The differential effects of IL-1 and TNF-α on proinflammatory cytokine and matrix metalloproteinase expression in human chondrosarcoma cells

Abstract

Objective and design:Interleukin-1 (IL-1), tumor necrosis factor-α (TNF-α), 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-α on gene expression in the human chondrosarcoma cell line, SW1353.

Materials and methods:SW1353 cells were stimulated singularly with IL-1α, TNF-α, 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-α. The expression pattern of proIL-1β, MMP-1, and MMP-13 in chondrocytes is differentially regulated when stimulated with proinflammatory cytokines. IL-1, but not TNF-α, 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-1β is produced in the proform in IL-1α-activated chondrosarcoma cells and that additional signals are required for its posttranslational processing/activation.

Conclusions:IL-1 and TNF-α 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-1α appears to serve an important role in the pathogenesis OA contributing to joint inflammation and cartilage destruction.