Lipopolysaccharide stimulates nitric oxide synthase-2 expression in murine skeletal muscle and C2C12 myoblasts via Toll-like receptor-4 and c-Jun NH2-terminal kinase pathways

Robert A. Frost, Gerald J. Nystrom, Charles H. Lang

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

The inducible form of nitric oxide synthase (NOS2) catalyzes the synthesis of nitric oxide (NO) from arginine in response to injury and infection. NOS2 is expressed predominantly by macrophages and lymphocytes. However, skeletal muscle also expresses NOS2 in response to inflammatory stimuli. The present study sought to determine whether lipopolysaccharide (LPS) stimulates NOS2 in skeletal muscle via Toll-like receptor-4 (TLR4). Intraperitoneal injection of LPS in wild-type mice (C3H/HeSnJ) increased NOS2 mRNA fourfold in skeletal muscle, while no change in NOS2 mRNA was observed in C3H/HeJ mice that harbored a mutation in the LPS receptor. NOS2 coimmunoprecipitated with the muscle-specific caveolin-3 protein, suggesting that myofibers per se respond to LPS in vivo. LPS stimulated NOS2 mRNA expression in C2C12 myocytes, and the regulation of NOS2 mRNA was comparable in myoblasts and differentiated myotubes. LPS transiently stimulated the phosphorylation of the interleukin-1 receptor-associated kinase (IRAK-1) in C2C12 cells and decreased the total amount of IRAK-1 both in vitro and in vivo over time. LPS stimulated the expression of an NF-κB reporter plasmid, and this was inhibited by the proteasomal inhibitor MG-132. Both myoblasts and myotubes expressed TLR2 and TLR4 mRNA. Expression of a dominant negative form of TLR4 in C2C12 cells blocked LPS-induced NF-κB reporter activity. SP-600125 [a c-Jun NH2-terminal kinase (JNK) inhibitor] also prevented LPS stimulation of NOS2 expression. Moreover, the JNK inhibitor prevented the LPS-induced increase in NO synthesis. These data indicate that LPS increases NOS2 mRNA expression in muscle via a TLR4-dependent mechanism.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume287
Issue number6 56-6
DOIs
StatePublished - Dec 1 2004

Fingerprint

Toll-Like Receptor 4
JNK Mitogen-Activated Protein Kinases
Myoblasts
Nitric Oxide Synthase
Lipopolysaccharides
Muscle
Skeletal Muscle
Messenger RNA
Caveolin 3
Inbred C3H Mouse
Skeletal Muscle Fibers
Nitric Oxide
Interleukin-1 Receptor-Associated Kinases
CD14 Antigens
Caveolins
Phosphorylation
Lymphocytes
Macrophages
Nitric Oxide Synthase Type II
Intraperitoneal Injections

All Science Journal Classification (ASJC) codes

  • Clinical Biochemistry
  • Cell Biology
  • Physiology

Cite this

@article{d1f017f404644397b26a7d288ff4a605,
title = "Lipopolysaccharide stimulates nitric oxide synthase-2 expression in murine skeletal muscle and C2C12 myoblasts via Toll-like receptor-4 and c-Jun NH2-terminal kinase pathways",
abstract = "The inducible form of nitric oxide synthase (NOS2) catalyzes the synthesis of nitric oxide (NO) from arginine in response to injury and infection. NOS2 is expressed predominantly by macrophages and lymphocytes. However, skeletal muscle also expresses NOS2 in response to inflammatory stimuli. The present study sought to determine whether lipopolysaccharide (LPS) stimulates NOS2 in skeletal muscle via Toll-like receptor-4 (TLR4). Intraperitoneal injection of LPS in wild-type mice (C3H/HeSnJ) increased NOS2 mRNA fourfold in skeletal muscle, while no change in NOS2 mRNA was observed in C3H/HeJ mice that harbored a mutation in the LPS receptor. NOS2 coimmunoprecipitated with the muscle-specific caveolin-3 protein, suggesting that myofibers per se respond to LPS in vivo. LPS stimulated NOS2 mRNA expression in C2C12 myocytes, and the regulation of NOS2 mRNA was comparable in myoblasts and differentiated myotubes. LPS transiently stimulated the phosphorylation of the interleukin-1 receptor-associated kinase (IRAK-1) in C2C12 cells and decreased the total amount of IRAK-1 both in vitro and in vivo over time. LPS stimulated the expression of an NF-κB reporter plasmid, and this was inhibited by the proteasomal inhibitor MG-132. Both myoblasts and myotubes expressed TLR2 and TLR4 mRNA. Expression of a dominant negative form of TLR4 in C2C12 cells blocked LPS-induced NF-κB reporter activity. SP-600125 [a c-Jun NH2-terminal kinase (JNK) inhibitor] also prevented LPS stimulation of NOS2 expression. Moreover, the JNK inhibitor prevented the LPS-induced increase in NO synthesis. These data indicate that LPS increases NOS2 mRNA expression in muscle via a TLR4-dependent mechanism.",
author = "Frost, {Robert A.} and Nystrom, {Gerald J.} and Lang, {Charles H.}",
year = "2004",
month = "12",
day = "1",
doi = "10.1152/ajpcell.00010.2004",
language = "English (US)",
volume = "287",
journal = "American Journal of Physiology",
issn = "0363-6143",
publisher = "American Physiological Society",
number = "6 56-6",

}

TY - JOUR

T1 - Lipopolysaccharide stimulates nitric oxide synthase-2 expression in murine skeletal muscle and C2C12 myoblasts via Toll-like receptor-4 and c-Jun NH2-terminal kinase pathways

AU - Frost, Robert A.

AU - Nystrom, Gerald J.

AU - Lang, Charles H.

PY - 2004/12/1

Y1 - 2004/12/1

N2 - The inducible form of nitric oxide synthase (NOS2) catalyzes the synthesis of nitric oxide (NO) from arginine in response to injury and infection. NOS2 is expressed predominantly by macrophages and lymphocytes. However, skeletal muscle also expresses NOS2 in response to inflammatory stimuli. The present study sought to determine whether lipopolysaccharide (LPS) stimulates NOS2 in skeletal muscle via Toll-like receptor-4 (TLR4). Intraperitoneal injection of LPS in wild-type mice (C3H/HeSnJ) increased NOS2 mRNA fourfold in skeletal muscle, while no change in NOS2 mRNA was observed in C3H/HeJ mice that harbored a mutation in the LPS receptor. NOS2 coimmunoprecipitated with the muscle-specific caveolin-3 protein, suggesting that myofibers per se respond to LPS in vivo. LPS stimulated NOS2 mRNA expression in C2C12 myocytes, and the regulation of NOS2 mRNA was comparable in myoblasts and differentiated myotubes. LPS transiently stimulated the phosphorylation of the interleukin-1 receptor-associated kinase (IRAK-1) in C2C12 cells and decreased the total amount of IRAK-1 both in vitro and in vivo over time. LPS stimulated the expression of an NF-κB reporter plasmid, and this was inhibited by the proteasomal inhibitor MG-132. Both myoblasts and myotubes expressed TLR2 and TLR4 mRNA. Expression of a dominant negative form of TLR4 in C2C12 cells blocked LPS-induced NF-κB reporter activity. SP-600125 [a c-Jun NH2-terminal kinase (JNK) inhibitor] also prevented LPS stimulation of NOS2 expression. Moreover, the JNK inhibitor prevented the LPS-induced increase in NO synthesis. These data indicate that LPS increases NOS2 mRNA expression in muscle via a TLR4-dependent mechanism.

AB - The inducible form of nitric oxide synthase (NOS2) catalyzes the synthesis of nitric oxide (NO) from arginine in response to injury and infection. NOS2 is expressed predominantly by macrophages and lymphocytes. However, skeletal muscle also expresses NOS2 in response to inflammatory stimuli. The present study sought to determine whether lipopolysaccharide (LPS) stimulates NOS2 in skeletal muscle via Toll-like receptor-4 (TLR4). Intraperitoneal injection of LPS in wild-type mice (C3H/HeSnJ) increased NOS2 mRNA fourfold in skeletal muscle, while no change in NOS2 mRNA was observed in C3H/HeJ mice that harbored a mutation in the LPS receptor. NOS2 coimmunoprecipitated with the muscle-specific caveolin-3 protein, suggesting that myofibers per se respond to LPS in vivo. LPS stimulated NOS2 mRNA expression in C2C12 myocytes, and the regulation of NOS2 mRNA was comparable in myoblasts and differentiated myotubes. LPS transiently stimulated the phosphorylation of the interleukin-1 receptor-associated kinase (IRAK-1) in C2C12 cells and decreased the total amount of IRAK-1 both in vitro and in vivo over time. LPS stimulated the expression of an NF-κB reporter plasmid, and this was inhibited by the proteasomal inhibitor MG-132. Both myoblasts and myotubes expressed TLR2 and TLR4 mRNA. Expression of a dominant negative form of TLR4 in C2C12 cells blocked LPS-induced NF-κB reporter activity. SP-600125 [a c-Jun NH2-terminal kinase (JNK) inhibitor] also prevented LPS stimulation of NOS2 expression. Moreover, the JNK inhibitor prevented the LPS-induced increase in NO synthesis. These data indicate that LPS increases NOS2 mRNA expression in muscle via a TLR4-dependent mechanism.

UR - http://www.scopus.com/inward/record.url?scp=8644226134&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=8644226134&partnerID=8YFLogxK

U2 - 10.1152/ajpcell.00010.2004

DO - 10.1152/ajpcell.00010.2004

M3 - Article

VL - 287

JO - American Journal of Physiology

JF - American Journal of Physiology

SN - 0363-6143

IS - 6 56-6

ER -