Insulin-Like Growth Factor I Suppresses Parathyroid Hormone (PTH)/PTH-Related Protein Receptor Expression via a Mitogen-Activated Protein Kinase Pathway in UMR-106 Osteoblast-Like Cells

Abstract

Insulin-like growth factor I (IGF-I) is important in skeletal growth and has been implicated in the maintenance of bone integrity. PTH stimulates bone resorption through the G protein-linked PTH/PTH-related protein (PTHrP) receptor in osteoblasts. Using a heterogeneous nuclear RNA assay and Northern blot analysis, we showed that IGF-I inhibited expression of the gene for PTH/PTHrP receptor in a dose- and time-dependent fashion, but did not alter the stability of the receptor messenger RNA (mRNA) in UMR-106 osteoblast-like cells. IGF-I treatment for 48 h also caused a decrease in the receptor number to 45% of that in controls without affecting receptor affinity and in functional receptor expression to 50-60% of that in controls as measured by PTH-stimulated cAMP production. In MC3T3-E1 murine nontransformed osteoblasts, IGF suppressed receptor mRNA expression dose dependently. In UMR-106 cells, IGF-I induced the mitogen-activated protein (MAP) kinase pathway. The effect of IGF-I was blocked by PD98059, a specific inhibitor of the MAP kinase-activating kinase, but not by wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase. IGF-I inhibition of PTH/PTHrP receptor mRNA expression in UMR-106 cells was abrogated completely by pretreatment with cycloheximide, an inhibitor of protein synthesis. These findings indicate that IGF-I suppresses gene expression for PTH/PTHrP receptor via the MAP kinase pathway, and this inhibition is required for new protein synthesis in UMR-106 osteoblast-like cells.

Pulsed electromagnetic fields rapidly modulate intracellular signaling events in osteoblastic cells: Comparison to parathyroid hormone and insulin

“…14,15 Second, the UMR cell line was originally derived from an induced paraosteal osteosarcoma of the Sprague-Dawley rat, 16 and the bone tissues in this breed of rat have been proven to respond anabolically to both PTH and PEMF. 4,17 Finally, our intent was to use the UMR cell line to complement other ongoing, parallel studies utilizing the effects of PEMF treatment in another osteoblastic cell line, MC3T3-E1.…”

Parathyroid hormone (PTH) down-regulates PTH/PTH-related protein receptor gene expression in UMR-106 osteoblast-like cells via a 3',5'-cyclic adenosine monophosphate-dependent, protein kinase A-independent pathway

“…Cells were then exposed to a test substance such as 100 nM rat (r) PTH(1-34) (Peninsula Laboratories, Belmont, CA, USA), human PTH(3-34) (Peptide Institute, Osaka, Japan), 100 nM human IGF-I (Austral Biologicals, San Roman, CA, USA), 100 µM 8-(4-chlorophenylthio)-adenine 3 ,5 -cyclic monophosphate (8-pCPTcAMP; Alexis Biochemicals, San Diego, CA, USA), 10 nM forskolin (Sigma Chemical Co., St Louis, MO, USA) and phorbol-12-myristate-13-acetate (TPA; Sigma) in DMEM with 0·1% BSA for the duration indicated. In studies of inhibitors, serum-deprived cells were treated with 10 µM H89 (Seikagaku Corp., Tokyo, Japan), a PKA inhibitor, 100 µM PD98059 (Alexis Biochemicals), an extracellularly regulated kinase 1/2 (ERK1/2) mitogen-activated protein (MAP) kinase pathway inhibitor or 35 µM cycloheximide (Sigma), a protein synthesis inhibitor, for 1 h preceding incubation for various time-periods with 10 7 M rPTH(1-34) (Kawane & Horiuchi 1999, Kawane et al 2001.…”

“…To assess the effect of rPTH(1-34) on the abundance of PTH/PTHrP receptor mRNA, Northern blot analysis was performed as previously described (Kawane & Horiuchi 1999). Briefly, total RNA was extracted from cells using guanidine thiocyanate.…”

“…For the PTH/ PTHrP receptor, down-regulation results from lower rates of receptor biosynthesis (Gonzalez & Martin 1996, Jongen et al 1996, Kawane et al 2001 or accelerates clearance of functional cell surface receptors via their internalization and subsequent proteolysis (Huang et al 1995, Ferrari et al 1999, Tawfeek et al 2001. Prolonged exposure to PTH and other substances such as insulin-like growth factor-I (IGF-I) and 1,25-dihydroxyvitamin D 3 (1,25(OH) 2 D 3 ) lowers the amounts of PTH/PTHrP receptor mRNA in osteoblastic cells (Gonzalez & Martin 1996, Jongen et al 1995, Kawane & Horiuchi 1999, Kawane et al 2001. Down-regulation of the cAMP response to PTH, and also PTH/PTHrP receptor downregulation has been observed in primary cultures of osteoblasts (Jongen et al 1995(Jongen et al , 1996 and in various osteoblastic cell lines (Mitchell & Goltzman 1990, Fukayama et al 1994, Gonzalez & Martin 1996.…”

Specific inhibitor of MEK-mediated cross-talk between ERK and p38 MAPK during differentiation of human osteosarcoma cells

“…Recent studies suggest that, in addition to its effect on apoptosis, the p38 pathway might also be involved in the differentiation of neural cells (Iwasaki et al 1999), adipocytes (Engelman et al 1998), and chondrocytes (Yoshimichi et al 2001;Shimo et al 2005). In osteoblast-like cells, activation of ERK has been reported in response to several growth factors including mitogens acting through receptor tyrosine kinases (RTKs) such as basic fibroblast growth factor (bFGF; Suzuki et al 2000), epidermal growth factor (EGF; Matsuda et al 1998), platelet-derived growth factor (PDGF; Chaudhary and Avioli 1997), and insulin-like growth factor-1 (IFG-1; Kawane and Horiuchi 1999). As reported to be its effect in other cell systems, activation of ERK in osteoblast-like cells by growth factors is associated with enhanced cell proliferation.…”