The differential effects of IL-1 and TNF-alpha on proinflammatory cytokine and matrix metalloproteinase expression in human chondrosarcoma cells.
OBJECTIVE
Interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-alpha), and matrix metalloproteinases (MMPs) play important roles in the pathogenesis of osteoarthritis (OA). In the present study, using Affymetrix oligonucleotide array technology and real-time quantitative RT-PCR we have investigated the molecular mechanisms underlying the differential effect of IL-1 and TNF-alpha on gene expression in the human chondrosarcoma cell line, SW1353.


METHODS
SW1353 cells were stimulated singularly with IL-1alpha, TNF-alpha, Phorbol 12-myristate 13-acetate (PMA), or treated with the combination of cytokine and PMA. Total RNA was collected at multiple time points over a 24-h period followed by biotinylated cRNA target preparation and hybridization onto the Affymetrix HG-U95Av2 array. The differential expression patterns of several cytokine and MMP genes were further confirmed by real time quantitative RT-PCR, Western blot, and ELISA.


RESULTS
Our microarray experiments have broadly confirmed previously published data on chondrocyte gene expression regulated by IL-1 and TNF-alpha. The expression pattern of proIL-1beta, MMP-1, and MMP-13 in chondrocytes is differentially regulated when stimulated with proinflammatory cytokines. IL-1, but not TNF-alpha, can induce IL-6, bone morphogenic protein 2 (BMP-2), and cyclooxygenase (COX-2) expression in SW1353 cells. Additionally, our Western blot results provide the first evidence that IL-1beta is produced in the proform in IL-1alpha-activated chondrosarcoma cells and that additional signals are required for its posttranslational processing/activation.


CONCLUSIONS
IL-1 and TNF-alpha each activate a distinct set of genes in chondrosarcoma cells, and gene expression in these cells is regulated by groups of genes related in part by their function. Chondrocyte IL-1alpha appears to serve an important role in the pathogenesis OA contributing to joint inflammation and cartilage destruction.
Shi
J
J
Department of Antibacterial, Immunology, and Inflammation, Pfizer Global Research and Development, Groton, CT 06340, USA.
Schmitt-Talbot
E
E
DiMattia
D A
DA
Dullea
R G
RG
D045744:Cell Line, Tumor; D002813:Chondrosarcoma / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D016000:Cluster Analysis; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics*; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D007249:Inflammation / Q000235:genetics / Q000378:metabolism; D018836:Inflammation Mediators / Q000097:blood / Q000378:metabolism; D007375:Interleukin-1 / Q000096:biosynthesis / Q000494:pharmacology*; D020782:Matrix Metalloproteinases / Q000096:biosynthesis / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D016207:Cytokines; D018836:Inflammation Mediators; D007375:Interleukin-1; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D020782:Matrix Metalloproteinases; D013755:Tetradecanoylphorbol Acetate