肝脏作为人体最大的代谢器官，通过糖原代谢、解毒功能及蛋白质合成等多重机制维持内环境的稳态。急性肝损伤（Acute Liver Injury，ALI）以短期内大量肝细胞坏死为特征，其中药物性肝损伤（Drug-Induced Liver Injury，DILI）占比高达50%，而对乙酰氨基酚过量更是DILI的主要诱因。尽管肝移植是目前终末期肝病唯一有效的治疗方法，但供体短缺及术后免疫排斥问题迫使研究者亟需开发基于病理机制的新型治疗策略。近年来，代谢重编程在器官保护中的作用逐渐受到关注，而糖原作为肝脏能量储备的核心分子，其在ALI中的动态变化及调控机制尚未明确。本研究旨在探索急性肝损伤中肝糖原耗竭的潜在原因及其对疾病发展的影响，并通过各种细胞和动物模型探讨促进肝糖原积累如何能够保护急性肝损伤，从而为该疾病提供全新的治疗思路。本研究采用实验模型构建、糖原代谢分析、分子机制解析与体内动物模型验证等方法进行深入探讨。

为探究肝糖原与ALI之间的调控机制，本研究建立对乙酰氨基酚（Acetaminophen，APAP）与硫代乙酰胺（Thioacetamide，TAA）诱导的ALI模型，通过血清生化检测相关指标以及组织病理学实验进行分析。结果显示，在急性肝损伤小鼠中观察到区域性的糖原耗竭现象。为进一步了解这一现象与实质细胞损伤之间的关系，本研究通过构建糖原磷酸化酶L（Glycogen phosphorylase L，Pygl）敲低的小鼠原代肝实质细胞糖原积累模型，并用APAP处理该细胞，以模拟氧化应激状态下的急性肝损伤过程。结果表明，糖原积累显著提高小鼠原代肝实质细胞在APAP引发氧化应激条件下的存活能力。通过RNA测序并进行通路富集分析发现，与对照组相比，该细胞模型中涉及蛋白质合成相关生物代谢进程发生显著差异。为进一步探究糖原积累与细胞蛋白质合成之间的联系，通过检测新生蛋白质含量，发现糖原积累导致新生蛋白质合成水平显著抑制，同时，多胺代谢相关蛋白转录水平也出现下降。这一系列实验表明，在APAP诱导氧化应激条件下抑制细胞蛋白质合成能够增强小鼠原代肝实质细胞对损伤的抵抗力。鉴于当前研究认为哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin，mTOR）通路通常调控着细胞蛋白质合成，因此进一步探究抑制mTOR通路与抵抗能力之间关系。在使用雷帕霉素(Rapamycin)处理小鼠原代肝实质细胞的后，有效地抑制mTOR通路激活，从而间接降低蛋白质合成水平，这也提升APAP处理状态下细胞存活率。这些结果提示，糖原积累可能会抑制小鼠肝实质细胞mTOR通路激活，从而抑制细胞蛋白质合成，起到损伤保护作用。在得出初步结论后，本研究进行进一步的体内实验验证。通过构建肝脏靶向敲低或全身敲除Pygl的小鼠模型，诱导其肝糖原积累，结果显示小鼠在肝脏糖原积累后，显著缓解由APAP与TAA引起的急性肝损伤。同时，与蛋白质合成相关的mTOR通路相关蛋白的激活也受到明显抑制。此外基于这一机制，本研究应用糖原磷酸化酶抑制剂Ingliforib，以抑制小鼠肝脏中的糖原分解，从而促进其糖原积累，并成功治疗由APAP与TAA诱导的急性肝损伤。这些发现为未来针对急性肝损伤的治疗提供新的思路。

总之，本研究发现在APAP与TAA诱导损伤的小鼠肝糖原出现明显的区域性耗竭的现象。进一步研究证明，糖原可通过抑制mTOR磷酸化，从而抑制细胞蛋白质合成，增强肝实质细胞抵御急性肝损伤能力。通过应用糖原分解抑制剂Ingliforib来模拟糖原的积累，发现其显著减轻APAP和TAA引起的肝脏损伤。本研究通过构建药物或化学诱导的急性肝损伤模型，探讨肝脏中糖原积累在此类损伤代谢途径中的作用，揭示糖原通过抑制蛋白质合成对药物或化学物质引发的急性肝损伤所具有的新型保护机制，并为临床提供潜在的治疗方案。

