PHYSIOLOGICAL EFFECTS OF Na+-Ca2+ EXCHANGE INHIBITION BY ANTISENSE OLIGODEOXYNUCLEOTIDES IN OSTEOBLASTS

초록

Physiological activity of osteoblast including osteogenesis is known to be related to the increase in intracellular Ca2+ activity ([Ca2+]i) of osteoblast. The cellular regulation of [Ca2+]i in osteoblasts is mediated by the transmembrane Ca2+ movement via Na+-Ca2+ exchange and store-operated Ca2+ influx, and intracellular Ca2+ movement through intracellular Ca2+ stores. In this study, we investigated the role of Na+-Ca2+ exchange transport in regulation of intracellular Ca2+ activity in osteoblasts, imploying antisense oligodeoxynucleotide (AS-oligo) technique. Osteoblast-like cells (OLC) were isolated from femur and tibia of neonatal rats, and cultured for 8 days. On the 7th day, antisense oligodeoxynucleotides (AS-oligo) 5'AAAAATGGTGAAGAGAGTG3' and 5'GTGAACATGACAAATGTCA3' were added into the culture media. Cultured OLCs were loaded with Fura-2, and fluorescence images were monitored with a cooled CCD camera. [Ca2+]i decreased as Ca2+ concentrations in the superfusing solutions were lowered, and this Ca2+-dependence was augmented in AS-oligo treated OLCs. The removal of Na+ and Ca2+ from the superfusing solutions elicited spike-like transient [Ca2+]i increase, which was inhibited in the AS-oligo treated cells. The Ca2+ influx following the emptying of internal Ca2+ stores was enhanced in the AS-oligo treated cells. The results from this study suggest that Na+-Ca2+ exchange plays a role in regulating the intracellular Ca2+ activity in osteoblasts.