title: Sirt1 overexpression protects murine osteoblasts against TNF-α-induced injury in vitro by suppressing the NF-κB signaling pathway.
abstract: OBJECTIVE
  Sirtuin 1 (Sirt1) is the class III histone/protein deacetylase that interferes with the NF-κB signaling pathway, thereby has anti-inflammatory function. This study was undertaken to investigate whether Sirt1 could protect osteoblasts against TNF-α-induced injury in vitro.
  
  
  METHODS
  Murine osteoblastic cell line, MC3T3-E1, was used. Overexpress of Sirt1 protein in MC3T3-E1 cells was made by transfection the cells with Sirt1-overexpressing adenovirus. The levels of mRNAs and proteins were determined with qRT-PCR and Western blotting, respectively. The activity of NF-κB was examined using NF-κB luciferase assay. The NO concentration was measured using the Griess method.
  
  
  RESULTS
  Treatment of MC3T3-E1 cells with TNF-α (2.5-10 ng/mL) suppressed Sirt1 protein expression in a concentration-dependent manner. TNF-α (5 ng/mL) resulted in an increase in apoptosis and a reduction in ALP activity in the cells. Overexpression of Sirt1 in the cells significantly attenuated TNF-α-induced injury through suppressing apoptosis, increasing ALP activity, and increasing the expression of Runx2 and osteocalcin mRNAs. Furthermore, overexpression of Sirt1 in the cells significantly suppressed TNF-α-induced NF-κB activation, followed by reducing the expression of iNOS and NO formation. Sirt1 activator resveratrol (10 μmol/L) mimicked the protection of the cells by Sirt1 overexpression against TNF-α-induced injury, which was reversed by the Sirt1 inhibitor EX-527 (5 μmol/L).
  
  
  CONCLUSIONS
  Overexpression of Sirt1 protects MC3T3-E1 osteoblasts aganst TNF-α-induced cell injury in vitro, at least in part, via suppressing NF-κB signaling. Sirt1 may be a novel therapeutic target for treating rheumatoid arthritis-related bone loss.
authors.affiliation: Department of Orthopaedic Surgery, the First Affiliated Hospital of the General Hospital of PLA, Beijing, China. huang2011341@hotmail.com
authors.forename: Wei
authors.initials: W
authors.lastname: Huang
authors.forename: Wei-lin
authors.initials: WL
authors.lastname: Shang
authors.forename: Hua-dong
authors.initials: HD
authors.lastname: Wang
authors.forename: Wen-wen
authors.initials: WW
authors.lastname: Wu
authors.forename: Shu-xun
authors.initials: SX
authors.lastname: Hou
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D002227:Carbazoles / Q000494:pharmacology; D045744:Cell Line, Tumor; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D019610:Cytoprotection; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D018836:Inflammation Mediators / Q000378:metabolism*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D010006:Osteoblasts / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology / Q000473:pathology; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D000077185:Resveratrol; D015398:Signal Transduction / Q000187:drug effects; D056564:Sirtuin 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013267:Stilbenes / Q000494:pharmacology; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: C550547:6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide; D002227:Carbazoles; D050659:Core Binding Factor Alpha 1 Subunit; D020536:Enzyme Activators; D056572:Histone Deacetylase Inhibitors; D018836:Inflammation Mediators; D016328:NF-kappa B; D012333:RNA, Messenger; C493716:Runx2 protein, mouse; D013267:Stilbenes; D014409:Tumor Necrosis Factor-alpha; D015675:Osteocalcin; D009569:Nitric Oxide; D000469:Alkaline Phosphatase; C470545:Sirt1 protein, mouse; D056564:Sirtuin 1; D000077185:Resveratrol