Protective effect of Shenge Formula against liver injury in alcoholic liver disease in mice and its related mechanisms

Huang Zhili; Feng Junhua; Cao Haixia; Li Qian; Ji Longshan; Liang Xiaohui; Gao Yueqiu; Li Man

doi:10.16305/j.1007-1334.2026.z20251216001

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Protective effect of Shenge Formula against liver injury in alcoholic liver disease in mice and its related mechanisms

更新时间：2026-07-03

- Protective effect of Shenge Formula against liver injury in alcoholic liver disease in mice and its related mechanisms
- Shanghai Journal of Traditional Chinese Medicine Vol. 60, Issue 7, Pages: 60-73(2026)
- 作者机构：
  
  1.上海中医药大学附属曙光医院细胞免疫实验室（上海 201203）
  2.上海市中医药研究院疫病研究所（上海 201203）
  3.上海中医药大学附属曙光医院肝病科（上海 201203）
  4.上海交通大学医学院附属新华医院（上海 200092）
- 作者简介：
- 基金信息：
- DOI：10.16305/j.1007-1334.2026.z20251216001
  CLC：
- Received：16 December 2025，
  
  Published：10 July 2026
- 稿件说明：
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黄知礼,冯俊华,曹海霞,等.参葛方对酒精性肝病小鼠肝脏损伤的保护作用及其相关机制[J].上海中医药杂志,2026,60(7):60-73.

Huang Zhili,Feng Junhua,Cao Haixia,et al.Protective effect of Shenge Formula against liver injury in alcoholic liver disease in mice and its related mechanisms[J].Shanghai Journal of Traditional Chinese Medicine,2026,60(7):60-73.
黄知礼,冯俊华,曹海霞,等.参葛方对酒精性肝病小鼠肝脏损伤的保护作用及其相关机制[J].上海中医药杂志,2026,60(7):60-73. DOI： 10.16305/j.1007-1334.2026.z20251216001.

Huang Zhili,Feng Junhua,Cao Haixia,et al.Protective effect of Shenge Formula against liver injury in alcoholic liver disease in mice and its related mechanisms[J].Shanghai Journal of Traditional Chinese Medicine,2026,60(7):60-73. DOI： 10.16305/j.1007-1334.2026.z20251216001.

摘要

目的

阐明参葛方（SGF）通过上调肝细胞脂联素受体1（AdipoR1）-钙调蛋白依赖性蛋白激酶激酶2（CaMKK2）-腺苷酸活化蛋白激酶（AMPK）信号通路治疗酒精性肝病（ALD）的作用机制。

方法

①采用网络药理学方法筛选SGF治疗ALD的活性成分及作用靶点，进而系统分析其潜在的治疗靶点与相关功能通路。②30只SPF级健康雄性C57BL/6J小鼠随机分为对照组（Control）、模型组（Model）、SGF低剂量组（SGF-L，7.95 mg/g）、SGF中剂量组（SGF-M，15.9 mg/g）、SGF高剂量组（SGF-H，31.8 mg/g），每组6只。除对照组外，其余各组建立ALD小鼠模型（NIAAA模型），用药组在造模的同时分别给予相应药物灌胃，而对照组与模型组则给予等体积0.9%氯化钠溶液，共干预10 d。采用生化分析仪检测小鼠血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）和甘油三酯（TG）水平及小鼠肝组织超氧化物歧化酶（SOD）、丙二醛（MDA）水平；实时荧光定量逆转录PCR（RT-qPCR）检测小鼠肝组织炎症和纤维化相关基因表达水平；Western blot法检测小鼠肝组织AdipoR1、CaMKK2、AMPK蛋白表达；苏木精-伊红（HE）、油红O染色和免疫组织化学法检测小鼠肝组织病理变化、脂质沉积及炎症细胞浸润情况。③以无水乙醇干预HepG2细胞，建立ALD细胞模型；并分别运用si-AdipoR1、Dorsomorphin（AMPK抑制剂）和SGF大鼠药物血清干预，验证SGF治疗ALD的作用靶点。

