Effect of rhBMP-2 on the osteogenic potential of bone marrow stromal cells from an osteogenesis imperfecta mouse

M. L. Balk, J. Bray, C. Day, M. Epperly, J. Greenberger, Christopher H Evans, Christopher Niyibizi

Research output: Contribution to journalArticle

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Abstract

To understand whether osteogenesis imperfecta (OI) could result from defective differentiation of osteoprogenitor cells, we investigated the osteogenic potential of bone marrow stromal cells from a mouse model of human OI (oim). Bone marrow was flushed from the femurs and tibias of oim and normal littermates using a syringe with Dulbecco's modified Eagle's medium, and cells were allowed to adhere to flasks. Adherent cells were trypsinized and passaged weekly at a 1:4 split. The established stromal cells were assessed for collagen synthesis, alkaline phosphatase, and osteocalcin production in the presence or absence of rhBMP-2. The stromal cells were also assessed for mineralization by Von-Kossa staining and for exogenous gene transfer using adeno-lacZ and a retroviral vector. The bone marrow stromal cells from oim mice synthesized α1(I) homotrimers as expected, whereas the stromal cells from the normal littermates synthesized α1(T)2α2(I) heterotrimers. The bone marrow stromal cells exhibited low levels of alkaline phosphatase activity under basal conditions; upon treatment with rhBMP-2, the level of the alkaline phosphatase activity increased approximately 40-fold. Cytochemical staining of the cells confirmed the expression of alkaline phosphatase by the oim stromal cells and its augmentation by rhBMP-2. Osteocalcin production in the stromal cells was also enhanced approximately threefold by rhBMP-2, oim stromal cells grown in the presence of β-glycerophosphate and ascorbic acid demonstrated Von-Kossa-positive solid deposits after 3 weeks in culture. Ten days after infection with adeno-lacZ, approximately 70% of oim stromal cells expressed the transgene product, and after infection with a retrovirus, approximately 20% of the cells expressed the transgene. These data indicate that bone marrow stromal cells have osteogenic potential, acid also the potential to be transduced with exogenous genes. Under basal conditions, however, the stromal cells from oim mice exhibited significantly lower levels of alkaline phosphatase activity than their normal littermates.

Original languageEnglish (US)
Pages (from-to)7-15
Number of pages9
JournalBone
Volume21
Issue number1
DOIs
StatePublished - Jul 1997
Externally publishedYes

Fingerprint

Osteogenesis Imperfecta
Stromal Cells
Mesenchymal Stromal Cells
Alkaline Phosphatase
Osteocalcin
Transgenes
Staining and Labeling
Glycerophosphates
Eagles
Syringes
Retroviridae
Infection
Tibia
Femur
Genes
Ascorbic Acid
Cell Differentiation
Collagen
Bone Marrow
Acids

Keywords

  • Bone
  • Bone marrow
  • Collagen
  • Oim
  • Osteogenesis imperfecta
  • Stromal cells

ASJC Scopus subject areas

  • Physiology
  • Hematology

Cite this

Effect of rhBMP-2 on the osteogenic potential of bone marrow stromal cells from an osteogenesis imperfecta mouse. / Balk, M. L.; Bray, J.; Day, C.; Epperly, M.; Greenberger, J.; Evans, Christopher H; Niyibizi, Christopher.

In: Bone, Vol. 21, No. 1, 07.1997, p. 7-15.

Research output: Contribution to journalArticle

Balk, M. L. ; Bray, J. ; Day, C. ; Epperly, M. ; Greenberger, J. ; Evans, Christopher H ; Niyibizi, Christopher. / Effect of rhBMP-2 on the osteogenic potential of bone marrow stromal cells from an osteogenesis imperfecta mouse. In: Bone. 1997 ; Vol. 21, No. 1. pp. 7-15.
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AU - Evans, Christopher H

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AB - To understand whether osteogenesis imperfecta (OI) could result from defective differentiation of osteoprogenitor cells, we investigated the osteogenic potential of bone marrow stromal cells from a mouse model of human OI (oim). Bone marrow was flushed from the femurs and tibias of oim and normal littermates using a syringe with Dulbecco's modified Eagle's medium, and cells were allowed to adhere to flasks. Adherent cells were trypsinized and passaged weekly at a 1:4 split. The established stromal cells were assessed for collagen synthesis, alkaline phosphatase, and osteocalcin production in the presence or absence of rhBMP-2. The stromal cells were also assessed for mineralization by Von-Kossa staining and for exogenous gene transfer using adeno-lacZ and a retroviral vector. The bone marrow stromal cells from oim mice synthesized α1(I) homotrimers as expected, whereas the stromal cells from the normal littermates synthesized α1(T)2α2(I) heterotrimers. The bone marrow stromal cells exhibited low levels of alkaline phosphatase activity under basal conditions; upon treatment with rhBMP-2, the level of the alkaline phosphatase activity increased approximately 40-fold. Cytochemical staining of the cells confirmed the expression of alkaline phosphatase by the oim stromal cells and its augmentation by rhBMP-2. Osteocalcin production in the stromal cells was also enhanced approximately threefold by rhBMP-2, oim stromal cells grown in the presence of β-glycerophosphate and ascorbic acid demonstrated Von-Kossa-positive solid deposits after 3 weeks in culture. Ten days after infection with adeno-lacZ, approximately 70% of oim stromal cells expressed the transgene product, and after infection with a retrovirus, approximately 20% of the cells expressed the transgene. These data indicate that bone marrow stromal cells have osteogenic potential, acid also the potential to be transduced with exogenous genes. Under basal conditions, however, the stromal cells from oim mice exhibited significantly lower levels of alkaline phosphatase activity than their normal littermates.

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