title: Multiple signaling pathways involved in stimulation of osteoblast differentiation by N-methyl-D-aspartate receptors activation in vitro.
abstract: OBJECTIVE
  Glutamate receptors are expressed in osteoblastic cells. The present study was undertaken to investigate the mechanisms underlying the stimulation of osteoblast differentiation by N-methyl-D-aspartate (NMDA) receptor activation in vitro.
  
  
  METHODS
  Primary culture of osteoblasts was prepared from SD rats. Microarray was used to detect the changes of gene expression. The effect of NMDA receptor agonist or antagonist on individual gene was examined using RT-PCR. The activity of alkaloid phosphotase (ALP) was assessed using a commercial ALP staining kit.
  
  
  RESULTS
  Microarray analyses revealed that 10 genes were up-regulated by NMDA (0.5 mmol/L) and down-regulated by MK801 (100 μmol/L), while 13 genes down-regulated by NMDA (0.5 mmol/L) and up-regulated by MK801 (100 μmol/L). Pretreatment of osteoblasts with the specific PKC inhibitor Calphostin C (0.05 μmol/L), the PKA inhibitor H-89 (20 nmol/L), or the PI3K inhibitor wortmannin (100 nmol/L) blocked the ALP activity increase caused by NMDA (0.5 mmol/L). Furthermore, NMDA (0.5 mmol/L) rapidly increased PI3K phosphorylation, which could be blocked by pretreatment of wortmannin (100 nmol/L).
  
  
  CONCLUSIONS
  The results suggest that activation of NMDA receptors stimulates osteoblasts differentiation through PKA, PKC, and PI3K signaling pathways, which is a new role for glutamate in regulating bone remodeling.
authors.affiliation: Department of Endocrine and Metabolic Diseases, Shanghai Institute of Endocrine and Metabolic Diseases, Shanghai Clinical Center for Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Jie-li
authors.initials: JL
authors.lastname: Li
authors.forename: Lin
authors.initials: L
authors.lastname: Zhao
authors.forename: Bin
authors.initials: B
authors.lastname: Cui
authors.forename: Lian-fu
authors.initials: LF
authors.lastname: Deng
authors.forename: Guang
authors.initials: G
authors.lastname: Ning
authors.forename: Jian-min
authors.initials: JM
authors.lastname: Liu
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D016202:N-Methylaspartate / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010012:Osteogenesis; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D016194:Receptors, N-Methyl-D-Aspartate / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D018691:Excitatory Amino Acid Antagonists; D000081082:Phosphoinositide-3 Kinase Inhibitors; D016194:Receptors, N-Methyl-D-Aspartate; D016202:N-Methylaspartate; D016291:Dizocilpine Maleate; D058539:Phosphatidylinositol 3-Kinase; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D000469:Alkaline Phosphatase