CCAAT/ENHANCER binding protein β mediates expression of matrix metalloproteinase 13 in human articular chondrocytes in inflammatory arthritis

Abstract

Objective. To determine the function of CCAAT/ enhancer binding protein ␤ (C/EBP␤) in the expression of matrix metalloproteinase 13 (MMP-13) in chondrocytes in inflammatory arthritis.Methods. Cartilage obtained from patients with rheumatoid arthritis and osteoarthritis was immunostained for expression of C/EBP␤ or MMP-13. Interleukin-1␤-or tumor necrosis factor ␣ (TNF␣)-stimulated chondrocytes were subjected to Western blotting and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). MMP-13 promoter assays were conducted, and the C/EBP␤ response element was characterized by deletion and mutation analysis. C-28/I2 cells were treated with TNF␣ and subjected to chromatin immunoprecipitation (ChIP) assays. Finally, C/EBP␤-liver-enriched activator protein (LAP) was overexpressed in C-28/I2 cells or cartilage tissues, and MMP-13 expression was analyzed.Results. C/EBP␤ and MMP-13 expression was colocalized in chondrocytes in arthritic cartilage. MMP-13 promoter activity was stimulated by C/EBP␤ overexpression in a dose-dependent manner. Luciferase assays revealed that a -981-bp promoter had the greatest activity, while deletion to -936 bp strongly diminished promoter activity. Luciferase activity was repressed to basal levels by mutations in potential C/EBP binding sites. The stimulatory effects of C/EBP␤ overexpression were diminished by mutation. ChIP assays revealed that TNF␣ treatment enhanced the binding of C/EBP␤ to the MMP-13 promoter. When C/EBP␤-LAP was overexpressed in C-28/I2 cells, endogenous MMP-13 expression was stimulated up to 32-fold as detected by real-time RT-PCR. Furthermore, following adenoviral overexpression of C/EBP␤-LAP in organ culture of articular cartilage, stimulation of MMP-13 was also detected by immunohistochemistry. Conclusion. C/EBP␤ directly binds to the MMP-13 promoter region and stimulates the expression of MMP-13 in chondrocytes in inflammatory arthritis.Degeneration of articular cartilage is the principal process causing disability in patients with inflammatory arthritis, including those with rheumatoid arthritis (RA) (1,2) and osteoarthritis (OA) (3,4). Normally, maintenance of the cartilage extracellular matrix (ECM) is strictly regulated by the balance of anabolic and catabolic factors secreted by synovial cells or by chondrocytes themselves. Arthritic cartilage is characterized by excessive production of proteinases, such as matrix metalloproteinases (MMPs) and aggrecanases, and reduced synthesis of new matrix by chondrocytes, which disturbs the balance of anabolic and catabolic factors. Among the proteinases, MMP-13 cleaves a broad range of substrates, including fibrillar type II collagen and type II procollagen as well as gelatin, types I, III, IV, IX, X, and XIV collagen, tenascin, fibronectin, fibronectin fragments, and aggrecan (5-8). It has been reported that MMP-13 has the highest degradation activity for type II collagen (5,9). A recent study showed that up-regulated transgenic expression of MMP-13 in mouse articular cartilage resulted in pathologic chan...

CCAAT/Enhancer-binding Protein β Regulates the Repression of Type II Collagen Expression during the Differentiation from Proliferative to Hypertrophic Chondrocytes

“…7). MMP13 is known to be a major degrading enzyme of COL2A1, and we previously reported that C/EBP␤ is a direct inducer of MMP13 (22). Increase in the expression of MMP13 could be involved in the degradation of COL2A1 in C/EBP␤-infected tibias.…”

“…C/EBP␤ is known to be induced by pro-inflammatory cytokines such as IL-1␤, IL-6, and TNF-␣, and it is expressed in arthritic cartilage (15,20,22). It has been suggested that C/EBP␤ also plays a role in the repression of COL2A1 in response to inflammatory signals (20).…”

“…Previously, it was reported that C/EBP␤, in response to IL-1␤, down-regulated cartilage-derived retinoic acid-sensitive protein (Cd-rap), a small secreted protein expressed in cartilage throughout chondrogenesis and in mature chondrocytes (20). C/EBP␤ induced by pro-inflammatory cytokines such as IL-1␤ and TNF-␣ directly binds to the MMP13 and MMP3 promoter regions and stimulates their expression in chondrocytes, resulting in degradation of cartilage in arthritis (21,22). Furthermore, it was reported that hypertrophic differentiation of chondrocytes was delayed in C/EBP␤ knock-out mice through transactivation of cell cycle factor p57 (23).…”

C/EBPβ and Nuclear Factor of Activated T Cells Differentially Regulate Adamts-1 Induction by Stimuli Associated with Vascular Remodeling

“…The C/EBP␤-binding sites in the Adamts1 promoter have also been involved in its induction by progesterone receptor-mediated activation of granulosa cells (29), further supporting the relevance of C/EBP␤ proteins in the regulation of Adamts-1 expression. Furthermore, active C/EBP␤ proteins have been linked to the expression of other metalloproteinase proteins, including MMP-3, MMP-13, and Adamts-5, during arthritic tissue remodeling (51,52).…”

GADD45β Enhances Col10a1 Transcription via the MTK1/MKK3/6/p38 Axis and Activation of C/EBPβ-TAD4 in Terminally Differentiating Chondrocytes

“…The Gadd45␤ and C/ebp␤ knock-out mice display similar phenotypic changes in the embryonic growth plate, characterized by a delay in terminal chondrocyte differentiation along with decreased expression of genes linked to this process, including Col10a1 and Mmp13 (8,37,69). In addition, the embryonic limbs of C/ebp␤ knock-out mice display impaired osteogenesis (53).…”