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Glutaminolysis is required for transforming growth factor-?1-induced myofibroblast differentiation and activation.

ABSTRACT: Myofibroblasts participate in physiological wound healing and pathological fibrosis. Myofibroblast differentiation is characterized by the expression of ?-smooth muscle actin and extracellular matrix proteins and is dependent on metabolic reprogramming. In this study, we explored the role of glutaminolysis and metabolites of TCA in supporting myofibroblast differentiation. Glutaminolysis converts Gln into ?-ketoglutarate (?-KG), a critical intermediate in the TCA cycle. Increases in the steady-state concentrations of TCA cycle metabolites including ?-KG, succinate, fumarate, malate, and citrate were observed in TGF-?1-differentiated myofibroblasts. The concentration of glutamate was also increased in TGF-?1-differentiated myofibroblasts compared with controls, whereas glutamine levels were decreased, suggesting enhanced glutaminolysis. This was associated with TGF-?1-induced expression of the glutaminase (GLS) isoform, GLS1, which converts Gln into glutamate, at both the mRNA and protein levels. The stimulation of GLS1 expression by TGF-?1 was dependent on both SMAD3 and p38 mitogen-activated protein kinase activation. Depletion of extracellular Gln prevented TGF-?1-induced myofibroblast differentiation. The removal of extracellular Gln postmyofibroblast differentiation decreased the expression of the profibrotic markers fibronectin and hypoxia-inducible factor-1? and reversed TGF-?1-induced metabolic reprogramming. Silencing of GLS1 expression, in the presence of Gln, abrogated TGF-?1-induced expression of profibrotic markers. Treatment of GLS1-deficient myofibroblasts with exogenous glutamate or ?-KG restored TGF-?1-induced expression of profibrotic markers in GLS1-deficient myofibroblasts. Together, these data demonstrate that glutaminolysis is a critical component of myofibroblast metabolic reprogramming that regulates myofibroblast differentiation.

SUBMITTER: Bernard K

PROVIDER: S-EPMC5787800 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Glutaminolysis is required for transforming growth factor-β1-induced myofibroblast differentiation and activation.

Bernard Karen K Logsdon Naomi J NJ Benavides Gloria A GA Sanders Yan Y Zhang Jianhua J Darley-Usmar Victor M VM Thannickal Victor J VJ

The Journal of biological chemistry 20171208 4

Myofibroblasts participate in physiological wound healing and pathological fibrosis. Myofibroblast differentiation is characterized by the expression of α-smooth muscle actin and extracellular matrix proteins and is dependent on metabolic reprogramming. In this study, we explored the role of glutaminolysis and metabolites of TCA in supporting myofibroblast differentiation. Glutaminolysis converts Gln into α-ketoglutarate (α-KG), a critical intermediate in the TCA cycle. Increases in the steady-s ...[more]

PMID: 29222329

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Project description:IntroductionIdiopathic pulmonary fibrosis (IPF) is a chronic progressive disease with very few effective treatments. The key effector cells in fibrosis are believed to be fibroblasts, which differentiate to a contractile myofibroblast phenotype with enhanced capacity to proliferate and produce extracellular matrix. The role of the lung epithelium in fibrosis is unclear. While there is evidence that the epithelium is disrupted in IPF, it is not known whether this is a cause or a result of the fibroblast pathology. We hypothesized that healthy epithelial cells are required to maintain normal lung homeostasis and can inhibit the activation and differentiation of lung fibroblasts to the myofibroblast phenotype. To investigate this hypothesis, we employed a novel co-culture model with primary human lung epithelial cells and fibroblasts to investigate whether epithelial cells inhibit myofibroblast differentiation.Measurements and main resultsIn the presence of transforming growth factor (TGF)-β, fibroblasts co-cultured with epithelial cells expressed significantly less α-smooth muscle actin and collagen and showed marked reduction in cell migration, collagen gel contraction, and cell proliferation compared to fibroblasts grown without epithelial cells. Epithelial cells from non-matching tissue origins were capable of inhibiting TGF-β induced myofibroblast differentiation in lung, keloid and Graves' orbital fibroblasts. TGF-β promoted production of prostaglandin (PG) E2 in lung epithelial cells, and a PGE2 neutralizing antibody blocked the protective effect of epithelial cell co-culture.ConclusionsWe provide the first direct experimental evidence that lung epithelial cells inhibit TGF-β induced myofibroblast differentiation and pro-fibrotic phenotypes in fibroblasts. This effect is not restricted by tissue origin, and is mediated, at least in part, by PGE2. Our data support the hypothesis that the epithelium plays a crucial role in maintaining lung homeostasis, and that damaged and/ or dysfunctional epithelium contributes to the development of fibrosis.

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Glutaminolysis is required for transforming growth factor-?1-induced myofibroblast differentiation and activation.

Publications

Glutaminolysis is required for transforming growth factor-β1-induced myofibroblast differentiation and activation.

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