dldhcri3 zebrafish exhibit altered mitochondrial ultrastructure, morphology, and dysfunction partially rescued by probucol or thiamine
Manuela Lavorato, … , Vernon E. Anderson, Marni J. Falk
Manuela Lavorato, … , Vernon E. Anderson, Marni J. Falk
Published August 20, 2024
Citation Information: JCI Insight. 2024;9(18):e178973. https://doi.org/10.1172/jci.insight.178973.
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Research Article Genetics Metabolism

dldhcri3 zebrafish exhibit altered mitochondrial ultrastructure, morphology, and dysfunction partially rescued by probucol or thiamine

Abstract

Dihydrolipoamide dehydrogenase (DLD) deficiency is a recessive mitochondrial disease caused by variants in DLD, the E3 subunit of mitochondrial α-keto (or 2-oxo) acid dehydrogenase complexes. DLD disease symptoms are multisystemic, variably manifesting as Leigh syndrome, neurodevelopmental disability, seizures, cardiomyopathy, liver disease, fatigue, and lactic acidemia. While most DLD disease symptoms are attributed to dysfunction of the pyruvate dehydrogenase complex, the effects of other α-keto acid dehydrogenase deficiencies remain unclear. Current therapies for DLD deficiency are ineffective, with no vertebrate animal model available for preclinical study. We created a viable Danio rerio (zebrafish) KO model of DLD deficiency, dldhcri3. Detailed phenotypic characterization revealed shortened larval survival, uninflated swim bladder, hepatomegaly and fatty liver, and reduced swim activity. These animals displayed increased pyruvate and lactate levels, with severe disruption of branched-chain amino acid catabolism manifest as increased valine, leucine, isoleucine, α-ketoisovalerate, and α-ketoglutarate levels. Evaluation of mitochondrial ultrastructure revealed gross enlargement, severe cristae disruption, and reduction in matrix electron density in liver, intestines, and muscle. Therapeutic modeling of candidate therapies demonstrated that probucol or thiamine improved larval swim activity. Overall, this vertebrate model demonstrated characteristic phenotypic and metabolic alterations of DLD disease, offering a robust platform to screen and characterize candidate therapies.

Authors

Manuela Lavorato, Donna Iadarola, Cristina Remes, Prabhjot Kaur, Chynna Broxton, Neal D. Mathew, Rui Xiao, Christoph Seiler, Eiko Nakamaru-Ogiso, Vernon E. Anderson, Marni J. Falk

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Figure 5

Liver pathology in dldh–/– zebrafish larvae.

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Liver pathology in dldh–/– zebrafish larvae. (A and B) WT and dldh–/– mo...
(A and B) WT and dldh–/– morphology at 7 dpf showed mutants had increased liver size (yellow line) and deflated swim bladder (arrow). Scale bar: 100 μm (×2.5 magnification of the middle fish in Figure 4B). (C and D) Oil Red O staining of 8 dpf WT and dldh–/– liver showed increased lipid accumulation in mutant larvae. Scale bar: 100 μm. (E) Liver area (μm2) was increased in 14 dldh–/– relative to 11 WT larvae by ImageJ analysis across 3 replicate experiments. ****P < 0.0001 by unpaired Student’s t test. (F and G) Thin sections of WT and dldh–/– liver stained with toluidine blue showed mutant larvae had increased frequency of lipid droplets (arrowheads). Scale bars: 10 µm. (H and I) Hepatocyte ultrastructure in 7 dpf WT and dldh–/– showed mutant larvae had mitochondrial damage (M) and large lipid droplets (L). Scale bars: 1 µm. (J) Ultrastructural analysis showed increased fractional lipid droplet to liver area analyzed in 4 WT and 3 dldh–/– larvae at 7 dpf. Areas determined using ImageJ. *P < 0.05 by unpaired Student’s t test.