Regulatory roles of zinc in matrix vesicle-mediated mineralization of growth plate cartilage.
Zinc (Zn2+) has long been known to play important roles in mineralization and ossification of skeletal tissues, but the mechanisms of Zn2+ action are not well understood. In this study we investigated the effects of Zn2+ on mineralization in a cell culture system in which terminal differentiation and mineralization of hypertrophic growth plate chondrocytes was induced by retinoic acid (RA) treatment. Addition of Zn2+ to RA-treated cultures decreased mineralization in a dose-dependent manner without affecting alkaline phosphatase (APase) activity. Characterization of matrix vesicles (MVs), particles that initiate the mineralization process, revealed that vesicles isolated from RA-treated and RA/Zn2+-treated cultures showed similar APase activity, but vesicles from RA/Zn2+-treated cultures contained significantly less Ca2+ and Pi. MVs isolated from RA-treated cultures were able to take up Ca2+ and mineralize in vitro, whereas vesicles isolated from RA/Zn2+-treated cultures were not able to do so. Detergent treatment, which ruptures the MV membrane and exposes preformed intravesicular Ca2+-Pi-phospholipid complexes, did not restore the Ca2+ uptake abilities of MVs isolated from RA/Zn2+-treated cultures, suggesting that vesicles from RA/Zn2+-treated cultures did not contain functional Ca2+-Pi-phospholipid complexes. Zn2+ treatment did not affect the content of annexins II, V, and VI in MVs or the Ca2+-dependent, EDTA-reversible binding of these molecules to the membrane surface. However, Zn2+ treatment did affect the EDTA-nonreversible binding of these molecules to the MV membrane, suggesting that Zn2+ interferes with the assembly of annexins in the MV membrane. In addition, Zn2+ inhibited annexin II-, V-, and VI-mediated Ca2+ influx into liposomes. In conclusion, Zn2+ inhibits the mineralizing competence of intravesicular Ca2+-Pi-phospholipid complexes and function of annexin channels, thereby controlling Ca2+ influx into MVs, the formation of the first crystal phase inside the vesicles and initiation of mineralization.
Kirsch
T
T
Department of Anatomy/Histology, School of Dental Medicine, University of Pennsylvania, Philadelphia 19104, USA.
Harrison
G
G
Worch
K P
KP
Golub
E E
EE
D000818:Animals; D017302:Annexins / Q000502:physiology; D002113:Calcification, Physiologic; D002214:Capsules; D002356:Cartilage / Q000166:cytology / Q000502:physiology*; D002454:Cell Differentiation / Q000502:physiology; D002642:Chick Embryo; D019902:Chondrocytes / Q000166:cytology / Q000502:physiology*; D005109:Extracellular Matrix / Q000502:physiology*; D015032:Zinc
D017302:Annexins; D002214:Capsules; D015032:Zinc