结果

①网络药理学筛选出CaMKK2是SGF治疗ALD的潜在靶点，主要涉及AMPK以及钙离子相关信号通路。②与模型组相比，各用药组小鼠血清ALT、AST、TG水平均明显降低，肝脏促炎性细胞因子和胶原相关因子的 mRNA水平均降低；肝脏炎症和脂质沉积均明显改善。③体外应用无水乙醇干预HepG2细胞可增加细胞脂质沉积，抑制AdipoR1、CaMKK2和p-AMPK蛋白表达，SGF药物血清干预可上调AdipoR1、CaMKK2和p-AMPK蛋白表达，减少脂质沉积；si-AdipoR1可抑制SGF对脂质沉积的改善作用以及SGF对AdipoR1、CaMKK2和AMPK的激活作用；Dorsomorphin可抑制SGF对脂质沉积的改善作用以及SGF对AMPK的激活作用。

结论

SGF通过激活AdipoR1-CaMKK2-AMPK信号通路，发挥治疗ALD的作用。

Abstract

Objective

To elucidate the mechanism of Shenge Formula （SGF） in the treatment of alcoholic liver disease （ALD） by up-regulating the adiponectin receptor 1 （AdipoR1）-calcium/calmodulin-dependent protein kinase kinase 2 （CaMKK2）-adenosine 5′-monophosphate-activated protein kinase （AMPK） signaling pathway in hepatocytes.

Methods

① Network pharmacology was employed to screen the active components and potential targets of SGF for ALD treatment， followed by systematic analysis of its core therapeutic targets and related functional pathways. ② Thirty specific pathogen-free （SPF） healthy male C57BL/6J mice were randomly divided into five groups （

=6 per group）： control group， model group， low-dose SGF group （SGF-L， 7.95 mg/g）， medium-dose SGF group （SGF-M， 15.9 mg/g）， and high-dose SGF group （SGF-H， 31.8 mg/g）. Except for the control group， a mouse model of ALD （NIAAA） was established in all other groups. Concurrently with modeling， the treatment groups were administrated with corresponding drugs via gavage， while the control and model groups were given an equal volume of 0.9% sodium chloride solution for 10 consecutive days. Serum levels of alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， and triglyceride （TG）， as well as hepatic levels of superoxide dismutase （SOD） and malondialdehyde （MDA）， were measured. The mRNA expression levels of inflammation- and fibrosis-related genes in liver tissues were detected by reverse transcription quantitative real-time polymerase chain reaction （RT-qPCR）. Protein expression levels of AdipoR1， CaMKK2，

and AMPK in liver tissues were determined by Western blot analysis. Hepatic pathological changes， lipid accumulation， and inflammatory cell infiltration were assessed by hematoxylin-eosin （HE） staining， Oil Red O staining， and immunohistochemistry.③ An

in vitro

ALD cell model was constructed by treating HepG2 cells with absolute ethanol. Cells were further treated with SGF-containing rat serum， combined with AdipoR1 siRNA （si-AdipoR1） and Dorsomorphin （an AMPK inhibitor）， to verify the key targets of SGF in ALD treatment.

Results

① Network pharmacology identified CaMKK2 as a putative target of SGF against ALD， which was mainly involved in the AMPK and calcium-related signaling pathways. ② Compared with the model group， serum ALT， AST， and TG levels were significantly decreased in all SGF-treated groups， accompanied by down-regulated mRNA levels of hepatic pro-inflammatory factors and collagen-related factors， as well as markedly alleviated hepatic inflammation and lipid accumulation.③

In vitro

stimulation of HepG2 cells with absolute ethanol promoted intracellular lipid deposition and suppressed the protein expression of AdipoR1， CaMKK2， and phosphorylated AMPK （p-AMPK）. Treatment with SGF-containing serum up-regulated AdipoR1， CaMKK2， and p-AMPK expression levels and reduced lipid accumulation. Transfection with si-AdipoR1 abolished the ameliorative effect of SGF on lipid deposition and its activation of AdipoR1， CaMKK2， and AMPK. Dorsomorphin abrogated SGF-mediated improvements in lipid deposition and SGF-triggered AMPK activation.