The liver, as the largest metabolic organ in the human body, maintains the homeostasis of the internal environment through multiple mechanisms such as glycogen metabolism, detoxification function, and protein synthesis. Acute liver injury (ALI) is characterized by massive hepatocyte necrosis within a short period of time. Among them, drug-induced liver injury (DILI) accounts for up to 50%, and excessive acetaminophen is the main cause of DILI. Although liver transplantation is currently the only effective treatment for end-stage liver disease, the shortage of donors and postoperative immune rejection problems have forced researchers to urgently develop new treatment strategies based on pathological mechanisms. In recent years, the role of metabolic reprogramming in organ protection has gradually attracted attention. However, the dynamic changes and regulatory mechanisms of glycogen, a core molecule of liver energy reserves, in ALI remain unclear. This study aims to explore the potential causes of liver glycogen depletion in acute liver injury and its impact on disease progression, and to investigate through various cell and animal models how promoting liver glycogen accumulation can protect against acute liver injury, thereby providing a new therapeutic approach for this disease. This study employs methods such as experimental model construction, glycogen metabolism analysis, molecular mechanism interpretation, and in vivo animal model validation for in-depth exploration.

To explore the regulatory mechanism between glycogen and ALI, we established ALI models induced by acetaminophen (APAP) and thioacetamide (TAA). We analyzed relevant indicators through serum biochemical tests and histopathological experiments. The results showed a regional depletion of glycogen in mice with acute liver injury. To further understand the relationship between this phenomenon and hepatocyte damage, we constructed a glycogen phosphorylase L (Pygl) knockdown model for primary mouse hepatocytes to assess glycogen accumulation, treating these cells with APAP to simulate the process of acute liver injury under oxidative stress conditions. The results indicated that glycogen accumulation significantly enhanced the survival capacity of primary mouse hepatocytes under oxidative stress induced by APAP. RNA sequencing followed by pathway enrichment analysis revealed significant differences in protein synthesis-related biological metabolic processes compared to the control group within this cell model. To further investigate the connection between glycogen accumulation and cellular protein synthesis, we measured newly synthesized protein levels, finding that glycogen accumulation led to a significant inhibition of new protein synthesis while also decreasing transcription levels of polyamine metabolism-related proteins. This series of experiments suggests that inhibiting cellular protein synthesis under oxidative stress conditions induced by APAP can enhance the resistance of primary mouse hepatocytes to damage. Given current research indicating that mammalian target of rapamycin (mTOR) pathway typically regulates cellular protein synthesis, we further explored the relationship between mTOR pathway inhibition and resistance capability. After treating primary hepatocytes from mice with Rapamycin, the activation of the mTOR pathway was effectively inhibited, thereby indirectly reducing protein synthesis levels and enhancing cell survival under APAP treatment conditions. These results suggest that glycogen accumulation may inhibit the activation of the mTOR pathway in mouse hepatocytes, thus suppressing cellular protein synthesis and providing a protective effect against damage. Following these preliminary conclusions, we conducted further in vivo experimental validation. By constructing liver-targeted knockdown or systemic knockout models of Pygl to induce hepatic glycogen accumulation, it was found that mice exhibited significant alleviation of acute liver injury induced by APAP and TAA after hepatic glycogen accumulation. Additionally, the activation of proteins related to the mTOR pathway associated with protein synthesis was also significantly suppressed. Furthermore, based on this mechanism, we applied the glycogen phosphorylase inhibitor Ingliforib to inhibit glycogen breakdown in mouse livers, thereby promoting glycogen accumulation and successfully treating acute liver injury induced by APAP and TAA. These findings provide new insights for future treatments targeting acute liver injury.

In summary, we found that in the livers of mice with APAP and TAA-induced injury, there was a significant regional depletion of liver glycogen. Further research demonstrated that glycogen can inhibit mTOR phosphorylation, thereby suppressing cellular protein synthesis and enhancing the ability of hepatocytes to resist acute liver injury. By applying the glycogenolysis inhibitor Ingliforib to simulate glycogen accumulation, it was found that it significantly alleviated liver injury caused by APAP and TAA. This study constructed acute liver injury models induced by drugs or chemicals to explore the role of glycogen accumulation in the metabolic pathways of such injuries, revealing a novel protective mechanism of glycogen against acute liver injury caused by drugs or chemicals through inhibiting protein synthesis, and providing potential therapeutic options for clinical practice.