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Objective. To analyze the differences in gene expression profiles of chondrocytes in intact and damaged regions of cartilage from the same knee joint of patients with osteoarthritis (OA) of the knee.Methods. We compared messenger RNA expression profiles in regions of intact and damaged cartilage (classified according to the Mankin scale) obtained from patients with knee OA. Five pairs of intact and damaged regions of OA cartilage were evaluated by oligonucleotide array analysis using a double in vitro transcription amplification technique. The microarray data were confirmed by real-time quantitative polymerase chain reaction (PCR) amplification and were compared with previously published data.Results. About 1,500 transcripts, which corresponded to 8% of the expressed transcripts, showed >2-fold differences in expression between the cartilage tissue pairs. Approximately 10% of these transcripts (n ‫؍‬ 151) were commonly expressed in the 5 patient samples. Accordingly, 114 genes (35 genes expressed in intact > damaged; 79 genes expressed in intact < damaged) were selected. The expression of some genes related to the wound-healing process, including cell proliferation and interstitial collagen synthesis, was higher in damaged regions than in intact regions, similar to the findings for genes that inhibit matrix degradation. Comparisons of the real-time quantitative PCR data with the previously reported data support the validity of our microarray data.Conclusion. Differences between intact and damaged regions of OA cartilage exhibited a similar pattern among the 5 patients examined, indicating the presence of common mechanisms that contribute to cartilage destruction. Elucidation of this mechanism is important for the development of effective treatments for OA.

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Objective. Membrane-associated prostaglandin E synthase (mPGES) is a recently identified terminal enzyme of the arachidonic acid cascade, which converts PGH 2 to PGE 2 in rheumatoid arthritis synovial fibroblasts (RASFs). This study was undertaken to investigate factors regulating the expression of mPGES.Methods. RASFs were treated with interleukin-1␤ (IL-1␤), indomethacin, NS-398, rofecoxib, or meloxicam. The effects of PGE 2 and selective agonists for PGE 2 receptor subtypes (EP1, EP2, EP3, and EP4) were also studied. Expression of mPGES messenger RNA (mRNA) and protein was measured by Northern and Western blot analysis, respectively. EP receptor mRNA expression in RASFs was determined by reverse transcriptase-polymerase chain reaction. Production of PGE 2 and cAMP was measured by enzyme-linked immunosorbent assay.Results. The enhanced expression of mPGES mRNA and protein in IL-1␤-stimulated RASFs was attenuated by the addition of indomethacin, NS-398, rofecoxib, or meloxicam. This reduction of expression was reversed by PGE 2 . IL-1␤-induced PGES activity, measured by conversion of PGH 2 to PGE 2 , was decreased by rofecoxib. EP2 and EP4 receptor mRNA was detected in RASFs. EP2 and EP4 agonists, as well as PGE 2 , restored the inhibitory effect of rofecoxib on mPGES expression. The effect of PGE 2 was mimicked by forskolin, a direct activator of adenylate cyclase. Intracellular cAMP was increased by IL-1␤ and was inhibited by rofecoxib. Conclusion. Enhancement of mPGES expression by PGE 2 via the EP2/EP4 receptors with an increase in cAMP may play an important role in articular inflammation in patients with RA. It also seems that cyclooxygenase 2 (COX-2) inhibitors decrease PGE 2 production not only by direct inhibition of COX-2, but also by reducing mPGES expression in activated RASFs.Prostaglandin E synthase (PGES) catalyzes the final step of PGE 2 production, converting PGH 2 to PGE 2 . PGES exists downstream of cyclooxygenase (COX) (1,2). At least 2 forms of PGES, membraneassociated PGES (mPGES) (3-5) and cytosolic PGES (cPGES) (6), have recently been cloned and characterized. In cotransfection experiments, functional coupling between inducible mPGES and COX-2, as well as between constitutive cPGES and COX-1, has been demonstrated (4,6).Originally called microsomal glutathione S-transferase 1-like 1 (MGST1-L1), mPGES is a glutathione-dependent enzyme that shows coordinated induction with COX-2 in various cells and tissues by inflammatory stimuli (3,4). We recently showed that mPGES and COX-2 were induced by interleukin-1␤ (IL-1␤) on the perinuclear membrane in rheumatoid arthritis synovial fibroblasts (RASFs), causing an increase in PGE 2 production at sites of inflammation (7).

Early apoptotic Jurkat T cells undergo capping of CD43, and its polylactosaminyl saccharide chains serve as ligands for phagocytosis by macrophages. This suggests the presence of a polylactosaminoglycan-binding receptor on macrophages. Here we show that this receptor is nucleolin, a multifunctional shuttling protein present in nucleus, cytoplasm, and on the surface of some types of cells. Nucleolin was detected at the surface of macrophages, and antinucleolin antibody inhibited the binding of the early apoptotic cells to macrophages. Nucleolin-transfected HEK293 cells expressed nucleolin on the cell surface and bound the early apoptotic cells but not phosphatidylserine-exposing late apoptotic cells. This binding was inhibited by anti-nucleolin antibody, by polylactosamine-containing oligosaccharides, and by anti-CD43 antibody. Deletion of the antibody binding region of nucleolin resulted in loss of the apoptotic cell-binding ability. Moreover, truncated recombinant nucleolin in solution containing this region blocked the apoptotic cell binding to macrophages, and the blocking effect was cancelled by the oligosaccharides. These results indicate that nucleolin is a macrophage receptor for apoptotic cells.Macrophages and other phagocytes recognize and ingest apoptotic cells in tissue, preventing their lysis and subsequent release of harmful or immunogenic intracellular components. Therefore, clearance of apoptotic cells by phagocytes is crucial in the maintenance of tissue turnover and homeostasis. Moreover, it has been suggested that the apoptotic cell-ingested phagocytes play a role in suppression or resolution of inflammation (1-5).Cells undergoing apoptosis display a variety of "eat me" signals, namely cell-surface changes to be recognized by phagocytes. These include externalization of phosphatidylserine (PS),3 as yet little identified alterations of carbohydrates, and unidentified alterations of other membrane components such as intercellular adhesion molecule-3 and "thrombospondin binding sites" (1-5). Among these, the most common and the best-characterized change is externalization of PS, although the mechanism of externalization has not been fully understood (6, 7).In contrast to the poor understanding of the nature of eat me signals, various proteins of phagocyte membrane or in extracellular fluid have been reported as receptors or bridging factors for apoptotic cells. For externalized PS on apoptotic cells, CD36 (8), CD68 (9), CLA-1 (10), LOX-1 (11), and PS receptor (12) have been reported as phagocyte receptors; a serum protein ␤ 2 -glycoprotein I (13), complement component C3bi (14), and milk fat globule-epidermal growth factor-factor 8 (15) have been reported to bridge apoptotic cells and phagocytes through exposed PS. For intercellular adhesion molecule-3 on apoptotic cells, CD14 was suggested to be a macrophage receptor (16). Thrombospondin is also known as a bridging protein between unidentified sites on apoptotic cells and CD36 or vitronectin receptor (␣ v ␤ 3 integrin) on phagocytes (17). In addi...

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