Conclusion

SGF exerts therapeutic effects against ALD via activation of the AdipoR1-CaMKK2-AMPK signaling pathway.

关键词

Keywords

references

Jophlin L L ， Singal A K ， Bataller R ， et al . ACG Clinical Guideline： Alcohol-Associated Liver Disease ［J］. Am J Gastroenterol ， 2024 ， 119 （ 1 ）： 30 - 54 .

Feng X ， Huang N ， Wu Y ， et al . Alcoholic liver disease in China： A disease influenced by complex social factors that should not be neglected ［J］. J Clin Transl Hepatol ， 2024 ， 12 （ 7 ）： 677 - 684 .

Seitz H K ， Bataller R ， Cortez-Pinto H ， et al . Alcoholic liver disease ［J］. Nat Rev Dis Primers ， 2018 ， 4 （ 1 ）： 16 .

Avila M A ， Dufour J F ， Gerbes A L ， et al . Recent advances in alcohol-related liver disease （ALD）： summary of a Gut round table meeting ［J］. Gut ， 2020 ， 69 （ 4 ）： 764 - 780 .

Gao B ， Bataller R . Alcoholic liver disease： pathogenesis and new therapeutic targets ［J］. Gastroenterology ， 2011 ， 141 （ 5 ）： 1572 - 1585 .

Hawley S A ， Pan D A ， Mustard K J ， et al . Calmodulin-dependent protein kinase kinase-beta is an alternative upstream kinase for AMP-activated protein kinase ［J］. Cell Metab ， 2005 ， 2 （ 1 ）： 9 - 19 .

Miao Z ， Sun Y ， Feng Z ， et al . CAMKK2-AMPK axis endows dietary calcium and phosphorus levels with regulatory effects on lipid metabolism in weaned piglets ［J］. J Anim Sci Biotechnol ， 2024 ， 15 （ 1 ）： 105 .

Hardie D G . AMP-activated protein kinase： maintaining energy homeostasis at the cellular and whole-body levels ［J］. Annu Rev Nutr ， 2014 ， 34 ： 31 - 55 .

Jiang Z ， Zhou J ， Zhou D ， et al . The adiponectin-SIRT1-AMPK pathway in alcoholic fatty liver disease in the rat ［J］. Alcohol Clin Exp Res ， 2015 ， 39 （ 3 ）： 424 - 433 .

Gopal T ， Ai W ， Casey C A ， et al . A review of the role of ethanol-induced adipose tissue dysfunction in alcohol-associated liver disease ［J］. Alcohol Clin Exp Res ， 2021 ， 45 （ 10 ）： 1927 - 1939 .

高月求，许小莉，孙学华，等 . 参葛方联合行为干预治疗非酒精性脂肪性肝病的临床研究［C］//中华中医药学会内科肝胆病专业委员会. 中华中医药学会第十五届内科肝胆病学术会议暨国家中医药管理局专科专病协作组（肝病组、传染病组）会议论文汇编 . 济南：中华中医药学会内科肝胆病专业委员会， 2012 ： 516 - 520 .

齐艳平，李曼，孙学华，等 . 参葛方防治大鼠非酒精性脂肪肝的实验研究［J］. 中国实验方剂学杂志， 2012 ， 18 （ 12 ）： 156 - 160 .

张云城，胡世平 . 《金匮要略》对黄疸病的认识及其在临床上的应用［J］. 中国医学创新， 2017 ， 14 （ 5 ）： 93 - 96 .

高斌，常彬霞，徐明江 . 慢性酒精喂养加急性酒精灌胃的酒精性肝病小鼠模型（NIAAA模型或Gao-Binge模型）［J］. 传染病信息， 2013 ， 26 （ 5 ）： 307 - 311 .

