MDPIActa Medica Okayama1422-00672352022Potential of a Novel Chemical Compound Targeting Matrix Metalloprotease-13 for Early Osteoarthritis: An In Vitro Study2681ENJunkoInagakiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAiriNakanoDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityOmer FarukHatipogluDepartment of Pharmacology, Faculty of Medicine, Kindai UniversityYukaOokaDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityYurinaTaniDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityAkaneMikiDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityKentaroIkemuraDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityGabrielOpokuDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityRyosukeAndoDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityShintaroKodamaDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityTakashiOhtsukiDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityHirosukeYamajiHeart Rhythm Center, Okayama Heart ClinicShuseiYamamotoDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityEriKatsuyamaDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityShogoWatanabeDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversitySatoshiHirohataDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityOsteoarthritis is a progressive disease characterized by cartilage destruction in the joints. Matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTSs) play key roles in osteoarthritis progression. In this study, we screened a chemical compound library to identify new drug candidates that target MMP and ADAMTS using a cytokine-stimulated OUMS-27 chondrosarcoma cells. By screening PCR-based mRNA expression, we selected 2-(8-methoxy-2-methyl-4-oxoquinolin-1(4H)-yl)-N-(3-methoxyphenyl) acetamide as a potential candidate. We found that 2-(8-methoxy-2-methyl-4-oxoquinolin-1(4H)-yl)-N-(3-methoxyphenyl) acetamide attenuated IL-1 beta-induced MMP13 mRNA expression in a dose-dependent manner, without causing serious cytotoxicity. Signaling pathway analysis revealed that 2-(8-methoxy-2-methyl-4-oxoquinolin-1(4H)-yl)-N-(3-methoxyphenyl) acetamide attenuated ERK- and p-38-phosphorylation as well as JNK phosphorylation. We then examined the additive effect of 2-(8-methoxy-2-methyl-4-oxoquinolin-1(4H)-yl)-N-(3-methoxyphenyl) acetamide in combination with low-dose betamethasone on IL-1 beta-stimulated cells. Combined treatment with 2-(8-methoxy-2-methyl-4-oxoquinolin-1(4H)-yl)-N-(3-methoxyphenyl) acetamide and betamethasone significantly attenuated MMP13 and ADAMTS9 mRNA expression. In conclusion, we identified a potential compound of interest that may help attenuate matrix-degrading enzymes in the early osteoarthritis-affected joints.No potential conflict of interest relevant to this article was reported.ElsevierActa Medica Okayama0022-22754392002ω-Carboxyl variants of 7-ketocholesteryl esters are ligands for β2-glycoprotein I and mediate antibody-dependent uptake of oxidized LDL by macrophages14861495ENQingpingLiuDepartment of Cell Chemistry, Okayama University Graduate School of Medicine and DentistryKazukoKobayashiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine and DentistryJun-ichiFurukawaDivision of Bioscience, Graduate School of Environment Earth Science, Hokkaido UniversityJunkoInagakiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine and DentistryNobuoSakairiDivision of Bioscience, Graduate School of Environment Earth Science, Hokkaido UniversityAkimasaIwadoGraduate School of Natural Science and Technology, Okayama UniversityTatsujiYasudaDepartment of Cell Chemistry, Okayama University Graduate School of Medicine and DentistryTakaoKoikeDepartment of Medicine II, Hokkaido University Graduate School of MedicineDennis R.VoelkerProgram in Cell Biology, Department of Medicine, National Jewish Medical and Research CenterEijiMatsuuraDepartment of Cell Chemistry, Okayama University Graduate School of Medicine and Dentistrybeta(2)-Glycoprotein I (beta(2)-GPI) is a major antigen for anticardiolipin antibodies (aCL, Abs) present in patients with antiphospholipid syndrome. We recently reported that beta(2)-GPI specifically binds to oxidized LDL (oxLDL) and that the beta(2)-GPI's major ligand, oxLig-1 is 7-ketocholesteryl-9-carboxynonanoate (Kobayashi, K, E. Matsuura, Q. P. Liu, J. Furukawa, K. Kaihara, J. Inagaki, T. Atsumi, N. Sakairi, T. Yasuda, D. R. Welker, and T. Koike. 