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基于烟酰胺磷酸核糖转移酶（NAMPT）探讨红景天苷在非酒精性脂肪性肝病小鼠模型中的保护作用
DOI: 10.12449/JCH240112
伦理学声明：本研究方案于2021年9月29日经由西南医科大学实验动物福利伦理委员会审批，批号：20210929-013，符合实验室动物管理与使用准则。
利益冲突声明：本文不存在任何利益冲突。
作者贡献声明：王缕、李荣军、张志红负责课题设计；王缕、张庭负责饲养动物，造模，处理动物获得标本；王缕负责Western Blot、ELISA、病理组织学相关试验；徐春霞负责资料分析；李荣军、张志红负责拟定写作思路；李荣军、王缕负责撰写论文，修改论文，最后定稿。
Role of salidroside in a mouse model of non-alcoholic fatty liver disease： A study based on nicotinamide phosphoribosyltransferase
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摘要:
目的 研究红景天苷对非酒精性脂肪性肝病（NAFLD）的保护作用，并探讨其作用机制。 方法 将24只雄性KM小鼠随机分为正常组、高脂饮食（HFD）组、HFD+空白对照组、HFD+红景天苷组，每组6只，正常组小鼠予以普通饲料喂养，其余各组高脂饮食，造模14周开始给予100 mg·kg-1·d-1红景天苷灌胃给药，22周末取材。酶联免疫法测定血清ALT、AST、TG、TC、HDL-C、LDL-C等相关生化指标；HE染色和NAFLD活动评分（NAS）观察小鼠肝组织病理。Western Blot检测肝组织内烟酰胺磷酸核糖转移酶（NAMPT）、Sirt1、AMPKα、SREBP1的表达变化。多组间比较采用单因素方差分析，进一步两两比较采用LSD-t检验。 结果 与正常组比较，HFD组小鼠肝组织存在广泛较大的脂肪滴，脂肪变性明显，NAS评分升高（P<0.01），外周血AST、ALT、TG、TC、LDL-C的含量明显增高（P值均<0.05），HDL-C含量降低（P<0.05），肝组织内NAMPT、AMPKα、Sirt1的表达降低（P值均<0.05），SERBP1的表达增加（P<0.01）。而HFD+红景天苷组较HFD组、HFD+空白对照组肝组织空泡状脂滴分布、小叶内炎症减少，肝细胞气球样变减轻，NAS评分下降（P<0.01），外周血AST、ALT、TG、LDL-C的含量明显降低（P值均<0.05），HDL-C含量升高（P<0.05），NAMPT、AMPKα、Sirt1的表达增加（P值均<0.05），SERBP1的表达减少（P<0.01）。 结论 红景天苷能明显改善高脂饮食诱导小鼠NAFLD的病理状态，并通过增加NAMPT、Sirt1、AMPKα的表达，减少SERBP1的表达，发挥其对 NAFLD的保护作用。 -
关键词:
- 烟酰胺磷酸核糖基转移酶 /
- 红景天苷 /
- 非酒精性脂肪性肝病 /
- 小鼠
Abstract:Objective To investigate the protective effect of salidroside against nonalcoholic fatty liver disease （NAFLD） and its mechanism of action. Methods A total of 24 male KM mice were randomly divided into normal group， HFD group， HFD+blank control group， and HFD+salidroside group， with 6 mice in each group. The mice in the normal group were given normal diet， and those in the other groups were given high-fat diet. After 14 weeks of modeling， the mice were given salidroside 100 mg/kg/day by gavage， and related samples were collected at the end of week 22. Enzyme-linked immunosorbent assay was used to measure the serum levels of related biochemical parameters including alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， triglyceride （TG）， total cholesterol （TC）， high-density lipoprotein cholesterol （HDL-C）， and low-density lipoprotein cholesterol （LDL-C）； HE staining and NAFLD activity score （NAS） were used to observe the liver histopathology of mice； Western blot was used to measure the changes in the expression of NAMPT， Sirt1， AMPKα， and SREBP1 in liver tissue. A one-way analysis of variance was used for comparison between multiple groups， and the least significant difference t-test was used for further comparison between two groups. Results Compared with the normal group， the HFD group had obvious steatosis and extensive large lipid droplets in liver tissue， with significant increases in NAS score （P<0.01） and the content of AST， ALT， TG， TC， and