Pro‐inflammatory IL‐1beta and/or TNF‐alpha up‐regulate matrix metalloproteases‐1 and ‐3 mRNA in chondrocyte subpopulations potentially pathogenic in osteoarthritis: in situ hybridization studies on a single cell level

Abstract Aim In osteoarthritis chondrocytes, matrix metalloproteases ( MMP s) and their inhibitors are induced by interleukin ( IL )‐1beta or tumor necrosis factor ( TNF )‐alpha and balanced by inhibitors, but their messenger RNA ( mRNA ) expression has not been studied in individual cells. Methods Normal articular chondrocytes (10 donors; age 50 ± 6 years, mean ± SEM ) were stimulated in a monolayer for 24 h with IL ‐1beta, TNF ‐alpha, or transforming growth factor ( TGF )‐beta1 (10 ng/mL each), alone or in combination. m RNA expression for MMP ‐1, MMP ‐3 and tissue inhibitor of metalloproteinase ( TIMP )‐1 was studied by in situ hybridization ( 35 S ‐c RNA ) and quantitative reverse transcription polymerase chain reaction ( RT ‐ PCR ) ( n ≥ 3 each). Results Whereas < 5% chondrocytes constitutively expressed MMP ‐1, a higher percentage expressed MMP ‐3 and TIMP ‐1 (31.1 ± 1.8%; 36.7 ± 2.8%, respectively). Upon stimulation with IL ‐1beta, TNF ‐alpha or IL ‐1beta/ TNF ‐alpha, the percentage of cells positive for MMP ‐1, MMP ‐3 and TIMP ‐1 rose significantly ( IL ‐1beta: 31.5%, 54.5% and 60.2%, respectively; TNF ‐alpha: 35.4%, 56.6%, 50.9%; IL ‐1beta/ TNF ‐alpha: 38.8%, 45.2%, 52.1%). In bulk population ( RT ‐ PCR ), m RNA for MMP ‐1 and MMP ‐3 was also induced by IL ‐1beta (11.9‐fold, 1.2‐fold, respectively), TNF ‐alpha (4.8‐fold, 1.0‐fold) or IL ‐1beta/ TNF ‐alpha (14.7‐fold, 1.4‐fold), an effect attenuated by TGF ‐beta1. TIMP ‐1 m RNA , in contrast, was down‐regulated by IL ‐1beta, TNF ‐alpha or IL ‐1beta/ TNF ‐alpha, an effect again partially reverted by TGF ‐beta1. Finally, collagen type II m RNA was down‐regulated by IL ‐1beta, TNF ‐alpha or IL ‐1beta/ TNF ‐alpha (by 90%, 50% and 98%, respectively) and that of collagen type I was up‐regulated (5.7‐fold, 3.0‐fold, 3.7‐fold). Conclusions Up‐regulation of MMP ‐1/ MMP ‐3 by IL ‐1beta and/or TNF ‐alpha in a fraction of chondrocytes in vitro suggests that a subpopulation of catabolic cells may also exist in osteoarthritis. These cells may undergo considerable dedifferentiation, as indicated by a decreased collagen‐ II /collagen‐ I ratio.