Shunsuke Fukuyo1, Kunihiro Yamaoka1, Koshiro Sonomoto1, Koichi Oshita2, Yosuke Okada1, Kazuyoshi Saito1, Yasuhiro Yoshida1, Tamotsu Kanazawa1, Yasuhiro Minami1, Yoshiya Tanaka3. 1. First Department of Internal Medicine, University of Occupational and Environmental Health, Kitakyushu, Pharmacology Research Laboratories I, Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Department of Immunology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu and Department of Physiology and Cell Biology, Graduate School of Medicine, School of Medicine, Kobe University, Kobe, Japan. 2. First Department of Internal Medicine, University of Occupational and Environmental Health, Kitakyushu, Pharmacology Research Laboratories I, Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Department of Immunology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu and Department of Physiology and Cell Biology, Graduate School of Medicine, School of Medicine, Kobe University, Kobe, Japan.First Department of Internal Medicine, University of Occupational and Environmental Health, Kitakyushu, Pharmacology Research Laboratories I, Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Department of Immunology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu and Department of Physiology and Cell Biology, Graduate School of Medicine, School of Medicine, Kobe University, Kobe, Japan. 3. First Department of Internal Medicine, University of Occupational and Environmental Health, Kitakyushu, Pharmacology Research Laboratories I, Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Department of Immunology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu and Department of Physiology and Cell Biology, Graduate School of Medicine, School of Medicine, Kobe University, Kobe, Japan. tanaka@med.uoeh-u.ac.jp.
Abstract
OBJECTIVE: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). METHODS: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. RESULTS: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1β accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while β-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. CONCLUSION: IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3-dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.
OBJECTIVE: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). METHODS: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. RESULTS: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1β accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while β-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. CONCLUSION:IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3-dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.
Authors: Ivan Lau; Ajay Potluri; Cliff-Lawrence Ibeh; Robert S Redman; Edina Paal; Bidhan C Bandyopadhyay Journal: Arch Oral Biol Date: 2017-06-07 Impact factor: 2.633
Authors: Elena Zoico; Elena Darra; Vanni Rizzatti; Simona Budui; Guido Franceschetti; Gloria Mazzali; Andrea P Rossi; Francesco Fantin; Marta Menegazzi; Saverio Cinti; Mauro Zamboni Journal: Oncotarget Date: 2016-04-12