Tadanobu Morimoto

Measurement of soluble Fcγ receptor type IIIa derived from macrophages in plasma : Increase in patients with rheumatoid arthritis

FcγRIII (CD16) is found in two alternative forms, a transmembrane FcγRIIIa expressed on NK cells and macrophages, and a glycosylphosphatidylinositol‐linked FcγRIIIb present on neutrophils. Previously, we measured soluble FcγRIIIa (sFcγRIIIa) in plasma of NA(1 +, 2‐) phenotyped donors with the anti‐FcγRIII monoclonal antibody (MoAb) GRM1, which recognizes NA2‐FcγRIIIb and FcγRIIIa. The level of sFcγRIIIa, as well as the total sFcγRIII (sFcγRIIIa plus sFcγRIIIb) in patients with rheumatoid arthritis (RA) was significantly higher than that in healthy controls. In this study, we measured sFcγRIIIaMφ in plasma with a newly developed anti‐FcγRIII MoAb, MKGR14 (mIgM), which recognizes FcγRIIIaMφ specifically. From the recovery of purified sFcγRIIIaMφ, the amount of sFcγRIIIaMφ present was about half that of sFcγRIIIaNK, and that of sFcγRIIIa was about 50 times lower than that of sFcγRIIIb in pooled plasma from healthy NA(1 +, 2‐) phenotyped donors. The level of sFcγRIIIaMφ in RA patients was about four times higher than that in healthy controls. In RA patients, both the sFcγRIIIaMφ and sFcγRIIIa levels were increased as proportionally as the Lansbury Index. The sFcγRIIIa, but not sFcγRIIIaMφ levels, were increased directly proportional to C‐reactive protein. sFcγRIIIaMφ may be a novel marker of disease activity in RA.

Participation of substance P distribution in the cytokine production of rheumatoid synovium

Abstract Based on findings which suggested the involvement of the neuropeptide substance P in the pathogenesis of rheumatoid arthritis (RA), we investigated the mechanism of synovial pannus formation in RA, and examined the interaction between the cytokine production of synovial tissues and the concentration of substance P in the cartilage–pannus junction (CPJ). The CPJ and other peripheral synovial tissues were separately obtained from each part of the synovium from the knee joints of seven RA patients. The concentrations of substance P and the cytokines interleukin (IL)-1β and IL-6 in the CPJ and peripheral synovial tissues were determined by enzyme-linked immunosorbent assays. In addition, synovial cells were isolated from the CPJ and peripheral synovial tissues and treated with substance P or neurokinin-1 receptor antagonist to analyze the changes in cytokine production. The substance P levels were 211.2 and 50.5 pg/mg protein in the CPJ and the peripheral synovium, respectively. The IL-1β and IL-6 levels in the CPJ were 24.6 and 12.8 pg/mg protein, respectively. In the peripheral synovium, these levels were 4.3 and 2.5 pg/mg protein, respectively. In the CPJ, the IL-1β and IL-6 levels in tissue containing a high concentration of substance P (>200 pg/mg protein) were 39.4 and 21.6 pg/mg protein, respectively, and those in tissue containing a low concentration of substance P (≤200 pg/mg protein) were 11.6 and 5.1 pg/mg protein, respectively. Synovial cells from the CPJ produced higher levels of IL-1β and IL-6 than those from peripheral tissues. In addition, treatment of the cells with an NK-1 antagonist significantly reduced the production of these cytokines by the synovial cells. The theory that substance P plays a role in the pathogenesis of RA via the upregulation of cytokine production should be considered in further studies on the immunomodulatory properties of substance P in arthritis.

Substance P-induced priming effects on synovial cells

We detected and analyzed an intracellular mechanism of a substance P-induced priming effect on cytokine production using human synovial cells. The synovial tissues were isolated from the knee joints of osteoarthritis patients. After the administration of a low dose of substance P (1 nM) without significant effect alone, the synovial cells were stimulated with substance P (30 μM), phorbol 12-myristate 13-acetate (PMA) (100 nM), and calcium ionophore (A23187), (1 μM). The total interleukin (IL)-1β and IL-6 levels in the supernatant was measured by an enzymelinked immunosorbent assay (ELISA) kit, and the changes in the intracellular calcium concentration ([Ca2+]i) and protein kinase C (PKC) activation were measured by the fura 2-AM fluorescence method and a radioimmunoassay, respectively. The substance P-induced cytokine production was accompanied by an elevation of [Ca2+]i and PKC activation. The amounts of cytokines produced from the substance P (1 nM)-primed synovial cells stimulated with 30 μM substance P were approximately 4 times as much as that observed in non-primed cells. In addition, the priming treatment with 1 nM substance P enhanced not only the subsequent substance P-induced cytokine production, but also the PMA-induced response. However, substance P (1 nM) priming treatment did not affect the A23187-induced response. Furthermore, in substance P-primed cells, substance P (30 μM) induced a significant activation of PKC without changing the [Ca2+]i elevation response. These results suggest that the substance P-priming effect on synovial cells contributed to changes in intracellular mechanisms such as PKC activation.

Relationship Between HLA-DRB1-DQB1 Haplotypes and the Effect of Chicken Cartilage Soup Containing Type II Collagen on Rheumatoid Arthritis

The HLA-DQB1 and HLA-DRB1 genotypes were determined by a PCR-SSO technique in 65 Japanese outpatients with rheumatoid arthritis (RA). Group A consisted of patients having one or two HLA-DRB1*0405-HLA-DQB 1*0401 haplotypes. Group B consisted of patients without the haplotype. Eighteen and 14 patients were given chicken cartilage soup containing type II collagen (32mg/160ml) daily for over 3 weeks in group A and B, respectively (CII group). Nineteen and 14 patients were not treated with the soup in groups A and B, respectively (non-CII group).