高血糖促单纯性脂肪肝向非酒精性脂肪性肝炎转变的线粒体机制及花色苷干预研究

Hyperglycemia will accelerate nonalcoholic fatty liver disease (NAFLD) progression from benign simple steatosis to steatohepatitis (NASH), the underlying mechanisms remain unclear. In our previous studies, anthocyanins were shown to inhibit high-glucose-induced reactive oxygen species generation, promote fatty acid oxidation, and reduce lipid deposition in hepatocytes. Here, we hypothesized that anthocyanin can protect against hyperglycemia-induced metabolic liver injury in NAFLD by improving mitochondrial function. Firstly, different NAFLD animal models will be used to determine the influence of anthoycanin on hepatic mitochondrial function and related biomarkers during progressive hyperglycemia. Then, the protective role of anthocyanin on peripheral blood mononuclear cell mitochondrial function and liver cell apoptosis will be evaluated in prediabetic NAFLD patients. Concerning the molecular action and mechanism, effects of anthocyanin on the signaling pathways required to maintain and repair mitochondrial function will also be investigated in primary hepatocytes from NAFLD mice and fatty aid-incubated human hepatocyte strain L-02, two cellular models of steatosis. This study will provide novel insights into the pathogenic mechanisms of hyperglycemia-induced transformation of simple steatosis to NASH, and lay a theoretical foundation for the clinical application of anthocyanins in the treatment of NAFLD.

高血糖会加速非酒精性脂肪性肝病（NAFLD）由单纯性脂肪肝向脂肪性肝炎转变，其中的机制尚不明确。我们前期的研究发现，花色苷可以抑制高糖孵育肝细胞内自由基产生，促进脂肪酸氧化，减少脂质沉积。本项目提出花色苷通过改善线粒体功能保护高血糖所致NAFLD代谢性肝损伤的科学假设。首先利用不同NAFLD动物模型，观察花色苷对渐进性血糖升高过程中肝脏线粒体功能及相关生物标志物的影响。进而在NAFLD伴糖耐量减低患者分析花色苷干预前后外周血单核细胞线粒体功能和肝细胞凋亡因子的变化，明确花色苷对线粒体功能和肝损伤的保护作用。然后利用NAFLD小鼠原代肝细胞和体外诱导脂肪变性肝细胞，研究花色苷对线粒体功能维持和修复关键信号通路的影响，探讨花色苷改善高糖所致线粒体功能障碍的作用机制。研究将对高血糖促单纯性脂肪肝向脂肪性肝炎转变的致病机理提出新的解释，为NAFLD的早期防治和花色苷的临床应用奠定理论基础。

高血糖会加速非酒精性脂肪性肝病（NAFLD）由单纯性脂肪肝向脂肪性肝炎（NASH）转变，其中的机制尚不明确。本项目旨在通过动物和细胞模型观察高糖应激引起的线粒体功能障碍对NAFLD病程进展的影响，结合人群干预试验探讨花色苷对线粒体功能障碍和肝损伤的保护作用。首先利用高脂饮食诱导建立小鼠NAFLD模型，分离原代肝细胞，高糖（35 mmol/L）孵育12 h后，细胞活力下降，凋亡显著增加，表现为染色质浓缩，线粒体膜电位下降，线粒体细胞色素C释放到细胞质，促进凋亡相关蛋白表达水平升高，而花色苷预处理能够减少线粒体自由基积聚，维持线粒体正常形态和功能，抑制胞浆JNK MAPK磷酸化并促进PI3K活化，保持细胞活力，减少凋亡。在高脂饲料和胆碱-蛋氨酸缺乏饲料喂养NAFLD小鼠，给予花色苷提取物（1g 花色苷/kg饲料）预防性干预16周或者4周，与对照组动物相比，花色苷能够有效改善小鼠肝脏线粒体氧化磷酸化功能，减轻肝脏的脂肪变性、气球样变和炎症细胞浸润等病变程度。此外，项目按照CONSORT临床研究规范，以花色苷提取物（花色苷320 mg/d）和安慰剂胶囊作为受试物，对74名中度NAFLD患者进行了为期12周的随机、双盲、对照干预试验，与对照组相比，花色苷干预组血浆丙氨酸转氨酶、角蛋白-18 M30 片段和髓过氧化物酶含量/活性显著降低，且葡萄糖耐量有所改善。综上所述，高糖应激会诱发肝细胞线粒体功能障碍，促使细胞凋亡，花色苷可以通过减少自由基积聚，维持线粒体氧化磷酸化功能，防止肝细胞凋亡和炎性细胞浸润，从而减缓单纯性脂肪肝向NASH转变。这些研究结果可以为NAFLD及其并发症的早期防治和花色苷的临床应用奠定理论基础。