Can altered production of interleukin-1β, interleukin-6, transforming growth factor-β and prostaglandin E2 by isolated human subchondral osteoblasts identify two subgroups of osteoarthritic patients

AbstractObjective To determine the capacity of human subchondral osteoarthritic osteoblasts (Ob) to produce interleukin (IL)-1β, IL-6, transforming growth factor-β (TGF-β) and prostaglandin E2 (PGE2), and determine if a relationship exists between IL-1β, TGF-β, PGE2 and IL-6 production.Methods We measured the abundance of IL-1β, IL-6, TGF-β and PGE2 using very sensitive ELISA in conditioned-media of human primary subchondral Ob from normal individuals and osteoarthritic patients. Selective inhibition of IL-6 or IL-6 receptor signaling was performed to determine its effect on PGE2 production whereas the inhibiton of PGE2 production was performed to determine its effect on IL-6 production. The expression of bone cell markers and urokinase plasminogen activator (uPA) activity was also determined.Results Osteoarthritic Ob produced all these factors with greater variability than normal cells. Interestingly, the production of IL-6 and PGE2 by osteoarthritic Ob separated patients into two subgroups, those whose Ob produced levels comparable to normal (low producers) and those whose Ob produced higher levels (high producers). In those cells classified as high osteoarthritic Ob, PGE2 and IL-6 levels were increased two- to three-fold and five- to six-fold, respectively, compared with normal. In contrast, while using their IL-6 and PGE2 production to separate osteoarthritic Ob into low and high producers, we found that IL-1β levels were similar in normal and all osteoarthritic Ob. Using the same criteria, TGF-β levels were increased in all osteoarthritic Ob compared with normal. Reducing PGE2 synthesis by Indomethacin [a cyclo-oxygenase (COX) -1 and -2 inhibitor] reduced IL-6 levels in all osteoarthritic Ob, whereas Naproxen (a more selective COX-2 inhbitor) reduced PGE2 and IL-6 levels only in the high osteoarthritic group. Conversely, PGE2 addition to osteoarthritic Ob enhanced IL-6 production in both groups. Moreover, the addition of parathyroid hormone also stimulated IL-6 production to similar normal levels in both osteoarthritic groups. In contrast, using an antibody against IL-6 or IL-6 receptors did not reduce PGE2 levels in either group. The evaluation of alkaline phosphatase activity, osteocalcin release, collagen type I and uPA activity in osteoarthritic Ob failed to show any differences between these cells regardless to which subgroup they were assigned.Conclusions These results indicate that IL-6 and PGE2 production by subchondral Ob can discriminate two subgroups of osteoarthritic patients that cannot otherwise be separated by their expression of cell markers, and that endogenous PGE2 levels influence IL-6 synthesis in osteoarthritic Ob. Copyright 2002 OsteoArthritis Research Society International. Published by Elsevier Science Ltd. All rights reserved

Peroxisome proliferator-activated receptor γ1 expression is diminished in human osteoarthritic cartilage and is downregulated by interleukin-1β in articular chondrocytes

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor involved in the regulation of many cellular processes. We and others have previously shown that PPARγ activators display anti-inflammatory and chondroprotective properties in vitro and improve the clinical course and histopathological features in an experimental animal model of osteoarthritis (OA). However, the expression and regulation of PPARγ expression in cartilage are poorly defined. This study was undertaken to investigate the quantitative expression and distribution of PPARγ in normal and OA cartilage and to evaluate the effect of IL-1β, a prominent cytokine in OA, on PPARγ expression in cultured chondrocytes. Immunohistochemical analysis revealed that the levels of PPARγ protein expression were significantly lower in OA cartilage than in normal cartilage. Using real-time RT-PCR, we demonstrated that PPARγ1 mRNA levels were about 10-fold higher than PPARγ2 mRNA levels, and that only PPARγ1 was differentially expressed: its levels in OA cartilage was 2.4-fold lower than in normal cartilage (p < 0.001). IL-1 treatment of OA chondrocytes downregulated PPARγ1 expression in a dose- and time-dependent manner. This effect probably occurred at the transcriptional level, because IL-1 decreases both PPARγ1 mRNA expression and PPARγ1 promoter activity. TNF-α, IL-17, and prostaglandin E2 (PGE2), which are involved in the pathogenesis of OA, also downregulated PPARγ1 expression. Specific inhibitors of the mitogen-activated protein kinases (MAPKs) p38 (SB203580) and c-Jun N-terminal kinase (SP600125), but not of extracellular signal-regulated kinase (PD98059), prevented IL-1-induced downregulation of PPARγ1 expression. Similarly, inhibitors of NF-κB signaling (pyrrolidine dithiocarbamate, MG-132, and SN-50) abolished the suppressive effect of IL-1. Thus, our study demonstrated that PPARγ1 is downregulated in OA cartilage. The pro-inflammatory cytokine IL-1 may be responsible for this downregulation via a mechanism involving activation of the MAPKs (p38 and JNK) and NF-κB signaling pathways. The IL-1-induced downregulation of PPARγ expression might be a new and additional important process by which IL-1 promotes articular inflammation and cartilage degradation