Original article
High concentration of branched-chain amino acids promotes oxidative stress, inflammation and migration of human peripheral blood mononuclear cells via mTORC1 activation

https://doi.org/10.1016/j.freeradbiomed.2017.01.009Get rights and content
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Highlights

  • High BCAA concentration induces ROS generation and mitochondrial dysfunction in PBMCs.

  • High BCAA concentration promotes the activation of NF-κB favoring the expression of cytokines and adhesion molecules and PBMCs migration.

  • The pro-oxidant and pro-inflammatory actions of BCAA are mediated by the Akt-mTORC1 axis.

Abstract

Leucine, isoleucine and valine are essential aminoacids termed branched-chain amino acids (BCAA) due to its aliphatic side-chain. In several pathological and physiological conditions increased BCAA plasma concentrations have been described. Elevated BCAA levels predict insulin resistance development. Moreover, BCAA levels higher than 2 mmol/L are neurotoxic by inducing microglial activation in maple syrup urine disease. However, there are no studies about the direct effects of BCAA in circulating cells. We have explored whether BCAA could promote oxidative stress and pro-inflammatory status in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors. In cultured PBMCs, 10 mmol/L BCAA increased the production of reactive oxygen species (ROS) via both NADPH oxidase and the mitochondria, and activated Akt-mTOR signalling. By using several inhibitors and activators of these molecular pathways we have described that mTOR activation by BCAA is linked to ROS production and mitochondrial dysfunction. BCAA stimulated the activation of the redox-sensitive transcription factor NF-κB, which resulted in the release of pro-inflammatory molecules, such as interleukin-6, tumor necrosis factor-α, intracellular adhesion molecule-1 or CD40L, and the migration of PBMCs. In conclusion, elevated BCAA blood levels can promote the activation of circulating PBMCs, by a mechanism that involving ROS production and NF-κB pathway activation. These data suggest that high concentrations of BCAA could exert deleterious effects on circulating blood cells and therefore contribute to the pro-inflammatory and oxidative status observed in several pathophysiological conditions.

Graphical abstract

Representative diagram of proposed mechanisms involved in BCAA signalling applied for PBMCs

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Abbreviations

AICAR
5-Aminoimidazole-4-carboxamide 1-β-D-ribofuranoside
AMPK
AMP-activated protein kinase
AP
alkaline phosphatase
BCAA
branched-chain amino acids
BCKDC
branched-chain alpha-ketoacid dehydrogenase complex
BSA/PBS
bovine serum albumin/phosphate-buffered saline
CD40L
CD40 ligand
DAPI
4',6-diamidino-2-phenylindole dihydrochloride
DPI
Diphenyleneiodonium chloride
GAPDH
Glyceraldehyde 3-phosphate dehydrogenase
HRP
Horseradish peroxidase
ICAM-1
intercellular adhesion molecule 1
IL-6
Interleukin-6
LPS
lipopolysaccharide
MAPK
Mitogen-activated protein kinase
Mito-TEMPO
2,2,6,6-tetramethyl-4-[[2-(triphenylphosphonio)acetyl]amino]-1-piperidinyloxy, monochloride, monohydrate
mTORC
mammalian target of rapamycin complex
NFκB
nuclear transcription factor-κB
Nrf2 or NFE2L2
Nuclear factor (erythroid-derived 2)-like 2
O2•−
Superoxide anion radical
PBMC
peripheral blood mononuclear cells
p-NPP
p-Nitrophenyl Phosphate
ROS
Reactive Oxygen Species
RT-PCR
Reverse transcription polymerase chain reaction
PI3K/Akt
phosphatydilinositol (3,4,5)-triphosphate
SDS-PAGE
sodium dodecyl sulfate-polyacrylamide gel electrophoresis
TMRM
Tetramethyl rhodamine methyl ester
TNFα
Tumor necrosis factor alpha
UCP-2
uncoupling protein 2
ΔΨm
mitochondrial membrane potential

Keywords

BCAA
Peripheral blood mononuclear cells
mTORC1
PI3K/Akt
Inflammation
Oxidative stress

Cited by (0)

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These authors have jointly directed this work.