C-terminal parathyroid hormone-related protein (PTHrP) (107-139) stimulates intracellular Ca2+ through a receptor different from the type 1 PTH/PTHrPreceptor in osteoblastic osteosarcoma UMR 106 cells

Studies were undertaken to determine whether PTH-related protein (PTHrP) (1 07-139) mobilizes [Ca2+], in osteoblastic osteosarcoma UMR 106 cells. PTHrP ( 107-139). in a manner similar to PTHrP (107-111). induced a rapid [Ca2+]( i) response in these cells that was dose dependent (EC50 of approximate to0 .1 pM) and more efficient than that of PTHrP (1-36) (EC50 of approximate to 1 nM). This effect of PTHrP (107-139) was abrogated by micromolar doses of verapamil or nifedipine. However, it was unaffected by 10 muM U73122 (a ph ospholipase C inhibitor), 100 mug/ml heparin (an inositol 1,4,5-trisphospha te receptor inhibitor), or 400 ng/ml pertussis toxin ia Gi inhibitor), whic h inhibited the [Ca2+](i) response to PTHrP (1-36), or by either 25 nM bisi ndolylmaleimide I (BIM). a protein kinase (PK)C inhibitor, or 1 muM phorbol -12-myri-state-13-acetate preincubation (22 h). PTHrP (107-139) and PTHrP ( 1-38), at 100 nM, desensitized the [Ca2+](i) response to a second challenge with the same peptide. but not with the other peptide in these cells. PTHr P (7-34), a type 1 PTH/PTHrP receptor (PTH1R) antagonist. decreased the eff ect of PTHrP (1-36) on [Ca2+](i),. In contrast, PTHrP (107-111), but neithe r PTHrP (109-138) nor PTHrP (7-34), abolished this effect of PTHrP (107-139 ). Both PTHrP (107-139) and PTHrP(1-36), added together at submaximal doses , induced a higher [Ca2+](i) response. Moreover. PTHrP (107-139) increased the efficacy of PTHrP (1-36) on [Ca2+](i), but decreased its induced increa se in PKA activity in these cells. Verapamil or nifedipine (at 50 muM) or 2 5 nM BIM, but not 25 muM adenosine 3 ' ,5 ' -cyclic monophosphorothioate, R p-isomer, a PKA inhibitor, abolished the PTHrP (107-139)-induced increase i n interleukin 6 messenger RNA (assessed by RT, followed by PCR) in UMR 106 cells. This peptide also increased c-fos messenger RNA in these cells; an e ffect inhibited by BIM, but unaffected by either verapamil or EGTA. These f indings support the existence of high-affinity receptors for PTHrP (107-139 ), associated with an induced Ca2+ influx, different from the PTH1R in UMR 106 cells. The present results suggest that PTHrP could affect bone turnove r by interacting with the PTH1R and other yet unknown receptors in bone cel ls through complex mechanisms.