋白质印迹法测定与脂肪生成和分解代谢有关的蛋白质水平。
结果:与对照组相比,HAS组小鼠血丙氨酸和天冬氨酸转氨酶水平升高,肝脏指数升高。病理结果为肝细胞坏死。血清和肝脏甘油三酯水平,总胆固醇,低密度脂蛋白胆固醇水平升高,而高密度脂蛋白胆固醇水平下降。ORO染色结果显示肝脏脂质水平升高。体外实验表明,HAS组甘油三酯和总胆固醇水平显着升高。ATP,呼吸链复合酶基因表达,线粒体膜电位,HAS组线粒体数量减少。脂质合成相关蛋白的水平(ACC,FASN,和SREBP-1c)增加,体内和体外脂质分解代谢相关蛋白水平(PPARα和CPT1)和p-AMPK/AMPK比值均降低。
结论:HAS具有肝毒性作用,通过抑制AMPK信号通路诱导脂肪酸合成和线粒体功能损伤,导致异常脂质增加。
BACKGROUND: With the increasing awareness of the safety of traditional Chinese medicine and food, as well as in-depth studies on the pharmacological activity and toxicity of Zanthoxylum armatum DC. (ZADC), it has been found that ZADC is hepatotoxic. However, the toxic substance basis and mechanism of action have not been fully elucidated. Hydroxy-α-sanshool (HAS) belongs to an amide compound in the fruits of ZADC, which may be hepatotoxic. However, the specific effects of HAS, including liver toxicity, are unclear.
OBJECTIVE: The objectives of this research was to determine how HAS affects hepatic lipid metabolism, identify the mechanism underlying the accumulation of liver lipids by HAS, and offer assurances on the safe administration of HAS.
METHODS: An in vivo experiment was performed by gavaging C57 BL/6 J mice with various dosages of HAS (5, 10, and 20 mg/kg). Biochemical indexes were measured, and histological analysis was performed to evaluate HAS hepatotoxicity. Hepatic lipid levels were determined using lipid indices and oil red O (ORO) staining. Intracellular lipid content were determined by biochemical analyses and ORO staining after treating HepG2 cells with different concentrations of HAS in vitro. Mitochondrial membrane potential, respiratory chain complex enzymes, and ATP levels were assessed by fluorescence labeling of mitochondria. The levels of proteins involved in lipogenesis and catabolism were determined using Western blotting.
RESULTS: Mice in the HAS group had elevated alanine and aspartate aminotransferase blood levels as well as increased liver index compared with the controls. The pathological findings showed hepatocellular necrosis. Serum and liver levels of triglycerides, total cholesterol, and low-density lipoprotein cholesterol levels were increased, whereas high-density lipoprotein cholesterol levels decreased. The ORO staining findings demonstrated elevated liver lipid levels. In vitro experiments demonstrated a notable elevation in triglyceride and total cholesterol levels in the HAS group. ATP, respiratory chain complex enzyme gene expression, mitochondrial membrane potential, and mitochondrial number were reduced in the HAS group. The levels of lipid synthesis-associated proteins (ACC, FASN, and SREBP-1c) were increased, and lipid catabolism-associated protein levels (PPARα and CPT1) and the p-AMPK/AMPK ratio were decreased in vivo and in vitro.
CONCLUSIONS: HAS has hepatotoxic effects, which can induce fatty acid synthesis and mitochondrial function damage by inhibiting the AMPK signaling pathway, resulting in aberrant lipid increases.