Abstract
BackgroundMatrix metalloproteinase-9 (MMP-9) has been shown to be induced by cytokines including TNF-α and may contribute to bone inflammatory diseases. However, the mechanisms underlying MMP-9 expression induced by TNF-α in MC3T3-E1 cells remain unclear.ResultsWe applied gelatin zymography, Western blot, RT-PCR, real-time PCR, selective pharmacological inhibitors of transcription (actinomycin D, Act.D), translation (cycloheximide, CHI), c-Src (PP1), MEK1/2 (U0126), p38 MAPK (SB202190), JNK1/2 (SP600125), and NF-κB (Bay11-7082), respective siRNAs transfection, promoter assay, immunofluorescence staining, and ELISA to investigate the MMP-9 expression and soluble ICAM-1 (sICAM-1) release induced by TNF-α in MC3T3-E1 cells. Here we demonstrated that TNF-α-induced MMP-9 expression was attenuated by Act.D, CHI, PP1, U0126, SB202190, SP600125, and Bay11-7082, and by the transfection with siRNAs for ERK2, p38 MAPK, and JNK2. TNF-α-stimulated TNFR1, TRAF2, and c-Src complex formation was revealed by immunoprecipitation and Western blot. Furthermore, TNF-α-stimulated NF-κB phosphorylation and translocation were blocked by Bay11-7082, but not by PP1, U0126, SB202190, or SP600125. TNF-α time-dependently induced MMP-9 promoter activity which was also inhibited by PP1, U0126, SB202190, SP600125, or Bay11-7082. Up-regulation of MMP-9 was associated with the release of sICAM-1 into the cultured medium, which was attenuated by the pretreatment with MMP-2/9i, an MMP-9 inhibitor.ConclusionsIn this study, we demonstrated that TNF-α up-regulates MMP-9 expression via c-Src, MAPKs, and NF-κB pathways. In addition, TNF-α-induced MMP-9 expression may contribute to the production of sICAM-1 by MC3T3-E1 cells. The interplay between MMP-9 expression and sICAM-1 release may exert an important role in the regulation of bone inflammatory diseases.
Highlights
Matrix metalloproteinase-9 (MMP-9) has been shown to be induced by cytokines including tumor necrosis factor-α (TNF-α) and may contribute to bone inflammatory diseases
We further investigated whether TNF-α-induced Matrix metalloproteinases (MMPs)-9 expression is mediated through transcription and translation, a transcription inhibitor Act.D and a translation inhibitor CHI were used for these purposes
TNF-α stimulates two independent pathways: c-Srcdependent mitogen-activated protein kinases (MAPKs) and Nuclear factorkappa B (NF-κB)-dependent cascades in MC3T3-E1 cells According to the above data, we have demonstrated that TNF-α induced MMP-9 expression via activation of cSrc, ERK1/2, p38 MAPK, JNK1/2, and NF-κB in
Summary
Matrix metalloproteinase-9 (MMP-9) has been shown to be induced by cytokines including TNF-α and may contribute to bone inflammatory diseases. Matrix metalloproteinases (MMPs) are a family of extracellular matrix (ECM)-degrading enzymes and induced by different stimuli including growth factors, cytokines, and tumor promoters [1]. The important roles of MMPs have been demonstrated in bone using various approaches for ossification, remodeling, and destruction. Co-culture of osteoarthritis (OA) subchondral bone osteoblasts with normal articular cartilage chondrocytes resulted in significantly increased the expression of MMP-2 and MMP-9 [8]. These studies have indicated that the expression of MMP-9 may be up-regulated during bone inflammation
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