Potential bone-inducing activity in vitro of recombinant human bone morphogenetic protein-7 from a CHO expression system 被引量：2

Potential bone-inducing activity in vitro of recombinant human bone morphogenetic protein-7 from a CHO expression system

暂未订购

导出

摘要 Objective: To express the recombinant human bone morphogenetic protein-7(rhBMP-7) in Chinese hamster ovary(CHO) cells, and to establish the in vitro biological activity assay of rhBMP-7.Methods: Human BMP-7 cDNA was subcloned into p114 mammalian expression vector and transfected to CHO cells by using the Lipofectamine 2000 transfection method. CHO cell supernatants were harvested and analyzed to identify the molecule mass of secreted rhBMP-7 and examine its biological activity in vitro to stimulate the synthesis of alkaline phophatase(ALP), a characteristic of osteoblast phenotypes. Results: rhBMP-7 was produced stably in CHO cells, as a processed mature disulfide-linked homodimer, with an apparent molecular mass of 36 000. Examination of the rhBMP-7 biological activity showed that rhBMP-7 specifically stimulated the production of ALP(4-fold increase at 100 ng of rhBMP-7/ml). Conclusion: The rhBMP-7 from CHO expression system has significant biological activity in induction of osteoblast phenotype, which demonstrates rhBMP-7 has the potential bone regeneration activity. Objective: To express the recombinant human bone morphogenetic protein-7(rhBMP-7) in Chinese hamster ovary(CHO) cells, and to establish the in vitro biological activity assay of rhBMP-7.Methods: Human BMP-7 cDNA was subcloned into p114 mammalian expression vector and transfected to CHO cells by using the Lipofectamine 2000 transfection method. CHO cell supernatants were harvested and analyzed to identify the molecule mass of secreted rhBMP-7 and examine its biological activity in vitro to stimulate the synthesis of alkaline phophatase(ALP), a characteristic of osteoblast phenotypes. Results: rhBMP-7 was produced stably in CHO cells, as a processed mature disulfide-linked homodimer, with an apparent molecular mass of 36 000. Examination of the rhBMP-7 biological activity showed that rhBMP-7 specifically stimulated the production of ALP(4-fold increase at 100 ng of rhBMP-7/ml). Conclusion: The rhBMP-7 from CHO expression system has significant biological activity in induction of osteoblast phenotype, which demonstrates rhBMP-7 has the potential bone regeneration activity.

作者李晓燕施伟伟王皓李博华杨扬谈岷薛静亚郭亚军

机构地区 International Joint Cancer Institute Department of Orthopaedics Shanghai CP Guojian Pharmaceutical Co. Ltd

出处《Journal of Medical Colleges of PLA(China)》 CAS 2005年第3期141-145,共5页 中国人民解放军军医大学学报（英文版）

基金 SupportedbyMilitaryFifteenthDictatedFundation(No.028002)

关键词 bone morphogenetic protein-7 CHO expression system activity assay in vitro alkaline phophatase 重组细胞 CHO 基因表达骨疾病骨遗传蛋白-7

分类号 R68 [医药卫生—骨科学]

引文网络
相关文献

参考文献11

1[1]Wang EA, Rosen V, D'Alessandro JS et al. Recombinant human bone morphogenetic protein induces bone formation [J].Proc Natl Acad Sci USA, 1990;87(6) :2220.
2[2]Jones WK, Richmond EA, White K et al. Osteogenic protein-1 (OP-1) expression and processing in Chinese hamster ovary cells: isolation of a soluble complex containing the mature and pro-domains of OP-1[J]. Growth Factors, 1994; 11 (3): 215.
3[3]Jaiswal N, Haynesworth SE, Caplan AI et al. Osteogenic differentiation of purified, culture-expanded human mesenchymal stem cells in vitro[J]. J Cell Biochem, 1997;64(2):295.
4[4]RosserMP, Xia W, Hartsell S et al. Transient transfection of CHO-K1-S using serum-free medium in suspension: a rapid mammalian protein expression system[J]. Preotein Expr Purif,2005; 40 (2): 237.
5[5]Thies RS, Bauduy M, Ashton BA et al. Recombinant human bone morphogenetic protein-2 induces osteoblastic differentiation in W-20-17 stromal cells [J]. Endocrinology, 1992; 130 (3):1318.
6[6]Kim JD, Yoon Y, Hwang HY et al. Efficient selection of stable Chinese hamaster ovary (CHO) cell lines for expression of recombinant proteins by using human interferon beta SAR element [J]. Biotechol Prog, 2005;21(3) :933.
7[7]Rutherford RB, Nussenbaum B, Krebsbach PH. Bone morphogenetic protein 7 ex vivo gene therapy[J]. Drug News Perspect,2003;16(1) :5.
8[8]Rutherford RB, Moalli M, Franceschi RT et al. Bone morphogenetic protein-transduced human fibroblasts convert to osteoblasts and form bone in vivo[J]. Tissue Eng, 2002;8(3):441.
9[9]Rutherford RB, Racenis P, Fatherazi S et al. Bone formation by BMP-7-transduced human gingival keratinocytes [J]. J Dent Res, 2003; 82 (4): 293.
10[10]Cheline AJ, Reddi AH, Martin RB. Bone morphogenetic protein-7 selectively enhances mechanically induced bone formation [J]. Bone, 2002;31(5):570.