Sunjoon Park

SAT0046 Regulation of Sirt1 Maybe A Pioneering Strategy in Treatment of Rheumatoid Arthritis

Background Monocyte maybe a dual role in rheumatoid arthritis (RA) development, because of differentiation to macrophages in joint and osteoclasts in bone so, blocking of monocyte differentiation maybe effective target in RA treatment. Objectives In this study, we interrupted monocyte differentiation to macrophages and osteoclasts via SIRT1 blocking and proposed pioneering treatment of RA. Methods In this study, monocytes from synovial fluid of RA patients (RAMCs), THP-1 monocytes, bone marrow–derived monocytes (BMDCs) from mice and RAW 264.7 cells were studied. PMA-induced expression of macrophages surface markers (CD11b, CD14 and CD36), proinflammatory cytokines (TNF-α, IL-1β and IL-6) secretion and differentiation of BMDCs from SIRT1 TG mice were measured after administration of resveratrol, which was reversed by sirtinol, an inhibitor of sirtuin proteins. In addition, RANKL [receptor activator of nuclear factor kappa B (RANK) ligand]-induced RANK expression in bone marrow–derived monocyte/macrophage precursors (BMMs) and RAW 264.7 cells were treated cilostazol and this was blocked by sirtinol (a SIRT1 inhibitor) in vitro and in a mouse model of collagen-induced arthritis (CIA). Results PMA-induced expression of macrophages surface markers (CD11b, CD14 and CD36) and proinflammatory cytokines (TNF-α, IL-1β and IL-6) secretion was inhibited by resveratrol. BMDCs from SIRT1 TG mice were inhibited differentiation by PMA than control BMDCs from C57BL/6 mice. These effects were associated with decrease in proinflammatory cytokines (TNF-α, IL-1β and IL-6) secreation by decreasing mRNA expression in BMDCs from SIRT1 TG mice. Further, resveratrol suppressed the PMA-induced PU.1 activation, which is critical transcription factor for macrophages differentiation. SIRT1 inhibits monocytes to macrophages differentiation through the down-regulates PU.1 activity and cilostazol elevated SIRT1 mRNA and protein levels in 12–24 h and increased SIRT1 activity, and these effects were also inhibited by sirtinol. Furthermore, the RANKL-induced nuclear expression of PU.1 was suppressed by cilostazol. In addition, marked RANKL-induced RANK immunofluorescence staining in Raw264.7 cells was strongly attenuated by cilostazol and by rSIRT1, and these attenuations were prevented by sirtinol. Extensive RANK staining of knee synovial tissues in a mouse model of collagen-induced arthritis (CIA) was also markedly reduced by cilostazol (30 mg/kg/day), and in BMMs both RANKL- and M-CSF-induced differentiation of BMMs to multinucleated TRAP+ giant cells and resorption pit formation were inhibited by cilostazol in association with a decrease in TRAP (a marker enzyme of osteoclasts) activity. Conclusions SIRT1 have dual a role in anti-osteoclast formation BMMs of RA bone and inhibiting differentiation monocyte to macrophage in RA synovium. so, SIRT1 maybe a perfect strategy for RA treatment. References Scott DL, Wolfe F, Huizinga TWJ (2010) Rheumatoid arthritis. Lancet 376(9746):1094–1108 Scott DL, Wolfe F, Huizinga TWJ (2010) Rheumatoid arthritis.Lancet 376(9746):1094–1108. Disclosure of Interest None declared

AB0055 Rho a protein and erk/jnk pathways regulate il-23 production by tlr2 in synovial macrophages from patients with rheumatoid arthritis

Background TLR2 expression upon application of TNF-a, IL-1b, and LPSin synovial fibroblasts from joints of RA patients (1, 2).IL-23 is involved in autoimmune diseases such as RA and psoriasis, in which the cellular mechanism of IL-23 is associated with self-reactive production of IL-17, IL-6, and TNF-a, thereby playing a critical role in development of autoimmune inflammation (3, 4). Objectives This study aimed to elucidate the signaling pathways of TLR2-mediated IL-23 production in synovial fluid macrophages from patients with rheumatoid arthritis (RA). Methods Expression of IL-23 was measured by ELISA and immunofluorescence in synovial fluid macrophages from RA patients. RhoA activity was assessed by pull-down assay. The role of MAP kinase was investigated using selective inhibitors and western blot. Results TLR2 ligand LTA-stimulated IL-23 production measured by ELISAwas elevated time- and concentration-dependently. TLR2 stimulation by LTA significantly increased not only GTP-bound RhoA activity but also phosphorylation of ERK and JNK in RA macrophages in association with increased nuclear translocation and DNA-binding activity of NF-kB. Inhibition of RhoA by Y27632, and of ERK and JNK phosphorylation by PD98059 and SP600125, resulted in suppression of LTA-induced NF-kB activation and IL-23 production. Image/graph Conclusions TLR2-mediated IL-23 production in synovial fluid macrophages was mediated through activation of RhoA GTPase and phosphorylation of ERK/JNK in association with NF-kB activation. References Brentano F, Kyburz D, Schorr O, Gay R, Gay S. The role of Toll-like receptor signalling in the pathogenesis of arthritis. Cell Immunol 2005, 233:90-96. Seibl R, Birchler T, Loeliger S, Hossle JP, Gay RE, Saurenmann T, Michel BA, Seger RA, Gay S, Lauener RP. Expression and regulation of Toll-like receptor 2 in rheumatoid arthritis synovium. Am J Pathol 2003, 162:1221-1227. Murphy CA, Langrish CL, Chen Y, Blumenschein W, McClanahan T, Kastelein RA, Sedgwick JD, Cua DJ. Divergent pro- and antiinflammatory roles for IL-23 and IL-12 in joint autoimmune inflammation. J Exp Med 2003, 198:1951-1957. Langrish CL, Chen Y, Blumenschein WM, Mattson J, Basham B, Sedgwick JD, McClanahan T, Kastelein RA, Cua DJ. IL-23 drives a pathogenic T cell population that induces autoimmune inflammation. J Exp Med 2005, 201: 233-240. Disclosure of Interest None Declared