Bertola A ， Mathews S ， Ki S H ， et al . Mouse model of chronic and binge ethanol feeding （the NIAAA model）［J］. Nat Protoc ， 2013 ， 8 （ 3 ）： 627 - 637 .

Gong P ， Long H ， Guo Y ， et al . Chinese herbal medicines： The modulator of nonalcoholic fatty liver disease targeting oxidative stress ［J］. J Ethnopharmacol ， 2024 ， 318 （ Pt B ）： 116927 .

方淼，张荣杰，房华，等 . 参葛方联合生活饮食运动干预治疗肝功能异常代谢相关脂肪性肝病的临床观察［J］. 上海中医药杂志， 2024 ， 58 （ 8 ）： 41 - 44 .

Mcaloon L M ， Muller A G ， Nay K ， et al . CaMKK2： bridging the gap between Ca 2+ signaling and energy-sensing ［J］. Essays Biochem ， 2024 ， 68 （ 3 ）： 309 - 320 .

Dewenter M ， Neef S ， Vettel C ， et al . Calcium/Calmodulin-dependent protein kinase II activity persists during chronic β‑ adrenoceptor blockade in experimental and human heart failure ［J］. Circ Heart Fail ， 2017 ， 10 （ 5 ）： e003840 .

Racioppi L ， Means A R . Calcium/calmodulin-dependent protein kinase kinase 2： roles in signaling and pathophysiology ［J］. J Biol Chem ， 2012 ， 287 （ 38 ）： 31658 - 31665 .

Chauhan A S ， Liu X ， Jing J ， et al . STIM2 interacts with AMPK and regulates calcium-induced AMPK activation ［J］. FASEB J ， 2019 ， 33 （ 2 ）： 2957 - 2970 .

Saikia R ， Joseph J . AMPK： a key regulator of energy stress and calcium-induced autophagy ［J］. J Mol Med （Berl）， 2021 ， 99 （ 11 ）： 1539 - 1551 .

Malik N ， Shaw R J . The AMPK pathway： Molecular rejuvenation of metabolism and mitochondria ［J］. Annu Rev Cell Dev Biol ， 2025 ， 41 （ 1 ）： 375 - 402 .

Zhou Q ， Gong L ， Bao G ， et al . β -Elemene improves morphine tolerance in bone cancer pain via N -methyl- D -aspartate receptor 2B subunit-mediated μ ‑opioid receptor ［J/OL］. Comput Math Methods Med ， 2022-09-17 . https：//pubmed.ncbi.nlm.nih.gov/36164617/ https://pubmed.ncbi.nlm.nih.gov/36164617/ .

Wang Y Z ， Lu J ， Li Y Y ， et al . microRNA-378b regulates ethanol-induced hepatic steatosis by targeting CaMKK2 to mediate lipid metabolism ［J］. Bioengineered ， 2021 ， 12 （ 2 ）： 12659 - 12676 .

Ortiz A R ， Nay K ， Stork B A ， et al . Myeloid-specific CAMKK2 deficiency protects against diet-induced obesity and insulin resistance by rewiring metabolic gene expression and enhancing energy expenditure ［J］. Mol Metab ， 2025 ， 101 ： 102250 .

Huang M Q ， Zhou C J ， Zhang Y P ， et al . Salvianolic acid B ameliorates hyperglycemia and dyslipidemia in db/db mice through the AMPK pathway ［J］. Cell Physiol Biochem ， 2016 ， 40 （ 5 ）： 933 - 943 .

Li Q ， Tan J X ， He Y ， et al . Atractylenolide Ⅲ ameliorates non-alcoholic fatty liver disease by activating hepatic adiponectin receptor 1-mediated AMPK pathway ［J］. Int J Biol Sci ， 2022 ， 18 （ 4 ）： 1594 - 1611 .

Chen J ， Yang S ， Luo H ， et al . Polysaccharide of atractylodes macrocephala Koidz alleviates NAFLD-induced hepatic inflammation in mice by modulating the TLR4/MyD88/NF‑κB pathway ［J］. Int Immunopharmacol ， 2024 ， 141 ： 113014 .

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