炎症信号通路及相关miRNA介导槐果碱抗非酒精性脂肪性肝炎的实验研究

The regulation of inflammatory signaling pathways and the related miRNA in NASH pathogenesis is current research focus. Our previous work confirmed that sophocarpine could inhibit the development of NASH in vivo through anti-inflammation. In this study, we would investigate the effect of sophocarpine on proliferation, apoptosis, fat degeneration in primary hepatocytes at first. The activity of important inflammatory cell signaling pathways, TLR4 and its downstream NF-κB, JNK/SAPK and its upstream regulatory element miR-140 would be also examined. We aim to prove the critical role of miR-140and TLR4 in the anti-NASH effect of sophocarpine. Thereafter, Adenovirus carrying miR-140 and TLR4 antagonist HTA 125 will be delivered into primary hepatocytes, and proliferation, apoptosis, steatosis and inflammation-related signaling pathways will be evaluated in vitro. In addition, we will comfirm that down-regulating miR-140 and activating TLR4 by LPS can alleviate the inhibiting effect of sophocarpine on NASH. Furthermore, sophocarpine or miR-140 will be delived into experimental NASH rats to evaluate their effects on NASH and inflammatory signaling pathway in vivo. Taken together, our research will further explore the mechanism of anti-NASH effect of sophocarpine and provide new therapeutic targets for NASH.

炎症信号通路及相关miRNA对非酒精性脂肪性肝炎（NASH）的调控作用是目前研究热点。前期工作证实槐果碱可在体内抑制NASH发展，其机制与抗炎作用有关。本课题观察槐果碱对原代肝细胞增殖、凋亡、脂肪变性的影响，检测TLR4及下游NF-κB和JNK/SAPK等炎症信号通路活性及其上游 miR-140表达变化，证实miR-140和TLR4在槐果碱抗NASH中发挥关键作用。利用腺病毒载体及特异性拮抗剂分别上调原代肝细胞中miR-140或抑制TLR4表达，观察肝细胞增殖、凋亡、脂肪变性及炎症相关信号通路的变化；下调miR-140或利用特异性激动剂激活TLR4通路，研究其对槐果碱抗NASH的抑制作用；将槐果碱和miR-140导入实验性NASH大鼠，体内研究其对NASH的治疗作用，以进一步明确槐果碱抗NASH作用的分子机制，并初步阐明miR-140在NASH发病中的作用，为NASH治疗提供新靶点和手段。

炎症信号通路及相关 miRNA 对非酒精性脂肪性肝炎（NASH）的调控作用是目前研究热点。前期工作证实槐果碱可在体内抑制 NASH 发展，其机制与抗炎作用有关。本研究中，我们将不同浓度的槐果碱加入原代分离并经过油酸诱导脂肪变性的肝细胞中，发现0.4mmol/L槐果碱对NASH有显著抑制作用，并可阻断炎症信号通路TLR4及其下游炎症信号通路NF-κB和JNK、ERK活性表达，该过程伴随miR-140上调。将miR-140 mimic导入油酸诱导脂肪变性的原代肝细胞中，发现TLR4表达下调，NF-κB和JNK、ERK信号通路活性受到抑制；将miR-140 inhibitor导入槐果碱治疗的油酸诱导脂肪变性的原代肝细胞中，发现TLR4 表达上调，NF-κB和JNK、ERK信号通路活性恢复。本课题研究证实槐果碱在体内外均对NASH有显著抑制作用，其主要作用机制为调控TLR4及其下游炎症信号通路NF-κB和JNK、ERK活性，炎症信号通路相关miRNA miR-140介导该过程。本研究进一步明确了槐果碱抗 NASH作用的分子机制，并初步阐明 miR-140 在 NASH 发病中的作用，为 NASH 治疗提供新靶点和手段。通过该研究，发表SCI收录论著3篇，培养研究生2名。