人参皂苷Rg1通过鞘氨醇激酶通路调控大鼠NAFLD的作用和机制研究

Nonalcoholic fatty liver disease (NAFLD) is one of diseases that threaten the health of people. In present, the pathogenesis of NAFLD is unclear, and there are oxidative stress and insulin resistance and so on. In our previous research has been found that ginsenoside Rg1 can effectively regulate the expression of sphingosine kinase signaling pathway-related proteins: activation of sphingosine kinase SPHK-1 and 1-phosphate sphingosine (S1P) 1 and 2 receptors, reducing S1P lysate (SPL), resulting in anti-cell apoptosis and cell proliferation, control the occurrence of NAFLD fibrosis, It is speculated that the therapeutic effect of Rg1 is mediated by this pathway. So in our ideas, There are three main purposes: 1. To investigated whether the sphingosine kinase signaling pathway is involved in the development of NAFLD; 2. Using RNA interference to SPHK-1 and S1P1, complete cytological validation of the mechanism on NAFLD treatment by Rg1;3. The protective effect of ginsenoside Rg1 on NAFLD rats and its molecular mechanism for the regulation of sphingosine kinase signaling pathway, provide the theoretical basis for the treatment of NAFLD and for develop potential drug.

非酒精性脂肪肝病（NAFLD）是威胁我国人民健康的一大疾病。本病发病机理不清，曾认为与氧化应激及胰岛素抵抗等有关。前期工作中发现：人参皂苷Rg1能明显改善NAFLD大鼠的肝功、血脂和肝细胞形态；同时调节鞘氨醇激酶信号通路相关蛋白的表达，即活化鞘氨醇激酶SPHK-1和1-磷酸鞘氨醇（S1P）1和2受体（S1P1/2），降低S1P裂解酶(SPL)，从而产生抗细胞凋亡和促细胞增殖, 控制NAFLD纤维化的发生，推测Rg1的治疗作用由该通路所介导。进一步结合大鼠和细胞实验：1、阐明鞘氨醇激酶信号通路是否参与NAFLD的发生与发展；2、使用RNA干扰SPHK-1和S1P1，完成Rg1对NAFLD治疗机制的细胞学验证；3、明确人参皂苷Rg1对NAFLD大鼠的保护作用及其对鞘氨醇激酶信号通路调控的分子机制，最终为治疗NAFLD提供理论依据，开发潜在的药物。

人参皂苷Rg1作为从三七中提取出的有效成分，在前期研究中，我们发现Rg1能够有效改善NAFLD大鼠的胰岛素抵抗情况，并降低LDL，同时促凋亡蛋白Bax下调和抗凋亡蛋白Bcl-2上调，由于Bcl-2、Bax受鞘氨醇激酶信号通路控制，我们推测人参皂苷Rg1是通过调控鞘氨醇激酶通路进而抑制细胞凋亡，从而到达治疗NAFLD的效果。因此本文旨在通过建立体内、体外NAFLD模型，再分别从体内、体外两个方向探究Rg1是否对NAFLD有治疗作用；鞘氨醇激酶信号通路和细胞凋亡是否参与NAFLD的发生与发展；并通过人参皂苷Rg1干预，探究其在鞘氨醇激酶信号通路中对抑制NAFLD的肝细胞凋亡作用；以及建立SPHK1和S1P1基因沉默模型后再次给予Rg1干预，检测鞘氨醇激酶通路和凋亡蛋白的表达，反向验证我们的猜想。结果显示随着人参皂苷Rg1剂量的提高，NAFLD大鼠肝脏的HE染色、油红O染色等结果表明，人参皂苷Rg1对NAFLD大鼠有着积极的治疗作用；其中SGPL、S1PR3表达下调， SPHK1、S1PR1表达上调，表明激活了鞘氨酸激酶通路；Bax表达下调、Bcl-2表达上调，表明Rg1抑制了NAFLD细胞的凋亡；且通过SGPL过表达细胞系的建立，再次给予Rg1干预治疗，发现情况没有好转，表明人参皂苷Rg1是通过鞘氨醇激酶通路治疗NAFLD的。综上所述人参皂苷Rg1对NAFLD有治疗作用，且这种治疗作用可能是通过调节鞘氨醇激酶通路的相关基因，进而抑制肝细胞的凋亡作用实现的。这对我们去探究NAFLD的发病机制以及开发地方特色药物都有相当积极的作用。