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Triacsin C reduces lipid droplet formation and induces mitochondrial biogenesis in primary rat hepatocytes

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Abstract

Intracellular long-chain acyl-CoA synthetases (ACSL) activate fatty acids to produce acyl-CoA, which undergoes β-oxidation and participates in the synthesis of esterified lipids such as triacylglycerol (TAG). Imbalances in these metabolic routes are closely associated with the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Triacsin C is one of the few compounds that inhibit TAG accumulation into lipid droplets (LD) by suppressing ACSL activity. Here we report that treatment of primary rat hepatocytes with triacsin C at concentrations lower than the IC50 (4.1 μM) for LD formation: (i) diminished LD number in a concentration-dependent manner; (ii) increased mitochondrial amount; (iii) markedly improved mitochondrial metabolism by enhancing the β-oxidation efficiency, electron transport chain capacity, and degree of coupling – treatment of isolated rat liver mitochondria with the same triacsin C concentrations did not affect the last two parameters; (iv) decreased the GSH/GSSG ratio and elevated the protein carbonyl level, which suggested an increased reactive oxygen species production, as observed in isolated mitochondria. The hepatocyte mitochondrial improvements were not related to either the transcriptional levels of PGC-1α or the content of mTOR and phosphorylated AMPK. Triacsin C at 10 μM induced hepatocyte death by necrosis and/or apoptosis through mechanisms associated with mitochondrial permeability transition pore opening, as demonstrated by experiments using isolated mitochondria. Therefore, triacsin C at sub-IC50 concentrations modulates the lipid imbalance by shifting hepatocytes to a more oxidative state and enhancing the fatty acid consumption, which can in turn accelerate lipid oxidation and reverse NAFLD in long-term therapies.

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Acknowledgments

This study was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, Brazil, grant # 2010/17259-9). F.H.Z and C.R.P.D received scholarships from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brazil). The authors thank Prof. Emy Luiza Ishii-Iwamoto, PhD, from State University of Maringá (Maringá, PR, Brazil) for teaching the hepatocyte isolation method, Mrs. Ieda Maria Razaboni Prado from School of Pharmaceutical Sciences of Ribeirão Preto (Ribeirão Preto, SP, Brazil) for technical assistance, and Mrs. Maria Antonieta Rissato Garófalo from Ribeirão Preto Medical School (Ribeirão Preto, SP, Brazil) for helping us to quantify palmitate oxidation.

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Author notes

  1. Carlos R. P. Dechandt and Felippe H. Zuccolotto-dos-Reis contributed equally to this work.

Authors and Affiliations

  1. Departamento de Física e Química, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, USP, Av. Café s/n, Ribeirão Preto, SP, 14040-903, Brazil

    Carlos R. P. Dechandt, Felippe H. Zuccolotto-dos-Reis, Bruno G. Teodoro, Carlos Curti & Luciane C. Alberici

  2. Laboratório Thomson de Espectrometria de Massas, Instituto de Química, Universidade Estadual de Campinas, Rua Sérgio Buarque de Holanda s/n, Campinas, SP, 13083-970, Brazil

    Anna Maria A. P. Fernandes & Marcos N. Eberlin

  3. Departamento de Bioquímica e Imunologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, USP, Av. Bandeirantes, n° 3900, Ribeirão Preto, SP, 14049-900, Brazil

    Isis C. Kettelhut

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  1. Carlos R. P. Dechandt
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Contributions

CRPD performed the cell death, oxygen uptake, fatty acid oxidation, citrate synthase, western blot, RT-PCR, glutathione and protein carbonyl levels, and mitochondrial staining assays in hepatocytes. FHZR conducted the experiments in isolated mitochondria and oxygen uptake by hepatocytes, and isolated hepatocytes for all the assays. BGT helped to assess mitochondrial staining and oxygen uptake assays. AMAPF, MNE, and LCA performed mass spectrometry analyses. ICK helped to analyze fatty acid oxidation. CC analyzed the data and drafted the paper. LCA designed the study, analyzed data, wrote the paper, and has primary responsibility for the final content. All authors read and approved the final manuscript.

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Correspondence to Luciane C. Alberici.

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Highlights

• Triacsin C induces mitochondrial biogenesis not related to PGC-1α in hepatocytes

• Triacsin C increases the fatty acid oxidation capacity in primary rat hepatocytes

• Triacsin C (> 10 μM) induces cell death and mitochondrial permeability transition

• Triacsin C induces mitochondrial ROS release and an oxidized state in hepatocytes

• Specific TAG synthesis inhibition elicits mitochondrial biogenesis in hepatocytes

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Dechandt, C.R.P., Zuccolotto-dos-Reis, F.H., Teodoro, B.G. et al. Triacsin C reduces lipid droplet formation and induces mitochondrial biogenesis in primary rat hepatocytes. J Bioenerg Biomembr 49, 399–411 (2017). https://doi.org/10.1007/s10863-017-9725-9

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