In this study, we have identified the source of nitric oxide (NO) produced in the human inflammatory joints by analyzing expression of inducible NO synthase. In ex vivo organ cultures, both inflammatory synovium and cartilage from patients with rheumatoid arthritis produced NO. The NO production was suppressed by NG-monomethyl-L-arginine, an inhibitor of NO synthase. The amount of NO produced by the synovium correlated with the proportion of CD14+ cells in the corresponding tissue (r = 0.8, P < 0.05). Immunohistochemical analysis as well as in situ hybridization showed that inducible NO synthase was predominantly expressed in synovial lining cells, endothelial cells, chondrocytes, and to a lesser extent, in infiltrating mononuclear cells and synovial fibroblasts. The synovial lining cells and the infiltrating cells expressing inducible NO synthase were identified where CD14+ cells were located. Together with morphological features, this suggests that they are type A synoviocytes. NO production from freshly isolated synoviocytes and chondrocytes was up-regulated by in vitro stimulation with a combination of IL-TNF-fB, TNF-a, and LPS.In summary, the present results suggest that NO is produced primarily by CD14+ synoviocytes, chondrocytes, and endothelial cells in inflammatory joints of arthritides. NO production can be upregulated by cytokines present in inflamed joints. The increased NO production may thus contribute to the pathological features in inflammatory arthriti-

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It is hypothesized that infectious prions are generated as the cellular form of the prion protein (PrP(C)) undergoes pronounced conformational change under the direction of an infectious PrP(Sc) template. Conversion to the infectious conformer is particularly associated with major structural rearrangement in the central portion of the protein (residues 90-120), which has an extended flexible structure in the PrP(C) isoform. Using a panel of recombinant antibodies reactive with different parts of PrP, we show that equivalent major structural rearrangements occur spontaneously in this region of PrP immobilized on a surface. In contrast, regions more towards the termini of the protein remain relatively unaltered. The rearrangements occur even under conditions where individual PrP molecules should not contact one another. The propensity of specific unstructured regions of PrP to spontaneously undergo large and potentially deleterious conformational changes may have important implications for prion biology.

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Objective. To investigate the expressions of costhulatory molecules CD86 (B7-2, B70) and CD28 by cells obtained from the synovial tissues (ST) and synovial fluids (SF) of patients with rheumatoid arthritis (RA).Methods. Monoclonal antibodies (MAb) against CD86 and CD28 were used for immunochemical study of synovia from 18 RA patients, 4 osteoarthritis (OA) patients, and 4 normal subjects. These MAb were also used for flow cytometry of isolated ST cells from 8 RA and 5 OA patients and of SF mononuclear cells from 5 RA and 5 OA patients.Results. Immunohistochemical examination revealed that CD86+ cells occurred in 11 of the 18 RA synovia, but in none of the 4 OA or 4 normal synovia. Most of the positive cells had macrophage-like morphology, and surrounded lymphoid aggregates. Most cells within lymphoid aggregates were stained positively for CD28. Flow cytometry showed that CD86+ cells comprised 2.9-33.4% (average 14.3%) of the total ST cells and 2.1-14.9% (average 6.1%) of the total SF mononuclear cells from RA patients. Approximately 40% of the CD86+ cells expressed CD14. A majority (mean 72%, range 57439%) of the T cells in ST and SF expressed CD28. RA synovia expressed more CD86 molecules than did OA synovia (mean frequency of positive cells 14.3% versus 2.8%; mean fluorescence intensity 104.6 versus 40.9).Conclusion. This study has shown that the exMing Fei Liu, MD: National Cheng Kung University, Tainan, Taiwan, Republic of China; Hitoshi Kohsaka, MD, Hiroshi Sakurai, BSc, Ichiro Saito, DDS, PhD, Nobuyuki Miyasaka, MD: Tokyo Medical and Dental University, Tokyo, Japan; Miyuki Azuma, PhD, KO Okumura, MD: School of Medicine, Juntendo University, Tokyo, Japan.Address reprint requests to Ming Fei Liu, MD, Department of Internal Medicine, National Cheng Kung University, Tainan, Taiwan, Republic of China.Submitted for publication February 17, 1995; accepted in revised form June 23, 1995. pression of the CD86 molecule was up-regulated in RA synovia. The CD86+ c e k appeared macrophage-like and surrounded CD28+ cells in lymphoid aggregates. The simultaneous presence and close localization of CD86+ and CD28+ cells in the RA synovial compartment suggests that their interaction potentially contributes to the sustained immune activation of RA.Optimal T cell activation requires a costimulatory signal in addition to the primary signal provided by T cell receptor cross-linkage (1). The earliest example of the costimulatory signals discovered was the interaction between the CD28 on T lymphocytes and the B7 on antigen-presenting cells (2-5). Recently, another molecule, CTLA-4, was found to have homologous structure with CD28 and to bind to B7 (6,7). There is evidence that ligands of CD28 and CTLA-4 consist of CD80 (B7-1) and CD86 (B70, B7-2) (8-13). It has been shown that CD86 is constitutively expressed on fresh monocytes, whereas CD80 requires further lipopolysaccharide (LPS) stimulation. Compared with CDHO, CD86 on monocytes and B cells are upregulated at much higher levels and more rapidly by activation with interferon...