TNFR1-activated reactive oxidative species signals up-regulate osteogenic Msx2 programs in aortic myofibroblasts

Chung Fang Lai, Jian Su Shao, Abraham Behrmann, Karen Krchma, Su Li Cheng, Dwight A. Towler

Research output: Contribution to journalArticle

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Abstract

In LDLR -/- mice fed high-fat diabetogenic diets, osteogenic gene-regulatory programs are ectopically activated in vascular myofibroblasts and smooth muscle cells that promote arteriosclerotic calcium deposition. Msx2-Wnt signaling pathways previously identified as important for craniofacial skeletal development are induced in the vasculature by TNF, a prototypic cytokine mediator of the low-grade systemic inflammation of diabesity. To better understand this biology, we studied TNF actions on Msx2 in aortic myofibroblasts. TNF up-regulated Msx2 mRNA 4-fold within 3 h but did not regulate Msx1. Although IL-1β could also induce Msx2 expression, TNF-related apoptosis inducing ligand, receptor activator of nuclear factor-κB ligand, and IL-6 were inactive. Inhibition of nicotinamide adenine dinucleotide phosphate oxidase (Nox) activity and genetically induced Nox deficiency (p47phox -/-) reduced Msx2 induction, indicating contributions of reactive oxygen species (ROS) and redox signaling. Consistent with this, rotenone, an antagonist of mitochondrial complex I, inhibited TNF induction of Msx2 and Nox2, whereas pyruvate, an anapleurotic mitochondrial metabolic substrate, enhanced induction. Moreover, the glutathione peroxidase-mimetic ebselen abrogated this TNF response. Treatment of aortic myofibroblasts with hydrogen peroxide up-regulated Msx2 mRNA, promoter activity, and DNA-protein interactions. In vivo, SM22-TNF transgenic mice exhibit increased aortic Msx2 with no change in Msx1. Dosing SM22-TNF mice with either 20 ng/g Nox1 + 20 ng/g Nox2 antisense oligonucleotides or low-dose rotenone reduced arterial Msx2 expression. Aortic myofibroblasts from TNFR1 -/- mice expressed levels of Msx2 that were 5% that of wild-type and were not inducible by TNF. Wnt7b and active β-catenin levels were also reduced. By contrast, TNF-inducible Msx2 expression was not reduced in TNFR2 -/-cells. Finally, when cultured under mineralizing conditions, TNFR1 -/- aortic myofibroblasts exhibited reduced calcification compared with wild-type and TNFR2 -/- cells. Thus, ROS metabolism contributes to TNF induction of Msx2 and procalcific responses in myofibroblasts via TNFR1. Strategies that reduce vascular Nox- or mitochondrially activated ROS signals may prove useful in mitigating arteriosclerotic calcification.

Original languageEnglish (US)
Pages (from-to)3897-3910
Number of pages14
JournalEndocrinology
Volume153
Issue number8
DOIs
StatePublished - Aug 1 2012

Fingerprint

Receptors, Tumor Necrosis Factor, Type I
Myofibroblasts
Up-Regulation
Receptors, Tumor Necrosis Factor, Type II
Rotenone
Reactive Oxygen Species
TNF-Related Apoptosis-Inducing Ligand Receptors
Catenins
Messenger RNA
Wnt Signaling Pathway
Antisense Oligonucleotides
High Fat Diet
Regulator Genes
Glutathione Peroxidase
Pyruvic Acid
Interleukin-1
NADP
Vascular Smooth Muscle
Hydrogen Peroxide
Transgenic Mice

ASJC Scopus subject areas

  • Endocrinology

Cite this

TNFR1-activated reactive oxidative species signals up-regulate osteogenic Msx2 programs in aortic myofibroblasts. / Lai, Chung Fang; Shao, Jian Su; Behrmann, Abraham; Krchma, Karen; Cheng, Su Li; Towler, Dwight A.

In: Endocrinology, Vol. 153, No. 8, 01.08.2012, p. 3897-3910.

Research output: Contribution to journalArticle

Lai, Chung Fang ; Shao, Jian Su ; Behrmann, Abraham ; Krchma, Karen ; Cheng, Su Li ; Towler, Dwight A. / TNFR1-activated reactive oxidative species signals up-regulate osteogenic Msx2 programs in aortic myofibroblasts. In: Endocrinology. 2012 ; Vol. 153, No. 8. pp. 3897-3910.
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abstract = "In LDLR -/- mice fed high-fat diabetogenic diets, osteogenic gene-regulatory programs are ectopically activated in vascular myofibroblasts and smooth muscle cells that promote arteriosclerotic calcium deposition. Msx2-Wnt signaling pathways previously identified as important for craniofacial skeletal development are induced in the vasculature by TNF, a prototypic cytokine mediator of the low-grade systemic inflammation of diabesity. To better understand this biology, we studied TNF actions on Msx2 in aortic myofibroblasts. TNF up-regulated Msx2 mRNA 4-fold within 3 h but did not regulate Msx1. Although IL-1β could also induce Msx2 expression, TNF-related apoptosis inducing ligand, receptor activator of nuclear factor-κB ligand, and IL-6 were inactive. Inhibition of nicotinamide adenine dinucleotide phosphate oxidase (Nox) activity and genetically induced Nox deficiency (p47phox -/-) reduced Msx2 induction, indicating contributions of reactive oxygen species (ROS) and redox signaling. Consistent with this, rotenone, an antagonist of mitochondrial complex I, inhibited TNF induction of Msx2 and Nox2, whereas pyruvate, an anapleurotic mitochondrial metabolic substrate, enhanced induction. Moreover, the glutathione peroxidase-mimetic ebselen abrogated this TNF response. Treatment of aortic myofibroblasts with hydrogen peroxide up-regulated Msx2 mRNA, promoter activity, and DNA-protein interactions. In vivo, SM22-TNF transgenic mice exhibit increased aortic Msx2 with no change in Msx1. Dosing SM22-TNF mice with either 20 ng/g Nox1 + 20 ng/g Nox2 antisense oligonucleotides or low-dose rotenone reduced arterial Msx2 expression. Aortic myofibroblasts from TNFR1 -/- mice expressed levels of Msx2 that were 5{\%} that of wild-type and were not inducible by TNF. Wnt7b and active β-catenin levels were also reduced. By contrast, TNF-inducible Msx2 expression was not reduced in TNFR2 -/-cells. Finally, when cultured under mineralizing conditions, TNFR1 -/- aortic myofibroblasts exhibited reduced calcification compared with wild-type and TNFR2 -/- cells. Thus, ROS metabolism contributes to TNF induction of Msx2 and procalcific responses in myofibroblasts via TNFR1. Strategies that reduce vascular Nox- or mitochondrially activated ROS signals may prove useful in mitigating arteriosclerotic calcification.",
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