| 1. |
- Bandaru, Manoj Kumar, et al.
(författare)
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Zebrafish larvae as a model system for systematic characterization of drugs and genes in dyslipidemia and atherosclerosis
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Tidskriftsartikel (refereegranskat)abstract
- Background: Hundreds of loci have been robustly associated with circulating lipids, atherosclerosis and coronary artery disease; but for most loci the causal genes and mechanisms remain uncharacterized.Methods: We developed a semi-automated experimental pipeline for systematic, quantitative, large-scale characterization of mechanisms, drugs and genes associated with dyslipidemia and atherosclerosis in a zebrafish model system. We validated our pipeline using a dietary (n>2000), drug treatment (n>1000), and genetic intervention (n=384).Results: Our results show that five days of overfeeding and cholesterol supplementation had independent pro-atherogenic effects, which could be diminished by concomitant treatment with atorvastatin and ezetimibe. CRISPR-Cas9-induced mutations in orthologues of proof-of-concept genes resulted in higher LDL cholesterol levels (apoea), and more early stage atherosclerosis (apobb.1).Conclusions: In summary, our pipeline facilitates systematic, in vivo characterization of drugs and candidate genes to increase our understanding of disease etiology, and can likely help identify novel targets for therapeutic intervention.
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| 2. |
- Gunja, Sethu Madhava Rao, 1986-, et al.
(författare)
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PARN acts as a maternal factor required for normal embryogenesis and telomere length maintenance in zebrafish
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Poly(A)-specific ribonuclease (PARN) is a 3’-5’ exoribonuclease that removes poly(A) tails of mRNAs and ncRNAs in eukaryotes. Mutations in the human PARN gene are associated with developmental delay and/or telomere biology disorders (TBDs). Here we have established a parn loss-of-function zebrafish model that recapitulates TBD symptoms and phenotypes displayed in human patients. Homozygous parn deficient zebrafish exhibited aberrant snoRNA profile, telomerase RNA maturation and shortening of telomere length over generations. In addition, we found that zygotic parn mutant embryos (Zparn) generated by crossing homozygous male and heterozygous females developed a spectrum of growth/developmental defects from embryonic stage to adult stage. The mutant embryos showed developmental defects with lethality during blastula and gastrulation where the maternal mRNAs need to be destabilized with the activation of zygotic transcription; larvae that surpassed the embryonic stage developed severe developmental defects like bent tail and cardiac edema; the fish that survived to adulthood have severe growth defects. Overall, the array of disease phenotypes observed in PARN mutant fish explains the importance of PARN at different stages of life and could provide a link to the mechanism of TBD penetrance in humans.
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| 3. |
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| 4. |
- von der Heyde, Benedikt, et al.
(författare)
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Translating GWAS-identified loci for cardiac rhythm and rate using an in vivo image- and CRISPR/Cas9-based approach
- 2020
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- A meta-analysis of genome-wide association studies (GWAS) identified eight loci that are associated with heart rate variability (HRV), but candidate genes in these loci remain uncharacterized. We developed an image- and CRISPR/Cas9-based pipeline to systematically characterize candidate genes for HRV in live zebrafish embryos. Nine zebrafish orthologues of six human candidate genes were targeted simultaneously in eggs from fish that transgenically express GFP on smooth muscle cells (Tg[acta2:GFP]), to visualize the beating heart. An automated analysis of repeated 30 s recordings of beating atria in 381 live, intact zebrafish embryos at 2 and 5 days post-fertilization highlighted genes that influence HRV (hcn4 and si:dkey-65j6.2 [KIAA1755]); heart rate (rgs6 and hcn4); and the risk of sinoatrial pauses and arrests (hcn4). Exposure to 10 or 25 mu M ivabradine-an open channel blocker of HCNs-for 24 h resulted in a dose-dependent higher HRV and lower heart rate at 5 days post-fertilization. Hence, our screen confirmed the role of established genes for heart rate and rhythm (RGS6 and HCN4); showed that ivabradine reduces heart rate and increases HRV in zebrafish embryos, as it does in humans; and highlighted a novel gene that plays a role in HRV (KIAA1755).
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| 5. |
- von der Heyde, Benedikt, et al.
(författare)
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Translating GWAS-identified loci for cardiac rhythm and rate using an in vivo, image-based, large-scale genetic screen in zebrafish
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Tidskriftsartikel (refereegranskat)abstract
- A meta-analysis of genome-wide association studies (GWAS) identified eight loci that are associated with heart rate variability (HRV) in data from 53,174 individuals. However, functional follow-up experiments - aiming to identify and characterize causal genes in these loci - have not yet been performed. We developed an image- and CRISPR-Cas9-based pipeline to systematically characterize candidate genes for HRV in live zebrafish embryos and larvae. Nine zebrafish orthologues of six human candidate genes were targeted simultaneously in fertilized eggs from fish that transgenically express GFP on smooth muscle cells (Tg(acta2:GFP)), to visualize the beating heart using a fluorescence microscope. An automated analysis of repeated 30s recordings of 381 live zebrafish atria at 2 and 5 days post-fertilization highlighted genes that influence HRV (hcn4 and kiaa1755); heart rate (rgs6 and hcn4) and the risk of sinoatrial pauses and arrests (hcn4). Hence, our screen confirmed the role of established genes for heart rate (rgs6 and hcn4), and highlighted a novel gene implicated in HRV (kiaa1755).
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