Synovial cell proliferation and activation is one of the major feature of rheumatoid arthritis, and articular tissue destruction is mediated by a granulation tissue (pannus) composed of macrophage like type A synoviocytes and fibroblast-like type B synoviocytes. Monokines such as IL 1, TNFα, and PDGF produced by type A synoviocytes stimulate the proliferation and activation of type B synoviocytes. We have employed antisense oligonucleotides to inhibit the production of cytokines including IL-1 and TNFα. And the inhibition of synoviocytes growth with vitamine D3 and a site-specific cAMP analogue was studied. Furthermore the regulation of transcription factor including AP-1, CREB and NF-κB in synoviocytes was discussed in this review.

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When human umbilical endothelial cells (HUVEC) were incubated for 8h in the presence of TNFa (0.1 ng/ml), they produced cellassociated IL-1α. However, in the conditioned medium, no detectable amount of IL-lα was produced by the TNFa treatment. IL 1β also was not detected in the cells or in the conditioned medium. Combined treatment with TNFα (0.1ng/ml) and the protein kiase C (PKC) activator TPA (1 ng/ml) suppressed the TNFα-induced IL-1α production. A significant inhibition of the TNFα-induced IL-1α production was observed when HUVEC were pretreated with TPA (1 ng/ml) for 1 to 15 min. Other PKC activators such as aplysiatoxin (AT) or teleocidin (TC) at a concentration of 1 ng/ml also inhibited the TNFα-induced IL-1α production when HUVEC were pretreated for 15 min prior to the stimulation with TNFα. IL-1β (0.1 ng/ml) or lipopolysaccharide (LPS) (5μg/ml) also stimulated the cell-associated IL-1α production. Pretreatment with TPA, AT, or TC (1ng/ml) for 15 min suppressed the IL-1β-, or the LPS-induced IL-1α production. However, the inhibition of the TNFα-induced IL-1α production by TPA was not counteracted by pretreatment with the PKC inhibitor H-7 or staurosporine (SS) . TNFα stimulated PGI₂ production, and pretreatment with TPA further increased PGI₂ production. The TPA-induced PGI₂ production was significantly suppressed by the pretreatment with H-7.
In conclusion, IL-1α production stimulated by TNFα, IL-1β, or LPS is inhibited by the PKC activators TPA, AT, or TC, but the inhibition of the IL-1α production might not be due to the activation of PKC, but by PKC-independent mechanisms.

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Using type IV collagen-Sepharose affinity chromatography, the binding proteins were isolated from ¹²⁵I surface-labeled human neutrophils. SDS-polyacrylamide gel electrophoresis analysis showed that the binding molecules were composed of the 28-, 49-, 67-, and 95-kDa proteins. Western blot analysis revealed that the 95-kDa proteins contained both L-selectin and nonspecific cross-reacting antigen 90 (NCA90), and that the 67-kDa protein was corresponded to 67-kDa elastin/laminin receptor (67BP) . When f-Met-Leu-Phe (fMLP) -stimulated neutrophils were analyzed using the affinity chromatography, the amounts of the binding proteins were increased about three-fold. However, western blot analysis demonstrated that L-selectin was significantly decreased by the fMLP stimulation. On the other hand, NCA90 and 67BP were increased by the stimulation. Together these observations indicate that neutrophils have several kinds of the adhesion molecules to type IV collagen, and that L-selectin is likely important for the binding of resting neutrophils to type IV collagen, whereas 67BP and NCA90 are likely important for the binding of stimulated neutrophils to type IV collagen.

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Neutrophils contain the antimicrobial cationic peptides, defensins in the granules. To understand the mechanism of gene expression, the cDNAs for the peptides were analyzed. From a guinea-pig bone marrow cell cDNA library, three kinds of cDNA clones encoding neutrophil cationic peptides, GNCP-1 and -2 were isolated. Two cDNAs coded for GNCP-1 (GNCP-1A and -1B), and one cDNA coded for GNCP-2. The nucleotide sequences of these cDNAs were highly homologous (>99%), and the deduced amino acid sequences showed that GNCP-1A and GNCP-1B differed by only one amino acid substitution in the propeptide region, and GNCP-1B and GNCP-2 differed by only one amino acid substitution in the mature peptide region.
Next, a genomic library was screened, and the GNCP gene clones were analyzed. Among four different GNCP gene clones isolated, two clones encoded GNCP-1A and-1B, and other two clones encoded GNCP-2. The sequence analyses revealed that the nucleotide sequences of GNCP-1 and -2 genes were highly homologous (>96%) in the region sequenced, and that the GNCP-1 and -2 genes spanned 3kb, and consisted of three exons and two introns. Exon 1 encoded the 5' untranslated region, exon 2 encoded the prepro-peptide region, and exon 3 encoded the mature peptide and 3' untranslated regions.
Finally, to evaluate the promoter activity, the 5' flanking regions (0.4 kb) of the GNCP-1 and -2 genes were subcloned into the promoterless PGV-B luciferase vector, and the plasmid constructs were transfected into guinea-pig 104C1 cells and GPC-16 cells. The GNCP-PGV-B constructs expressed> 10-fold more luciferase activity than the promoterless PGV-B vector in both cells. Interestingly, the promoter activity of GNCP-1 gene was 2-fold more than that of GNCP-2 gene. Together these observations indicate that GNCP-1 and -2 are encoded by the homologous but different genes, and the expression of GNCP -1 and -2 genes are likely different at the level of transcription.

