Stimulation by parathyroid hormone of 45Ca2+ uptake in osteoblast-like cells: possible involvement of alkaline phosphatase.
We have investigated the actions of human PTH [hPTH-(1-34)] on the association of 45Ca2+ with two human (SaOS-2 and MG-63) and two rat (ROS 17/2.8 and UMR-106) osteoblast-like cell types. In SaOS-2 cells, hPTH-(1-34) binds to specific membrane receptors to activate adenylate cyclase. Treatment of SaOS-2 cells with hPTH-(1-34) resulted in an increase in 45Ca2+ uptake, in a dose-dependent fashion, up to 2- to 4-fold above control values. The increase was first evident at 10 min and persisted for at least 30 min. Treatment with nimodipine, a calcium channel antagonist, was without effect on the stimulatory action of PTH. A similar enhancement of cell-associated 45Ca2+ was observed when the cells were incubated with vasoactive intestinal peptide, which acts via different receptors to activate adenylate cyclase in SaOS-2 cells. Treatment with (Bu)2cAMP also induced an increase in cell-associated 45Ca2+. Pretreatment of SaOS-2 cells with hPTH-(1-34) for 4 h, which induced homologous desensitization to a second challenge with the same peptide for stimulation of cAMP production, did not attenuate the further enhancement of cell-associated 45Ca2+ by a second treatment with hPTH-(1-34). We then examined a possible relationship between alkaline phosphatase (ALPase) and 45Ca2+ uptake. SaOS-2 cells contained high levels of alkaline phosphatase activity and continuously released the enzyme into the medium. Release was enhanced by treatment with hPTH-(1-34) for 10 min. Incubation of cells with levamisole (an inhibitor of the liver/bone/kidney type of ALPase) resulted in a rapid decrease in basal and PTH-stimulated 45Ca2+ uptake, while treatment with L-Phe-Gly-Gly (an inhibitor of human placental ALPase) was without effect. Treatment of the cells with ALPase (bovine kidney) enhanced 45Ca2+ uptake. In MG-63 cells, a stimulatory effect of hPTH-(1-34) on cell-associated 45Ca2+ was also observed; however, hPTH-(1-34) did not stimulate cAMP production in MG-63 cells. In ROS 17/2.8 cells, neither hPTH-(1-34) nor rat PTH-(1-34) stimulated an increase in cell-associated 45Ca2+, while in UMR-106 cells, rat PTH-(1-34) and (Bu)2cAMP did enhance 45Ca2+ uptake, although hPTH-(1-34) was without effect. We conclude that PTH can stimulate an increase in cell-associated 45Ca2+ in several osteoblast-like cell lines, possibly by modulating local ALPase activity; however, this action of PTH does not appear to be obligatorily dependent on the adenylate cyclase-stimulating action of PTH.
Fukayama
S
S
Laboratory of Toxicology, Harvard School of Public Health, Boston, Massachusetts 02115.
Tashjian
A H
AH
D000469:Alkaline Phosphatase / Q000037:antagonists & inhibitors / Q000502:physiology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002132:Calcium Radioisotopes; D006801:Humans; D007978:Levamisole / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D010006:Osteoblasts / Q000378:metabolism* / Q000473:pathology; D010281:Parathyroid Hormone / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
D002132:Calcium Radioisotopes; D009842:Oligopeptides; D010281:Parathyroid Hormone; C043963:phenylalanyl-glycyl-glycine; D007978:Levamisole; D014660:Vasoactive Intestinal Peptide; D000469:Alkaline Phosphatase; D002118:Calcium