title: Potential of l-thyroxine to differentiate osteoblast-like cells via Angiopoietin1.
abstract: Angiogenesis is closely associated with osteoblast differentiation. Previously, we demonstrated that bone formation can be accelerated by treatment with COMP-Angiopoietin1, a known angiogenic factor. Angiopoietin1 (Ang1) is a specific growth factor that generates stable and mature vasculature through the Tie2 receptor. In this study, we aimed to identify a novel drug that can activate endogenous Ang1 expression as a pharmacological treatment for bone formation. Therefore, Ang1 expression was examined in U2OS osteoblast-like cells treated with 770 drugs from a library of Food and Drug Administration (FDA)-approved drugs by using ELISA for Ang1. l-thyroxine was selected as a novel drug candidate. l-Thyroxine is a synthetic form of the hormone thyroxine, which is used to treat patients with hypothyroidism. Enzyme-linked immunosorbent assays (ELISAs) were performed to test whether Ang1 is induced in a dose-dependent manner in human osteoblast-like cell lines, U2OS and MG63. The effects of l-thyroxine on osteoblast differentiation and mineralization were evaluated by alkaline phosphatase (ALP) activity and Alizarin red s staining. To determine the molecular mechanism, the expression of proteins related to bone formation and differentiation, such as type I collagen (COL1A1), osteocalcin (OC), bone sialoprotein (BSP), distal-less homeobox 5 (Dlx5), Runt-related transcription factor 2 (Runx2), osterix (OSX), and ALP, was tested by Western blotting analysis. Consequently, l-thyroxine induced Ang1 expression in a dose-dependent manner in both U2OS and M63 cells, which was confirmed by ELISA and Western blotting. Also, l-thyroxine activated ALP activity in U2OS and MG63 cells as well as ALP expression. Furthermore, l-thyroxine enhanced the expression of COL1A1, Runx2, OC, BSP, Dlx5, and OSX mRNA and proteins. Taken together, we demonstrated that l-thyroxine increased Ang1 expression and induces bone formation, differentiation, and mineralization in U2OS and MG63 cell lines, which suggests that l-thyroxine could be a potential bone production agent.
authors: Department of Bio and Chemical Engineering, Hongik University, 2639 Sejong-ro, Sejong 30016, Republic of Korea.
authors: See-Hyoung
authors: SH
authors: Park
authors: Department of Genetic Engineering, Sungkyunkwan University, Suwon, Republic of Korea.
authors: Jongsung
authors: J
authors: Lee
authors: Department of Orthopaedic Surgery, Chonbuk National University Medical School, Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital and Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea.
authors: Mi-Ae
authors: MA
authors: Kang
authors: Department of Biochemistry, Chonbuk National University Medical School, Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea.
authors: Young Jae
authors: YJ
authors: Moon
authors: Department of Orthopaedic Surgery, Chonbuk National University Medical School, Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital and Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea.
authors: Sung Il
authors: SI
authors: Wang
authors: Department of Pathology, Chonbuk National University Medical School, Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital and Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea.
authors: Kyoung Min
authors: KM
authors: Kim
authors: Department of Biochemistry, Chonbuk National University Medical School, Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea.
authors: Byung-Hyun
authors: BH
authors: Park
authors: Department of Pathology, Chonbuk National University Medical School, Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital and Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea. Electronic address: kyjang@chonbuk.ac.kr.
authors: Kyu Yun
authors: KY
authors: Jang
authors: Department of Orthopaedic Surgery, Chonbuk National University Medical School, Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital and Research Institute for Endocrine Sciences, 567 Baekje-daero, Deokjin-gu, Jeonju 54896, Republic of Korea. Electronic address: jrkeem@jbnu.ac.kr.
authors: Jung Ryul
authors: JR
authors: Kim
mesh_terms: D042683:Angiopoietin-1 / Q000378:metabolism*; D015415:Biomarkers / Q000378:metabolism; D002113:Calcification, Physiologic / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D013974:Thyroxine / Q000737:chemistry / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D042683:Angiopoietin-1; D015415:Biomarkers; D013974:Thyroxine
keywords: Angiopoietin1; Differentiation; Osteoblast; l-Thyroxine