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Excess branched-chain amino acids alter myotube metabolism and substrate preference which is worsened by concurrent insulin resistance

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Abstract

Purpose

Branched-chain amino acids (BCAA) have been shown to enhance several cellular signaling pathways including protein synthesis and mitochondrial biogenesis, yet population data demonstrate a correlation between circulating BCAA and severity of insulin resistance which has been hypothesized to be, in part, a byproduct of BCAA inhibition of mitochondrial function. The purpose of this study is to examine the effect of a BCAA mixture on muscle metabolism and related gene expression in vitro.

Methods

C2C12 myotubes were treated with a BCAA mixture containing leucine:isoleucine:valine at a ratio of 2:1:1 at 0.2, 2, or 20 mM (based on leucine content) for 6 days. qRT-PCR was used to measure metabolic gene expression. Oxygen consumption and extracellular acidification were used to assess mitochondrial and glycolytic metabolism, respectively. Mitochondrial content was determined via mitochondrial-specific staining.

Results

Despite significantly elevated mitochondrial staining, 6-day BCAA treatment reduced basal mitochondrial metabolism at a supraphysiological concentration (20 mM) in both insulin sensitive and resistant cells. Peak mitochondrial capacity was also reduced in insulin-resistant (but not insulin sensitive) cells. Conversely, basal glycolytic metabolism was elevated following 20 mM BCAA treatment, regardless of insulin resistance. In addition, insulin-resistant cells treated with 20 mM BCAA exhibited reduced gene expression of Ppargc1a, Cytc, Atp5b, Glut4, and several glycolytic enzymes versus insulin sensitive cells treated with 20 mM BCAA.

Conclusions

Collectively, these findings suggest BCAA at supraphysiologically high levels may negatively alter mitochondrial metabolism, and concurrent insulin resistance may also diminish peak mitochondrial capacity, as well as impede molecular adaptations that support a transition to a glycolytic preference/compensation.

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Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Abbreviations

BCAA:

branched-chain amino acid

BCKDH:

branched-chain alpha-keto acid dehydrogenase

CS:

citrate synthase

ECAR:

extracellular acidification rate

FCCP:

carbonyl cyanide p-[trifluoromethoxy]-phenyl-hydrazone

GLUT4:

glucose transporter 4

IRS-1:

insulin receptor substrate 1

LDHa:

lactate dehydrogenase A

LDHb:

lactate dehydrogenase B

mTOR:

mammalian target of rapamycin (mTOR)

NRF1/2:

nuclear respiratory factor 1/2

OCR:

oxygen consumption rate

PGC-1α:

peroxisome proliferator-activated receptor gamma coactivator 1 alpha

PPARα:

peroxisome proliferator-activated receptor alpha

PPARβ/δ:

peroxisome proliferator-activated receptor beta/delta

PK:

pyruvate kinase

TBP:

TATA binding protein

TFAM:

mitochondrial transcription factor A

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Acknowledgements

The authors would like also to thank the Department of Physical Therapy (Congdon School of Health Sciences) for the use of shared lab space and equipment. The mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the authors, contributors, or publisher.

Funding

Support for this work was provided by the Department of Exercise Science within the Congdon School of Health Sciences.

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  1. Department of Exercise Science, High Point University, High Point, NC, USA

    Madison E. Rivera, Caroline N. Rivera & Roger A. Vaughan

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M.E.R. and C.N.R. conducted experiments and assisted with manuscript preparation. R.A.V. conceived the study, conducted and oversaw experiments, performed all statistical analyses, and oversaw manuscript preparation. All authors have read and approved the final manuscript.

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Correspondence to Roger A. Vaughan.

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Rivera, M.E., Rivera, C.N. & Vaughan, R.A. Excess branched-chain amino acids alter myotube metabolism and substrate preference which is worsened by concurrent insulin resistance. Endocrine 76, 18–28 (2022). https://doi.org/10.1007/s12020-021-02939-z

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