Optimizing methods for the study of intravascular lipid metabolism in zebrafish

Abstract

Abstract. The zebrafish (Danio rerio) is a useful vertebrate model for use in cardiovascular drug discovery. The present study aimed to construct optimized methods for the study of intravascular lipid metabolism of zebrafish. The lipophilic dye, Oil Red O, was used to stain fasting zebrafish one to eight days post-fertilization (dpf) and to stain 7-dpf zebrafish incubated in a breeding system containing 0.1% egg yolk as a high-fat diet (HFD) for 48 h. Three-dpf zebrafish were kept in CholEsteryl boron-dipyrromethene (BODIPY) 542/563 C11 water for 24 h which indicated the efficiency of CholEsteryl BODIPY 542/563 C11 intravascular cholesterol staining. Subsequently, 7-dpf zebrafish were incubated in water containing the fluorescent probe CholEsteryl BODIPY 542/563 C11 and fed a high-cholesterol diet (HCD) for 10 d. Two groups of 7-dpf zebrafish were incubated in regular breeding water and fed with a regular or HCD containing CholEsteryl BODIPY 542/563 C11 for 10 d. Finally, blood lipids of adult zebrafish fed with regular or HFD for seven weeks were measured. Oil Red O was not detected in the blood vessels of 7-8-dpf zebrafish. Increased intravascular lipid levels were detected in 7-dpf zebrafish incubated in 0.1% egg yolk, indicated by Oil Red O staining. Intravascular cholesterol was efficiently stained in 3-dpf zebrafish incubated in breeding water containing CholEsteryl BODIPY 542/563 C11; however, this method was inappropriate for the calculation of intravascular fluorescence intensity in zebrafish >7-dpf. In spite of this, intra-aortic fluorescence intensity of zebrafish fed a HCD containing CholEsteryl BODIPY 542/563 C11 was significantly higher (P<0.05) than that of those fed a regular diet containing CholEsteryl BODIPY 542/563 C11. The serum total cholesterol and triglyceride levels of adult zebrafish fed a HFD were markedly increased compared to those of the control group (P<0.05). In conclusion, the use of Oil Red O staining and CholEsteryl BODIPY 542/563 C11 may have applications in zebrafish intravascular lipid metabolism research and screens for novel lipid-regulating drugs.

High fat plus high cholesterol diet lead to hepatic steatosis in zebrafish larvae: a novel model for screening anti-hepatic steatosis drugs

“…The ability to conduct high-throughput drug screening is an outstanding advantage of larvae model for NAFLD studies. Unlike adult fish, lipids can be visualized directly in transparent larvae and juveniles using whole-mount oil red O staining method [ 13 , 38 – 41 ]. This simple method provides a high-throughput means to detect the incidence of steatosis in larvae, enabling us to examine steatosis across a large number of fish.…”

Evaluation of Metal‐Organic Framework MIL-89 nanoparticles toxicity on embryonic zebrafish development

“…Zebrafish embryos have been extensively used to evaluate the effect of drugs, chemicals, and molecules on the hepatic function through well-established functional assays [70] , [71] , [72] , [73] , as they highly conserve genetic pathways which underlay liver disease [74] . Around 70% of the ZF embryos yolk sac is composed of lipids, which are primarily metabolized by hepatocellular uptake [75] , [76] , [77] .…”

ATF4 overexpression induces early onset of hyperlipidaemia and hepatic steatosis and enhances adipogenesis in zebrafish

“…Zebrafish lipids are normally stored as TGs 34 , and the main storage sites include visceral, intramuscular, and subcutaneous lipid depots, however their storage is poor in blood vessels 35 , 36 and the liver 37 – 39 . The neutral lipid stain ORO was recently proposed for monitoring endotrophic lipid consumption during zebrafish embryonic and larval stages 35 , 40 , 41 . We observed minimal ORO staining in the swim bladder, heart, head, and vasculature of control larvae (ATs + Dox and WT).…”