2001. A specific ligand for beta(2)-glycoprotein I mediates autoantibody-dependent uptake of oxidized low density lipoprotein by macrophages. J Lipid Res. 42: 697-709). In the present study, we demonstrate that omega-carboxylated 7-ketocholesteryl esters are critical for beta(2)-GPI binding. A positive ion mass spectrum of a novel ligand, designated oxLig-2, showed fragmented ions at m/z 383 and 441 in the presence of acetone, which share features of oxLig-1 and 7-ketocholesterol. In the negative ion mode, ions at m/z 627, 625, and 243 were observed. oxLig-2 was most likely 7-ketocholesteryl-12-carboxy (keto) dodecanoate. These ligands were recognized by beta(2)-GPI. Liposome binding to macrophages was significantly increased depending on the ligand's concentration, in the presence of beta(2)-GPI and an anti-beta(2)-GPI Ab. Synthesized variant, 7-ketocholesteryl-13-carboxytxidecanoate (13-COOH-7KC), also showed a significant interaction with beta(2)-GPI and a similar binding profile with macrophages. Methylation of the carboxyl function diminished all of the specific ligand interactions with beta(2)-GPI. Thus, omega-carboxyl variants of 7-ketocholesteryl esters can mediate anti-beta(2)-GPI Ab-dependent uptake of oxLDL by macrophages, and autoimmune atherogenesis linked to beta(2)-GPI interaction with oxLDL.No potential conflict of interest relevant to this article was reported.ElsevierActa Medica Okayama2405-84406122020Antioxidative attributes of rice bran extracts in ameliorative effects of atherosclerosis-associated risk factorse05743ENWen TanXianDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesKazukoKobayashiCollaborative Research Center for OMIC, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesShenLianhuaDepartment of Pathophysiology, Zunyi Medical UniversityJunkoInagakiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesMasahiroIdeDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesSiaw SanHwangSchool of Chemical Engineering and Science, Faculty of Engineering, Computing and Science, Swinburne University of Technology Sarawak CampusEijiMatsuuraDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesOxidative stress, chronic inflammation, dyslipidemia, hyperglycemia, and shear stress (physical effect) are risk factors associated with the pathogenesis of atherosclerosis. Rice bran, a by-product of rice milling process, is known to house polyphenols and vitamins which exhibit potent antioxidant and anti-inflammatory properties. Through recent emerging knowledge of rice bran in health and wellness, the present study was aimed to assess the ameliorative effects of rice bran extracts (RBE) derived from Japanese colored rice varieties in modulating risk factors of atherosclerosis via in vitro and in vivo study models. Pre-treatment of lipopolysaccharide (LPS)-stimulated murine J774A.1 macrophage-like cells with RBE alleviated nitric oxide (NO) overproduction and downregulated gene expressions of pro-inflammatory modulators: tumor necrosis factor-α (TNF-α), interleukin (IL)-α (IL-1α), IL-1β, IL-6, and inducible nitric oxide synthase (iNOS). In addition, RBE also significantly attenuated LPS-stimulated protein expressions of iNOS, TNF-α, IL-1α, and IL-6 in J774A.1 macrophage-like cells as compared to non-treated LPS control group. In in vivo, 12 weeks of RBE dietary supplementations significantly reduced (p < 0.05) total cholesterol, triglycerides, and pro-atherogenic oxidized LDL/β2-glycoprotein I (oxLDL/β2GPI) complexes at plasma levels, in high fat diet (HFD) induced low density lipoprotein receptor knockout (Ldlr−/-) mice. En face pathological assessments of murine aortas also revealed significant reductions by 38% (p < 0.05) in plaque sizes of RBE-supplemented HFD mice groups as compared to non RBE-supplemented HFD control mice group. Moreover, gene expressions of aortic (iNOS, TNF-α, IL-1β) and hepatic (TNF-α, IL-1α, IL-1β) pro-inflammatory modulators were also downregulated in RBE-supplemented mice groups. Present study has revealed the potent health attributes and application of RBE as a dietary supplement to attenuate risks of inadvertent oxidative damage and chronic inflammation underlying the pathogenesis of atherosclerosis. Intrinsically, present preliminary findings may provide global health prospects for future dietary implementation of RBE in management of atherosclerosis.No potential conflict of interest relevant to this article was reported.MDPIActa Medica Okayama1422-00672192020Induction of CEMIP in Chondrocytes by Inflammatory Cytokines: Underlying Mechanisms and Potential Involvement in Osteoarthritis3140ENTakashiOhtsukiDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityOmer F.HatipogluDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityKeiichiAsanoDepartment of Molecular Biology and Biochemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesJunkoInagakiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKeiichiroNishidaDepartment of Orthopaediac Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSatoshiHirohataDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityIn patients with osteoarthritis (OA), there is a decrease in both the concentration and molecular size of hyaluronan (HA) in the synovial fluid and cartilage. Cell migration-inducing hyaluronidase 1 (CEMIP), also known as hyaluronan (HA)-binding protein involved in HA depolymerization (HYBID), was recently reported as an HA depolymerization-related molecule expressed in the cartilage of patients with OA. However, the underlying mechanism of CEMIP regulation is not well understood. We found that CEMIP expression was transiently increased by interleukine-1 beta (IL-1 beta) stimulation in chondrocytic cells. We also observed that ERK activation and NF-kappa B nuclear translocation were involved in the induction of CEMIP by IL-1 beta. In addition, both administration of HA and mechanical strain attenuated the CEMIP induction in IL-1 beta-stimulated chondrocytes. In conclusion, we clarified the regulatory mechanism of CEMIP in chondrocytes by inflammatory cytokines and suggested the potential involvement in osteoarthritis development.No potential conflict of interest relevant to this article was reported.ElsevierActa Medica Okayama0014482738322019Mechanical strain attenuates cytokine-induced ADAMTS9 expression via transient receptor potential vanilloid type 1111556ENTakashiOhtsukiDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityAkiraShinaokaDepartment of Human Morphology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKanaeKumagishi-ShinaokaDepartment of Human Morphology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKeiichiAsanoDepartment of Molecular Biology and Biochemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesOmer FarukHatipogluDepartment of Medical Technology, Graduate School of Health Sciences, Okayama UniversityJunkoInagakiDepartment of Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKenTakahashiDepartment of Cardiovascular Physiology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesToshitakaOohashiDepartment of Molecular Biology and Biochemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKeiichiroNishidaDepartment of Orthopaedic Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKeijiNaruseDepartment of Cardiovascular Physiology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSatoshiHirohataDepartment of Medical Technology, Graduate School of Health Sciences, Okayama University The synovial fluids of patients with osteoarthritis (OA) contain elevated levels of inflammatory cytokines, which induce the expression of a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) and of the matrix metalloproteinase (MMP) in chondrocytes. Mechanical strain has varying effects on organisms depending on the strength, cycle, and duration of the stressor; however, it is unclear under inflammatory stimulation how mechanical strain act on. Here, we show that mechanical strain attenuates inflammatory cytokine-induced expression of matrix-degrading enzymes. Cyclic