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We used flow cytometry to investigate the function and the surface membrane protein expression by platelet derived microparticles from normal individuals. Microparticles were detected by both forward scatter and side scatter using flow cytometry. The binding of coagulation factor on microparticles was investigated by using monoclonal anti Factor X (Xa) antibody. Microparticles released from platelets after activation with the calcium ionophore A23187 and the binding of Factor Xa to microparticles increased markedly. These microparticles contained not only glycoprotein IIb/IIIa or I b but also P-selectin. On the other hand, microparticles were also released by the activation of complement system. Furthermore, these releases increased by the incubation with normal leukocytes.
Our findings suggest that platelet-derived microparticles participate the inflammatory thrombosis and hemostasis.

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In vitro proliferative responses of T lymphocytes to bacterial lipopolysaccharides (LPS) were examined by determining the uptake of tritiated thymidine (³H-TdR) into cells. T lymphocyte populations were taken and purified from proteose peptoneinduced peritoneal exudate cell (PEG), spleen and thymus of C3H/HeN or C3H/HeJ mice. LPS were prepared from Salmonella typhimurium, Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum by the phenolwater proceduce.
The T lymphocytes taken from PEG (PEG-T) of C3H/HeN mice showed an increased ³H-TdR uptaken in response to LPS from S. typhimurium (S. t-LPS), while PEG-T of G3H/HeJ, splenic T and thymic T lymphocyte of C3H/HeN mice did not respond to S. t-LPS. Other LPS also have a mitogenic ability to PEG -T lymphocytes too, while those of S. t-LPS and P. i-LPS were higher than that of F. n-LPS or P. g-LPS. The response of PEG from C3H/HeN mice to S. t-LPS was abolished, when the PEG-T were pretreated with anti-Thy-1 orγδTCR antibody plus complement (c), but not anti-αβTCR antibody plus C. The thymic T lymphocytes did not show any increase of ³H-TdR uptake in response to LPS or anti-γδ TCR antibody. However, obvious uptake did occur when the cells were stimulated with LPS and anti-γδ TCR antibody together.
The results suggest that LPS has a mitogenic ability to a T lymphocytes population bearing γδ TCR in PEG that had been stimulated previously by proteose peptone.

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T-614 is a chemical compound which has been shown to have clinical efficacy as an antirheumatic drug in patients with rheumatoid arthritis (RA) . In the present study, we investigated the effects of T-614 and its derivatives M1 and M2 on mononuclear cell (MNC) function.
MNC obtained from the peripheral blood of RA patients were cultured with varying concentrations of T-614 for 24 h or 7 days. and the supernatants were measured for substances produced from monocytes and T and B lymphocytes. T-614 inhibited IL 1β, IL -6 and TNF-α production in MNC.
These results suggest that T-614 exerts immunosuppressive effects on monocytes and these effects may be one of the mechanisms by which T-614 shows therapeutic effects in patients with RA.

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In the current treatment of rheumatoid arthritis (RA), to achieve the high quality of life (QOL) of patients became important. Therefore, we assessed QOL as well a the efficacy and safety in the combined therapy of diclofenac sodium slow release capsule (Voltaren SR capsule) and suppository (Voltaren SUPPO) .
To 70 RA patients, slaw release capsule 37.5 mg was given orally twice daily I capsule each after breakfast and dinner, and 25 mg p.r. of suppository before sleep for 8 weeks, respectively.
Among the inflammatory findings, significant improvement was seen in the number of painful joints and painful/swollen joints after week 4 and 8 of the treatment.
There was no change in QOL evaluated using modified health assessment questionnaire (HAQ), quality of well-being score face scale and analogue scale evaluation method developed by us The results of HAQ stratified by the various factors showed improvement in the groups of“No treatment history with NSAIDs”in week 4 and more serious cases of“Stage”and“Pre-administration MHAQ value”.
The final general improvement rate was 20.8% (10/48) for“moderate improvement”or better, and 60.4% (29/48) for“slight improvement”or better.
Adverse effects were seen in 7 patients among 61 (11, 5%) . Symptoms were mild except for moderate epigastric pain.
From the above, the usefulness of the present therapy was confirmed as high as that of the double blind test of slow release capsule. The short-term efficacy of the present therapy was also suggested by some QOL measures, especially in serious RA patients. However, we think that further studies using controlled test with sufficient number of natients are necessary to confirm these findings.

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