title: The optimal dose of recombinant human osteogenic protein-1 enhances differentiation of mouse osteoblast-like cells: an in vitro study.
abstract: OBJECTIVE
  There is no certain conclusion on the effect of recombinant human Osteogenic Protein-1 (OP-1, BMP-7) on the proliferation of the osteoblast-like cell line, MC3T3-E1. Furthermore, the optimal dose of rhOP-1 on cell differentiation still needs to be elucidated. This investigation aims to delineate the biofunctional characteristics of rhOP-1 in inducing osteoblastogenesis of MC3T3-E1 through in vitro time-course and dose-response studies.
  
  
  METHODS
  MC3T3-E1 cells were cultured for 1, 4, 7 days with the addition of different rhOP-1 concentrations (0, 10, 20, 50, 100, 200, 400 ng/ml), and cell proliferation and cell differentiation were examined.
  
  
  RESULTS
  MC3T3-E1 cell proliferation was stimulated by rhOP-1 in a dose-dependent manner (0-400 ng/ml) on day 1, whereas on day 4 and 7, it was still stimulated at low concentrations (10, 20, 50 ng/ml) but inhibited at high ones (200, 400 ng/ml). The alkaline phosphatase (ALP) activity, osteocalcin (OC) production, collagen deposition and extracellular matrix mineralization were dramatically elevated by rhOP-1 treatment, as a function of culture time and rhOP-1 concentration, and all of them reached a plateau at the concentration of 200 ng/ml. Real-time quantitative RT-PCR results showed Runx2, AKP-2, OC and Nog mRNA expressions increased in a dose- and time-dependent manner, and their expressions were significantly higher at high rhOP-1 concentrations than that of low ones. No significant differences were found between the effects of 200 ng/ml rhOP-1 and 400 ng/ml rhOP-1 on the differentiation of MC3T3-E1 cells, except the expression of Nog mRNA, whose expression level was much higher at 400 ng/ml than that at 200 ng/ml.
  
  
  CONCLUSIONS
  These results suggest that cell proliferation of MC3T3-E1 is depended on culture time and rhOP-1 concentration, rhOP-1 could stimulate the differentiation of MC3T3-E1 cells and the optimal concentration could be 200 ng/ml.
authors: Taizhou Hospital of Zhejiang Province, Linhai, PR China.
authors: Feng
authors: F
authors: Zhang
authors: Ling-fei
authors: LF
authors: Ren
authors: Hai-sheng
authors: HS
authors: Lin
authors: Mei-nv
authors: MN
authors: Yin
authors: Yong-qing
authors: YQ
authors: Tong
authors: Geng-sheng
authors: GS
authors: Shi
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000008:administration & dosage* / Q000494:pharmacology*; D002352:Carrier Proteins / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism; D017931:DNA Primers; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000378:metabolism; D066298:In Vitro Techniques; D051379:Mice; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects*; D015675:Osteocalcin / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D013194:Staining and Labeling; D013997:Time Factors; D000099176:Noggin Protein
chemical_list: D000469:Alkaline Phosphatase; D019485:Bone Morphogenetic Proteins; D002352:Carrier Proteins; D003094:Collagen; D050659:Core Binding Factor Alpha 1 Subunit; D017931:DNA Primers; D015675:Osteocalcin; D000099176:Noggin Protein; C493716:Runx2 protein, mouse; C511401:osteogenic protein 1 putty, recombinant human; C510878:ALPL protein, mouse