AB0056 Hmgb1 regulates hypoxia-inducible factor 1alpha expression and function in synovial fibroblasts from patients with rheumatoid arthritis

Background High mobility group box chromosomal protein 1 (HMGB1) is a nuclear DNA binding protein that is secreted into extracellular medium by cellular activation and proinflammatory response, and promotes inflammation. HMGB1 is known as ligand for TLR4 that produces TNF-a and IL-8 (1, 2). Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor which is composed of an alpha and beta subunit. HIF-1 has been reported as regulator of angiogenesis in rheumatoid arthritis (RA) (3, 4). Objectives We investigated the role and molecular mechanism of HMGB1 in the expression and function of HIF-1α in synovial fibroblasts from patients with RA. Methods HIF-1 activity and VEGF expression were measured by ELISA. Signaling and molecular events were analyzed by immunoblotting. Results The expression of extracellular HMGB1 was increased in synovium of RA patients than OA patients. Treatment of RA synovial fibroblasts with HMGB1 increased HIF-1αexpression and HIF-1 activity. HMGB1 mediated production of VEGF was dependent upon HIF-1. When TLR4 was blocked by TLR4 siRNA or anti-TLR4 antibody, HIF-1αdidn’t be increased, leading to abolish the increase in VEGF production. Image/graph Conclusions Our results suggest that HMGB1stimulation of HIF-1α expression and activity results in VEGF expression that is dependent upon TLR4. References Bustin M, Reeves R. High-mobility-group chromosomal proteins: architectural components that facilitate chromatin function. Prog Nucleic Acid Res Mol Biol 1996, 54:35-100. Hamada T, Torikai M, Kuwazuru A, et al. Extracellular high mobility group box chromosomal protein 1 is a coupling factor for hypoxia and inflammation in arthritis. Arthritis Rheum 2008, 58:2675-2685, Imtiyaz HZ, Simon MC. Hypoxia-inducible factors as essential regulators of inflammation. Curr Top Microbiol Immunol 2010, 345:105-120. Giatromanolaki A, Sivridis E, Maltezos E, et al. Upregulated hypoxia inducible factor-1alpha and -2alpha pathway in rheumatoid arthritis and osteoarthritis. Arthritis Res Ther 2003, 5:R193–201. Disclosure of Interest None Declared

Particle Removal on Buffing Process After Copper CMP

Copper (Cu) had been attractive material due to its superior properties comparing to other metals such as aluminum or tungsten and considered as the best metal which can replace them as an interconnect metal in integrated circuits. CMP (Chemical Mechanical Polishing) technology enabled the production of excellent local and global planarization of microelectronic materials, which allow high resolution of photolithography process. Cu CMP is a complex removal process performed by chemical reaction and mechanical abrasion, which can make defects of its own such as a scratch, particle and dishing. The abrasive particles remain on the Cu surface, and become contaminations to make device yield and performance deteriorate. To remove the particle, buffing cleaning method used in post-CMP cleaning and buffing is the one of the most effective physical cleaning process. AE(Acoustic Emission) sensor was used to detect dynamic friction during the buffing process. When polishing is started, the sensor starts to be loaded and produces an electrical charge that is directly proportional to the applied force. Cleaning efficiency of Cu surface were measured by FE-SEM and AFM during the buffing process. The experimental result showed that particles removed with buffing process, it is possible to detect the particle removal efficiency through obtained signal by the AE sensor.

Development of Multiple CMP Monitoring System for Consumable Designs

Consumables used in Chemical Mechanical Polishing (CMP) have been played important role to improve quality and productivity. Since the properties of consumables constantly change with various reasons, such as shelf time, manufactured time, lot to lot variation from supplier and so on, CMP results are not constant during the process. Also, CMP process results are affected by multiple sources from wafer, conditioner, pad and slurry. Therefore, multiple sensing systems are required to monitor CMP process variation. In this paper, the authors focus on development of monitoring system for CMP process which consist of force, temperature and displacement sensor to measure the signal from CMP process. With monitoring systems mentioned above, complex CMP phenomena can be investigated more clearly.

Fabrication of Real 3D Shape Components Using Bi-Sn Alloys

In this paper, new replication techniques fur a metal microcomponent having a real 3D shape were introduced. Helical gear was selected as one of a real 3D microcomponents for this study. The helical gear, which was made of photo-curable resin, was fabricated as a master pattern by microstereolithography technology. Then, a silicone rubber mold was fabricated from the master pattern. Lastly, a final bismuth alloy pattern was transferred from the silicone rubber mold by the microcasting process. In this paper, the replication technique is described in detail from the master pattern to the final pattern with some investigation on factors related to the technique.