title: Multiple Transduction Pathways Mediate Thyrotropin Receptor Signaling in Preosteoblast-Like Cells.
abstract: It has been shown that the TSH receptor (TSHR) couples to a number of different signaling pathways, although the Gs-cAMP pathway has been considered primary. Here, we measured the effects of TSH on bone marker mRNA and protein expression in preosteoblast-like U2OS cells stably expressing TSHRs. We determined which signaling cascades are involved in the regulation of IL-11, osteopontin (OPN), and alkaline phosphatase (ALPL). We demonstrated that TSH-induced up-regulation of IL-11 is primarily mediated via the Gs pathway as IL-11 was up-regulated by forskolin (FSK), an adenylyl cyclase activator, and inhibited by protein kinase A inhibitor H-89 and by silencing of Gαs by small interfering RNA. OPN levels were not affected by FSK, but its up-regulation was inhibited by TSHR/Gi-uncoupling by pertussis toxin. Pertussis toxin decreased p38 MAPK kinase phosphorylation, and a p38 inhibitor and small interfering RNA knockdown of p38α inhibited OPN induction by TSH. Up-regulation of ALPL expression required high doses of TSH (EC50 = 395nM), whereas low doses (EC50 = 19nM) were inhibitory. FSK-stimulated cAMP production decreased basal ALPL expression, whereas protein kinase A inhibition by H-89 and silencing of Gαs increased basal levels of ALPL. Knockdown of Gαq/11 and a protein kinase C inhibitor decreased TSH-stimulated up-regulation of ALPL, whereas a protein kinase C activator increased ALPL levels. A MAPK inhibitor and silencing of ERK1/2 inhibited TSH-stimulated ALPL expression. We conclude that TSH regulates expression of different bone markers via distinct signaling pathways.
authors.affiliation: Laboratory of Endocrinology and Receptor Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Alisa
authors.initials: A
authors.lastname: Boutin
authors.affiliation: Laboratory of Endocrinology and Receptor Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Susanne
authors.initials: S
authors.lastname: Neumann
authors.affiliation: Laboratory of Endocrinology and Receptor Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Marvin C
authors.initials: MC
authors.lastname: Gershengorn
mesh_terms: D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D006801:Humans; D017370:Interleukin-11 / Q000378:metabolism*; D007546:Isoquinolines / Q000494:pharmacology; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011989:Receptors, Thyrotropin / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013449:Sulfonamides / Q000494:pharmacology; D013972:Thyrotropin / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology*
chemical_list: D017370:Interleukin-11; D007546:Isoquinolines; D047428:Protein Kinase Inhibitors; D011989:Receptors, Thyrotropin; D013449:Sulfonamides; D005576:Colforsin; D013972:Thyrotropin; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide