Nicotine suppresses bone sialoprotein gene expression.
OBJECTIVE
Tobacco smoking is a risk factor for periodontitis and osteoporosis. Nicotine is a major component of tobacco, and has been reported to inhibit proliferation and differentiation of osteoblasts. Bone sialoprotein (BSP) is a mineralized tissue-specific protein expressed by differentiated osteoblasts that appears to function in the initial mineralization of bone. The purpose of this study was to determine the effects of nicotine on bone metabolism.


METHODS
We used rat osteobast-like UMR106 and ROS 17/2.8 cells and rat stromal bone marrow RBMC-D8 cells. To determine the molecular basis of the transcriptional regulation of the BSP gene by nicotine, we conducted Northern hybridization, transient transfection analyses with chimeric constructs of the BSP gene promoter linked to a luciferase reporter gene and gel mobility shift assays.


RESULTS
Nicotine (250 microg/mL) decreased the BSP mRNA levels at 12 and 24 h in UMR106 and ROS 17/2.8 cells. From transient transfection assays using various sized BSP promoter-luciferase constructs, nicotine decreased the luciferase activities of the construct, including the promoter sequence nucleotides -116 to +60, in UMR106 and RBMC-D8 cells. Nicotine decreased the nuclear protein binding to the cAMP response element (CRE), fibroblast growth factor 2 response element (FRE) and homeodomain protein-binding site (HOX) at 12 and 24 h.


CONCLUSIONS
This study indicates that nicotine suppresses BSP transcription mediated through CRE, FRE and HOX elements in the proximal promoter of the rat BSP gene.
Nakayama
Y
Y
Department of Periodontology, Tianjin Medical University, Tianjin, China.
Mezawa
M
M
Araki
S
S
Sasaki
Y
Y
Wang
S
S
Han
J
J
Li
X
X
Takai
H
H
Ogata
Y
Y
D000818:Animals; D015152:Blotting, Northern; D001854:Bone Marrow Cells / Q000187:drug effects; D002460:Cell Line; D002678:Chimera / Q000235:genetics; D017362:Cyclic AMP Response Element-Binding Protein / Q000187:drug effects; D024202:Electrophoretic Mobility Shift Assay; D016222:Fibroblast Growth Factor 2 / Q000235:genetics; D017930:Genes, Reporter / Q000235:genetics; D018398:Homeodomain Proteins / Q000187:drug effects; D058574:Integrin-Binding Sialoprotein; D008156:Luciferases / Q000235:genetics; D009538:Nicotine / Q000494:pharmacology*; D018722:Nicotinic Agonists / Q000494:pharmacology*; D009687:Nuclear Proteins / Q000187:drug effects; D010006:Osteoblasts / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics; D051381:Rats; D020218:Response Elements / Q000187:drug effects; D012795:Sialoglycoproteins / Q000187:drug effects* / Q000235:genetics; D017154:Stromal Cells / Q000187:drug effects; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D017362:Cyclic AMP Response Element-Binding Protein; D018398:Homeodomain Proteins; C546873:Ibsp protein, rat; D058574:Integrin-Binding Sialoprotein; D018722:Nicotinic Agonists; D009687:Nuclear Proteins; D012333:RNA, Messenger; D012795:Sialoglycoproteins; D016222:Fibroblast Growth Factor 2; D009538:Nicotine; D008156:Luciferases