| Background and Aims: Hyperlipidemia is the main cause of cardiovascular diseases.At present,a large number of studies have proved that high-fat diet can cause hyperlipidemia throughout the world,and the hyperlipidemia model has been established by this method for therapy drug screening,toxicity assessment and mechanism research of hyperlipidemia and related cardiovascular diseases.The model animal used in the establishment of hyperlipidemia model in this experiment is zebrafish larvae.Zebrafish is characterized by small size,high spawning rate,rapid development,transparent embryo and larva,and high homology with human genome.The complete genome sequencing of zebrafish has been completed,and it has been widely used in the research fields of organ development,neurological diseases,tumors,cardiovascular diseases,obesity,diabetes,lipid metabolism,inflammation,therapeutic drug screening and toxicity evaluation.Compared with the model animals commonly used in the past,such as mice,chickens,small pigs and rabbits,zebrafish has the advantages of low cost,short cycle,and can be used for large-scale experiments and in vivo observation.Bisphenol S(BPS)is a typical substitute for bisphenol A(BPA),which isrecognized environmental endocrine disruptor.At present,many studies have found that BPS has toxic effects on animal reproductive,immune system,nervous and metabolic systems.It has been reported that BPS can cause abnormal lipid metabolism,promote fat generation and cause obesity.High fat diet and environmental factors in life do not exist alone,and it is not clear how the combined effect of high fat diet and BPS exposure could affect animal lipid metabolism.Contents and Methods: This study mainly used 5dpf wild-type AB zebrafish and transgenic Fli1-EGFP zebrafish as experimental animals.Firstly,the zebrafish model of hyperlipidemia was constructed,and the hyperlipidemia of zebrafish could be caused by the high cholesterol diet prepared in this experiment.Then,the exposure of environment-related concentration BPS was conducted to clarify the effect of environment-related concentration BPS on the lipid metabolism of high-fat diet zebrafish,and to determine the effect on the key lipid metabolism genes(HMGCR and LDLR)and the related pathway genes(PPAR?,RXR and ROR)of zebrafish.Results:(1)Cholesterol tracer in vivo assay was used in this study from two aspects: A.It was explored that increasing the percentage of cholesterol in the feed could cause significant increase of blood lipid in juvenile fish(P<0.05).B.Gradually increasing the feed amount of cholesterol feedcan lead to a significant increase in juvenile lipids at first(P<0.05)and then a flattening trend,but the higher the feed amount,the lower the survival rate(P<0.05).The results showed that the establishment of hyperlipidemia model could be accelerated by increasing the percentage of cholesterol in the feed and the amount of feed.(2)5% high cholesterol diet(HCD)was used as high cholesterol diet in the experiment,and cholesterol tracer fluorescence microscope in vivo observation,body length and body width measurement,and biochemical indexes such as total cholesterol(TC),triglyceride(TG)and low-density lipoprotein cholesterol(LDL-C)were used as evaluation methods.The results showed that the level of vascular lipid accumulation in the 5%high cholesterol diet group was significantly higher than that in the Control group(P<0.01).The body length of 5%HCD was not significantly different from that of the Control group(P>0.05),but the body width was significantly greater than that of the Control group(P<0.01).TC and TG indexes of 5%HCD were significantly higher than those of the Control group(P<0.01),and LDL-C indexes were very significantly higher than those of the Control group(P<0.05).These manifestations were similar to those of other hyperlipidemic animals.Therefore,the modeling of hyperlipidemia zebrafish in the 5% cholesterol diet model group was successful.The results showed that the high cholesterol diet could induce hyperlipidemia in zebrafish.(3)The cholesterol tracer in vivo method was used in the experiment to study from two aspects: A,To explore the influence of environmental related concentration BPS exposure on vascular lipid accumulation of high-fat diet zebrafish,and the results showed that 1or10 ?g/L BPS exposure was significantly lower than the solvent control group(DMSO control group)(P<0.01).B,We explored the effect of environmental related concentration BPS exposure on vascular lipid accumulation of high cholesterol diet zebrafish under the condition of reducing or increasing feeding amount on the basis of experiment A,and the results showed that increased feeding amount could increase the susceptibility of environmental related concentration BPS exposure to lipid metabolism of zebrafish.(4)The current experiment explored the mechanism of the effect of environment-related concentration BPS exposure on lipid metabolism of high cholesterol diet zebrafish.The results showed that 10 ?g/L BPS exposure could promote the endogenous cholesterol synthesis of high-fat diet zebrafish(P<0.05),namely the expression of key enzyme HMGCR.100 ?g/L BPS exposure inhibited the expression of LDLR gene(P<0.05).The effect of BPS exposure on lipid metabolism of high cholesterol diet zebrafish may be related to the RXR /ROR related signaling pathway,which can inhibit the expression of RXR /ROR gene(P<0.05),but has no effect on PPAR? gene(P>0.05).Conclusion: This study explored a stable and reliable method to establish a model of hyperlipidemia in zebrafish,and provided an important tool for drug screening,toxicity assessment and mechanism research of hyperlipidemia and related cardiovascular diseases.The effects of BPS exposure and high cholesterol diet on the lipid metabolism of zebrafish were also investigated.It was clear that the combined effect of BPS and high cholesterol diet could not promote the further increase of blood lipid,and the mechanism might be related to the RXR /ROR related signaling pathway.This study thus provides novel data according to the study of BPS exposure combined toxicity. |
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