Parathyroid hormone(1-34) regulates the expression of osteoclast inhibitory lectin via multiple signaling pathways in UMR106 osteoblast-like cells

Abstract

Objective To investigate the regulation of parathyroid hormone(1-34) on mRNA expression of osteoclast inhibitory lectin (OCIL) gene in UMR106 osteoblastic-like cells and involved signaling pathway.Methods Rat UMR106 osteoblastic-like cells were cultured and treated with various concentration of PTH(1-34) and specific agonists or inhibitors of PKA,PKC,Ca2+/calmodulin-dependent protein kinase (CaMK) and mitogen-activated protein kinase (MAPK) signal pathways for indicated time intervals.Then the cells were gathered at indicated time points and total RNA were extracted.OCIL mRNA expression was analyzed using real-time PCR technique.Results PTH(1-34) stimulated OCIL mRNA expression in a time-and dose-dependentmanner.A dose of 10 nmol/L PTH(1-34) started to induce OCIL mRNA from 6 h,with a highest increase of about 2.8-fold vs.control group (without PTH treatment) at 24 h.The up-regulation of OCIL mRNA began and reached maximum later than RANKL induction and OPG suppression effected by PTH(1-34).Protein Kinase A (PKA) signaling activators forskolin(FSK) and dibutyryl cAMP (db-cAMP),as well as calcium ionophore A23187 all up-regulated OCIL mRNA with the maximal induction of about 4.2-fold,4.5-fold and 5.1-fold.Protein Kinase C (PKC) activator phorbol-12-myristate-13-acetate(PMA) reduced OCIL mRNA expression at the early stage(2-6 h),with the highest down-regulation of 50％ at 6 h.However,the inhibitory effect on OCIL mRNA turned into slightly stimulatory effect later (24 h).PKA inhibitor KT5720,calmodulin antagonist W-7,CaMK Ⅱ inhibitor KN-62 and mitogen-activated protein kinase (MAPK) inhibitor PD98059 all blocked PTH(1-34)-induced OCIL mRNA expression by the maximal reduction of 56％,61％,63％ and 50％ respectively.There also exist cross-talks between different signal pathways.MAPK inhibitor PD98059 blocked the expression of OCIL mRNA which was stimulated by PKA activators FSK or db-cAMP,with the reduction of 98％ and 63％ respectively,while the OCIL mRNA expression stimulated by A23187 remained unaffected.Conclusion PTH(1-34) increased OCIL mRNA expression in vitro through cAMP/PKA,Ca2+/CaMK and MAPK signaling pathways. Key words: Parathyroid hormone; Osteoclasts; Gene expression; Agglutinins