title: Effects of tensile forces on serum deprivation-induced osteoblast apoptosis: expression analysis of caspases, Bcl-2, and Bax.
abstract: BACKGROUND
  Apoptosis is involved in the adaptive responses of bone to mechanical loading. The purpose of this study was to investigate the effects of tensile forces on osteoblast apoptosis and the related mechanism by analyzing the expression of caspases, Bcl-2, and Bax.
  
  
  METHODS
  Primary osteoblasts were harvested from neonatal rat calvaria and were subjected to cyclic tensile forces for 72 hours using Flexcell 4000 strain unit in Minimum Essential Medium (MEM) with 10% fetal calf serum (FCS) or with serum deprivation. Apoptosis was tested by flow cytometry using annexin V/PI staining. Caspase-3 activity was analyzed via Elisa. The gene expression of caspase-8, -9, Bcl-2, and Bax was quantified by reverse transcription (RT)-PCR.
  
  
  RESULTS
  In 10% FCS condition, no significant difference in cell apoptosis was found between the stretched and non-stretched osteoblast cultures. Serum withdrawal resulted in higher apoptosis rate in the osteoblasts with increased caspase-3 activity, and elevated expression of caspase-9 and Bax. Six-percent elongation of stretch attenuated the cell apoptosis induced by serum starvation, concurrent with a decrease in caspase-3 activity, a decline of caspase-8 expression, and an elevation of Bcl-2 level. On the contrary, 12% elongation of stretch increased caspase-3 activity and promoted the apoptosis with an elevated expression of caspase-8 and Bax. No significant change of caspase-9 expression was identified upon force application.
  
  
  CONCLUSIONS
  These results suggested that tensile forces regulate cell apoptosis of primary rat osteoblasts through caspase-3 and caspase-8 signaling cascade. Light forces rescue the cells from serum deprivation-induced apoptosis by elevating Bcl-2 expression, while heavy forces promote the apoptotic insult by inducing Bax expression.
authors.affiliation: Department of Orthodontics, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai 200011, China.
authors.forename: Xuan
authors.initials: X
authors.lastname: Li
authors.forename: Xiao-Ling
authors.initials: XL
authors.lastname: Zhang
authors.forename: Gang
authors.initials: G
authors.lastname: Shen
authors.forename: Guo-Hua
authors.initials: GH
authors.lastname: Tang
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053181:Caspase 8 / Q000235:genetics / Q000378:metabolism; D053453:Caspase 9 / Q000235:genetics / Q000378:metabolism; D020169:Caspases / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases