Receptors for the carboxyl-terminal region of pth(1-84) are highly expressed in osteocytic cells.
PTH is a potent systemic regulator of cellular differentiation and function in bone. It acts upon cells of the osteoblastic lineage via the G protein-coupled type-1 PTH/PTH-related peptide receptor (PTH1R). Carboxyl fragments of intact PTH(1-84) (C-PTH fragments) are cosecreted with it by the parathyroid glands in a calcium-dependent manner and also are generated via proteolysis of the hormone in peripheral tissues. Receptors that recognize C-PTH fragments (CPTHRs) have been described previously in osteoblastic and chondrocytic cells. To directly study CPTHRs in bone cells, we isolated clonal, conditionally transformed cell lines from fetal calvarial bone of mice that are homozygous for targeted ablation of the PTH1R gene and transgenically express a temperature-sensitive mutant SV40 T antigen. Cells with the highest specific binding of the CPTHR radioligand (125)I-[Tyr(34)]hPTH(19-84) exhibited a stellate, dendritic appearance suggestive of an osteocytic phenotype and expressed 6- to 10-fold more CPTHR sites/cell than did osteoblastic cells previously isolated from the same bones. In these osteocytic (OC) cells, expression of mRNAs for CD44, connexin 43, and osteocalcin was high, whereas that for alkaline phosphatase and cbfa-1/osf-2 was negligible. The CPTHR radioligand was displaced completely by hPTH(1-84), hPTH(19-84) and hPTH(24-84) (IC(50)s = 20-50 nM) and by hPTH(39-84) (IC(50) = 500 nM) but only minimally (24%) by 10,000 nM hPTH(1-34). CPTHR binding was down-regulated dose dependently by hPTH(1-84), an effect mimicked by ionomycin and active phorbol ester. Human PTH(1-84) and hPTH(39-84) altered connexin 43 expression and increased apoptosis in OC cells. Apoptosis induced by PTH(1-84) was blocked by the caspase inhibitor DEVD. We conclude that osteocytes, the most abundant cells in bone, may be principal target cells for unique actions of intact PTH(1-84) and circulating PTH C-fragments that are mediated by CPTHRs.
Divieti
P
P
Endocrine Unit, Massachusetts General Hospital, and Harvard Medical School, Boston Massachusetts 02114, USA. divieti@helix.mgh.harvard.edu
Inomata
N
N
Chapin
K
K
Singh
R
R
Jüppner
H
H
Bringhurst
F R
FR
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002478:Cells, Cultured; D006801:Humans; D008024:Ligands; D051379:Mice; D008822:Mice, Transgenic / Q000235:genetics; D010011:Osteocytes / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D010281:Parathyroid Hormone / Q000235:genetics* / Q000378:metabolism* / Q000494:pharmacology; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D018016:Receptors, Parathyroid Hormone / Q000378:metabolism*
D008024:Ligands; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D018016:Receptors, Parathyroid Hormone; C023443:carboxyl-terminal parathyroid hormone