| Summary: | 碩士 === 國立臺灣海洋大學 === 生物科技研究所 === 99 === In recent years, non-alcoholic fatty liver disease (NAFLD) has become the highest prevalence of chronic liver disease, and its also been highly correlated with some other human chronic disease, e.g. obesity, diabetes or hypertension. High-fat diet and alcohol intake will produce fatty acid accumulation in liver, the main reason for fatty acid abnormal accumulate in the liver might be hepatic lipid metabolism dysfunction. Carbohydrate response element-binding protein (ChREBP) is a transcription factor, which is regulated by hepatic glucose concentration, will promote the rate-limiting enzyme of glycolysis, L-PK, and lipogenesis, ACC and FAS. On the other hand, cannabinoid receptor type 1 (CB1R) has been considered its relationship with energy homeostasis and fatty acid synthesis, which expression of CB1R will induce hepatic steatosis. According to this, we designed a regulatable complex mosaic vector system with L-FABP promoter, which can specifically expressed in liver. With inducing substances added, this vector system will subsequently make gene express. We use this verctor system to carry ChREBP and CB1R gene, therefore we inject this regulatable complex mosaic construct into the wild-type zebrafish zygotes. After that we screened the F0 zebrafish. Then we crossed the F0 zebrafish with the wild type zebrafish to obtain the offspring (F1). Finally, induced by Oil Red O stain of the 5dpf embryo for two days to observed the fat accumulation in the liver. We also designed another special inducible complex mosaic vector system, once this vector system been induced, it will produces its own transcription factors and overexpressed the interested genes in a short time. By the characteristics of this vector system, we carried lipogenesis-related genes, ChREBP, CB1R, and PPARγ on this vector system. We injected this inducible complex mosaic constructs into the wild-type zebrafish zygotes, then add the inducing substances in 5dpf stage for 2 days. By using Nile Red lipid fluorescent stain, we can observe the situation of fatty acids accumulated in liver. According to our research, we found that to regulate the gene of fatty acid synthesis by these two vector systems can induce steatosis in early stage, and may be useful in the research of non-alcoholic fatty liver disease.
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