title: Mouse limb skeletal growth and synovial joint development are coordinately enhanced by Kartogenin.
abstract: Limb development requires the coordinated growth of several tissues and structures including long bones, joints and tendons, but the underlying mechanisms are not wholly clear. Recently, we identified a small drug-like molecule - we named Kartogenin (KGN) - that greatly stimulates chondrogenesis in marrow-derived mesenchymal stem cells (MSCs) and enhances cartilage repair in mouse osteoarthritis (OA) models. To determine whether limb developmental processes are regulated by KGN, we tested its activity on committed preskeletal mesenchymal cells from mouse embryo limb buds and whole limb explants. KGN did stimulate cartilage nodule formation and more strikingly, boosted digit cartilaginous anlaga elongation, synovial joint formation and interzone compaction, tendon maturation as monitored by ScxGFP, and interdigit invagination. To identify mechanisms, we carried out gene expression analyses and found that several genes, including those encoding key signaling proteins, were up-regulated by KGN. Amongst highly up-regulated genes were those encoding hedgehog and TGFβ superfamily members, particularly TFGβ1. The former response was verified by increases in Gli1-LacZ activity and Gli1 mRNA expression. Exogenous TGFβ1 stimulated cartilage nodule formation to levels similar to KGN, and KGN and TGFβ1 both greatly enhanced expression of lubricin/Prg4 in articular superficial zone cells. KGN also strongly increased the cellular levels of phospho-Smads that mediate canonical TGFβ and BMP signaling. Thus, limb development is potently and harmoniously stimulated by KGN. The growth effects of KGN appear to result from its ability to boost several key signaling pathways and in particular TGFβ signaling, working in addition to and/or in concert with the filamin A/CBFβ/RUNX1 pathway we identified previously to orchestrate overall limb development. KGN may thus represent a very powerful tool not only for OA therapy, but also limb regeneration and tissue repair strategies.
authors.affiliation: Translational Research Program in Pediatric Orthopaedics, Division of Orthopaedic Surgery, The Children׳s Hospital of Philadelphia, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA. Electronic address: DeckerR@email.chop.edu.
authors.forename: Rebekah S
authors.initials: RS
authors.lastname: Decker
authors.affiliation: Translational Research Program in Pediatric Orthopaedics, Division of Orthopaedic Surgery, The Children׳s Hospital of Philadelphia, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA.
authors.forename: Eiki
authors.initials: E
authors.lastname: Koyama
authors.affiliation: Translational Research Program in Pediatric Orthopaedics, Division of Orthopaedic Surgery, The Children׳s Hospital of Philadelphia, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA.
authors.forename: Motomi
authors.initials: M
authors.lastname: Enomoto-Iwamoto
authors.affiliation: Department of Reconstructive Sciences, University of Connecticut Health Center School of, Dental Medicine, 263 Farmington Ave, Farmington, CT 06030, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Maye
authors.affiliation: Department of Reconstructive Sciences, University of Connecticut Health Center School of, Dental Medicine, 263 Farmington Ave, Farmington, CT 06030, USA.
authors.forename: David
authors.initials: D
authors.lastname: Rowe
authors.affiliation: California Institute for Biomedical Research, 11119 North Torrey Pines Road, Suite 100, La Jolla, CA 92037, USA.
authors.forename: Shoutian
authors.initials: S
authors.lastname: Zhu
authors.affiliation: The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
authors.forename: Peter G
authors.initials: PG
authors.lastname: Schultz
authors.affiliation: Translational Research Program in Pediatric Orthopaedics, Division of Orthopaedic Surgery, The Children׳s Hospital of Philadelphia, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA.
authors.forename: Maurizio
authors.initials: M
authors.lastname: Pacifici
mesh_terms: D000813:Anilides / Q000494:pharmacology*; D000818:Animals; D020219:Chondrogenesis / Q000187:drug effects*; D017931:DNA Primers / Q000235:genetics; D005121:Extremities / Q000196:embryology*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D007091:Image Processing, Computer-Assisted; D015151:Immunoblotting; D017403:In Situ Hybridization; D017746:Joint Capsule / Q000187:drug effects / Q000196:embryology*; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D008648:Mesoderm / Q000187:drug effects*; D051379:Mice; D018613:Microscopy, Confocal; D010795:Phthalic Acids / Q000494:pharmacology*; D011509:Proteoglycans / Q000378:metabolism; D012038:Regeneration / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D000071676:Zinc Finger Protein GLI1; D000097573:Red Fluorescent Protein; D000099158:Proteoglycan 4
chemical_list: D000813:Anilides; D017931:DNA Primers; D051741:Kruppel-Like Transcription Factors; D008164:Luminescent Proteins; D010795:Phthalic Acids; D011509:Proteoglycans; D053773:Transforming Growth Factor beta1; D000071676:Zinc Finger Protein GLI1; D000097573:Red Fluorescent Protein; C498520:Gli1 protein, mouse; C498837:Prg4 protein, mouse; D000099158:Proteoglycan 4; C572342:kartogenin
keywords: BMP and hedgehog signaling; Joint interzone; Kartogenin; Limb development; Lubricin; Skeletogenesis; Synovial joint formation; TGFB