Cytosolic free Ca2+ concentration exhibits a characteristic temporal pattern during in vitro cartilage differentiation: a possible regulatory role of calcineurin in Ca-signalling of chondrogenic cells.
We measured changes of cytosolic Ca2+ concentration during chondrogenesis, which occurs in high-density cultures (HDC) of chondrifying chicken mesenchymal cells. A significant, transient elevation was detected in Fura-2-loaded cells on day 3 of culturing, when majority of chondrogenic cells of HDC become differentiated. This 140 nM peak of cytosolic Ca2+ concentration is a result of increased Ca-influx and is indispensable to proper chondrogenesis, because addition of 0.8mM EGTA to culture medium on day 2 or 3 significantly decreased the intracellular Ca2+ concentration abolishing the Ca2+-peak of day 3 and inhibited cartilage formation. Uncontrolled Ca2+ influx evoked by a Ca2+ ionophore exerted dual effects on chondrogenesis in a concentration-dependent manner; 0.1mg/L A23187 increased, whereas 5 mg/L A23187 almost totally blocked cartilage formation. Intracellular Ca-stores seemed not to have any significant participation in the regulation of changes of cytosolic Ca2+ concentration of chondrifying cells. Activity of Ca-calmodulin-dependent protein phosphatase, calcineurin responded to changes of intracellular Ca2+ concentration induced by EGTA or A23187 in a differentiation stage-dependent manner. Since inhibition of calcineurin with cyclosporine A eliminated the peak in the cytosolic Ca2+ concentration, an active regulatory role of calcineurin on Ca2+ influx of chondrifying cells can be supposed.
Matta
Csaba
C
Department of Anatomy, Histology and Embryology, Medical and Health Science Centre, University of Debrecen, Nagyerdei krt. 98, H-4032 Debrecen, Hungary.
Fodor
János
J
Szíjgyártó
Zsolt
Z
Juhász
Tamás
T
Gergely
Pál
P
Csernoch
László
L
Zákány
Róza
R
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D019703:Calcineurin / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D002356:Cartilage / Q000166:cytology* / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D002642:Chick Embryo; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D055758:SOX9 Transcription Factor / Q000378:metabolism
D055758:SOX9 Transcription Factor; D000001:Calcimycin; D004533:Egtazic Acid; D019703:Calcineurin; D002118:Calcium