NOTCH reprograms mitochondrial metabolism for proinflammatory macrophage activation
Jun Xu, … , Samuel W. French, Hidekazu Tsukamoto
Jun Xu, … , Samuel W. French, Hidekazu Tsukamoto
Published March 23, 2015
Citation Information: J Clin Invest. 2015;125(4):1579-1590. https://doi.org/10.1172/JCI76468.
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Research Article Immunology

NOTCH reprograms mitochondrial metabolism for proinflammatory macrophage activation

Abstract

Metabolic reprogramming is implicated in macrophage activation, but the underlying mechanisms are poorly understood. Here, we demonstrate that the NOTCH1 pathway dictates activation of M1 phenotypes in isolated mouse hepatic macrophages (HMacs) and in a murine macrophage cell line by coupling transcriptional upregulation of M1 genes with metabolic upregulation of mitochondrial oxidative phosphorylation and ROS (mtROS) to augment induction of M1 genes. Enhanced mitochondrial glucose oxidation was achieved by increased recruitment of the NOTCH1 intracellular domain (NICD1) to nuclear and mitochondrial genes that encode respiratory chain components and by NOTCH-dependent induction of pyruvate dehydrogenase phosphatase 1 (Pdp1) expression, pyruvate dehydrogenase activity, and glucose flux to the TCA cycle. As such, inhibition of the NOTCH pathway or Pdp1 knockdown abrogated glucose oxidation, mtROS, and M1 gene expression. Conditional NOTCH1 deficiency in the myeloid lineage attenuated HMac M1 activation and inflammation in a murine model of alcoholic steatohepatitis and markedly reduced lethality following endotoxin-mediated fulminant hepatitis in mice. In vivo monocyte tracking further demonstrated the requirement of NOTCH1 for the migration of blood monocytes into the liver and subsequent M1 differentiation. Together, these results reveal that NOTCH1 promotes reprogramming of mitochondrial metabolism for M1 macrophage activation.

Authors

Jun Xu, Feng Chi, Tongsheng Guo, Vasu Punj, W.N. Paul Lee, Samuel W. French, Hidekazu Tsukamoto

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Figure 5

NOTCH-dependent glucose oxidation and mtROS augment M1 activation.

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NOTCH-dependent glucose oxidation and mtROS augment M1 activation. (A) F...
(A) FACS analysis of mtROS using MitoSox Red in WT primary HMacs pretreated with or without dTTP or MitoQ for 1 hour followed by LPS for 24 hours. dTTP served as a pharmacologic control for MitoQ (n = 4–6). *P < 0.05 vs. control, #P < 0.05 vs. LPS plus dTTP, 1-way ANOVA. (B) FACS analysis of mtROS using MitoSox Red in WT or Notch1 KO HMacs treated with or without LPS for 24 hours (n = 6–8). *P < 0.05 vs. WT, #P < 0.05 vs. WT+LPS, 1-way ANOVA. (C) 2-DG reduces the expression of M1 genes in HMacs from OF+Alc mice (n = 3). *P < 0.05 vs. control, #P < 0.05 vs. OF+Alc, 1-way ANOVA. (D) 2-DG attenuates mtROS in LPS-induced M1 Raw 264.7 cells (n = 3). *P < 0.05 vs. control, #P < 0.05 vs. LPS, 1-way ANOVA. (E) MitoQ suppresses the expression of M1 genes in M1 Raw 264.7 cells treated with LI for 4 hours (n = 3). The dashed line indicates the mRNA levels of control cells, which are set to 1. #P < 0.05 vs. LI+dTTP, t test. (F and G) Pdp1 silencing abrogates the expression of (F) M1 genes or (G) mtROS in Raw 264.7 cells infected with scrambled or Pdp1 shRNA with or without LPS treatment (n = 4–6). *P < 0.01 vs. Scr control, #P < 0.01 vs. Scr LPS, 2-way ANOVA. (H) Expression of M1 genes in WT HMacs cultured in glucose/pyruvate-free medium (control) or supplemented with either glucose (5.5 mM) or pyruvate (10 mM) (n = 6). *P < 0.05 vs. control, t test.