TY - JOUR
T1 - Peptidoglycan enhances proinflammatory cytokine expression through the TLR2 receptor, MyD88, phosphatidylinositol 3-kinase/AKT and NF-kappaB pathways in BV-2 microglia
AU - Lin, Hsiao Yun
AU - Tang, Chih Hsin
AU - Chen, Yi Hung
AU - Wei, I. Hua
AU - Chen, Jia Hong
AU - Lai, Chih Ho
AU - Lu, Dah Yuu
PY - 2010/8
Y1 - 2010/8
N2 - In this study, we investigated the signaling pathways involved in inflammatory production caused by peptidoglycan (PGN), a cell wall component of the gram-positive bacterium, in BV-2 microglia. PGN caused a concentration- and time-dependent increase in inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA and protein levels. In addition, PGN also induced IL-1β, TNF-α and IL-6 mRNA up-regulation in a concentration-dependent manner. Moreover, PGN also increased Toll-like receptor 2 (TLR2) expression in BV-2 microglia. Administration of TLR2 neutralizing antibody effectively inhibited PGN-induced iNOS and COX-2 expression. On the other hand, PGN-induced iNOS and COX-2 up-regulation were attenuated by PI3-kinase inhibitors (LY294002 and wortmannin), and an AKT inhibitor. Treatment of BV-2 microglia with PGN caused a time-dependent activation of PI3-kinase (p85) and AKT. PGN-induced PI3-kinase/AKT activation, iNOS and COX-2 expression were also inhibited by MyD88 inhibitory peptide. Treatment of cells with NF-κB inhibitor (pyrrolidine dithiocarbamate), IκBα phosphorylation inhibitor (Bay 117082), or IκB protease inhibitor (l-1-tosylamido-2-phenylethyl chloromethyl ketone) inhibited PGN-induced iNOS and COX-2 expression. Furthermore, stimulation of cells with PGN also activated IKKκ/α, IκBα phosphorylation, IκBα degradation, p65 phosphorylation at Ser536, and increased κB-luciferase activity. PGN-induced IKKα/κ phosphorylation, IκB7α phosphorylation, and IκBα degradation were further inhibited by pre-treatment with PI3-kinase inhibitors. Moreover, PGN-mediated increase of κB-luciferase activity was also inhibited by pre-transfection with dominant-negative mutants of p85, AKT, IKKα or IKKβ. Our data demonstrate that PGN-induced iNOS, COX-2 and proinflammatory cytokine expression was mediated through the TLR2/MyD88/PI3-kinase/AKT pathway, which in turn initiates IKKα/β and NF-κB activation in BV-2 microglia.
AB - In this study, we investigated the signaling pathways involved in inflammatory production caused by peptidoglycan (PGN), a cell wall component of the gram-positive bacterium, in BV-2 microglia. PGN caused a concentration- and time-dependent increase in inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA and protein levels. In addition, PGN also induced IL-1β, TNF-α and IL-6 mRNA up-regulation in a concentration-dependent manner. Moreover, PGN also increased Toll-like receptor 2 (TLR2) expression in BV-2 microglia. Administration of TLR2 neutralizing antibody effectively inhibited PGN-induced iNOS and COX-2 expression. On the other hand, PGN-induced iNOS and COX-2 up-regulation were attenuated by PI3-kinase inhibitors (LY294002 and wortmannin), and an AKT inhibitor. Treatment of BV-2 microglia with PGN caused a time-dependent activation of PI3-kinase (p85) and AKT. PGN-induced PI3-kinase/AKT activation, iNOS and COX-2 expression were also inhibited by MyD88 inhibitory peptide. Treatment of cells with NF-κB inhibitor (pyrrolidine dithiocarbamate), IκBα phosphorylation inhibitor (Bay 117082), or IκB protease inhibitor (l-1-tosylamido-2-phenylethyl chloromethyl ketone) inhibited PGN-induced iNOS and COX-2 expression. Furthermore, stimulation of cells with PGN also activated IKKκ/α, IκBα phosphorylation, IκBα degradation, p65 phosphorylation at Ser536, and increased κB-luciferase activity. PGN-induced IKKα/κ phosphorylation, IκB7α phosphorylation, and IκBα degradation were further inhibited by pre-treatment with PI3-kinase inhibitors. Moreover, PGN-mediated increase of κB-luciferase activity was also inhibited by pre-transfection with dominant-negative mutants of p85, AKT, IKKα or IKKβ. Our data demonstrate that PGN-induced iNOS, COX-2 and proinflammatory cytokine expression was mediated through the TLR2/MyD88/PI3-kinase/AKT pathway, which in turn initiates IKKα/β and NF-κB activation in BV-2 microglia.
KW - COX-2
KW - INOS
KW - Microglia
KW - NF-κB
KW - Peptidoglycan
UR - http://www.scopus.com/inward/record.url?scp=77954956788&partnerID=8YFLogxK
U2 - 10.1016/j.intimp.2010.04.026
DO - 10.1016/j.intimp.2010.04.026
M3 - 文章
C2 - 20451669
AN - SCOPUS:77954956788
SN - 1567-5769
VL - 10
SP - 883
EP - 891
JO - International Immunopharmacology
JF - International Immunopharmacology
IS - 8
ER -