tensile strain (CTS), as a mechanical stressor, attenuated interleukin (IL)-1β and tumor necrosis factor (TNF)-α-induced mRNA expression of ADAMTS4, ADAMTS9, and MMP-13 in normal chondrocytes (NHAC-kn) and in a chondrocytic cell line (OUMS-27). This effect was abolished by treating cells with mechano-gated channel inhibitors, such as gadolinium, transient receptor potential (TRP) family inhibitor, ruthenium red, and with pharmacological and small interfering RNA-mediated TRPV1 inhibition. Furthermore, nuclear factor κB (NF-κB) translocation from the cytoplasm to the nucleus resulting from cytokine stimulation was also abolished by CTS. These findings suggest that mechanosensors such as the TRPV protein are potential therapeutic targets in treating OA.No potential conflict of interest relevant to this article was reported.Wiley-BlackwellActa Medica Okayama1347-9032103102012Tumor growth inhibitory effect of ADAMTS1 is accompanied by the inhibition of tumor angiogenesis18891897ENMasanariObikaHirokoOgawaKatsuyukiTakahashiJiayiLiOmer FarukHatipogluMehmet ZeynelCilekToruMiyoshiJunkoInagakiTakashiOhtsukiShozoKusachiYoshifumiNinomiyaSatoshiHirohataAngiogenesis plays an important role in tumor progression. Several reports have demonstrated that a disintegrin and metalloproteinase with thrombospondin motifs1 (ADAMTS1) inhibited angiogenesis via multiple mechanisms. The aim of this study was to investigate the effect of ADAMTS1 on endothelial cells in vitro and on tumor growth with regard to angiogenesis in vivo. We examined the effects of the transfection of ADAMTS1 using two constructs, full-length ADAMTS1 (full ADAMTS1) and catalytic domain-deleted ADAMTS1 (delta ADAMTS1). Transfection of both the full ADAMTS1 and delta ADAMTS1 gene constructs demonstrated the secretion of tagged-ADAMTS1 protein into the conditioned medium, so we examined the effects of ADAMTS1-containing conditioned medium on endothelial cells. Both types of conditioned media inhibited endothelial tube formation, and this effect was completely abolished after immunoprecipitation of the secreted protein from the medium. Both types of conditioned media also inhibited endothelial cell migration and proliferation. We then examined the impact of ADAMTS1 on endothelial cell apoptosis. Both conditioned media increased the number of Annexin V-positive endothelial cells and caspase-3 activity and this effect was attenuated when z-vad was added. These results indicated that ADAMTS1 induced endothelial cell apoptosis. We next examined the effects of ADAMTS1 gene transfer into tumor-bearing mice. Both full ADAMTS1 and delta ADAMTS1 significantly inhibited the subcutaneous tumor growth. Collectively, our results demonstrated that ADAMTS1 gene transfer inhibited angiogenesis in vitro and in vivo, likely as a result of the induction of endothelial cell apoptosis by ADAMTS1 that occurs independent of the protease activity.No potential conflict of interest relevant to this article was reported.岡山大学農学部Acta Medica Okayama0474-0254 1002011放線菌Streptomyces sp.590由来l-メチオニン脱炭酸酵素の精製および性質検討37ENTomomiMaemuraKumikoUchitomiChikaKusakaJunkoInagakiTakashiTamuraKenjiSodaKenjiInagakiL-Methionine decarboxylase [EC 4.1.1.57] catalyzes the decarboxylation of L-methionine and is a pyridoxal 5’-phosohate(PLP)-dependent enzyme. L-Methionine decarboxylase has been purified 630-fold by DEAE-Toyopearl 650M, Phenyl-Toyopearl 650M and Sephacryl S-300 column chromatographies from Streptomyces sp.590. The enzyme has a dimeric structure with identical subunits of Mr 60,000. This enzyme shows optimum activity at pH7.0 and 45°C, and is stable between pH5.7 and pH9.0. L-Methionine decarboxylase has antitumor activity against RERF-LC-AI and HeLa cells. Ten N-terminal amino acid sequence of L-methionine decarboxylase was determined, and the sequence showed no homology with other reported proteins.No potential conflict of interest relevant to this article was reported.Acta Medica Okayama1994STUDIES ON TRANSPORT OF THE NUCLEUS-ENCODED 30 kDa PROTEIN OF PHOTOSYSTEM Ⅱ INTO EUGLENA CHLOROPLASTSENJunkoInagakiNo potential conflict of interest relevant to this article was reported.