PUBLICATION
Investigating the role of lipid genes in liver disease using fatty liver models of alcohol and high fat in zebrafish (Danio rerio)
- Authors
- Shihana, F., Cholan, P.M., Fraser, S., Oehlers, S.H., Seth, D.
- ID
- ZDB-PUB-230831-56
- Date
- 2023
- Source
- Liver international : official journal of the International Association for the Study of the Liver 43(11): 2455-2468 (Journal)
- Registered Authors
- Keywords
- lipid droplet biology, risk genes, steatosis, zebrafish
- MeSH Terms
-
- Animals
- Ethanol/toxicity
- Fatty Acids/metabolism
- Humans
- Lipid Metabolism/genetics
- Liver/pathology
- Non-alcoholic Fatty Liver Disease*/pathology
- Triglycerides/metabolism
- Zebrafish*/genetics
- Zebrafish*/metabolism
- PubMed
- 37650211 Full text @ Liver Int.
Citation
Shihana, F., Cholan, P.M., Fraser, S., Oehlers, S.H., Seth, D. (2023) Investigating the role of lipid genes in liver disease using fatty liver models of alcohol and high fat in zebrafish (Danio rerio). Liver international : official journal of the International Association for the Study of the Liver. 43(11):2455-2468.
Abstract
Background Accumulation of lipid in the liver is the first hallmark of both alcohol-related liver disease (ALD) and non-alcohol-related fatty liver disease (NAFLD). Recent studies indicate that specific mutations in lipid genes confer risk and might influence disease progression to irreversible liver cirrhosis. This study aimed to understand the function/s of lipid risk genes driving disease development in zebrafish genetic models of alcohol-related and non-alcohol-related fatty liver.
Methods We used zebrafish larvae to investigate the effect of alcohol and high fat to model fatty liver and tested the utility of this model to study lipid risk gene functions. CRISPR/Cas9 gene editing was used to create knockdowns in 5 days post-fertilisation zebrafish larvae for the available orthologs of human cirrhosis risk genes (pnpla3, faf2, tm6sf2). To establish fatty liver models, larvae were exposed to ethanol and a high-fat diet (HFD) consisting of chicken egg yolk. Changes in morphology (imaging), survival, liver injury (biochemical tests, histopathology), gene expression (qPCR) and lipid accumulation (dye-specific live imaging) were analysed across treatment groups to test the functions of these genes.
Results Exposure of 5-day post-fertilisation (dpf) WT larvae to 2% ethanol or HFD for 48 h developed measurable hepatic steatosis. CRISPR-Cas9 genome editing depleted pnpla3, faf2 and tm6sf2 gene expression in these CRISPR knockdown larvae (crispants). Depletion significantly increased the effects of ethanol and HFD toxicity by increasing hepatic steatosis and hepatic neutrophil recruitment ≥2-fold in all three crispants. Furthermore, ethanol or HFD exposure significantly altered the expression of genes associated with ethanol metabolism (cyp2y3) and lipid metabolism-related gene expression, including atgl (triglyceride hydrolysis), axox1, echs1 (fatty acid β-oxidation), fabp10a (transport), hmgcra (metabolism), notch1 (signalling) and srebp1 (lipid synthesis), in all three pnpla3, faf2 and tm6sf2 crispants. Nile Red staining in all three crispants revealed significantly increased lipid droplet size and triglyceride accumulation in the livers following exposure to ethanol or HFD.
Conclusions We identified roles for pnpla3, faf2 and tm6sf2 genes in triglyceride accumulation and fatty acid oxidation pathways in a zebrafish larvae model of fatty liver.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping