RRC ID 37492
Author Masuda E, Shirai K, Maekubo K, Hirai Y.
Title A newly established culture method highlights regulatory roles of retinoic acid on morphogenesis and calcification of mammalian limb cartilage.
Journal BioTechniques
Abstract During mammalian embryogenesis, sclerotome-derived chondrocytes in the limb bud are arranged into a complicated bone shape with specific areas undergoing hypertrophy and calcification, creating a region-specific mineralized pattern in the cartilage. To follow chondrogenesis progression in vitro, we isolated limb cartilage from mice on embryonic day 13 (E13) and cultured it at the air-liquid interface after microsurgical removal of the ectoderm/epidermis. Explants underwent proper morphogenesis, giving rise to complete templates for limb bones in vitro. We found that region-specific calcification patterns resembling limbs of prepartum mature embryos could be induced in explants using culture medium containing high concentrations of CaCl2 (Ca), ascorbic acid (AA), and β-glycerophosphoric acid (BGP). In this culture system, excess amounts of all-trans retinoic acid (RA) severely disrupted morphogenesis and calcification patterns in limb cartilage. These effects were more pronounced in forearms than in phalanges. Although dissociated, the nascent chondrocytes in culture did not give rise to cartilage units even though augmented calcification was induced in these cell aggregates in the presence of RA. Taken together, our newly established culture system revealed that RA independently regulates three-dimensional morphogenesis and calcification.
Volume 58(6)
Pages 318-24
Published 2015-6
DOI 10.2144/000114300
PII 000114300
PMID 26054768
MeSH Animals Ascorbic Acid / metabolism Calcification, Physiologic* Cartilage / embryology* Cartilage / physiology Cell Differentiation Cell Line Extremities / embryology* Extremities / physiology Glycerophosphates / metabolism Mice Morphogenesis Tissue Culture Techniques / methods* Tretinoin / metabolism*
IF 1.659
Times Cited 1
WOS Category BIOCHEMICAL RESEARCH METHODS BIOCHEMISTRY & MOLECULAR BIOLOGY
Resource
Human and Animal Cells MC3T3-E1(RCB1126)