title: A comparative study of mechanical strain, icariin and combination stimulations on improving osteoinductive potential via NF-kappaB activation in osteoblast-like cells.
abstract: BACKGROUND
  The combination of drugs and exercise was the effective treatment in bone injure and rebuilding in clinic. As mechanical strain has potential in inducing the differentiation of osteoblasts in our previous study, the further research to investigate the combination of mechanical strain and icariin stimulation on inducing osteoblast proliferation, differentiation and the possible mechanism in MC3T3-E1 cell line.
  
  
  METHODS
  A whole cell enzyme-linked immunosorbent assay that detects the bromodeoxyuridine incorporation during DNA synthesis was applied to evaluate the proliferation. The mRNA expression of alkaline phosphatase (ALP), osteocalcin (OCN), type I collagen (Col I), bone morphogenetic protein-2 (BMP-2) and BMP-4 was detected by real-time reverse-transcription polymerase chain reaction. The activity of ALP was analyzed by ELISA and the protein expression of OCN, Col I and BMP-2 was assessed by western blot. Moreover, the activity of nuclear transcription factor kappa-B (NF-κB) signaling pathway was investigated with the expression of inhibitor of κB (IκB) α, phosphorylation of IκB-α (P-IκB-α), p65, P-p65 by western blot.
  
  
  RESULTS
  We observed that compared to single mechanical strain or icariin stimulation, the mRNA and protein expressions of ALP (P < 0.05 or P < 0.01), OCN (P < 0.01) and Col I (P < 0.05 or P < 0.01) were increased significantly by the combination of mechanical strain and icariin stimulation. Moreover, the combination of mechanical strain and icariin stimulation could up-regulate the expression of BMP-2 (P < 0.01) and BMP-4 compared to single mechanical strain or icariin stimulation. The combination of mechanical strain and icariin stimulation could activate NF-κB signaling pathway by increasing the expression of IκB α, P-IκB-α, p65, P-p65 (P < 0.01).
  
  
  CONCLUSIONS
  The combination of mechanical strain and icariin stimulation could activate the NF-κB pathway to improve the proliferation, differentiation of osteoblast-like cells.
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. wangqiangsong2008@163.com.
authors.forename: Qiang-Song
authors.initials: QS
authors.lastname: Wang
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. xinchangzhang@163.com.
authors.forename: Xin-Chang
authors.initials: XC
authors.lastname: Zhang
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. limxinxin@163.com.
authors.forename: Rui-Xin
authors.initials: RX
authors.lastname: Li
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. sunjg@sina.vip.com.
authors.forename: Jing-Gong
authors.initials: JG
authors.lastname: Sun
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. 13512062095@126.com.
authors.forename: Wei-Hua
authors.initials: WH
authors.lastname: Su
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. guoyong74@163.com.
authors.forename: Yong
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. lihao2006011653@sina.com.
authors.forename: Hao
authors.initials: H
authors.lastname: Li
authors.affiliation: Tianjin Institute of Medical Equipment, Academy of Military Medical Sciences, No. 106 Wandong Road, Hedong District, Tianjin, 300162, People's Republic of China. z56717@sohu.com.
authors.forename: Xi-Zheng
authors.initials: XZ
authors.lastname: Zhang
mesh_terms: D016475:3T3 Cells; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics / Q000378:metabolism; D055415:Bone Morphogenetic Protein 4 / Q000235:genetics / Q000378:metabolism; D016723:Bone Remodeling / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013314:Stress, Mechanical
chemical_list: C525719:Bmp2 protein, mouse; C525810:Bmp4 protein, mouse; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D024042:Collagen Type I; D005419:Flavonoids; D016328:NF-kappa B; D015675:Osteocalcin; D000469:Alkaline Phosphatase; C056599:icariin