基于斑马鱼胚胎肝损伤模型的蒙药德都红花-7味散护肝机制研究

Liver fibrosis, fatty liver, alcoholic liver, liver cancer and other liver diseases is one of the major diseases of the threat to human health in today's society. Liver injury lesions of various liver diseases, incasing by external factors, such as Drinking, drugs. Establishment of experimental animal models of liver injury is good ways that study the mechanism of liver disease. The Animal models helps screening hepatoprotective drugs, explore the hepatoprotective effect principle. The existing research is based on the application of rats or mice to establish liver injury model. In this study, we will establish zebrafish embryos animal models by known mammalian liver hepatotoxic drugs inducing. Liver development, apoptosis, pathological changes, and the expression liver fibrosis-related gene will be analyzed in the new liver injury model. The appropriate detection indicators will be used for high-throughput screening of drug screening, that Zebrafish transgenic technology play an important role. On the other hand, Mongolian medicine Deduhonghua-7 pill on liver protection mechanism of liver injury model will be studied by Analyzing augmenter of liver regeneration (ALR) expression. Knockdown of alr through morpholino antisense oligonucleotide (MO) used positioning liver regeneration targets of Deduhonghua-7 pill. The liver regeneration factor expression ,such as NF-κB、TNF-α, will is useful for analyzing the intervention of the signal transduction in liver regeneration process after liver damage. In this study, the use of modern biotechnology research of the mechanism of action of traditional Mongolian medicine and to explore Mongolian medicine efficacy.

肝纤维化、脂肪肝、酒精肝、肝癌等肝病是当今社会威胁人类健康的主要疾病之一。肝损伤是各种肝脏疾病的病变结果，通过建立实验性肝损伤动物模型，研究肝病的发生机制，筛选保肝药物，探索保肝作用原理，具有重要的现实意义。现有研究主要是以应用大鼠或小鼠建立肝损伤模型。本研究通过用能够引起肝损伤药物诱导的斑马鱼胚胎，分析其肝部发育、细胞凋亡、病理学变化、肝纤维化脂肪化相关基因表达等建立肝损伤模型，通过转基因技术筛选适当检测指标以达到高通量筛选药物的目的。在此基础上，研究蒙药德都红花-7味散对肝损伤模型肝保护作用机制，重点分析德都红花-7味散对肝再生增强因子（ALR）表达影响，通过基因敲低（Knockdown）对ALR基因进行功能阻断，检测NF-κB、TNF-α等肝再生相关因子的表达，分析德都红花-7味散在肝损伤后肝细胞再生过程中信号转导的干预方式。本研究利用现代生物技术研究传统蒙药的作用机理，发掘蒙药功效

肝损伤是各种肝脏疾病的病变结果，通过建立实验性肝损伤动物模型，研究肝病的发生机制，筛选保肝药物，探索保肝作用原理，具有重要的现实意义。.本课题完成了以下主要研究目标：（1）建立了斑马鱼胚胎肝损伤药物诱导模型，研究了硫代乙酰胺（TAA）、对乙酰氨基酚（APAP）、乙醇引起的肝脏脂肪化、细胞凋亡等变化情况。（2）分析了德都红花-7味散、伊赫-汤对斑马鱼胚胎肝损伤的保护性作用及其作用机制。（3）对斑马鱼胚胎肝损伤模型（TAA诱导模型、APAP诱导模型）及德都红花-7味散保护模型进行了转录组测序分析，筛选出一批相关基因。（4）建立了简便的斑马鱼护肝药物大通量筛选模型，可用于快速筛选护肝药物。. 将对乙酰氨基酚（APAP）处理组和对照组的斑马鱼进行转录组测序分析，结果发现APAP处理组与对照组的差异表达基因的数量为72个，其中上调差异表达基因的数量为40个，下调差异表达基因的数量为32个，其中与肝脏损伤有关的基因主要有prdx1、gstp1、fkbp5、txn、ucp3、tm4sf4、mt-cyb、ddx5、g3bp2、anxa4和try；与色素沉着有关的基因是Rho、opn1lw2和opn1mw1。结果证明APAP不仅引起斑马鱼肝脏损伤，同时还会引起色素沉着减少。 . 应用德都红花-7味散对硫代乙酰胺（TAA）诱导的斑马鱼胚胎肝损伤模型进行治疗，建立保护模型。对德都红花-7味散治疗组和TAA诱导的肝损伤组进行转录组测序分析，发现了7个可能参与非酒精性脂肪肝的致病基因，为研究非酒精性脂肪肝发病的分子调控机制打下坚实的基础。研究证明德都红花-7味散对TAA诱发的非酒精性脂肪肝具有一定的保护作用。. 此外，本研究使用斑马鱼为实验动物模型，研究酒精导致斑马鱼胚胎脂肪肝脏病变及“伊赫-汤”的防护作用，实验结果表明“伊赫-汤”对酒精导致斑马鱼胚胎肝损伤具有修复作用。.