LDL-C in peripheral blood （all P<0.05） and a significant reduction in the content of HDL-C （P<0.05）， as well as significant reductions in the expression levels of NAMPT， AMPKα， and Sirt1 in liver tissue （all P<0.05） and a significant increase in the expression level of SERBP1 （P<0.01）. Compared with the HFD group and the HFD+blank control group， the HFD+salidroside group had reductions in the distribution of vacuolar lipid droplets and intralobular inflammation in liver tissue， alleviation of the ballooning degeneration of hepatocytes， significant reductions in NAS score （P<0.01） and the content of AST， ALT， TG， and LDL-C in peripheral blood （all P<0.05）， and a significant increase in the content of HDL-C （P<0.05）， as well as significant increases in the expression levels of NAMPT， AMPKα， and Sirt1 in liver tissue （all P<0.05） and a significant reduction in the expression level of SERBP1 （P<0.01）. Conclusion Salidroside can significantly improve the pathological state of mice with NAFLD induced by high-fat diet and exert a protective effect against NAFLD by increasing the expression of NAMPT， Sirt1， and AMPKα and reducing the expression of SERBP1. -
图 1 各组小鼠肝组织病理形态学变化（HE染色，×100）
Figure 1. Pathological changes of liver tissue in mice of each group（HE staining，×100）
图 3 各组小鼠肝组织NAMPT、Sirt1、AMPKα、SREBP1蛋白表达
Figure 3. The hepatic protein expression of NAMPT、Sirt1、AMPKα、SREBP1 in mice of each group
图 4 红景天苷对NAFLD小鼠肝脏NAMPT、Sirt1、AMPKα、SREBP1表达的影响
Figure 4. The effect of salidroside on the hepatic protein expression of NAMPT、Sirt1、AMPKα、SREBP1 in mice of each group
表 1 NAS评分标准
Table 1. NAS scoring criteria for liver tissue
组织学标志 评分（分） 评分标准 脂肪变性 0 <5％ 1 5％～33％ 2 33％～66％ 3 >66％ 小叶炎症 0 无 1 <2 2 2～4 3 >4 气球样变 0 无 1 很少；不明显 2 大量；显著 
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表 2 红景天苷对KM小鼠血清生化指标的影响
Table 2. The effect of Salidroside on serum biochemical indicators in KM mice
项目 正常组 （n=6） HFD组 （n=6） HFD+空白对照组（n=6） HFD+红景天苷组（n=6） F值 P值 ALT（U/L） 60.48±11.03 83.17±9.611） 85.17±9.291） 67.21±7.202）3） 9.940 <0.001 AST（U/L） 243.78±60.49 532.7±80.551） 575.25±114.491） 403.91±80.171）2）3） 17.970 <0.001 TG（mmol/L） 1.92±0.17 3.51±0.701） 3.90±0.851） 2.64±0.772）3） 5.180 <0.010 TC（mmol/L） 0.42±0.08 0.81±0.301） 0.88±0.271） 0.70±0.161） 10.270 <0.001 HDL-C（mmol/L） 4.29±0.95 2.39±0.631） 2.64±0.661） 3.63±0.562）3） 9.050 <0.05 LDL-C（mmol/L） 0.36±0.12 0.65±0.081） 0.67±0.111） 0.45±0.122）3） 11.510 <0.001 注：与正常组比较，1）P<0.05；与HFD组比较，2）P<0.05；与HFD+空白对照比较，3）P<0.05。
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表 3 各组小鼠肝组织NAS评分
Table 3. The NAS score of liver tissue in mice of each group
组别 动物数（只） 评分（分） 正常组 6 0.33±0.52 HFD组 6 6.17±0.981） HFD+空白对照组 6 5.5±1.051） HFD+红景天苷组 6 2.5±0.551）2）3） F值 67.07 P值 <0.001 注：与正常组比较，1）P<0.01；与HFD组比较，2）P<0.01；与HFD+空白对照组比较，3）P<0.01。
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